Physiol Rev 93: 1803–1845, 2013
doi:10.1152/physrev.00039.2012
PHYSIOLOGY AND PATHOPHYSIOLOGY OF
CARNOSINE
Alexander A. Boldyrev, Giancarlo Aldini, and Wim Derave
International Biotechnological Center, M. V. Lomonosov Moscow State University, Moscow, Russia; Research Center
of Neurology, Russian Academy of Medical Sciences, Moscow, Russia; Department of Pharmaceutical Sciences,
University of Milan, Milan, Italy; and Department of Movement and Sports Sciences, Ghent University, Ghent, Belgium
Boldyrev AA, Aldini G, Derave W. Physiology and Pathophysiology of Carnosine.
Physiol Rev 93: 1803–1845, 2013; doi:10.1152/physrev.00039.2012.—
Carnosine (␤-alanyl-L-histidine) was discovered in 1900 as an abundant non-protein nitrogen-containing compound of meat. The dipeptide is not only found in skeletal muscle, but
also in other excitable tissues. Most animals, except humans, also possess a methylated
variant of carnosine, either anserine or ophidine/balenine, collectively called the histidine-containing
dipeptides. This review aims to decipher the physiological roles of carnosine, based on its biochemical
properties. The latter include pH-buffering, metal-ion chelation, and antioxidant capacity as well as the
capacity to protect against formation of advanced glycation and lipoxidation end-products. For these
reasons, the therapeutic potential of carnosine supplementation has been tested in numerous diseases in which ischemic or oxidative stress are involved. For several pathologies, such as diabetes and
its complications, ocular disease, aging, and neurological disorders, promising preclinical and clinical
results have been obtained. Also the pathophysiological relevance of serum carnosinase, the enzyme
actively degrading carnosine into L-histidine and ␤-alanine, is discussed. The carnosine system has
evolved as a pluripotent solution to a number of homeostatic challenges. L-Histidine, and more specifically its imidazole moiety, appears to be the prime bioactive component, whereas ␤-alanine is mainly
regulating the synthesis of the dipeptide. This paper summarizes a century of scientific exploration on
the (patho)physiological role of carnosine and related compounds. However, far more experiments in
the fields of physiology and related disciplines (biology, pharmacology, genetics, molecular biology, etc.)
are required to gain a full understanding of the function and applications of this intriguing molecule.
L
I.
II.
III.
IV.
V.
VI.
VII.
VIII.
IX.
INTRODUCTION: CARNOSINE AND...
DISTRIBUTION OF CARNOSINE AND ITS...
CHEMICAL AND BIOCHEMICAL...
ENZYMES AND TRANSPORTERS...
PHYSIOLOGICAL ROLE OF CARNOSINE
CARNOSINE AND DISEASE
CARNOSINASE AND DISEASE
APPLICATIONS TO SPORT AND...
CONCLUSIONS AND FUTURE...
1803
1804
1808
1815
1820
1824
1828
1830
1833
I. INTRODUCTION: CARNOSINE AND
RELATED COMPOUNDS
Carnosine is the archetype of a family of naturally occurring
derivatives (see FIGURE 1). It is a dipeptide, composed of
␤-alanine and L-histidine. The most common variants are
the methylated analogs, anserine and ophidine (balenine),
in which the imidazole ring of L-histidine is methylated at,
respectively, the nitrogen atom closest to the side chain (Npi)
and away from the side chain (Ntau). There is considerable
confusion with regard to the nomenclature of the methylated
nitrogen atoms, with older literature designating anserine
(Npi methylated) as ␤-alanyl-N1-methyl-histidine, whereas
according to IUPAC this is ␤-alanyl-N3-methyl-histidine (and
vice versa for ophidine/balenine). Another carnosine analog is
homocarnosine, in which ␤-alanine is replaced by GABA.
Other related compounds include the acetylated form of carnosine (acetylcarnosine), in which the ␤-alanine is acetylated,
and carcinine, in which the L-histidine residue of carnosine is
replaced by histamine. The metabolic pathways linking the
different related compounds to carnosine are displayed in
FIGURE 2. The enzymes responsible for the synthesis, degradation, and methylation of carnosine will be discussed in detail in
section IV.
Carnosine was first discovered in 1900 by V.S. Gulewitch
(154), a Russian chemist, during his search for unidentified
nitrogen-containing not-protein compounds in Liebig’s
meat extract [for historical tribute to this recovery, see
Boldyrev (64)]. It was noticed that most bird muscles do not
contain carnosine (87). Instead, the new compound anserine was identified in goose (5) and chicken (366) muscles
and was named after the former (anser is Latin for goose).
Another methylated carnosine analog was found in snake
muscle and whale muscle and, respectively, named ophidine
and balenine. The two metabolites appeared to be the same
0031-9333/13 Copyright © 2013 the American Physiological Society
1803
BOLDYREV, ALDINI, AND DERAVE
H
N
O
N
NH
O
N
HO
NH
O
N
NH2
carcinine
anserine
NH2
O
H
N
O
HO
N
N
NH
O
carnosine
O
O
NH2
H
N
HO
HO
N
NH
NH2
O
N
NH
Ophidine/
Balenine
O
H
N
NH2
HO
homocarnosine
O
N
NH
NH
acetyl-carnosine
O
FIGURE 1.
Chemical structures of carnosine and its naturally occurring derivatives.
molecule (␤-alanyl-Ntau-methyl-histidine), and preference
is given to the name ophidine (348). Carnosine, anserine,
and ophidine will be collectively called the histidine-containing dipeptides (HCD) in this paper.
Carcinine
Histamine
Homocarnosine
CN
HDC
β-alanine
Histidine
CS
CS
γ-aminobutyric acid
(GABA)
CN
Carnosine
II. DISTRIBUTION OF CARNOSINE AND ITS
ANALOGS
A. Distribution of HCD in Animal Species
The distribution of HCD in various animal species has been
the subject of study for nearly 100 years. Almost all studies
have been conducted on skeletal muscle tissue. Therefore,
we will mainly limit our overview of the literature to skeletal muscle, and discuss below the distribution of HCD in
mammals and other vertebrates.
1. HCD in skeletal muscles of mammals
CMT
CMT
Anserine
Ophidine
Acetylcarnosine
Acetylanserine
FIGURE 2. Metabolic pathways of carnosine. The dipeptides carnosine and homocarnosine are synthesized by carnosine synthase (CS) and
degraded by carnosinase (CN). Solid lines represent well-described
reactions occurring in humans. Dashed lines represent reactions that
either do not occur in humans or are poorly understood at present.
HDC, L-histidine decarboxylase; CMT, carnosine-N-methyltransferase.
1804
Based on a large number of comparative studies (64, 80, 91,
164, 405) and studies on individual species (20, 21, 86, 120,
193, 261, 309, 337) that have been conducted on the presence and distribution of carnosine, anserine, and ophidine
in mammalian muscles, we have composed FIGURE 3. Data
were available on 39 different species from only 9 (out of
21) mammalian orders, meaning that many orders of mammals, e.g., the Chiroptera (bats etc.), Insectivora, Proboscidea (elephants) etc., still remain to be studied.
Still, several conclusions can be drawn from FIGURE 3.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
Class
Infra-class
Order
Family
Artiodactyla
Bovidae
Camel. Cervidae
Perisso.
Mammalia
Placentalia
Cetacea
Carnivora
Primates Lag. Rodentia
Metatheria
Marsupalia
Gir. Sui. Equid. Phy. Ziph Baleanopt. Delphinidae Can Felidae Pho Monkey Ape Lep. Das Muridae Chi. Macrop Pha Did
14
12
Ophidine
Anserine
Carnosine
10
8
6
4
2
0
Antilope
Cattle
Buffalo
Gnu
Sheep
Camel
Lama
Red deer
Water deer
White-tailed deer
Giraffe
Pig
Donkey
Horse
Spermwhale
Bottlenosed whale
Blue whale
Fin whale
Sei whale
Bottlenosed dolphin
Dolphin
Pygmy killer whale
Risso’s dolphin
Dog
Cat
Lion
Seal
Owl monkey
Patas monkey
Human
Rabbit
Agouti
Rat
Mouse
Viscacha
Kangaroo
Wallaby
Phalanger
Opossum
HCD concentration (g/kg wet muscle)
16
Species
FIGURE 3. Mammals: concentration of the HCD (carnosine, anserine, and ophidine) in mg/kg wet muscle.
Each bar represents the average of the retrieved values for the different muscle types and different publications
on one species. See text for references. The full names of the orders (and their families) are as follows:
Artiodactyla (Bovidae, Camelidae, Cervidae, Giraffidae, Suidae), Perissodactyla (Equidae), Cetacea (Physeteridae, Ziphidae, Balaenopteridae, Delphinidae), Carnivora (Canidae, Felidae, Phocidae), Primates (Monkeys,
Apes), Lagomorpha (Leporidae), Rodentia (Dasyproctidae, Muridae, Chinchillidae), Marsupialia (Macropodidae, Phalangeridae, Didelphidae).
There is a large range in concentrations in total HCD content between different mammals, with some of the highest
values reported in the wallaby and blue whale [50 – 60
mmol/kg wet weight (ww) or 12–14 g/kg ww, or more than
1% of the muscle’s ww] and some of the lowest reported in
mice and opossum (1–3 mmol/kg ww or ⬃0.04% of the
muscle’s ww).
As a rule, almost all mammals have both carnosine along
with just one of the methylated carnosine analogs (either
anserine or ophidine). The only mammalian species known
not to have any of the two methylated carnosine analogs is
in fact the Homo sapiens. Another rare exception is that
some animals express all three HCD, such as the pig and the
buffalo, but in these cases the methylated forms are in far
minority compared with carnosine.
In most cases, the methylated form is more abundant than
the nonmethylated carnosine itself. Entire mammalian orders follow this pattern, such as the Cetacea (marine mammals), Rodentia, and Marsupalia. In general, there is a good
similarity in the HCD expression pattern between the different species within one family and the different families
within one order.
Of the two methylated forms, anserine is more frequently
observed than ophidine. Whereas anserine is found in
nearly all mammalian orders, ophidine is only found in
significant amounts in marine mammals (Cetacea) and in
smaller amounts in some hoofed animals: the even-toed
ungulates (Artiodactyla) and odd-toed ungulates (Perissodactyla). These three orders all belong to the same superorder of mammals, the Laurasiatheria.
2. HCD in muscles of other animals
In nature, HCD are probably only present in animals. At
least nobody has ever detected them in plants, fungi, or
other eukaryotes, even though it must be said that it is not
clear how good the techniques were and how thorough the
search was for the presence of known or possibly unknown
HCD in non-animal eukaryotes. Within the animal kingdom, HCD have only been detected in vertebrates and not
in invertebrates, except from the reported presence of high
concentrations of anserine and carnosine in prawn, but this
has not been confirmed in a peer-reviewed paper (202).
The distribution of HCD in fish species are shown in FIGURE 4,
a compilation of data based on References 2, 4, 20, 21, 61,
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1805
BOLDYREV, ALDINI, AND DERAVE
Class
Infra-class Chond
Order Acip. Anguilliformes Clupeiformes
Family Acip. Ang. Con. Clupeidae Cob.
Actinopterygii
Telostei
Gad. Osm.
Cypriniformes
Cyprinidae
Gad.
Perciformes
Osm. Cent. Cichl.
Pleur. Salmoniformes
Istio. Malac. Sciae. Scombridae
Sol.
Salmonidae
18
Ophidine
Anserine
Carnosine
14
12
10
8
6
4
Trout
Salmon
Japanese Char
Sole
Tuna
Mackerel
Croaker
Whiting
Marlin
Tilapia
Bluegill sunfish
Smelt
Cod
Roach
Gudgeon
Carp
Barbel
Pond Loach
Sardine
Herring
Conger eel
0
Japanese Eel
2
Sturgeon
HCD concentration (g/kg wet muscle)
16
Species
FIGURE 4. Fishes: concentration of the HCD (carnosine, anserine, and ophidine) in mg/kg wet muscle.
See text for references. The full names of the orders (and their families) are as follows: Acipenseriformes
(Acipenseridae), Anguilliformes (Anguilidae, Congridae), Clupeiformes (Clupeidae), Cypriniformes (Cobitidae, Cyprinidae), Gadiformes (Gadidae), Osmeriformes (Osmeridae), Perciformes (Centrarchidae, Cichlidae, Istiophoridae, Malacanthidae, Sciaenidae, Scombridae), Pleuronectiformes (Soleidae), Salmoniformes (Salmonidae).
64, 80, 91, 201. It becomes clear that some of the fish
species (such as herring, sardine, carp, sole, etc.) contain
very little, if any, HCD. However, several of these fishes
contain high or very high concentrations of free L-histidine
instead. This would suggest that the role of HCD is replaced
by L-histidine and that this therefore mainly relates to the
imidazole ring of L-histidine (e.g., the pH buffering function). Of all the examined fishes (FIGURE 4), it seems that
they nearly all express only one type of HCD, either carnosine or anserine.
likely signifies that the enzymes for ophidine and anserine
synthesis have evolved at least two times in evolution independently of each other, a phenomenon which is called
convergent evolution (113).
B. Distribution of HCD in Tissues
The HCD concentrations found in muscles of amphibians,
birds, and reptiles are shown in FIGURE 5, a compilation
based on References 2, 4, 21, 64, 80, 91, 201. All the investigated species in these classes consistently express at least
one type of HCD: the amphibians carnosine, the birds anserine (sometimes along with carnosine), and the reptiles
primarily ophidine (sometimes along with carnosine).
In mammals, only two tissues have carnosine concentrations
as high as in the millimolar range, i.e., skeletal muscle and the
olfactory bulb. As only the former represents a considerable
percentage of the total body mass, it can be calculated that
over 99% of the carnosine present in an organism is found in
the skeletal muscle tissue. Carnosine is measurable in other
brain regions and other tissues and body fluids as well, but in
concentrations 10- to 1,000-fold lower than in muscle (see
FIGURE 6). A detailed discussion of the distribution of carnosine in the brain is given in section V.
When considering the occurrence of the three HCD in all
investigated vertebrate species (FIGURES 3–5), ophidine is
almost exclusively seen in two unrelated lineages, i.e., in
some snake families (order of Squamata) and some specific
marine mammalian families (order of Cetacean). This most
There is only fragmentary and sometimes conflicting literature on the distribution of HCD in non-muscle tissues.
There are reports of relatively high concentrations of HCD
in the skin of reptiles (snake and skink) and frogs (91).
Interestingly, the same HCD type that was present in the
1806
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
HCD concentration (g/kg wet muscle)
Class Amphibia
Infra-class Lissamphib.
Order Anura
Family Buf. Ran.
Aves
Galloanserae
Anseriformes
Galliformes
Anatidae
Phasianidae
Reptilia
Lepidosauromorpha
Squamata
Neogn. Neoav. Arch.
Colum. Pass. Croc.
Colum. Corv. Croc. Boidae Colubridae
Elapidae
Anap.
Test.
Scinc.
Viperidae
Chel.
12
Ophidine
Anserine
Carnosine
10
8
6
4
Sea turtle
Habu snake
Black Rattlesnake
Cotton-mouthed moccasin
Species
Fernandi’s skink
Sea snake
Many-banded krait
Gobra
Natrix tigrina
Elaphe carinata
Boa constrictor
Crocodile
Rook
Pigeon
Turkey
Red jungle fowl
Chicken
Mallard
Goose
Toad
Frog
0
Duckling
2
FIGURE 5. Amphibians, birds, and reptiles: concentration of the HCD (carnosine, anserine, and ophidine) in
mg/kg wet muscle. See text for references. The full names of the orders (and their families) are as follows:
Anura (Bufonidae, Ranidae), Anseriformes (Anatidae), Galliformes (Phasianidae), Columbiformes (Columbidae), Passeriformes (Corvidae), Crocodilia (Crocodylidae), Squamata (Boidae, Colubridae, Elapidae, Scincidae,
Viperidae), Testudines (Cheloniidae).
muscle was present in skin (e.g., cobra snake has only ophidine as HCD in muscle and has only ophidine in skin tissue)
(91). Some authors have reported the absence of HCD from
liver (95, 96), whereas Purchas and Busboom (307) have
reported very high bovine liver carnosine concentrations in
Carnosine concentration
(µmol/L or µmol/kg; logarithmic)
10000
1000
100
10
a
m
as
al
in
A recent profiling of HCD in rat tissues by the high-precision method of liquid chromatography/electrospray ionization tandem mass spectroscopy (ESI-MS)(13), revealed that
carnosine was detectable in the millimolar range in skeletal
muscle (2– 6 mmol/kg wet tissue), in the micromolar range
in brain, cerebellum, and myocardium (25–70 ␮mol/kg wet
tissue) and nondetectable in kidney, liver, lung, and plasma.
A similar profiling of carnosine content in human tissues is
currently lacking in the literature. The only frequently measured human tissue carnosine concentration is that of skeletal muscle, because of the high concentration and the relative ease to sample muscle tissue invasively (biopsy; Ref.
243) or noninvasively (magnetic resonance spectroscopy;
Ref. 286).
C
er
eb
er
ro
eb
sp
Pl
flu
id
en
le
Sp
dn
ra
Ki
or
lc
ey
x
te
us
ex
pl
id
C
C
ho
ro
al
et
el
Sk
O
lfa
ct
or
m
y
us
bu
cl
lb
e
1
the low end of the concentration range found in skeletal
muscles (⬃4 mmol/kg). Others found trace amounts of carnosine in tissues such as the bovine lens (77). Part of the
contradiction in the literature may have arisen from the fact
that 1) carnosine concentrations in various tissues are at the
limit of detection, 2) there are significant interspecies differences, and/or 3) there is a large difference in the accuracy of
HCD quantification between the different analytic methodologies.
FIGURE 6. Endogenous tissue and fluid carnosine concentrations in
mice. Note the logarithmic scale. [Based on data from Kamal et al. (206).]
Mammalian cardiac muscle has only a relatively low concentration of carnosine (⬃0.1 mmol/kg wet tissue). How-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1807
BOLDYREV, ALDINI, AND DERAVE
ever, the total content of carnosine derivatives is high and in
the order of 2–10 mmol/kg wet tissue, as the concentrations
of N-acetylated carnosine, anserine, and homocarnosine
are reportedly present in the millimolar range in cardiac
tissue (279, 280).
very close to 7.0, their proton sequestering capacity is high
(322). The contribution of carnosine to the buffering capacity of skeletal and cardiac muscle will be discussed in more
detail in section V.
B. Metal Ion Chelating Activity
III. CHEMICAL AND BIOCHEMICAL
PROPERTIES OF CARNOSINE
A. Acid-Base Properties and Buffering
Activity
Carnosine is a very water-soluble compound (1 g in 3.1 ml
of water at 25°C), characterized by three ionizable groups:
the carboxylic group (pKa 2.76), the amino group of the
␤-alanine residue (pKa 9.32), and the nitrogens of the imidazole ring (pKa 6.72) (384). Based on the pKa values, at
physiological pH, carnosine is mainly present in the zwitterionic form with both the carboxyl and amino group of
␤-alanine in their ionized states. The chemistry of the imidazole ring in carnosine is quite interesting and deserves
some detailed explanation since it regulates the buffering
activity of carnosine. By using Raman spectroscopy, Torreggiani et al. (371) reported that at pH 7 the bands caused
by the positively charged and neutral forms of the imidazole
ring are both visible, which indicates that at this pH value,
both forms are present. The two tautomeric forms of carnosine’s imidazole ring in its neutral form are the N␶ protonated (tautomer I) and the N␲ protonated (tautomer II)
(FIGURE 7). Raman studies (371) indicate that both tautomers exist at neutral pH, thus indicating an equilibrium between the two tautomeric forms. Furthermore, at pH 7 and
9, the tautomer I is the most predominant species (75%), in
accordance the tautomer II being energetically less stable
than the tautomer I (23). The tautomeric equilibrium is
affected by metal chelation. Binding of a Cu(II) ion and
dimer complex formation (vide infra) makes the tautomer II
dominate since N␶ is involved in the metal chelation (370).
The nitrogen atoms of the imidazole ring of carnosine regulate the buffering activity of the dipeptide. It is suggested
that because the pKa values of carnosine and anserine are
O
O
H
N
N
4 π
HO
O
5 τ
NH
NH2
Tautomer I
FIGURE 7.
carnosine.
1808
2
N
H
4 π 2
HO
O
5 τ
N
NH
NH2
Tautomer II
Tautomeric forms of the imidazolic ring of
Several analytical studies have been reported on the ability
of carnosine to form complexes with metals of the first
transition metal series such as Cu2⫹, Co2⫹, Ni2⫹, Cd2⫹, and
Zn2⫹. Copper and zinc complexes have been the most investigated ones, due to their biological relevance and are
described below.
1. Copper-carnosine complex
Several models have been proposed for the structure of the
Cu2⫹-carnosine complex in aqueous solution since 1955.
Dobbie and Kermack (108) first proposed the structure of
Cu2⫹-carnosine complex in which Cu2⫹ is bound to the
imidazole, amino, and peptide nitrogen atoms, forming two
six-membered chelate rings (FIGURE 8). The model was then
confirmed by other authors who agreed on a monomeric
form where Cu(II) is initially bound to the N␶ atom of
imidazole and that, as the pH is increased, the amino and
peptide nitrogen atoms deprotonate and bind to the
metal (44).
Later, Freeman and Szymanski (44) have determined, by
X-ray crystallography, the structure of the complex in the
solid state which was not a monomer but a dimer in which
the amino group, peptide nitrogen atom, and one carboxyl
oxygen atom of one peptide molecule, and the N␲ atom of
the imidazole ring of the second peptide molecule are
bonded to Cu2⫹. It was then proposed that the dimeric form
exists in equilibrium with the monomer in solution and that
the equilibrium is shifted towards the dimer at neutral pH.
Later, by using different analytical techniques including
ESI-MS (256) and Raman and IR spectroscopy (370 –372),
it has been found that, at neutral and basic pH and when
Cu(II) and carnosine are mixed in a 1:1 molar ratio, the
most relevant species formed is the dimer [Cu2H-2L2]0
which exists in equilibrium with the monomeric species. In
the dimeric form, the ligand can scavenge the metal through
the carboxylate, amide, and amino donor atoms. In the
imidazole rings (tautomer II), the N␶ nitrogen atoms bind
the copper(II) ions. The monomeric complexes [CuLH]2⫹
and [CuL]1⫹ are present at a somewhat lower pH. Consistently, the four chelation sites bound the Cu(II) ions and the
complexes of Cu(II) and carnosine presumably have a
square-planar coordination sphere (371).
Most of the studies on the characterization of Cu2⫹-carnosine complexes have been carried out in aqueous solutions,
but the existence of the dimer in living organisms was considered improbable due to the low copper concentration
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
A
O
OH
N
N
N
Cu
O
2HN
B
0
N
2HN
H2O
Cu
N O
H
O O
O
N
O
Cu
NH2
+
N
O
O
O
N
[CuL]+
O
Cu
N
H2O
H2O
O
H
N
N
H
NH
[Cu2H-2L2]0
O
2HN
Cu
N
2+
O
NH
NH3
N
H
[CuHL]2+
FIGURE 8. Proposed structures of the Cu(II)-carnosine complex. A: Cu(II)-carnosine complex as first proposed by Dobbie and Kermack (108). B: molecular structures of the dimeric Cu(II) carnosine complex
[Cu2H-2L2]0 and of the two monomers [CuL]⫹ [CuHL]2⫹ as reported by Tamba et al. (359).
and the presence of competing ligands that would shield
monomeric species with mixed ligands (75). Such consideration was confirmed by using localized 1H-NMR spectroscopy of human calf muscle. Schröder et al. (329) demonstrated that the dimer complex is insignificant in vivo,
whereas the presence of a monomeric species was confirmed, although in a low amount. Finally, the paper by
Velez et al. (382) should be carefully considered, because
they have directly compared the metal ion binding properties of carnosine and L-histidine. Interestingly, carnosine, at
pH 7.84 ⫾ 0.18, was found to possess a much lower binding constant with Cu(II) ions (1.1 M⫺1) compared with that
of L-histidine (71 M⫺1), as determined by Benesi-Hildebrand method.
troscopy by Torreggiani et al. (370). The dimer is the most
prevalent species at neutral pH, especially if the ligand/
metal ratio is 4, along with the charged monomeric complex, [ZnHL]2⫹, in which the N␲-imidazole nitrogen contributes to the coordination. In more acidic conditions, the
latter is also formed with monomeric complex, [ZnL]⫹ as
the most predominant. The existence at neutral pH of a
dimeric form was confirmed by ESI-MS experiments (256).
In conclusion, most of the studies on carnosine as a chelator
of Cu2⫹ and Zn2⫹ have been carried out in vitro and using
conditions very far from the physiological ones. Hence,
more studies are required not only to understand whether
carnosine is an effective physiological chelator of metal ions
but also to clarify the biological roles of such mechanisms.
2. Zinc-carnosine complex
There has been a recent research interest in the Zn(II)-carnosine complex because of its interesting and important
pharmacological application (250). In fact, a zinc-carnosine
complex (Polaprezinc or Z-103) was shown to protect gastric mucosa from experimental ulcerations in vivo (172,
218), and to be effective against Helicobacter pylori-associated gastritis (157, 195). Polaprezinc is a zinc complex
that is dissolvable in acid and appears to have the characteristic to protect against mucosal lesions. It has a stable
presence in the stomach and can adhere to ulcerous sites in
a much better way than either zinc or carnosine alone (141).
It is only possible to perform solution studies on the Zn(II)/
carnosine system in a narrow pH range (129). At a pH of 6,
there is an appreciable complex formation, but at higher pH
(above 7.5) precipitation will take place even when the ligand is in excess (129).
C. Antioxidant Activity
The zinc(II)-carnosine complex was investigated at different
pH values and metal/ligand ratios by Raman and IR spec-
At physiological concentrations, carnosine was found to
directly react with superoxide anion like superoxide dismu-
The antioxidant activity of carnosine was initially studied
by Boldyrev et al. since the 1980s (65– 67). Later on, several
studies were reported, showing a direct and indirect antioxidant activity of carnosine, as demonstrated in several in
vitro studies (see below). The antioxidant activity of carnosine is mediated by different mechanisms involving metal
ion chelation, and scavenging reactive oxygen species
(ROS) and peroxyl radicals. The metal ion chelating properties of HCD have been discussed above and below the
radical scavenging effect of HCD towards ROS and peroxyl
radical is described.
1. Reaction of HCD with ROS, peroxynitrite, and
hypochlorous acid
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1809
BOLDYREV, ALDINI, AND DERAVE
tase (SOD) and the constant for interaction of carnosine
with O2·⫺ was calculated to be 105 M⫺1·s⫺1 and not significantly different in respect to that of ascorbic acid and ␣-tocopherol (217). The mechanism of interaction of carnosine
with superoxide radicals was studied by Pavlov et al. (291)
in water solutions and is based on the ability of carnosine to
form a charge-transfer complex with the superoxide radical
which changes the reactivity of O2·⫺. Carnosine is also an
effective quencher of other ROS such as hydroxyl radicals
(·OH). In particular, evidence was provided through pulseradiolysis (358) indicating that carnosine is an adequate
scavenger of ·OH radicals. The reaction results in a reasonably stable intermediate that is less reactive than ·OH itself
towards other molecules. This transient intermediate is
obtained by a base-catalyzed water elimination reaction
(FIGURE 9).
reaction has been studied by Fontana et al. (133). The results showed that carnosine, at concentrations similar to
those found in human tissues, efficiently protects tyrosine
against nitration, ␣1-antiproteinase against inactivation
and human low-density lipoprotein against modification by
peroxynitrite. A similar protective effect was shown for
anserine, and the scavenging activity of carnosine and anserine was ascribable to the imidazole moieties of the molecules, since L-histidine but not ␤-alanine is an effective
inhibitor of peroxynitrite-induced tyrosine nitration (133).
Carnosine is also a highly effective protective agent against
hypochlorite (HOCl). Hipkiss et al. (184) reported the ability of carnosine to dose-dependently inhibit protein crosslinking and high-molecular-weight oligomers induced by
hypochlorite. The mechanism of reactions has been later
studied by Pattison et al. (290) who found that carnosine, as
well as other compounds containing imidazole and free amine
sites, rapidly form imidazole chloramines. In the case of carnosine, a selective intramolecular chlorine transfer reaction
occurs, leading a long-lived chloramine which, in contrast to
chloramine formed by amino compounds, is inefficient at inducing further chlorination, thus limiting HOCl-mediated oxidation in vivo. The mechanism for the peculiar intramolecular Cl shift in carnosine from the imidazole ring (the kinetically
favored site) to the terminal amine has been more recently
studied by Karton et al. (210) who found that the intramolecular Cl shift in carnosine occurs due to a unique structural
relationship between three adjacent functional groups,
namely, the imidazole ring, the carboxylic group, and the ␤-alanyl primary amine functions.
In the presence of copper, the fast oxidation of carnosine by
OH still takes place (at 6.5 the formation rate constant is
7.9 ⫻ 109 M⫺1·s⫺1)(369) and yields adducts of ·OH radicals at the imidazole ring (359). Thus the imidazole group is
also the preferred site for ·OH binding in the presence of
copper.
·
Imidazole compounds are known to react with singlet oxygen forming endoperoxide products. Accordingly, carnosine was reported to react with singlet oxygen two- to fourfold faster than free L-histidine and approximately twofold
faster than the NH2-terminal L-histidine dipeptides tested
(167). The rate constants of 1O2 quenching (Kq) by histidine
containing peptides towards singlet oxygen was then determined by monitoring the 1O2-phosphorescence (1270 nm).
The Kq values of carnosine and anserine were shown to be
similar [(3 ⫾ 1) ⫻ 107 M⫺1·s⫺1] and slightly higher in
respect to that of imidazole [Kq ⫽ (2 ⫾ 1) ⫻ 107 M⫺1·s⫺1].
The nonaromatic amino acids taurine and ␤-alanine
showed very low quenching activities (Kq ⬍ 3 ⫻ 103
M⫺1·s⫺1) (124).
2. Reaction with peroxyl radicals
Kohen et al. (219) reported the ability of carnosine and its
analogs such as anserine and homocarnosine to scavenge
peroxyl radicals. They further demonstrated that the L-histidine moiety is required for this activity. The antioxidant
and in particular the radical scavenging activity of carnosine was then confirmed by several other in vitro studies
which also reported a reducing ability of carnosine that
The ability of carnosine and related peptides to act as protective agents by counteracting the peroxynitrite-dependent
O
H
N
OH
O
NH
R
·OH
H
N
·
N
N
R
H
OH
N
H
·
+ -OH
N
NH2
- -OH
R
N
·
FIGURE 9.
1810
OH
·
N
Carnosine as a scavenger of OH radicals. [Based on Tamba and Torreggiani (358).]
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
would explain the radical scavenging mechanism (408).
However, it should be noted that most of these studies
overestimated the true antioxidant property of carnosine
and in particular its reducing ability, since they were performed by using synthetic carnosine containing hydrazine
as common contaminant which acts as a powerful reducing
agent that can deactivate free radicals (408). The antioxidant activity of carnosine was then reevaluated by Decker et
al. (100) using a purified carnosine. They confirmed that
purified carnosine was capable of scavenging peroxyl radicals at a millimolar concentration range, with a potency
similar to that shown by L-histidine, while ␤-alanine was
ineffective, thus suggesting that the radical scavenging activity of carnosine is largely due to the L-histidine residue.
Purified carnosine was then found not able to reduce MbFe(III) to MbFe(II)O2, clearly indicating that the radical
scavenging activity of carnosine is not due to reducing properties. The fact that carnosine does not act as an electrotransfer agent is also confirmed by the inability of carnosine
to scavenge the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals (at least up to 1 mM), in contrast to the well-known
reducing agents ascorbic acid or glutathione, that have this
effect in the micromolar concentration range (14).
3. Overall antioxidant activity: in vitro models
The antioxidant activity of carnosine and related peptides
has been demonstrated in several in vitro models, based on
metal ion-dependent and -independent inducers of oxidative damage (metal ions/H2O2, radical inducers), with proteins, nucleic acids, and lipids as substrates, and based on a
variety of methods for assessing the antioxidant capacity.
When metal ions were used to generate free radicals, carnosine was observed to have a greater antioxidant activity
when oxidation was promoted by copper than by iron
(100). This is due to the ability of carnosine to chelate and
deactivate copper ions while it can only chelate iron in a
manner that it does not reduce its pro-oxidant activity (99).
Overall, the efficacy of carnosine was usually found quite
similar to that of the other HCD (anserine, homocarnosine), spanning the millimolar range. In some studies, the
antioxidant effect of carnosine was compared with that of
the constitutive amino acids, ␤-alanine and L-histidine, and
the results depend on the models used, although it should be
considered that ␤-alanine was usually found ineffective.
4. Antioxidant activity: in vivo models
The in vivo antioxidant activity of carnosine has been demonstrated in several studies which can be divided into two
groups: the first, reporting the antioxidant efficacy of carnosine in physiological conditions, and the second, in animal models where oxidative stress was caused by xenobiotics acting as radical inducers, such as ethanol, thioacetamide, and anticancer agents.
In physiological conditions, carnosine was found to reduce, on
one hand, the oxidative damage and on the other hand to
improve the enzymatic and nonenzymatic antioxidant activity. More specifically, carnosine treatment for 1 mo at a daily
dose of 250 mg/kg (ip) was found effective in aged (but not in
young) rats to reduce some markers of lipid oxidation and to
restore the depleted levels of blood glutathione (GSH) and
basal activities of antioxidant enzymes such as SOD and glutathione peroxidase (GPX) (26). Accordingly, Kim et al. (215)
reported that 6 wk of carnosine supplementation at a dose of
0.5% in the diet of rats was found to increase the activities of
enzymatic antioxidants such as SOD and GPX. Furthermore,
the supplementation induced a decrease in lipid peroxidation
in serum, liver, and skin, and positively modified blood lipid
profiles. In pigs, the addition of 100 mg/kg diet increased GPX,
SOD, and catalase activities in plasma, liver, and muscle as
well as SOD and GPX expression in muscle (242). In addition
to affecting antioxidant enzymes, carnosine was also found to
impact on low-molecular-weight antioxidants. Aydin et al.
(26) reported that liver vitamin E was found to be significantly
elevated in carnosine-treated aged rats. A sparing or regenerating effect of carnosine towards endogenous antioxidants
was demonstrated in the liver of rats treated with carnosine or
L-histidine. Both compounds increased the liver content of
GSH and ␣-tocopherol (398). In contrast to the abovementioned studies, Ibrahim et al. (192) reported that carnosine
treatment for 15 days at levels of 200 or 1,000 mg/kg diet did
not significantly affect the antioxidant and oxidative status in
skeletal muscle, liver, and blood of rats.
Many studies have been reported on the antioxidant activity of
carnosine in animal models where the oxidative stress condition in different organs was induced by xenobiotics. The protective effect of carnosine in liver was studied in a rat model of
ethanol-induced chronic liver injury (237). Carnosine, as well
as L-histidine, was found to dose-dependently inhibit the
ethanol-induced oxidative damage and inflammatory mediators, and to increase catalase expression, GSH content, and
GPX activity (237). The antioxidant activity in the liver and
the reduction in plasma transaminases activities by carnosine
in ethanol-treated rats was confirmed by Artun et al. (22) and
Baykara et al. (55). The hepatic antioxidant activity of carnosine was also confirmed in a model of thioacetamide-induced
liver cirrhosis (27).
Supplemented carnosine was also found to act as an antioxidant in the brain, as evidenced by decreased MDA and
protein carbonyls induced by ethanol ingestion (284) and
by protecting cerebral cytosolic SOD in two in vivo models
of oxidative stress: hypobaric hypoxia in rats (150 mg/kg
carnosine in drinking water before the hypoxic exposure)
and in senescence-accelerated mice (100 mg/kg carnosine in
drinking water for 12 mo) (347).
The antioxidant activity of carnosine has been shown in skeletal muscle. A 2-wk L-histidine supplementation period in rats
can markedly increase the skeletal muscle carnosine content.
When subjecting these rats to a Fe-nitrilotriacetate administra-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1811
BOLDYREV, ALDINI, AND DERAVE
tion treatment, carnosine loading leads to a reduction in protein carbonyl content and lipid peroxidation (267).
propagation of several oxidative-based diseases such as diabetes, atherosclerosis, and Alzheimer’s disease.
In addition to inhibiting oxidative damage and regulating
the antioxidant enzymes, some studies have found a strong
relationship between the antioxidant activity induced by
carnosine treatment and the protection of the organ injury
and dysfunction induced by the oxidative damage. In particular, carnosine pretreatment was found to prevent bleomycin-induced lung injury, fibrosis and, most notably, a
complete abrogation of the mortality (92). The protective
effect of carnosine was attributed to its ability to scavenge
superoxide and peroxinitrite which are induced by bleomycin treatment. A beneficial effect of carnosine treatment was
also reported in a model of cardiomyopathy induced by
adriamycin which is a chemotherapeutic agent able to induce hydroxyl radical formation and lipid peroxidation
(121). Finally, the antioxidant activity of carnosine was
demonstrated in a model of kidney damage caused by cisplatin which is known to induce ROS in renal epithelial cells
primarily by decreasing the activity of antioxidant enzymes
and by depleting intracellular concentrations of GSH. Carnosine prevented the cisplatin-induced kidney oxidative
damage, together with a protection of the organ histology
and function (135, 274).
1. In vitro models
It can be concluded that there is much evidence indicating
that, at least in rodents, supplemented carnosine acts as an
antioxidant in both physiological conditions and in models
of induced oxidative damage by xenobiotics. The antioxidant effect can be explained by considering both a direct
scavenging effect towards ROS and by an increase and/or
sparing and/or regenerating effect on enzymatic and nonenzymatic antioxidants. In some cases, such an antioxidant
effect was found to be linked to a reduction in tissue damage
and functional impairment. However, some questions remain unanswered, in particular regarding the mechanism
through which carnosine potentiates the enzymatic antioxidants. Moreover, most of the studies showing the in vivo
antioxidant activity of carnosine were carried out by supplementing carnosine at pharmacological doses (⬃100 mg/
kg) for several weeks. It remains to be established whether
endogenous carnosine (in nonsupplemented conditions) is
contributing to the antioxidant system. Furthermore, it
should be clarified in what body compartments this contribution is situated and whether the system is of quantitative
relevance in the human organism.
D. Inhibiting Protein Carbonylation and
Glycoxidation
Several in vitro and in vivo studies have reported the ability of
carnosine and related peptides to prevent the formation of
advanced lipoxidation end-products (ALEs) and advanced
glycoxidation end-products (AGEs). These compounds are
both involved in the aging process as well as in the onset and
1812
Hipkiss first reported a series of papers showing the ability
of carnosine to inhibit AGEs and ALEs formation induced
by different precursors, including reducing sugars such as
glucose (181), deoxyribose, ribose, fructose (180), and reactive carbonyls such as malondialdehyde (183, 184),
glyoxal, methylglyoxal (179), acetaldehyde, and formaldehyde (182). The ability of carnosine to inhibit AGEs and
ALEs formation has then been confirmed and extended in
different in vitro models, as recently reviewed by Hipkiss
(174). The ability of carnosine to prevent AGEs and ALEs
formation is mediated by different mechanisms, and this is
due to the fact that AGEs and ALEs formation is a quite
complicated pathway, involving different reaction mechanisms and several catalyzing agents, including transition
metal ions. Glycation represents the first step of AGEs formation, involving the reaction of the carbonyl group of
reducing sugars with the amino group of lysine, followed by
several oxidative steps that are catalyzed by metal ions. An
alternative pathway consists of a direct reaction between
reactive carbonyls generated by sugar auto-oxidation or
AGEs decomposition and the nucleophilic sites of protein.
A detailed description of the pathways of AGEs and ALEs
formation is beyond the scope of the present review, but the
reader is referred to some extensive reviews published on this
aspect (11, 268, 285).
The different sites of the AGEs and ALEs cascade where carnosine has been proposed to act is shown in FIGURES 10 and 11,
respectively. Carnosine was found to be effective already in
the first step of AGEs formation and in particular in inhibiting protein glycation as well as by reversing glycated protein through a transglycation mechanism. In more detail,
carnosine was found to protect glycation-induced loss of
enzyme activity, and this effect paralleled the adduct formation between carnosine and sugar aldehyde such as glyceraldehyde 3-phosphate (330). The antiglycating property of
carnosine has been found in several other in vitro methods
(15, 295, 305, 312, 331, 397). Carnosine and anserine were
also reported to act as transglycating agents, thus reversing
the formation of the Schiff base, which is the first product of
protein glycation (353). The mechanism is based on the
nucleophilic attack of carnosine and/or anserine on the preformed aldosamine such as glucosyl-lysine with formation
of glucosyl carnosine or anserine. The transglycation properties of carnosine have been confirmed in different in vitro
models, the most striking being their ability to reverse preexisting glycated proteins (332). Another potential mechanism of carnosine concerns its ability to detoxify reactive
carbonyl species (RCS), precursor of AGEs, but up to now
no direct evidence has been reported to confirm such mechanism. Carnosine can also inhibit AGEs formation through an
antioxidant mechanism and more likely by a metal ion chelat-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
1
CARNOSINE
Transglycating
effect
O
H
OH
HO
H
H
OH
H
OH
H2N
+
Lys
N
Protein
H
HO
2
OH
Lys
Protein
OH
H
H
OH
H
OH
Schiff base
Glucose
OH
Oxidation
3
Reactive carbonyl
species (RCS)
H
N
Oxidation
AGEs
4
H
O
HO
H
H
OH
H
OH
Lys
Protein
Amadori product
OH
FIGURE 10. Carnosine as an inhibitor of advanced glycoxidation end products (AGEs) formation. Carnosine
can act as AGEs inhibitor through the following mechanisms: 1) restoring the native protein from the Schiff
base by a transglycating effect; 2) by a hypoglycemic effect; 3) by detoxifying RCS; and 4) by inhibiting the
oxidative conversion of glycated proteins to AGEs.
ing mechanism as suggested by Nagai et al. (266), who showed
that AGEs inhibitors and breakers act primarily as chelators,
inhibiting metal-catalyzed oxidation reactions that lead to
AGEs formation.
Carnosine is also a potent inhibitor of ALEs formation.
There is currently enough evidence to indicate that this
relates to the ability of carnosine to act by a direct RCS
quenching mechanism (10). The ability of carnosine to react
with ␣,␤-unsaturated aldehydes, such as HNE, which represent the most abundant class of ALEs precursors, was
firstly reported by Zhou et al. (407), who demonstrated the
disappearance of the reactive aldehydes (measured through
HPLC) when incubated in the presence of carnosine. By
means of MS and NMR, the reaction mechanism was further elucidated by showing the quenching effect of carnosine and by identifying the structure of the reaction products
(9, 12). Particularly, the N-acetylation of the ␤-alanine
amine group was found to preclude the quenching activity.
Moreover, the ␤-alanine and L-histidine were less efficient
than carnosine itself, indicating that 1) the amino acids act
synergistically in the dipeptide and 2) the reaction involves
the amine group of ␤-alanine. As depicted in FIGURE 12, the
multistep reaction mechanism between carnosine and HNE
involves both the amino group of ␤-alanine and the imidazolic ring as reported by two independent groups (9, 238).
In more detail, the reaction starts with the formation of a
Schiff base that is formed by the reaction of the aldehyde
with the amino group, followed by an intramolecular Michael addition, yielding the macrocyclic adduct which finally hydrolyzes to form the stable hemiacetal derivative
(FIGURE 12, compound 5a). The reaction was then elucidated also for acrolein, which is another reactive ␣,␤-unsaturated aldehyde (78). In addition to test tube experiments,
the formation of the Michael adducts between carnosine
and anserine with HNE were also demonstrated by LCESI-MS in biological matrices and in particular in oxidized
skeletal muscle (283).
In addition to acting as an inhibitor of AGEs and ALEs
formation, carnosine was also hypothesized to act on preformed AGEs/ALEs. Hipkiss found that carnosine is not
only able to prevent protein carbonylation but also to react
directly with protein carbonyl groups, producing protein-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1813
BOLDYREV, ALDINI, AND DERAVE
effects were reversed by carnosine treatment. Carnosine
was found to protect human peritoneal mesothelial cells
against the toxic effect of glucose degradation products
through the protection of cellular protein from modification and ROS-mediated oxidative damage (15). In addition
to inhibiting AGEs formation, carnosine was also found
able to protect cells against AGE-induced cell damage. In
human skin fibroblasts, carnosine was more effective than
N-acetylcysteine in avoiding cell damage induced by AGEs
(302).
CARNOSINE
Lipids
1
Oxidation
2
Reactive carbonyl species (RCS)
Nu
Protein
3
Carbonylated proteins
ALEs
Cell
damage
FIGURE 11. Carnosine as an inhibitor of advanced lipoxidation end
products (ALEs) formation. Nu represents the protein nucleophilic
sites undergoing RCS adduction. Carnosine can act as ALEs inhibitor
through the following mechanisms: 1) inhibiting lipid oxidation and
breakdown to RCS; 2) detoxifying RCS; and 3) reacting with carbonylated proteins (“carnosinylation”).
carbonyl-carnosine adducts or “carnosinylated” proteins,
thereby preventing cross-linking to other, unmodified protein (76, 176, 178). It was also suggested that since oxidized
proteins can inhibit proteasome function, carnosine may
suppress these inhibitory effects on proteasome activity and
possibly facilitate their proteolytic elimination, through reacting with protein carbonyls in oxidized proteins (177). It
must be stressed that the latter hypotheses (protein carnosinylation and proteasome modulation) are based on chemical experiments only, and no evidence for its involvement in
biological events is available at present.
2. In vivo models
The ability of carnosine to prevent AGEs and ALEs formation has also been demonstrated in some cellular models. By
using a prokaryotic model (Escherichia coli), Pepper et al.
(295) reported the ability of carnosine to suppress methylglyoxal- and glucose-mediated toxicity and the formation
of AGEs such as carboxymethyl lysine (CML). In cultured
neurons, Cheng et al. (82) demonstrated that MDA caused
protein cross-linking and cytotoxicity, and that both these
1814
With respect to ALEs, carnosine has the capacity to quench
HNE in vivo as demonstrated by identifying the HNE-carnosine Michael adduct and related metabolites in the urine
of Zucker obese rats, which is an animal model of oxidative
stress and carbonylation damage (282). Aldini et al. (14)
recently reported that carnosine supplementation for 24 wk
in Zucker rats significantly reduced the urinary AGEs and
protein carbonylation in the kidney, and improved collagen
solubility as an indicator of the extent of collagen crosslinking. However, more studies regarding the in vivo effect
of carnosine on AGEs and ALEs formation should be carried out, and it remains to be understood whether the reaction of carnosine towards electrophilic aldehyde is catalyzed by an enzyme. Recently, some studies have shown that
oral carnosine supplementation was not effective in reducing the levels of methylglyoxal-, CML-, and N-acetylglucosamine (304, 317).
E. Carnosine and Aberrant Proteins
Carnosine has been shown to stimulate neutral (nonlysosomal) protease activity in cell-free extracts from rat brain
and in cultured cardiomyocytes (57, 71). Intracellular elimination of aberrant polypeptides mostly, though not exclusively, involves the proteasomes, and recent evidence suggests that nitric oxide can stimulate proteasomal activity
(223, 363) and that carnosine can interfere with NO production (see below). Moreover, carnosine may accelerate
protein turnover in two steps: a first rapid step which involves a localized thermal unfolding, and then a proteolytic
event, which is slower (199). Possibly, the dipeptide can
initiate the first step in protein removal, through assistance
in damaged proteins unfolding. Carnosine enhances the
thermal unfolding of glycated protein as evidenced by a
decreased Gibbs free energy barrier and, additionally, decreases the enthalpy of denaturation, suggesting that carnosine may promote hydration during heat denaturation of
glycated protein (399).
Carnosine was also reported to impact on aggregates and
fibrils by disassembling/disaggregating them. This effect
was observed by the group of Rizzarelli (24) showing that
the dipeptide can prevent fibril formation in ␣-crystallin as
well as in a more complex model of cultured lens organ. The
lens transparency could be almost completely restored by
disassembling already formed fibrils (24).
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
OH
O
NH2
O
HO
NH
NH
O
OH
NH
O
O
NH
NH
HNE
OH
N
OH
+
N
NH2
O
N
O
N
O
O
NH2
NH
N
O
5a
O
OH
N
OH
N
2a
OH
N
O
NH
OH
N
H2O
O
N
OH
4a
3a
FIGURE 12. Reaction mechanism of carnosine with 4-hydroxy-trans-2,3-nonenal (HNE). [Adapted from Aldini
et al. (9), with permission from Elsevier.]
F. Carnosine and Nitric Oxide
Several studies have so far been reported on the ability of
carnosine to impact on nitric oxide (NO) release, metabolism, and activity. There currently is an apparent contradiction in the literature, as some studies have shown a stimulating effect of carnosine towards NO production, while
some others point to a direct or indirect inhibitory mechanism.
With regard to the stimulating effect, Alaghban-Zadeh (8)
reported that carnosine, in the presence of NADPH, induced NO synthase (NOS) activity as measured by soluble
guanylate cyclase. They identified that the effect was higher
than that induced by arginine and ␤-alanine and comparable to that of L-histidine (8). More recently, Takahashi et al.
(354) reported the ability of carnosine to stimulate NO
production in the endothelium at concentrations higher
than 5 mM, while neither the constituent amino acids of
carnosine nor anserine had an effect on NO production. It
was suggested that the mechanisms of carnosine action on
eNOS activation may include carnosine-induced Ca2⫹ mobilization from intracellular stores. Carnosine was found to
induce hyperactivity in chicks, and this effect was linked to
NO generation via NOS in the brain, as demonstrated by
using selective inhibitors as L-NAME (368).
The inhibitory effect of carnosine on NO-dependent signaling was first reported by Severina et al. (335), who showed
that carnosine is a selective inhibitor of NO-dependent activation of guanylate cyclase. Later a direct scavenging effect of carnosine was demonstrated. In particular, in cellfree experiments based on competitive assays, the group of
Rizzarelli (271) showed that carnosine significantly reduced
NO concentration and that such a scavenging effect was not
reproduced using the constituent amino acid of carnosine or
their mixture, thus suggesting that the two amino acids act
synergistically only when combined in the dipeptide. The
scavenging activity of carnosine was then confirmed by ESIMS, showing the formation of carnosine/NO and carnosine/NO2 adducts at different ratios and the binding of L-histidine with NO (271). The scavenging activity of carnosine
was then tested in a cell system, by examining the protection
that carnosine provides against NO-induced cell death in
primary rat astroglial cell cultures treated with lipopolysaccharide and interferon-␥. A correlation was found between
cellular protection and NO free radical scavenging activity
of carnosine. Also N-acetyl carnosine was demonstrated to
directly react with NO (132). In conclusion, there is available evidence for both an inhibitory and stimulatory effect
of carnosine on NO production and metabolism, and it
remains to be further determined which factors (concentration dependence, tissue specificity, etc.) drive the effect in
one way or the other.
IV. ENZYMES AND TRANSPORTERS
INVOLVED IN CARNOSINE METABOLISM
This section discusses the enzymes involved in the direct
formation and degradation of carnosine and its methylated
analogs. Other enzymatic reactions, such as decarboxylation of carnosine (carcinine production) and acetylation of
carnosine, are poorly understood and will not be treated.
The proteins capable of transmembrane transport of carnosine will be described at the end of the section. The functional groups of the carnosine molecule required for recognition by carnosinases and carnosine transporters are displayed in FIGURE 13.
A. Carnosine Synthase
Soon after the discovery of carnosine (154) it became clear
that ␤-alanine and L-histidine were the two constituent
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1815
BOLDYREV, ALDINI, AND DERAVE
CN1
O
PepT1
PepT1
CN1
PepT1
H
N
HO
O
N
NH
CN1
PepT1
NH2
PepT1
CN1
FIGURE 13. Structure activity relationships as derived by analyzing the reported carnosine derivatives. Notice that the considerations described for PepT1 hold reasonably true also for PepT2,
PHT1, and PHT2. [Adapted from Vistoli et al. (383), with permission
from Springer Science ⫹ Business Media.]
amino acids (153) required for synthesis. Indeed, during
ontogeny, the appearance of dipeptides is accompanied by
an abrupt decrease in the content of L-histidine and ␤-alanine (61). Bauman and Ingvaldsen (54) showed that they
were linked in a ␤-alanyl-L-histidine and not histidyl-␤-alanine formation. In the 1950s, the biochemical properties of
the carnosine synthesis reaction were further established by
in vitro experiments on partially purified enzyme preparations of chicken muscle, and the enzyme was called carnosine synthase (CS). It became clear that in addition to the
constituent amino acids, Mg2⫹, and ATP are required to
synthesize carnosine (205, 391). CS appeared to have broad
substrate specificity, and various other dipeptides can be
formed by ␤-alanine or L-histidine replacement (205).
Over the next 50 years, several attempts were made to further purify the enzyme (189, 339, 345), but it was not until
2010 that CS was finally molecularly identified by Drozak
et al. (114). This breakthrough was necessary to resolve and
correct the reaction properties that were derived from previous experiments with less purified preparations. A 1,000fold purification of chicken CS was shown to stoichiometrically form ADP ⫹ Pi, rather than AMP ⫹ pyrophosphate,
as was previously suggested (205, 345). Characterization of
CS as an ADP-producing ligase then led to the identification
as a member of the ATP-grasp superfamily and more specifically as ATPGD1 (ATP-Grasp domain containing protein 1; Gencode: NP_001159694.1) (114). The enzyme is
thought to be a homotetramer with a molecular mass of
⬃430 kDa for the native enzyme. Its gene is located on
chromosome 11q13 in humans and contains 9 coding exons
and expresses a 950-amino acid sequence.
Although early experiments (390) suggested that liver slices
can synthesize carnosine, it later became clear that carnosine synthase (ATPGD1) is mainly present in skeletal and
heart muscle and certain brain regions (114). The olfactory
neurons display very high CS expression, which is in agreement with its very high carnosine content (189, 248). In
other brain structures, however, homocarnosine rather
than carnosine is the main dipeptide. CS is now believed to
be responsible for the homocarnosine synthesis in the brain
1816
(269, 339). Oligodendrocytes, but not astrocytes, have been
shown to possess carnosine synthase activity (51, 186).
The enzyme is localized intracellularly, and studies on brain
subcellular fractions indicate that it is a cytosolic enzyme
(161, 270). The targetP calculation of the protein sequence,
which predicts the subcellular location of eukaryotic proteins, confirms the expected cytosolic localization (114).
Little is known about the regulatory mechanism of CS activity and expression. The highly conserved cysteine residues in the carnosine synthase protein sequence suggest that
the enzyme is potentially redox regulated (114).
Recently, Tsubone et al. (375) reported a potential novel
imidazole dipeptide synthase, which they purified from eel
muscle. They propose that this enzyme is different from the
above-mentioned CS (EC 6.3.2.11), as the synthesis would
not require ATP.
B. Carnosine N-methyltransferase
There are two possible pathways for the synthesis of the
methylated carnosine analogs, anserine (␤-alanyl-Npimethyl-histidine) and ophidine/balenine (␤-alanyl-Ntaumethyl-histidine). The first enzymatic pathway is through
carnosine N-methyltransferase (CMT), which catalyzes the
transfer of a methyl group of S-adenosylmethionine (SAM)
on carnosine to form anserine (or ophidine). A second pathway is the enzymatic condensation of ␤-alanine with Npimethylhistine (or Ntau-methylhistidine) through CS. Several studies suggest that the former pathway, i.e., through
CMT, is of more physiological importance.
The first evidence for the existence of a carnosine methylation process was provided by Winnick and Winnick (391),
who demonstrated in vitro in a chick pectoral muscle preparation that a methyl group from SAM can be transferred to
carnosine. Subsequently, McManus (254) partially purified
CMT (EC 2.1.1.22) from this chick pectoral muscle preparation. The Km for carnosine was found to be 4 mM, which
is a concentration comparable to the endogenous muscle
levels. The enzyme is confined to the soluble fraction of
muscle homogenate, and it does not seem to have specific
ion requirements. Interestingly, L-histidine was only found
to be a substrate of this enzyme preparation, when bound
into carnosine, and not as a free amino acid. Raghavan et al.
(310) showed that CMT can also methylate L-histidine residues in proteins, in conjunction with another unidentified protein, histidine-N-methyltransferase (EC 2.1.1.85). L-Histidine
methylation in proteins is a rare but evolutionarily conserved
posttranslational modification that specifically occurs in muscle proteins actin and myosin.
The second possible pathway for anserine (ophidine) synthesis is through direct condensation of ␤-alanine with
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
methyl-L-histidine. The ability of CS to use methyl-L-histidine as a substrate has been demonstrated by in vitro enzymatic assays with chicken muscle preparations by two independent groups in 1959 (205, 391). This was recently
confirmed using recombinant CS (114). Yet, methyl-Lhistidine was consistently found to be a poorer substrate
than nonmethylated L-histidine, making anserine synthesis
slower than carnosine synthesis. Furthermore, a study by
Bauer and Schulz (53) in primary culture myocytes indicated that anserine is not directly formed by condensation
of radiolabeled ␤-alanine with Npi-methyl-L-histidine, but
rather as a consequence of carnosine methylation, secondary to carnosine synthesis. Thus CMT and not CS is the
major enzyme involved in anserine synthesis.
1. Human serum carnosinase
Drozak et al. (113) recently molecularly identified the avian
CMT as the histamine N-methyltransferase (HNMT)-like
protein. Although the enzyme has high specificity for carnosine as a methyl acceptor, it probably evolved as a paralog of a histamine-methyltransferase, which acquired new
enzymatic activity (113). Surprisingly, the mammalian genomes do not contain orthologs of the CMT gene (HNMTlike), discovered in chicken, meaning that the mammalian
version evolved in a different manner and still needs to be
discovered.
CN1 is highly present in the serum of humans, but absent
from nonprimate mammals, except for the Syrian golden
hamster (196). For its presence in human serum, the enzyme
may be produced by the brain, but is presumably mainly
produced and secreted from the liver into the circulation,
which is supported by the finding that patients with liver
cirrhosis have low CN1 activity (300). By analogy with
other secreted proteins, CN1 is synthesized as a precursor
containing an NH2-terminal signal peptide sequence. There
is a (CTG)n polymorphism in this signal peptide which
appears to determine the rate of CN1 secretion. Riedl et al.
(315) recently showed that a high number of leucine repeats
enhances the secretion efficiency of carnosinase to the serum (see sect. VII). In addition, CN1 is N-glycosylated at
three different sites, and the glycosylation appears to be
essential for secretion of the enzyme (316). Anserine, ophidine, and homocarnosine are found to be more resistant to
carnosinase (293) and to be potent inhibitors of carnosinase
activity towards carnosine as a substrate (300, 301).
There is little or no insight into whether there are further
processes regulating anserine rather than balenine/ophidine
synthesis, but it would make sense that there is a different
enzyme for each compound. Likewise, it is unclear whether
the acetylation of carnosine, anserine, and homocarnosine
requires a specific enzyme to make the specific products
(acetylcarnosine, acetylanserine, etc.) present in cardiac tissue (279, 280).
Human serum carnosinase or CN1 (EC 3.4.13.20) was first
described in relation to carnosinemia, a disorder associated
with nutrition-independent elevated circulating carnosine
levels and a mental defect in children (297) (see sect. VII).
Under nonpathological conditions, serum carnosinase activity is high in the adult human, leading to almost undetectable levels of circulating carnosine in the postabsorptive
state (143). The activity of serum carnosinase (CN1) varies
greatly between individuals. CN activity is very low shortly
after birth and gradually increases to adult level during
adolescence, and attains higher levels in females than males
(126, 301).
2. Cytosolic nonspecific dipeptidase
C. Carnosinases
Carnosine and related compounds are not degraded by regular (di)peptidases, but their metabolism is characterized by
its own hydrolytic enzymes, named carnosinases. Carnosinase was first described and partially purified by Hanson
and Smith in 1949 from swine kidney (158). Two forms
have since been molecularly identified as CN1, or serum
carnosinase, and CN2, or tissue carnosinase or cytosolic
nonspecific dipeptidase (229, 360). They are members of a
family of M20/M28 metalloproteases. CN1 and CN2 show
53% sequence identity in humans, and both enzymes exist
as homodimers. A recent crystallographic study revealed
that each subunit of the carnosinases consist of two domains, an A domain with catalytic and metal binding activity and a B domain for dimerization (376). The genes of
these carnosinases are CNDP1 and CNDP2, and they are
localized side by side on human chromosome 18 in a headto-tail position.
According to Teufel et al. (360), carnosine is only a substrate of cytosolic nonspecific dipeptidase or CN2 (EC
3.4.13.18) under nonphysiological conditions (pH optimum of 9.5). Therefore, the authors propose that CN2 is
renamed as cytosolic nonspecific dipeptidase. However, a
more recent report suggests that, even though the catalytic
rate of CN2 is markedly lower than that of CN1, carnosine
can still be hydrolyzed by CN2 under physiological conditions (287). It remains to be determined whether CN2 is
involved in the degradation of carnosine in tissues with
abundant carnosine content and in the intestinal wall.
3. Substrate specificity and functional differences of
CN1 and CN2
summarizes the main functional differences between CN1 and CN2. CN2 has a broader substrate specificity than CN1, but it does not hydrolyze homocarnosine.
In contrast to CN1, CN2 activity is inhibitable by bestatin,
TABLE 1
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1817
BOLDYREV, ALDINI, AND DERAVE
Table 1. Comparative table of characteristics of dipeptidases with carnosinase activity
Serum Carnosinase
Other names
Expression in human tissues
Carnosinase-1, ␤-Ala-His dipeptidase
Brain, liver, serum, kidney glomerulus
Subcellular localization
Metal ion cofactor
Inhibitable by bestatin
pH optimum for carnosine hydrolysis
Substrate specificity
Secreted
Zn2⫹, Cd2⫹
No (only in presence of Cd2⫹)
8.5
Carnosine ⬎ anserine ⬎ ophidine ⬎ homocarnosine
Cytosolic Nonspecific Dipeptidase
Carnosinase-2
Ubiquitously expressed; not in serum or
cerebrospinal fluid
Cytosolic
Mn2⫹
Yes
9.5
Anserine/ophidine ⬎ Xaa-His ⬎
carnosine; not homocarnosine
Based on data from Peters et al. (301), Pegova et al. (293), and Lenney et al. (230).
which is a substrate mimetic of carnosine. Another marked
difference with other family members of M20/M28 metallopeptidases, which are all activated by Zn2⫹, is that CN2
requires Mn2⫹ for it catalytic activity.
4. Other dipeptidases with carnosinase activity
Anserinase or Xaa-methyl-His dipeptidase (EC 3.4.13.5) is
a dipeptidase found in fish, with a broad substrate specificity, including carnosine, anserine, and homocarnosine. It
was named anserinase because it was first described in codling muscle (203), which uniquely contains anserine as
HCD. Yamada et al. (394) recently identified anserinase
molecularly and proposed a phylogenetic tree with three
separate families of carnosinase-like enzymes, being serum
carnosinase like (CN1), cytosolic nonspecific dipeptidaselike (CN2), and anserinase-like. Other carnosinases have
been described within the EC 3.4.13 family of dipeptidases,
but their enzyme classification (such as EC 3.4.13.3) is now
deleted because its activity is covered by CN1 and CN2.
A handful of carnosinases/␤-peptidyl aminopeptidases have
now been described in bacteria, and this new peptidase
family is rapidly expanding (145). One of the first described
bacterial enzymes that can hydrolyze carnosine is the dinuclear zinc aminopeptidase (PepV), characterized in Lactobacillus delbrueckii (385). Similarly, L-aminopeptidase
D -Ala-esterase/amidase (DmpA) from Ochrobactrum
anthropi, ␤-alanyl-Xaa dipeptidase (Ps BapA) from Pseudomonas sp. and BapF from Pseudomonas aeruginosa
show high specificities towards carnosine (138, 145). Aminoacyl-histidine dipeptidase (also named Xaa-His dipeptidase, carnosinase, or PepD) has recently been cloned from
the bacterium Porphyromonas gingivalis (18). The recombinant protein displays carnosinase activity, which is dependent on transition metals, such as Cu2⫹, Mn2⫹, and
Ni2⫹. P. gingivalis is associated with periodontal disease
and rheumatoid arthritis, for which carnosine has been suggested to have therapeutic potential (110). It remains to be
determined whether the carnosinase activity of these bacte-
1818
ria is of pathogenic relevance and how this interacts with
the carnosine content in human tissues.
D. Carnosine Transporters
Carnosine can be transported across the cellular membrane
through a number of transporters from the proton-coupled
oligopeptide transporter family (POT-family or SLC15).
The mammalian members of this family are PEPT1 and
PEPT2 (oligopeptide transporter 1 and 2) and PHT1 and
PHT2 (peptide/histidine transporter 1 and 2). All of these
POTs have a broad specificity and are able to transport
carnosine and its methylated analogs (395). In addition,
they can transport a high number of other peptides. PEPT1
and PEPT2 are believed to have the capability to transport
400 different dipeptides and 8,000 different tripeptides, but
not longer peptides or single amino acids. Whether the specificity of PHT1 and PHT2 is equally broad is less clear, but
the main difference with the PEPTs is that the PHTs can
transport L-histidine, in addition to di/tripeptides. The
physiology and pharmacology of the SLC15 transporter
family has been reviewed by Daniel et al. (93, 94). The
following section will mainly review the literature on the
functional transport of carnosine by POTs in human tissues.
1. PEPT1 and PEPT2
PEPT1 is characterized as a high-capacity, low-affinity transporter and plays a major nutritive role in the intestinal absorption of peptides (FIGURE 14). Carnosine can be absorbed in the
small intestine, and at least part of it enters the blood intactly
(not hydrolyzed) upon oral ingestion (49, 143). Carnosine
uptake across the brush-border membrane is accomplished by
PEPT1. When carnosine enters the enterocytes, it is either hydrolyzed, presumably by carnosinase (CN2), or further transported across the basolateral membrane. PEPT1 is a peptide/
proton cotransporter, and its activity is therefore increased
with a lower luminal/apical pH.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
Carnosine
Carnosine
Intestinal lumen
Brush border
membrane
PEPT1
Enterocyte
H+
Carnosine
CN2
Enterocyte
Beta-alanine
Histidine
Basolateral
membrane
Peptide
transporter
Carnosine
Amino acid
transporters
CN1
Beta-alanine
Blood
Histidine
FIGURE 14. Possible pathways of intestinal absorption of HCD. In humans, most of the carnosine is
hydrolyzed upon oral ingestion, either inside the enterocyte (CN2) or upon appearance in the circulation (CN1).
The transcellular transport is quantitatively the most important, and the involvement of the paracellular
transport is uncertain.
PEPT2 is characterized as a low-capacity, high-affinity
transporter and contributes to reabsorption of filtered peptides in the renal tubule, where it is mainly localized in the
apical membrane of epithelial cells. Insight into the role of
PEPT2 in carnosine metabolism has been advanced by the
development of PEPT2 knockout mice (320). With respect
to kidney function, PEPT2 ablation markedly impedes the
tubular reabsorption of carnosine, as the renal carnosine
clearance is close to the glomerular filtration rate in the null
mice, whereas it is 20-fold lower in the wild-type mice
(206). Even though the carnosine uptake is high in the apical side of the epithelial cells, its further transport to the
blood through the basolateral membrane is very limited
(198). This means that carnosine, once arrived in the in the
epithelial cytosol partly accumulates but is mainly hydrolyzed by carnosinase, after which the constituent amino
acids are transported to the blood over the basolateral
membrane by amino acid transporters. Thus, even though
the dipeptide transporter (PEPT2) plays a determining role
in renal carnosine reabsorption, the transepithelial flux of
carnosine is limited and the integrity of the dipeptide is
probably lost during the renal passage.
In addition to its designation as, respectively, the intestinal
and renal isoform, PEPT1 and PEPT2 are expressed in several other (human) tissues. In the choroid plexus, PEPT2 is
responsible for over 90% of the carnosine uptake from the
CSF over the apical membrane into the epithelial cells (361,
362). This suggests that PEPT2 plays a critical role in the
homeostasis of carnosine (and other peptides) in the CSF.
Also other glia, such as astrocytes, seem to primarily depend
on PEPT2 for their uptake of carnosine (393). Equally,
other cell types such as cardiomyocytes (233) and epithelial
tissues (human nasal epithelium) have been shown to express PEPT2, along with the other POTs, and to possess the
ability to transport carnosine (6). The latter finding may
have importance as a potential route to administer carnosine or carnosine-related drugs.
2. PHT1 and PHT2
Whereas good biochemical and physiological information
is available for the PEPTs, relatively little is known about
the more recently discovered PHTs. The first PHT (PHT1)
was cloned from rat brain by Yamashita et al. (395). When
expressed in Xenopus laevis oocytes, carnosine transport
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1819
BOLDYREV, ALDINI, AND DERAVE
activity of PHT1 was confirmed. Bhardwaj et al. (58) similarly cloned human PHT1 (hPHT1) and transfected it in
COS-7 cells, in which it elicited pH-dependent carnosine
transport. The second peptide/L-histidine transporter (PHT2)
was identified by Botka et al. (72) and Sakata et al. (321).
Recombinant rPHT2 protein reconstituted into liposomes displays transport activity towards carnosine (321).
tractions. This anti-fatigue effect of carnosine was subsequently adopted as the “Severin’s phenomenon,” which
also holds true for anserine (64). Below we summarize the
currently proposed physiological roles of carnosine in skeletal muscle, depicted on FIGURE 15.
Northern analysis indicated that hPHT1 is primarily expressed in skeletal muscle and spleen, whereas hPHT2 is found
in spleen, placenta, lung, leukocytes, and heart (72). As the
majority of carnosine in the human body is found in the skeletal muscle compartment, it is expected that carnosine transporters are functionally expressed in skeletal muscle cells.
Everaert et al. (125) recently demonstrated that mRNA transcripts of the PHTs but not the PEPTs were found in mouse
and human skeletal muscle samples. It is currently unclear
what their physiological function is in muscle and whether
inward or outward transport is the main direction.
The role of carnosine as a pH buffer in skeletal muscle was
first proposed in 1938 by two independent groups (47,
107). Bate Smith (47) proposed that carnosine would account for up to 25% of the buffering capacity in rigor
muscle and as much as 40% in vivo. During high-intensity
muscle contractions, the anaerobic glycolysis leads to the
production of lactic acid which immediately dissociates into
protons (H⫹) and lactate ions at physiological pH values.
The resulting acidosis can reach pH values of 6.5 and lower
(191) and has often been associated with muscle contractile
fatigue. Even though the role of acidosis in muscular fatigue
remains a matter of debate (16, 292), it can be expected that
the reason for the abundance of carnosine in muscle is at
least partly related to the proton-sequestering property of
the carnosine molecule (see sect. III).
V. PHYSIOLOGICAL ROLE OF CARNOSINE
In this section, we focus on the putative roles of carnosine
and related compounds in different organs and organ systems. We limit our discussion to tissues in which carnosine
content is high or at least evidence for its involvement in
normal physiological function is available, and this seems to
mainly be the excitable tissues, skeletal and cardiac muscle,
and neuronal tissue. It is evident that carnosine does not
have the same function in different tissues, and even may
fulfill several different functions within one tissue. There is
little clarity about the role of endogenous carnosine in tissues, other than the nervous system and skeletal and cardiac
muscle. This probably relates to the fact that there is still
controversy as to whether carnosine is endogenously present in organs such as the liver and kidney (see sect. II).
A. Carnosine in Skeletal Muscle Function
From the very beginning, the research on the physiological
role of carnosine was directed towards skeletal muscle, as
the molecule was initially discovered in meat extract. Yet,
over a century of research has not yet led to the establishment of the definitive role of this compound in the tissue in
which it is so abundant. Several hypotheses were proposed,
and some were subsequently refuted, such as the idea of
carnosine functioning as a high-energy phosphate system,
similar to the creatine/phosphocreatine system (148).
Severin, one of Gulewitch’ pupils (334), suggested that carnosine plays a role in the contractile function of skeletal
muscle. In their experiments they demonstrated in nervemuscle preparations of frogs that the addition of 10 mM
carnosine to the surrounding medium can offset the fatigue
that occurs during rhythmic nerve-stimulated muscle con-
1820
1. Intramyocellular homeostasis during contractions
In addition to HCD, several compounds, such as proteins,
bicarbonate, and inorganic phosphate, contribute to the
cellular buffering system, and their relative contribution
markedly depends on the muscle types and animal species
(281, 381)(see sect. II). For instance, the HCD may account
for more than half of the total muscle buffering capacity
(␤%) in marine animals, such as whale, who are involved in
prolonged hypoxic dives (3). HCD is typically high in white
(fast-twitch) muscle of animals in which anaerobic exercise
types such as prolonged breath-hold diving and sprint
swimming, running, flying, and hopping are required to
catch prey or escape predators (3).
In humans, muscle contains a rather small amount of HCD
(see sect. II), which would mean that they contribute relatively little to ␤tot. The relative buffering capacity ␤carnosine/
␤tot% in the human vastus lateralis muscle was determined
to be 4.5 and 9.4% in fibers I and II, respectively (243).
When muscle carnosine content is increased by nutritional
intervention, the degree of acidosis in the blood during
high-intensity exercise will be attenuated in humans (38).
Possibly more important than it contribution to ␤tot, carnosine is a mobile buffer, freely dissolved in the myoplasm,
as opposed to proteins, which are fixed buffers (204, 340).
This means that its contribution to pH homeostasis and
prevention of local pH gradients is probably greater than
would be expected from its calculated proportion of ␤tot.
It seems likely that the physiological role of carnosine in contracting muscle relates to homeostasis, but this may include
more than only protection against acidosis. One such alternative perturbation may be the protection against exerciseinduced production of ROS in muscle. The evidence for car-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
Sarcoplasmic reticulum
[Carnosine + Cu]2+
Ca2+
Cu2+
2
4
?
CARNOSINE
Sarcomere
Carnosine
5
ATP
H+
1
3
Histidine
Histamine
[Carnosine + H]+
Glycolysis
Lactate ion
ROS
Antioxidant
Mitochondrion
Skeletal muscle cell
Vascular smooth
muscle cell
Blood
vessel
FIGURE 15. Potential roles of carnosine in skeletal muscles cells: 1) proton buffering capacity; 2) regulator
of calcium release and calcium sensitivity; 3) protection against reactive oxygen species (ROS); 4) chelation of
transition metal ions; and 5) extracellular provider of histidine/histamine.
nosine’s antioxidative function in muscle in vivo is very limited
at present. A study on 90-min downhill running in rats showed
that a supplementation-induced increase in muscle HCD levels
could diminish the production of thiobarbituric acid reactive
substances (TBARS, lipid peroxidation products) (97). This
study is in line with earlier Russian studies showing a protecting role of carnosine on lipid peroxidation in working muscles
(67). Another putative role of carnosine in skeletal muscle
tissue is as a physiological chelator of Cu2⫹. Schröder et al.
(329) have demonstrated the presence of Cu2⫹-carnosine chelates in human calf muscle by proton MR spectroscopy in
vivo. However, there are currently no data available to confirm or refute the hypothesis put forward by Brown in 1981 on
the regulation of anaerobic and aerobic metabolism through
Cu2⫹-carnosine complexation (74).
2. Regulation of excitation-contraction coupling
The excitation-contraction coupling in skeletal muscle cells
is a complexly regulated chain of events in which calcium is
released from the sarcoplasmic reticulum (SR) through ryanodine receptors (RyR), the released calcium will bind to
troponin C to allow cross-bridge formation and force production and ultimately the reuptake of calcium in the SR
will terminate contraction (150). Carnosine has been suggested to be involved in the regulation of several of these
steps (FIGURE 15).
Batrukova and Rubtsov (48) demonstrated in SR vesicles
from rabbit skeletal muscle that carnosine can induce Ca2⫹
release, whereas its constituent amino acids L-histidine and
␤-alanine, added alone or in combination, were not effective. Similar evidence for carnosine’s potentiating effect on
SR calcium release was presented in a study on chemically
skinned fibers of human vastus lateralis muscle (404). However, these conclusions were contested by Dutka and Lamb
(122), arguing that all previous studies were performed
with abnormally low Mg2⫹ levels. They instead demonstrated that carnosine does not play a role in stimulating the
RyRs in the presence of physiological levels of Mg2⫹ (122).
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1821
BOLDYREV, ALDINI, AND DERAVE
More recently, however, Dutka et al. (123) demonstrated in
mechanically skinned human muscle fibers that carnosine in
a physiological concentration can sensitize the RyRs to
Ca2⫹-induced Ca2⫹ release in type I, but not type II, fibers.
A step that is probably even more important for carnosine’s
effect on excitation-contraction coupling is the calcium sensitivity of the contractile apparatus. Dutka and Lamb (122)
demonstrated that physiological carnosine concentrations
(4 –16 mM) improve Ca2⫹ sensitivity of rat extensor digitorum longus (EDL) fibers in a concentration-dependent
manner, hereby confirming previous work from Lamont
and Miller (227) on frog sartorius muscles. More recently,
these observations were also demonstrated in skinned human muscle fibers (123). In the fast-twitch fibers, the improved calcium sensitivity probably entirely explains the
force-potentiating effect of carnosine, as this fiber type did
not show improvement in calcium release (123). Not only in
skinned fiber preparations, but also in whole incubated
skeletal muscles of mice, force potentiation in response to
nutritionally elevated muscle carnosine levels was observed,
based on a leftward shift of the force-frequency relationship
(127).
3. Muscle as a depot and donor of carnosine
It is tempting to speculate on the possible role of skeletal
muscle as a production, storage, and release organ for carnosine. Indeed, muscle contains the vast majority of the
carnosine present in the organism, and it is one of the few
tissues where the expression of the enzyme responsible for
carnosine synthesis is very high. Early in the scientific exploration of carnosine’s function, Russian scientists came to
the conclusion that tissues that are capable of synthesizing
carnosine should not be able to provide for its degradation
(64). Indeed, tissues and cell types either have a high capacity to synthesize carnosine (through CS) or to degrade carnosine (through CN), but never combine both capacities.
This suggests that tissues, such as skeletal muscle, that synthesize carnosine can also release it to other places where it
can be degraded/utilized, e.g., for delivery of L-histidine or
␤-alanine. Thus muscle carnosine could play an autocrine,
paracrine, or even an endocrine role.
There is some evidence to believe that especially during
exercise, thus when skeletal muscles contract, they release
carnosine into the interstitium and the circulation. This has
been shown in rats by Nagai et al. (262), who observed a
doubled carnosine concentration in plasma of rats who had
access to a running wheel compared with sedentary rats.
Similar, though smaller, changes were observed in running
horses (119). Also in human skeletal muscle, carnosine content markedly increases in the interstitial fluid during knee
extension exercise at 20W, determined by microdialysis
(275). However, it is not known whether this carnosine
release is substantial enough to elevate human plasma carnosine levels during and following exercise. The fact that
1822
not only carnosine, but also the L-histidine, concentration
increases in skeletal muscle interstitium in humans (evaluated by microdialysis; Ref. 156) would indicate that a portion of the released carnosine is promptly hydrolyzed by
serum CN1. There is controversy as to whether carnosine
release by muscle cells is a result of either sarcolemmal
rupture and microtrauma, or of a functional transportermediated process. In belief of the former, some researchers
have interpreted the release of carnosine by exercising muscles as a measure of exertional rhabdomyolysis and muscle
damage (119, 156, 275). Others, however, believe that
myocytes actively release/secrete carnosine through dipeptide transporter, which are expressed in muscle (see sect. IV)
as part of a physiological regulatory process. The physiological function of such contraction-induced release mechanism is still elusive but could be related to the carnosinehistidine-histamine pathway (151) (FIGURE 15).
The above-described functions are most likely the primary
roles of carnosine in skeletal muscle. A number of other
potential roles, e.g., activation of myosin ATPase, effects on
glycolysis, mitochondrial respiration, and the neuromuscular junction, have been sporadically suggested, but remain
poorly understood. A complete description of these studies
is provided in the monograph of Boldyrev (61).
B. Carnosine in Brain Function
Homocarnosine is a more prevalent dipeptide than carnosine in the mammalian brain. This probably relates to the
fact that next to L-histidine the other constituent amino acid
of homocarnosine is GABA, which is the most important
and widespread inhibitory neurotransmitter in the mammalian brain. However, also ␤-alanine, GABA’s equivalent in
carnosine, is found in mammalian brain in concentrations
ranging from 40 to 100 ␮M in various rat brain structures
(103), and has been proposed to possess the properties of a
neurotransmitter (364). The highest carnosine concentrations in the brain, found in the olfactory system, are comparable to those usually found in skeletal muscle.
In humans, brain homocarnosine concentration is rather
high (0.5–1 mM) and detectable by proton magnetic resonance spectroscopy (MRS) in vivo (303). Carnosine is probably not detectable in the human brain by MRS, and its
presence is presumably mainly limited to the olfactory system.
1. Distribution
The first evidence for the presence of carnosine in the brain
was provided by Margolis, who demonstrated that the olfactory tract of mice contains significantly higher carnosine
concentrations (1–2 mM) than other brain structures,
where its concentration is usually ⬃0.1 mM or lower (246).
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
The olfactory system of many animal species, including humans, is highly enriched with carnosine as well as with its
principal enzymes. Carnosine synthase activity is 50- to
100-fold higher in the olfactory epithelium than in brain
structures and 10-fold higher than in skeletal muscle (160,
216).
Carnosine is located in the olfactory receptor neurons, more
specifically in the perikaria and cell processes, including the
axons and synaptic terminals in the olfactory bulb (59).
When radiolabeled ␤-alanine and L-histidine are administered to the nasal mucosa, carnosine is rapidly formed and
directed by axoplasmic transport to the olfactory bulb
(247). Carnosine synthase activity in primary olfactory neurons decreases upon denervation and recovers upon regeneration (159, 160). These data indicate that the sensory
neurons are primarily responsible for the presence of carnosine in olfactory epithelium and the olfactory bulb (98).
Carnosinase, on the other hand, is mainly present in other
cells, such as pillar cells and Bowman gland cells, of the
nasal mucosa, which confirms the dogma that the enzymes
of carnosine synthesis and degradation are located in different cell types and compartments (81, 159).
The olfactory neurons seem to be the only neurons that
express carnosine. Other cells of the neuronal lineage, the
neuroblasts of the subependymal layer (SEL) of the forebrain, have been reported to display carnosine-like immunoreactivity. However, the immunohistochemical results
when using anti-carnosine serum may not allow to distinguish between the presence of either carnosine or homocarnosine (70, 98).
Carnosine-like immunoreactivity is widespread in glial cells
throughout the brain. Carnosine synthesis activity was first
demonstrated in cultured rat C-6 glioma cell lines (52). In
mouse glial primary cultures, Bauer et al. (51) demonstrated
that ␤-alanine is effectively incorporated into carnosine,
whereas GABA was rapidly degraded. Similar carnosine
synthesis activity could not be observed in neuronal cell
cultures (50), but seems to be specifically confined to oligodendrocytes (186). Not only do glial cells display carnosine
synthesis activity, they also possess the ability to transport
carnosine, as observed by the release of significant amounts
of synthesized carnosine from cultured glia into the medium. This carnosine transport activity seems to be primarily observed in astrocytes (186) and is probably operated by
PEPT2 (112). Bauer (50) recently proposed that PEPT2,
expressed in choroid plexus epithelial cells, is also involved
in the removal of carnosine and other peptides from the
cerebrospinal fluid (CSF).
2. Putative functions of carnosine in the brain
It is fair to say that we currently have a poor understanding
of the role of carnosine in the brain. However, a number of
potential roles have been proposed, and the evidence for
these is summarized below.
On the basis of its high concentrations found in the olfactory neurons, carnosine was hypothesized to be involved in
sensory neurotransmission, either as a neurotransmitter or
a neuromodulator. As for most other sensory pathways,
glutamate is the main excitatory neurotransmitter involved
in the synapse between olfactory neurons and target cells
(mitral and periglomerular cells) in the olfactory bulb. Interestingly, immunohistochemical studies have identified
colocalization of carnosine and glutamate in the synaptic
terminals of mouse olfactory neurons (324), as well as bipolar cells of the frog retina (288). This supports the hypothesis for a putative role of the dipeptide in neuromodulation of glutamatergic sensory neurons. However, there
are a number of findings arguing against a direct role of
carnosine in glutamatergic neurotransmission, which are
summarized by Bonfanti et al. (70) and De Marchis et al.
(98). Possibly, interaction between carnosine and glutamate
involves glia. Bakardjiev (40) showed that oligodendrocytes
exhibit glutamate-receptor mediated release of carnosine.
More recently, Baslow (46) proposed that carnosine, homocarnosine, and other brain dipeptides such as N-acetylaspartylglutamate (NAAG) serve as neurotransmitters in neuron-to-glia communication.
In addition to its involvement in modulation of neurotransmission, the presence of carnosine in certain brain structures has been hypothesized to be homeostatic and protective, mainly through antioxidant, metal chelating, and antiglycative properties. It has been argued that the presence
of other endogenous antioxidant systems (glutathione, vitamins C and E) availability is very low in the nervous
system, which is compensated for by (homo-)carnosine
(70). It seems clear that this aspect is of therapeutic value
(see further in sect. VI), but it remains to be established
whether this is a true physiological role in normal function
as well. Carnosine has also been suggested to be indirectly
involved in neurotransmission through its capacity to chelate transition metals such as copper and zinc, which in
itself have been identified as modulators of neurotransmission (60, 373).
C. Carnosine in Cardiovascular Function
One of the first described physiological effects of carnosine
was the transient decrease in systemic blood pressure upon
intravenous injection of carnosine in dogs (313). The hypotensive and antihypertensive effect has since been frequently
confirmed in various other mammalian species (115, 272).
The lowering effect of carnosine on blood pressure is probably realized by a systemic arterial vasodilation. The direct
vasorelaxing effect of carnosine has been demonstrated on
isolated rat aortic rings (318). Interestingly, carnosine can
also potently provoke sustained contractures in rabbit sa-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1823
BOLDYREV, ALDINI, AND DERAVE
phenous vein rings (278). Given the potency of carnosine in
the regulation of the vascular tone, it is thought that not
only exogenously administered carnosine, but also endogenously available carnosine takes part in normal hemodynamic regulation. The formation in vivo of a zinc/carnosine
complex and the involvement of histamine H1 receptors
appear to be involved in vascular smooth muscle contraction/relaxation (255, 277). However, alternative vasoactive
mechanisms of carnosine may be equally involved, such as
the carnosine-histidine-histamine pathway (151), a NO/
cGMP mechanism (318), and the modulation of the autonomic nervous system (265).
The metabolic instability of carnosine in human serum due
to the presence of carnosinase has prompted a great interest
in the design and chemical synthesis of novel carnosine
derivatives, as well as in research towards the therapeutic
potential of the natural carnosine derivatives, such as anserine (224). These compounds are more resistant to carnosinase, while maintaining or even improving some beneficial effects of the parent compound, such as the antioxidant
and metal ion chelating ability or the reactivity with RCS.
Various reviews have recently been published regarding the
carnosine derivatives stable to carnosinases (56, 152, 383).
Carnosine can markedly potentiate the cardiac contractility, as demonstrated in experiments with isolated rat hearts
(319). In parallel with the mechanisms underlying improvement of contractility in skeletal muscle, it is believed that
also for the cardiac muscle the main mechanisms are improved calcium handling (excitation-contraction coupling)
and increased pH buffering capacity. In chemically skinned
cardiac myocytes, carnosine was able to both release calcium from the SR as well as improve the tension of the
contractile proteins in response to calcium (403). Besides
modulating intracellular calcium, it is likely that carnosine
and other HCD play a role in the pH regulation and contribute to the mobile buffering system in the cardiac cell
(380). In this respect, an interesting novel hypothesis was
recently proposed by Swietah et al. (352), in which they
describe carnosine as a diffusible Ca2⫹/H⫹ exchange
“pump,” creating functional Ca2⫹ gradients in response to
local pH changes in cardiomyocytes. Despite the abovementioned studies, it is surprising to note that still so little
scientific knowledge has been collected on the role of HCD
in the cardiovascular system.
B. Diabetes
VI. CARNOSINE AND DISEASE
A. Extrapolation of Animal Models of
Disease to Humans
With the exception of a limited number of small-scale clinical trials that have not been independently confirmed, such
as on patients with Parkinson disease (62), schizophrenia
(83), autistic spectrum disorder (84), and ocular diseases
(30), the vast majority of research on the therapeutic potential of carnosine is conducted in animal models of human
diseases, mainly in rodents. One should be aware, however,
that there currently still is a potential problem with the
transposability of these findings to humans. The limitation
relates to the low levels of serum carnosinase activity in
rodents compared with the high levels of carnosinase activity in humans. It is possible that a least part of the beneficial
and therapeutic effects of carnosine administration is related to elevated plasma carnosine levels in animals, which
are more difficult to obtain in humans (128, 400).
1824
A growing body of evidence indicates a protective role of
carnosine in diabetes due to its ability to affect glycemic
control and to prevent/ameliorate diabetic complications
such as nephropathy and ocular damage. An overall protective effect of carnosine in diabetic animal models was first
published by Lee et al. (228) using Balb/cA mice. An oral
supplementation with carnosine for 4 wk dose-dependently
reduced plasma glucose and fibronectin levels, increased the
insulin levels, and reduced the oxidative damage. A similar
protective effect was induced by L-histidine, which was
found to increase the carnosine content in plasma and organs to a similar extent to that reached by the carnosine
treatment. The beneficial effects of carnosine on diabetic
deterioration were confirmed some years later by Soliman et
al. (341) in a streptozotocin diabetic-induced model. Carnosine treatment (100 and 200 mg/kg) dose-dependently
reduced hyperglycemia, normalized dyslipidemia, and reduced liver damage. In addition to these beneficial effects, it
should be noted that the papers by Lee and Solima both
demonstrate a hypoglycemic effect of carnosine in diabetic
rats. The ability of carnosine to control blood glucose levels
was thoroughly investigated by Nagai et al. (262) who proposed that such effect occurs through the ability of carnosine, which is a precursor of histamine, to regulate the autonomic nervous system via the H3 receptor. A confirmation
of such a receptor-mediated mechanism comes from the
similar hypoglycemic effects evoked by L-histidine and histamine. In more detail, carnosine lowers neuronal activities
of sympathetic nerves and facilitates those of parasympathetic nerves causing both an increase in the insulin secretion and suppression in the glucagon secretion from the
pancreas, resulting in a hypoglycemic effect. A similar hypoglycemic effect was more recently reported for anserine,
which was also found to reduce blood glucose levels in
humans during oral glucose tolerance testing (224). A direct
correlation of carnosine serum levels with insulin levels was
also detected by Sauerhoefer et al. (326) in db/db and hCN1
transgenic db/db mice, whereas peripheral insulin sensitivity was unaltered. The mechanism by which serum carnosine levels affect insulin secretion was addressed to the ability
of carnosine to preserve or increase ␤-cells within the pancreas.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
Carnosine was found to be effective in inhibiting the development of diabetic complications including ocular diseases
and nephropathy (304, 317). Carnosine treatment was
found to reduce diabetes-induced nephropathy and in particular to restrain glomerular apoptosis, to prevent podocytes loss and to reduce the expression of Bcl-2-associated X
protein (bax) and cytochrome c (317), well-known proapoptotic proteins. Carnosine was also found to ameliorate
diabetic neuropathies, especially abnormal sensory perception (207).
Taken together, the protective effects of carnosine in diabetic animal models could be related to a hypoglycemic
effect, which is possibly a consequence of the ability of
carnosine to regulate the autonomic nervous system or by
preserving or increasing ␤-cells within the pancreas. Carnosine was also found to be effective in reducing diabetesrelated diseases. The underlying mechanism has been partially addressed to the ability of carnosine to reverse protein
glycation, to inhibit AGEs and ALEs formation (173). The
involvement of the carnosine/carnosinase system in diabetes and diabetes-related diseases has been also supported by
genetic experiments, which will be discussed in detail in
section VII.
C. Ischemia/Reperfusion Damage
Since the 1990s, several in vivo studies have reported the
ability of carnosine to reduce ischemia/reperfusion (I/R)
damage in different animal models and organs including
brain, liver, kidney, and testis. At the beginning, the rationale to use carnosine as a protecting agent in I/R animal
models was considered to be due to its antioxidant activity,
since oxidative stress induced by I/R is known to be involved in cell damage. Such a rational approach was then
sustained by several studies reporting that the protection
afforded by carnosine against I/R tissue damage paralleled a
reduction of tissue oxidative damage and a sparing effect on
enzymatic and nonenzymatic antioxidants. For example,
Dobrotvorskaya (109) reported that carnosine at a dose of
100 mg/kg in rats prevented the disturbances of the cognitive functions induced by brain ischemia under conditions
of hypercysteinemia as well as the development of oxidative
stress and the depletion of the antioxidant system. The antioxidant activity of carnosine was also proposed by Zhang
et al. (406) to explain the protective effect of the dipeptide
pretreatment (250 mg/kg) against I/R damage in the neonatal rat brain. In a model of permanent focal cerebral ischemia in mice, carnosine treatment (100, 500, and 1,000
mg/kg before ischemia) significantly decreased infarct size
and neuronal damage, and this was accompanied by a decrease of ROS levels in ischemic brains and a sparing effect
on the depletion of glutathione (311). A similar study in
rats, using intravenous carnosine administration (500 –
2,000 mg/kg) in a model of permanent or transient middle
cerebral artery occlusion, confirmed its therapeutical poten-
tial in stroke (33). More recently, carnosine was found to
act as an antioxidant in an experimental I/R injury in
testis (1).
In addition to the antioxidant effect, other mechanisms
have been proposed to explain the well-established antiischemic activity of carnosine, and among these, the activation of H3 receptor and the consequent inhibitory effect on
the neural activity of the renal sympathetic nerve is one
of the most studied. Fujii and co-workers (139, 140) reported that carnosine at doses of 1–10 ␮g/kg, concentrations far below those able to induce an antioxidant response, efficiently prevented I/R-induced acute renal failure. Such a mechanism was then further examined by
Kurata et al. (226) demonstrating that even at much lower
doses (intravenous injection 15 nmol/rat), carnosine efficiently prevented the I/R-induced renal injury and that a
similar protective effect was achieved by an intracerebroventricular injection of the peptide (1.5–5 pmol/rat), thus
suggesting its action on the central nervous system. Such
histamine-mediated mechanism was supported by the lack
of activity of N-acetylcarnosine, which is a carnosine derivative stable to carnosinases and by the suppression of the
protective effect of carnosine by thioperamide which is a H3
receptor antagonist (226).
On the other hand, Shen et al. (338) demonstrated that the
mechanism of carnosine action in a model of permanent
middle cerebral artery occlusion may not be mediated by
the histaminergic pathway. This pathway was also ruled
out by Ma et al. (241) to explain the protective effect of
carnosine in subcortical ischemic vascular dementia induced by carotid arteries occlusion. Carnosine (200, 500
mg/kg) but not L-histidine was found to ameliorate white
matter lesions and cognitive impairment in both wild-type
mice and histidine decarboxylase knockout mice (241).
More recently, Wang et al. (386) showed that carnosine
influences the mRNA expression of the signal transducer
and activator of transcription 3 (STAT3) which would explain the inhibition of neuronal apoptosis after acute
ischemic stroke.
D. Cancer
Carnosine has generated particular interest for its antiproliferative properties. As pointed out by Gaunitz and Hipkiss
in a perspective on carnosine and cancer (144), the first
report on the antineoplastic effect of carnosine was described in 1986 by Nagai and Suda (263), who reported
that carnosine treatment (50 mg·kg⫺1·day⫺1) inhibited tumor growth and mortality in mice. Ten years later, Holliday
and McFarland (187) reported that human fibroblasts grew
normally in the presence of carnosine while it selectively
inhibited the growth of neoplastic cell lines. A first hypothesis on the mechanism of action was then proposed, which
is based on the ability of carnosine to deplete glycolysis
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1825
BOLDYREV, ALDINI, AND DERAVE
intermediates (glyceraldehydes phosphate and dihydroxyacetone phosphate) by its carbonyl quenching ability and
therefore to reduce the generation of ATP. Such a mechanism well agrees with the metabolic differences between
these two cell types (Warburg effect) and in particular that
most tumor cells are predominantly glycolytic for ATP supply, whereas in differentiated fibroblasts, ATP synthesis is
predominantly aerobic and mitochondrial in origin. Cartwright et al. (79) showed that when yeast cells depend on
glycolysis for energy generation, L-carnosine slows growth
rates and increases cell death, but when they are reliant on
oxidative phosphorylation, they become resistant to L-carnosine’s inhibitory effects.
More recently, the antiproliferative activity of carnosine
was demonstrated in HCT16 colon cancer cells where carnosine was found to dose-dependently inhibit the ATP production (from 5 to 300 mM). In addition to targeting ATP,
the authors also demonstrated the ability of carnosine to
inhibit mitochondrial ROS generation, which is induced by
KRAS oncogene and which is involved in cell proliferation
(194).
The ability of carnosine to inhibit tumor growth in vivo was
demonstrated by Renner et al. (314) in a nude mouse model
in which NIH3T3 fibroblasts expressing the human epidermal growth factor receptor 2 were implanted in the dorsal
skin. More recently, the ability of carnosine to inhibit tumor
proliferation was confirmed in an animal model where human colon cancer (HCT116) cells were implanted in
BALB/c athymic mice (188).
E. Aging
Carnosine is described as an anti-aging compound, affecting a wide range of age-related processes and functions. In
1994, McFarland and Holliday first reported the ability of
carnosine to extend the maximum cell division capacity of
cultured human fibroblasts converting senescent cells to juvenile phenotypes (252). Some years later, the same authors
confirmed and extended such data, demonstrating that the
addition of carnosine in the fibroblasts medium switched
their phenotype from senescent to juvenile and that the
effect was reversed by removing the peptide (253). The
anti-aging effect of carnosine has also been described in
animal models. The group of Boldyrev reported that carnosine supplemented to a standard diet attenuates the development of senile features and increases life span by 20%
in senescence-accelerated mice (SAM) (68, 142, 402). Carnosine was also found to significantly increase the number
of spermatogonia and Sertoli cells in mice prone to accelerated aging (149) and to increase the lifespan of male Drosophila flies (401). Although some have ascribed the antiaging effect of carnosine to its antioxidant activity (26, 68),
others proposed that the geroprotective effect of carnosine
is mediated by its ability to prevent protein carbonylation
1826
and to modulate the degradation of aged proteins (175–
177). Further studies are definitely required to investigate
this important and popular potential application of carnosine, as well as the underlying mechanisms.
F. Neurological Disorders
The possible protective effect of carnosine in Alzheimer’s
disease (AD) was first proposed in the late 1990s when in
vitro studies demonstrated the ability of the dipeptide to
inhibit, on one hand the formation of ␤-amyloid polymerization and on the other hand the cell toxicity. Preston et al.
(306) reported that carnosine provided some protection
against the effect of the neurotoxin ␤-amyloid peptide
(A␤25–35) on rat brain vascular endothelial cells. The cell
protective effect of carnosine against amyloid-␤ (1– 42) mediated toxicity was then confirmed in other cell models.
Amyloid-␤ caused a dose-dependent loss of rat cerebellar
granule cell neurons viability which was significantly inhibited by carnosine. The protective effect of carnosine was
more recently demonstrated on amyloid-␤ (1– 42) impairment of differentiated PC12 cells.
Only a few in vivo studies on the effect of carnosine in
animal models of AD have thus far been reported. Corona
et al. (89) studied the effects of carnosine treatment in triple-transgenic AD mice, who develop an age-related neurodegenerative phenotype that is driven by intraneuronal deposition of amyloid-␤ and accumulation of h-tau. It was
found that carnosine had a strong effect in restoring mitochondrial functioning and in counteracting amyloid-␤ aggregation but had no effect in tau pathology and only a
trend toward the amelioration of cognitive deficits (89).
More recently, Herculano et al. (169) reported a prevention
of cognitive decline by carnosine supplementation in transgenic AD mice fed a high-fat diet. Some supportive evidence
in humans comes from a metabolomic study, consisting of
screening the changes in free amino acid and dipeptide concentrations in AD patients compared with controls. It was
found that carnosine levels were significantly lower in the
plasma of AD patients with respect to controls (134).
Regarding the mechanism of the therapeutic action of carnosine in AD, the carnosine-histidine-histamine pathway
was ruled out by using histamine receptor antagonists and
histidine decarboxylase inhibitors. Likewise, the antioxidant activity of carnosine is unlikely as amyloid-␤ induced
cell damage is not induced by generation of ROS. Other
hypotheses were proposed, considering the antiglycating
activity (63), the involvement in the regulation of glutamate
release and NMDA receptor trafficking (137), and the transient and weak interactions between carnosine and the
charged groups surrounding the central hydrophobic cluster of amyloid-␤ (1– 42) thus inhibiting the self-assembly
(25). In vitro and in vivo studies also indicate that a possible
mechanism of neuroprotection of carnosine is mediated by
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
its metal ion chelating activity. The rationale for addressing
Zn2⫹ dyshomeostasis in AD is substantial since Zn2⫹ may
induce aggregation and oligomerization of amyloid-␤ and
also cell toxicity. By using suitable fluorescent probes, Corona et al. (89) demonstrated the ability of carnosine to
chelate Zn2⫹ in cultured glia cells. The cytoprotective effect
was then supported by Horning et al. (190), demonstrating
that physiologically relevant concentrations of carnosine
provide a protective effect against toxicity induced by zinc
and copper in cultured neurons.
In relation to Parkinson’s disease, carnosine was found to
inhibit oligomerization of ␣-synuclein in model systems
(208, 214), and to suppress some changes in animal models
of Parkinson’s disease (374). In a small clinical trial, Boldyrev et al. (62) found that carnosine treatment (1.5 g/day) in
combination with L-DOPA therapy in Parkinson’s disease
patients significantly improved a number of neurological
symptoms, including decreased rigidity of the hands and
legs, and increased hand movement and leg agility. At a
biochemical level, a decrease in protein carbonyls in blood
plasma and an increase of SOD have been found (62).
Another small clinical trial on the effect of carnosine on
neurological disorder is a double-blinded placebo-controlled study on the effect of carnosine treatment (800 mg
daily for 8 wk) on the observable changes in 31 children
with autistic spectrum disorders. After 8 wk of carnosine
but not placebo treatment, children showed statistically significant improvements on the Gilliam Autism Rating Scale
and the Receptive One-Word Picture Vocabulary test (84).
Interestingly, a recent metabolomic study revealed decreased levels of urinary carnosine, ␤-alanine, and L-histidine in children with autistic spectrum disorder (257). Unfortunately, no further clinical trials on this aspect have
been reported, nor recorded in the database at www.
clinicaltrials.gov.
Recently, the effect of carnosine treatment (2 g daily for 3
mo) in adults with chronic schizophrenia was studied in a
randomized, double-blind, placebo-controlled trial (83).
Carnosine but not placebo was found to improve some
cognitive tests such as the strategic target detection test. The
authors concluded that carnosine merits further consideration as adjunctive treatment to improve executive dysfunction in persons with schizophrenia (83). A mild positive
effect of carnosine treatment (1.5 g daily for 12 wk) on
cognitive function has been also reported in a small randomized double-blind placebo controlled trial involving involving Persian Gulf war veterans affected by cognitive dysfunction termed Gulf War illness (GWI) (45).
G. Ocular Diseases
Carnosine has been reported to positively affect some ageand diabetes-related ocular diseases. Most of the data are
reported on cataract, and more specifically on the insoluble
aggregates (predominantly ␣-crystallin) which induce lens
opacity. In vitro studies reported the ability of carnosine to
inhibit ␣-crystallin aggregation induced by methylglyoxal
as well as to disaggregate glycated ␣-crystallin. The latter
effect referred to the ability of carnosine to increase peptide
chain mobility resulting in an unfolding of the glycated
protein (332). The ability of carnosine to modulate the aggregation-disaggregation dynamics was then confirmed and
extended by Attanasio et al. (24), demonstrating the prevention of fibril formation of ␣-crystallin alone or in a more
complex lens model. Moreover, they found a disassembling
effect of carnosine on ␣-crystallin amyloid fibrils. The ability of carnosine to prevent cataract formation and development was evaluated in different animal models, and some
controversial results were reported. Babizhayev (28) first
reported that carnosine significantly prevented cataracts induced in the rabbit eye by the administration of lipid peroxidation products. In contrast, carnosine was found ineffective in preventing cataracts in streptozotocin-induced diabetic rats (234) or in an in vivo model of cataracts induced
by sodium selenite (239). More recently, carnosine was
found to delay the progression of lens opacification in diabetic rats, but only during the earlier stages of the disease
(396). Clinical studies have been reported on the effect of
N-acetylcarnosine on cataracts. Topical eye application of
N-acetylcarnosine reportedly leads to a better accumulation of carnosine in the aqueous humor compared with a
direct application of carnosine (32). In volunteers with senile cataracts, N-acetylcarnosine-treated eyes demonstrated
visual improvement in general (30) and in particular an
improvement in visual acuity and glare sensitivity (29, 31).
H. Wound Healing
Since the beginning of the 1980s, the enhancing effect of
L-histidine and carnosine on wound healing was observed in
animal models (130). The mechanism is probably related to
the activity of both constitutive amino acids: L-histidine and
␤-alanine. On one hand, carnosine acts as L-histidine precursor, thus regulating histamine synthesis during trauma
(130). Faster wound healing through histamine is associated with the activity of basic fibroblast growth factor, leading to macrophage recruitment and angiogenesis (276). On
the other hand, the enhancement by carnosine of wound
healing may also be ascribed to the stimulation of collagen
biosynthesis by ␤-alanine (264). The efficacy of carnosine
on wound healing was demonstrated in several animal
models. Perel’man et al. (296) established that carnosine
was able to double the rate of wound reparative processes in
the injured lung of guinea pigs, by activating fibroblast proliferation, connective tissue generation, and intracellular regeneration. More recently, carnosine has been shown to
accelerate healing of bleomycin-induced and irradiationinduced pulmonary wounds (92, 155).
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1827
BOLDYREV, ALDINI, AND DERAVE
A large body of evidence exists on the healing effect of
Polaprezinc (a zinc-carnosine complex, see sect. III) in gastric ulcers, an effect which is partially mediated by the stimulating effect of the metal complex on the production of
insulin-like growth factor I (211, 222, 273, 333, 388).
More recently, the effect of carnosine on wound healing
was demonstrated in db/db mice, a model for type 2 diabetes. Carnosine treatment (100 mg/kg) significantly improved wound healing, and tissue analysis revealed higher
expression of cytokine genes and growth factors involved in
wound healing (17).
5-5 CNDP1 genotype
Low carnosinase activity and secretion
Partial protection against carnosine
hydrolysis in circulation
VII. CARNOSINASE AND DISEASE
In this section we discuss some diseases that have been
linked to the carnosinase enzyme. This implies that their
pathological features are in some way influenced by a dysregulation of carnosine or homocarnosine degradation.
Protection against AGEs, ROS, ....
A. CNDP1 Genotype and Disease Risk
Decreased risk of developing diabetic nephropathy
Several genome-wide linkage scans for diabetic nephropathy have mapped susceptibility loci to chromosome
18q22.3-23 (73, 379), a large region which also includes
the carnosinase genes CNDP1 and CNDP2. In 2005, Janssen et al. (197) managed to further fine map this region and
provided the first evidence for a genetic link between serum
carnosinase (CNDP1) and diabetic nephropathy. A trinucleotide repeat in exon 2 of CNDP1 determines the number
of leucine residues (5, 6 or 7) in the leader peptide of the
CN1 precursor. European diabetic patients homozygous
for the five leucine allele were less susceptible to diabetic
nephropathy and had lower serum carnosinase activity. A
later study on Cos-7 transfected cells (315) clarified that the
five leucine alleles impair the efficiency of secretion of serum
carnosinase, which may explain why serum carnosinase activity is linked to CNDP1 genotype. The disease-protecting
effect of lower serum carnosinase activity probably relates
to higher stability of carnosine in the circulation (128) or
alternatively to a local effect of enhanced carnosine availability in the kidney (221). The current working hypothesis
for the link between genetic predisposition and pathology is
shown in FIGURE 16.
The link between the (CTG)n polymorphism of the CNDP1
gene and diabetic nephropathy was confirmed by an independent study on European Americans with type 2 diabetes
(136), but could not be found in patients with diabetic
nephropathy due to type 1 diabetes (387) nor in AfricanAmericans (251). Mooyaart et al. (260) demonstrated that
the protection against development of diabetic nephropathy is only present in women, but not in men with the 5–5
leucine polymorphism. Recently, other single-nucleotide
polymorphisms (SNPs) in the CNDP1 and CNDP2 genes
have also been associated with diabetic nephropathy (7,
225, 251).
1828
FIGURE 16. Proposed pathogenetic link between the leucine-repeat variation in the CNDP1 gene and the predisposition to develop
diabetic nephropathy.
It now needs to be established how broad the influence of
polymorphisms in the carnosinase gene is in the susceptibility of diseases. Because the protective effects of carnosine
apply to many diseases (see sect. VI), it would be somewhat
surprising if the genetic association of CNDP1 will remain
limited to diabetic nephropathy alone. It has been suggested
that CNDP1 polymorphisms are also linked to end-stage
renal disease in nondiabetic patients (259). Only a single
study is currently available on the possible relationship between CNDP1 genotype and longevity and coronary heart
disease (409), and the results were negative.
B. Serum Carnosinase Deficiency and
Homocarnosinosis
There exist two types of inherited disorders that are characterized by the impaired ability to hydrolyze HCD: serum
carnosinase deficiency and homocarnosinosis. It is still uncertain whether or not the two metabolic disorders are in
fact the same disease. Nevertheless, the disorders share very
similar biochemical and clinical symptoms and are very
rare. The latter probably explains why the insight in the
disorders is still so limited.
Serum carnosinase deficiency has been described in 23 children, of which 16 cases have been proven (146). The disease
was initially called carnosinemia, because the high presence
of carnosine in the blood and enhanced excretion of carnosine in the urine (carnosinuria) were the prime biochemical
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
characteristics of the disorder (298). Upon ingestion of
meat, the patients’ blood and urine is enriched with carnosine and anserine, but contains little if any Npi-methyl-histidine, indicative of a malfunction of the dipeptide hydrolysis enzyme. Later studies have confirmed the absence or
low activity of serum carnosinase (131, 297). When a meatfree diet is ingested, carnosinemia and carnosinuria are suppressed but nonetheless still present, in contrast to healthy
individuals (392).
Reported symptoms of patients with serum carnosinase deficiency are mental retardation, hypotonia, myoclonic seizures, and spasticity, which usually manifest in the first year
of life (298, 377, 392). However, it appears that the severity
of symptoms is not related to the residual activity of serum
carnosinase (88), and serum carnosinase deficiency can occur in the absence of neurological symptoms. Dietary meat
restriction will partially reduce carnosinemia, but will not
relieve clinical symptoms.
Homocarnosinosis has been described in only one Norwegian family and is characterized by elevated concentrations
of homocarnosine in brain and CSF (147, 299). The mother
as well as the two sons and one daughter all show these
metabolic changes, whereas only the children have neurological symptoms: motor dysfunction, retinal pigmentation,
and mental deterioration. Similar to patients with serum
carnosinase deficiency, the homocarnosinosis patients have
carnosinuria and reduced capacity to hydrolyze dietary
HCD. Chronic dietary restriction of L-histidine, which is
one of the precursors of (homo-)carnosine, did markedly
reduce CSF homocarnosine concentration and carnosinuria, but did not relieve the symptoms (240).
Initially, it was proposed that homocarnosinosis patients
lack homocarnosinase activity in the brain in contrast to
healthy individuals (299). However, Lenney et al. (231)
indicated that homocarnosinase was not present in normal
human brain tissue either. The ability of brain extracts to
hydrolyze homocarnosine therefore appeared to be attributable to the carnosinase that is still present in the serum
trapped in a brain sample. As patients with homocarnosinosis have extremely low serum carnosinase activity, it appears that homocarnosinosis is in fact also attributable to
serum carnosinase deficiency (231). Despite this probably
common biochemical cause, there remain some dissimilarities between homocarnosinosis and serum carnosinase deficiency, e.g., with respect to the age of onset of symptoms
and the type of symptoms.
Our current knowledge fails to understand a possible link
between serum carnosinase on the one hand and the neurological symptoms on the other hand. It seems likely that the
accumulation itself of the carnosine and homocarnosine in
various body compartments is not posing the problem.
They are not to be considered as neurotoxins at high con-
centrations. Therefore, it is more likely that their functional
utilization, possibly as a source of GABA, ␤-alanine, or
L-histidine production, is causing the symptoms in affected
patients. A better insight in the pathogenesis in these patients would allow us to better understand the normal physiological role of the (homo-)carnosine/carnosinase system.
The scientific literature on the genetic basis of carnosinase
deficiency is still very scarce at present. The activity of serum carnosinase is usually suppressed in parents and siblings of patients and shows the pattern of an autosomal
recessive disorder. Willi et al. (389) reported the case of a
30-mo-old girl with serum carnosinase deficiency. The long
arm of her paternal chromosome 18 had a terminal deletion
at breakpoint q21.3. The father had normal but the mother
had reduced serum carnosinase activity. Therefore, the patient was likely hemizygous for the defect, having received
the deficiency allele from her mother and, as a result of a de
novo generated deletion, no allele from her father (389).
Another recent progress on this topic was made by
Zschocke et al. (409), who identified a null mutation in the
CNDP1 gene. This newly discovered frameshift mutation
(L17fsX20) would introduce a stop codon at position 36 of
the amino acid chain of CN1. Individuals homozygous for
L17fsX20 are predicted to have a complete absence of serum carnosinase activity, and therefore carnosinemia. The
null mutation was found to have a carrier frequency of
1.4% in a German population, and the prevalence for individuals with homozygosity for L17fsX20 is on this basis
estimated to be 1:20,000 (409). No such patients have yet
been identified.
In addition to the above-described inherited metabolic disorders, there are other diseases characterized by lowered
serum carnosinase activity. Young patients with progressive muscular dystrophy or myopathy and patients with
hypothyroidism have been shown to have very low serum
carnosinase activity (41, 43). Whether this enzymatic abnormality is causally related or just coincidental to the condition is not known. Because carnosinase is a serum enzyme
that is mainly produced and secreted by the liver, it is not
surprising that patients with chronic liver disorders also
display markedly reduced serum carnosinase activity (42),
attributable to low serum CN1 content (300).
C. The Human Serum Carnosinase Paradox
An unbalanced activity of serum carnosinase (CN1), either
too high (increased risk of diabetic nephropathy) or too low
(carnosinase deficiency), is apparently leading to diseased
state. These clinical cues may help us to decipher the enigmatic role of serum carnosinase. One could think that the
role of serum carnosinase is to regulate the serum carnosine
concentration (i.e., keep it within a certain homeostatic
range). However, this is somewhat unlikely. First, a large
number of mammals, including carnivores and omnivores
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1829
BOLDYREV, ALDINI, AND DERAVE
with large dietary HCD intake, have very low serum carnosinase activity, compared with humans and other hominids.
So, serum carnosinase probably did not evolve to avoid
high serum carnosine concentrations, even though it may
help in the digestion of HCD and the availability of ␤-alanine and L-histidine in meat-eating early hominids. Second,
the findings of dietary interventions in carnosinase deficiency patients do not support the idea that high carnosine
concentrations in itself would be causing the mental retardation and other clinical signs (88). Finally, it would be
somewhat strange that we would develop a system with
the aim to avoid the high presence of a molecule like carnosine
that has low toxicity and has been documented to have
health-promoting effects in rodents (who lack serum carnosinase activity). The latter has been termed by some researchers as the human carnosinase paradox (173): Why
did such an active serum enzyme evolve in humans, when
this degrades and prevents the presence of a molecule,
which has protective properties for a number of diseases
(see sect. VI). For these reasons, we currently believe that
serum carnosinase’s primary role is not to prevent excessive
serum carnosine concentrations. This does, however, not
exclude that too high serum carnosinase activity can still be
a disadvantage because it too actively limits the presence of
circulating carnosine.
If not for the avoidance of hypercarnosinemia, then for
what other function did hominids develop such an active
version of the serum carnosinase system? The most logical
alternative would be the deliverance of the hydrolysis products: L-histidine, ␤-alanine, and GABA (in the case of homocarnosine). As carnosine and homocarnosine are normally not present in the bloodstream, it is likely that serum
carnosinase would get in contact with these substrates in the
interstitium of certain tissues, where they have become
available through the release by cells. This proposal is still
hypothetical at present, but it makes teleological sense, and
there are some examples of tissues that would make use of
such system (see sect. V). Much work still needs to be performed before we can fully understand the, at first instance,
paradoxically presence of serum carnosinase in humans.
VIII. APPLICATIONS TO SPORT AND
NUTRITION SCIENCE
A. Determinants of Muscle Carnosine
Content
There is a considerable variation in muscle carnosine content among humans, as three- to fourfold differences have
been demonstrated between the lowest (⬃10 mmol/kg dry
wt) and highest (⬃40 mmol/kg dry wt) reported levels in
humans (126, 163, 355). Despite the large interindividual
variation, the intraindividual variation is rather limited. Repeated measurements of muscle carnosine over time in the
same person displays only small fluctuations (39), and very
high correlations have been found for muscle carnosine
content within monozygotic twin pairs (35). Below, we describe the most important determinants that are responsible
for the remarkable interindividual variation in muscle carnosine concentrations.
1. Fiber type
There is a clear difference in the carnosine content of different fiber types. It has repeatedly been reported that human
fast-twitch (type II, white) fibers contain 30 –100% more
carnosine compared with slow-twitch (type I, red) fibers,
based on HPLC analysis of pooled single fibers (162, 171,
213, 356). Similar fiber type differences have been documented in a wide range of mammals (116, 117, 336, 337).
Within the human population, individuals that have fasttwitch fibers as the predominant fiber type in their muscles
will have higher muscle carnosine levels compared with
those who mainly possess slow-twitch fibers (245, 336,
349). Examples of the former type of people are athletes
who excel in sprint-type and high-intensity exercise, such as
100-m runners, short-track skaters, etc., whereas examples
of the latter include endurance-type athletes such as marathon runners, triathletes, etc. Baguet et al. (37) have used a
noninvasive technique based on proton magnetic resonance
spectroscopy (1H-MRS) to quantify muscle carnosine content in a diverse group of elite athletes. They observed
markedly higher carnosine content in the sprinters compared with the endurance athletes, in agreement with previous reports using biopsies (289). Baguet et al. (37) thus
proposed the use of 1H-MRS as a noninvasive estimation
method for muscle fiber type composition.
2. Sex
As discussed in section V, an important aspect of the physiological role of carnosine in skeletal muscle is related to
contractile function in general and more specifically to excitation-contraction coupling (calcium handling) as well as
to the protection against exercise-induced acidosis (pH
buffering). Thus possessing high muscle carnosine content
may be advantageous for high-intensity exercise capacity.
Below we will discuss the determinants of muscle carnosine
content in humans, its implications for exercise capacity, as
well as the nutritional strategies that influence muscle carnosine content.
1830
Both biopsy (243) and MRS (126) studies have shown that
men display considerably (22– 82%) higher carnosine levels
compared with women. At prepubertal age however, this
sexual dimorphism is not yet present (35) (see FIGURE 17).
These findings suggest that androgens have a positive effect
on the muscle carnosine synthesis and content. Bodybuilders, likely using anabolic steroids, have indeed been shown
to hold very high muscle carnosine concentrations (355).
An animal study by Penafiel et al. (294) provided supportive
data for this, as they observed that castration of mice re-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
[carnosine] gastrocnemius (mM)
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
7
B. Dietary Manipulation of Muscle
Carnosine Content
Females
Males
6
5
4
3
2
1
0
0
10
20
30
40
50
60
70
80
Age (years)
FIGURE 17. Carnosine content in the gastrocnemius medialis
muscle of healthy men (n ⫽ 271) and women (n ⫽ 217) of different
ages, measured by NMR spectroscopy (data compiled from Refs.
35, 126). Each symbol represents the average of 10 healthy subjects, ranked by age.
duced muscle carnosine levels by 40%, while testosterone
administration to female mice increased muscle carnosine
content by 268%. A similar study on orchidectomy with or
without testosterone replacement in mice confirmed these
observations and clarified that the mechanism of testosterone’s effect is not based on enhanced transcription of the
carnosine synthase enzyme but more likely through the increased expression of the taurine/␤-alanine transporter
(125).
3. Age
There are no longitudinal data available, but cross-sectional
studies demonstrate that muscle carnosine levels are negatively correlated with age (104, 200, 346, 356). In a study
on 263 subjects of ages 9 – 83 yr, Baguet et al. (35) reported
that the lower carnosine content at old age is mainly the
result of a decrease that occurs during adulthood, even
shortly after puberty, rather than a decrease from adulthood to elderly (see FIGURE 17). The cause for the decline in
muscle carnosine with advancing age is unclear, but a decline in androgens/estrogens and fiber type changes may be
contributing factors.
4. Other
It has been suggested that exercise training can also alter
muscle carnosine content (350). However, most short-term
training intervention studies seem to agree that a few weeks
of training will not stimulate carnosine accumulation in
muscle (36, 212, 213, 244). It remains to be established
whether long-term (months to years) training intervention
is able to modulate muscle carnosine content, especially
when this chronic exercise intervention is possibly accompanied by a shift in fiber type composition (327). Other
determinants of muscle carnosine content, such as diet,
which will be discussed in the following part.
The synthesis rate of carnosine in muscle cells appears to be
rate-limited by the availability of ␤-alanine, rather than
L-histidine (118). L-Histidine, an essential amino acid, is
present in the blood in sufficient concentrations and will
probably only impose a limitation to muscle carnosine synthesis in the case of a L-histidine-free diet (357). The endogenous supply of ␤-alanine is presumably dependent on the
hepatic synthesis from uracil degradation. However, this
metabolic route is poorly understood and possibly inadequate for the total ␤-alanine demand. Therefore, exogenous
and thus dietary supply of ␤-alanine seems to be of relevance as well. The average daily intake of ␤-alanine from an
omnivore Western diet has been calculated to amount to
⬃330 mg/day (126), nearly all of which is derived from
animal products (red meat, white meat, fish, etc.) in the
form of HCD.
As indicated in section II, there is a large variation in muscle
HCD content between animal species. Therefore, the choice
of animal-derived dietary ingredients will markedly determine the daily HCD intake (166). Examples of food sources
in the human diet specifically rich in HCD are chicken,
turkey, tuna, horse, and whale, containing the equivalent of
2–7 g ␤-alanine per kg ww. Beef, pork, deer and rabbit meat
have more intermediate concentrations (1–2 g/kg ww ␤-alanine). Some fishes, such as mackerel and sole, contain virtually no HCD. As there exist large differences in HCD
content between muscle types within a species, there is also
large variability between the various meat cuts, which can
easily differ by a factor of 3 or more, in favor of the white
meat portions (containing more type II fibers and thus
HCD) (3, 261). Although carnosine has a good chemical
stability, small losses may occur in the HCD content of the
meat during the production process. Cooking of beef muscle will decrease muscle carnosine content by 5–20% (308).
Dry-curing of ham from pork biceps femoris muscle will
decrease HCD content by 35% over a 10-mo period (249),
but these losses are less pronounced than those for creatine.
Dairy and vegetable food products do not contain HCD nor
␤-alanine, except from the reported presence of ␤-alanine in
vegetable oils, albeit in concentrations 100- to 1,000-fold
lower than in meat (323). As shown in FIGURE 18, vegetarians, almost exclusively relying on endogenous sources of
␤-alanine, have been shown to possess ⬎20% lower muscle
carnosine content compared with omnivores (126). This
confirms the initial hypothesis that endogenous ␤-alanine
production is probably insufficient for unrestricted muscle
carnosine synthesis. Only short-term intervention studies (5
wk) are available at present and do not seem to indicate a
significant reduction in muscle carnosine content when
transiently switching omnivores onto a vegetarian diet (36).
However, the latter finding probably is caused by the long
half-life of carnosine in tissues. Possibly longer duration
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1831
BOLDYREV, ALDINI, AND DERAVE
Camosine content (% vs. control)
300
250
Human
200
BA suppl.
1.2%, 8 wks
BA suppl.
8 wks
150
BA suppl.
0.6%, 8 wks
Veg.
5 wks
Mouse
100
Ominv. diet
Veg.
50 > 8 yrs
0
0
~4
~60
~700
~1300
Daily ingested beta-alanine (mg/kg BW)
FIGURE 18. Dietary manipulation of muscle carnosine content. Relationship between daily ingested ␤-alanine
equivalent (ingested as meat or supplements) and muscle carnosine content (% of control diet). Human data
(black circles) are based on Everaert et al. (126) (⬎8 yr vegetarian diet and ⬃4 mg/kg body wt), Baguet et al.
(36) (5 wk vegetarian diet), and Stellingwerff et al. (344) (BA suppl. ⫽ ␤-alanine supplementation) and
calculated for a 70-kg subject. Mouse data are adapted from Everaert et al. (127) (open circles, extensor
digitorum longus muscle).
vegetarianism is required to longitudinally demonstrate a
decline in muscle carnosine content. On the other hand,
raising the dietary intake of ␤-alanine has been shown to be
an effective and powerful means to induce muscle carnosine
loading (166).
The regular ingestion of a chicken breast extract (CBEX),
which is high in HCD content (325) and is a popular dietary
product in Asia, can elevate muscle carnosine content in rats
(258, 351). In 2006, Harris et al. (165) have shown that the
daily ingestion of a very high dose of L-carnosine (13 g/day)
continued for 4 wk could elevate muscle carnosine content
of healthy volunteers by 65%. Interestingly, a similar increase was obtained when ingesting pure ␤-alanine, rather
than carnosine, in an isomolar dose, which lends further
credit to the conclusion that (dietary) ␤-alanine is rate limiting for muscle carnosine content (165). A number of independent groups (39, 102, 343) have since confirmed this
finding of muscle carnosine loading through chronic ␤-alanine supplementation (1.6 – 6.4 g/day for several weeks)
and have further optimized this procedure [reviewed in
Harris et al. (166) and Stellingwerff et al. (344)]. The muscle carnosine loading effect of chronic ␤-alanine supplementation has also been shown in mice (127), although
much higher doses are required to obtain the same effects
(FIGURE 18).
So far, the highest reported increases in human muscle carnosine content are 80 – 85% upon 10 –12 wk of supplementation (102, 171). Stellingwerff et al. (344) indicated that
the major determinant of supplementation-induced carnosine loading is the total ingested dose over a supplementation
period, rather than the daily amount. Coingestion of ␤-alanine supplements with meals seems to promote carnosine
synthesis, which points to a possible role of insulin in this
process (342). Once accumulated in the muscle, the elevated
carnosine content remains present for an extended period
1832
upon cessation of supplementation. The effective washout
period (return to presupplementation baseline) apparently
amounts to 10 –20 wk (39, 344). Little side-effects have
been reported for ␤-alanine ingestion in the concerned
doses, except from transient and unpleasant itching/flushing sensations on the skin, called parasthesia (165). To circumvent this, slow-release ␤-alanine tablets have been developed which result in slower absorption kinetics and
which reduce the incidence of parasthesia to the level of
placebo (101). The mechanism of the itching/tingling
sensation upon oral ␤-alanine has recently been ascribed
to Mas-related G protein-coupled receptor member D
(MrgprD), a G protein-coupled receptor, expressed in
dorsal root ganglion neurons involved in cutaneous
mechanosensation (236).
Several questions with regard to dietary manipulation of
muscle carnosine content remain to be answered. It still has
to be determined whether or not a normal (omnivore) variation in the type and amount of meat ingestion will influence the muscle carnosine concentration between individuals. An initial comparison of high versus low meat eaters did
not show a difference in muscle carnosine content (126).
Furthermore, the upper safe limit of muscle carnosine loading is still unknown. Finally, it is unclear whether dietary
intake of ␤-alanine has also consequences for HCD content
of nonmuscle tissues. In this respect, Tomonaga et al. (367)
recently showed that acute CBEX administration in rats can
elevate the HCD concentrations in certain brain regions.
C. Ergogenic Potential of Carnosine Loading
The above-described ␤-alanine supplementation-induced
muscle carnosine loading strategy has been shown to be
beneficial for certain types of exercise performance. In
2007, Hill et al. (172) reported that the capacity to cycle at
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
110% of maximal power (Wmax) was improved by 13 and
16% following, respectively, 4 and 10 wk of chronic ␤-alanine supplementation in healthy untrained volunteers.
Later studies confirmed that ergogenic effects can also be
obtained in well-trained athletes, such as cyclists and rowers (34, 378). However, not all studies have been able to
demonstrate beneficial effects of carnosine loading on exercise performance (105, 212), which probably relates to the
type of exercise. In a recent meta-analysis based on 15 experimental studies on ␤-alanine and exercise performance,
Hobson et al. (185) came to the conclusion that ␤-alanine is
beneficial in exercise types lasting 1– 4 min, whereas the
effect was less pronounced (P ⫽ 0.046) in longer duration
exercises (⬎4 min) and was nonsignificant for shorter duration exercise (⬍1 min). More well-designed studies are
definitely needed to better define the precise performanceenhancing effects of carnosine loading. In the meanwhile,
␤-alanine is rapidly gaining world-wide popularity as an
ergogenic nutritional supplement in the sports community.
D. Application in Food Science
There are many applications of the measurement of the
concentration of carnosine and other HCD to gain information about the composition of food products. For example, carnosine and anserine seem to be missing from several
industrially designed chicken soups, which indicates that no
chicken meat is used for its production (170). Another example is the analysis of HCD to detect products of animal
origin in feed for ruminants, which is often forbidden, because of the increased risk of bovine spongiform encephalopathy (BSE) (328). Also, the source of meat (chicken,
pork, beef) can be traced by the ratio of carnosine to anserine in a meat product (365).
Human urinary concentrations of carnosine, anserine, and
especially Npi-methyl-histidine (the degradation product of
anserine) have been proposed to be good biomarkers of
dietary meat intake, which can be used as a compliance tool
in dietary intervention studies (111) or as an estimate in
epidemiological studies investigating the effects of meat intake on health and disease (90).
Given the multitude of potential health-enhancing effects,
carnosine has been identified as one of the promising bioactives in meat (19). Several approaches can be developed to
take advantage of this characteristic. The functional or enriched meat approach aims to increase the total HCD content of meat. Different dietary strategies, such as carnosine
and L-histidine supplementation (168, 220, 242), are currently explored in farm animals to increase the quality of
the meat as well as the HCD content, with potential health
benefit for the consumer. Alternatively, extraction procedures may be applied to meat products to obtain a HCDrich extract, such as chicken essence, as a functional food
(232). Clearly, the potential applications of HCD in the
field of meat and food science continue to evolve.
IX. CONCLUSIONS AND FUTURE
PERSPECTIVES
A. Why Did the Carnosine System Evolve?
The above sections have summarized a large body of literature on the biochemical, physiological, and therapeutic
properties of carnosine. A complex and energy-consuming
molecular system has been established in the course of evolution to synthesize, modulate, transport, and degrade carnosine across the organism. In this section, we now aim to
combine these individual findings into a global vision on the
role of HCD in the function of organisms. A number of
questions can be asked related to why this system evolved
and why it evolved in the way it did: Why carnosine and not
free L-histidine? Why does L-histidine need to be combined
with ␤-alanine, and not with another amino acid? Why the
methylated carnosine analogs?
To resolve these questions, it is necessary to compare and
summarize the biochemical properties of carnosine with its
constituent amino acids, and with the other HCD in one
table (TABLE 2). From TABLE 2, it can be deduced that most
of the bioactive functions of carnosine relate to histidine
and its imidazole moiety, except from its role in AGEs and
ALEs formation and calcium regulation, which is accomplished much better by the dipeptide. From this information, we propose the structure-function relationship of carnosine, depicted in FIGURE 19, which will be used as a basis
for the discussion below. The methylated carnosine analogs
are mostly similar in functionality compared with carnosine, although there are some slight differences in effectiveness.
1. Why carnosine and not free L-histidine?
From the comparison of the biochemical properties of carnosine versus L-histidine (TABLE 2), it can be deduced that
most of the functional properties of carnosine are related to
the L-histidine part (mostly the imidazole moiety), of which
the dipeptide is composed. In other words, as roughly only
1 of 20 amino acids is L-histidine in proteins, HCD are a
way to more effectively store histidine in peptide form. Yet,
it can be questioned why nature did bother to create an
ATP-consuming carnosine synthesis system, if nearly the
same roles can be accomplished by the simple free L-histidine itself. In fact, it seems that having free L-histidine,
instead of the HCD, is probably a good solution to a certain
extent, concerning most of the homeostatic challenges for
which an organism is in need of the imidazole-related properties. As described in section II, several families of fishes
have a low concentration of HCD, but instead high millimolar concentrations of L-histidine, whereas mammals and
other animal classes with high HCD content, will only have
a micromolar concentration of free L-histidine. Thus free
L-histidine was a reasonably good solution earlier in verte-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1833
BOLDYREV, ALDINI, AND DERAVE
Table 2. Table comparing the biochemical properties of carnosine and related molecules
Activity
Carnosine
Anserine
Homocarnosine
Ophidine
␤-Alanine
Histidine
**
**
**
**
**
**
**
**
**
ND
**
**
ND
ND
ND
**
ND
ND
ND
ND
/
/
/
*
/
**
**
**
*
*
**
**
ND
**
ND
*
**
ND
ND
ND
ND
***
382
**
**
ND
ND
*
*
353
**
**
*
ND
*
*
9
Antioxidant (scavenging effect)
Peroxyl radicals
Hydroxyl radicals
Peroxynitrite
Singlet oxygen
Nitric oxide
Buffering activity
pKa
Metal ion chelating
Cu2⫹ chelation
AGEs inhibition
transglycation
ALEs inhibition
HNE scavenging
Reference Nos.
100, 209, 219
69, 85, 209
133
121
271
3
Marks summarize the activity of carnosine derivatives and of the constitutive amino acids in respect to
carnosine: **, equal; *, lower: ***, greater; /, no activity; ND, not determined.
brate evolution, but the system has further evolved, became
clearly more complex, and improved. So what are the additional advantages of a L-histidine-containing dipeptide
over free L-histidine?
A first probable reason is improvement and better suitability to the functions. As an example, the pKa value of the
imidazole of free L-histidine is 6.0. This is close to the physiological pH range, but not exactly in it. By combining
L-histidine with ␤-alanine, the pKa of the imidazole alters to
6.72, which is simply a better version of the buffer (even
though the pH buffering property in carnosine is still attributable to the L-histidine part). Similar fine-tuning can be
observed for other biochemical/physiological properties as
well (TABLE 2). More importantly, some functions, such as
the inhibiting effect of the formation of ALEs (see sect. III),
H
N
N
O
· AGEs and
ALEs inhibition
HO
· Antioxidant
· Metal ion chelation
· pH-buffering
· Calcium
NH
regulation
O
H 2N
· Regulates histidine
metabolic fate
FIGURE 19. Structure-activity relationship of carnosine. The imidazolic ring is required for the antioxidant, metal ion chelating and
buffering activity. The ␤-alanine moiety regulates the metabolic fate
of the L-histidine residue and synthesis of carnosine. Both ␤-alanine
and L-histidine residues synergistically act in the inhibiting effect
towards AGEs and ALEs formation.
1834
seem to be specific to the dipeptide, and can poorly be
replicated by L-histidine alone.
A second probable reason is guaranteeing a stable tissue
concentration. L-Histidine is involved in many pathways,
such as protein synthesis and histamine formation; pathways with high physiological priority. To avoid highly fluctuating tissue L-histidine content, it has probably proven
beneficial to capture L-histidine into a metabolically more
inert dipeptide. Even though there is a system to degrade
carnosine (see sect. IV on carnosinases), the hydrolyzing
enzyme is usually not present in the compartments where
carnosine is synthesized and subsequently stored.
A third possible reason may relate to reduced toxicity. It has
been suggested that dipeptides are less toxic than single
amino acids, for example, to neurons in the brain. Also,
single amino acids probably more easily initiate and activate unwanted metabolic pathways than dipeptides.
2. Why does L-histidine need to be combined with
␤-alanine?
It seems that in the “choice” for an ideal amino acid to bind
with L-histidine in the dipeptide, preference is given to a rare
and nonproteinogenic amino acid (a ␤-amino acid). A first
advantage of this is to stabilize the dipeptide by reducing the
affinity towards hydrolysis by (di)peptidases. If L-histidine
would be bound to another proteinogenic amino acid, it
would be readily hydrolyzed by the peptidases. However,
carnosine is not at all a good substrate to peptidases, which
thereby allows a separately regulated and specific metabolism (carnosinase) compared with other dipeptides. Not
only for hydrolysis, but also for synthesis, choosing a rare
(beta) amino acid that is not involved in many more path-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
ways like protein synthesis seems more easy to control.
Indeed, ␤-alanine is clearly the rate-limiting precursor in the
synthesis of carnosine. So to regulate the carnosine concentration, it is sufficient to regulate the availability of ␤-alanine. It has recently been proposed that there is a specific
enzyme in muscle, glutamate decarboxylase-like protein 1
(GADL1), directly synthesizing ␤-alanine from aspartate by
decarboxylation (235), which could be a demonstration
that ␤-alanine availability is controlled by a specific and
unique set of pathways to indirectly regulate tissue carnosine content.
3. Why the methylated carnosine analogs?
The distribution of HCD in muscles from different species
throughout the animal kingdom, as described in section II,
may help us to understand the reason and advantage for
having the methylated versus the nonmethylated carnosine
and for having one methylated form over the other (anserine versus ophidine). It is very likely that these typical patterns found in animal families are not a mere coincidence
and have a specific reason. As indicated in TABLE 2, the
differences in the chemical properties between carnosine,
anserine, and ophidine are only minor. However, there are
slight differences in the pH buffering capacity (pKa is 6.72
for carnosine, 7.01 for anserine, and 7.03 for ophidine),
and also the antioxidant capacity. When certain animals
predominantly express one type of HCD, this could mean
that the chemical property, for which this HCD is optimal,
is the primary reason to have this HCD present in the organism.
However, it seems more likely that the reason for carnosine
methylation is related to the fact that anserine and ophidine
are less easily hydrolyzed by carnosinase than carnosine
itself. The methylation of carnosine could therefore be a
way to protect HCD from hydrolysis and be a better way to
store and ascertain constant HCD concentrations in certain
cells and tissues. Yet why do humans, as the unique species
of all mammals (see FIGURE 3), only have carnosine and not
the methylated forms? One reason could be that the protection against carnosinase-induced hydrolysis through carnosine methylation is not required in humans. Even the
contrary, the absence of methylated carnosine analogs in
combination with the high expression of serum carnosinase
in humans compared with other animals even suggests that
the HCD pool in human tissues is less stable and should be
more mobile to contribute to other physiological processes.
This hypothesis, however, needs further testing.
an informal Carnosine Consortium (http://users.unimi.it/
carnosine_co/) was recently established, with the objective to organize regular international meetings on this topic and to
facilitate international collaboration. Some enzymes (e.g.,
mammalian CMT) still need to be discovered, transgenic
animals are waiting to be developed (e.g., knockout of CS),
and systematic efforts in the fields of genetics, pharmacology, etc. would greatly enhance the required scientific progress in the field.
Carnosine has pleiotropic effects. This is probably one of
the reasons why this system evolved in the first place. One
molecule can fix several problems: a broad-spectrum solution. At the same time, this pleiotropy imposes great challenges to the researchers. Most studies have been focusing
only on a single aspect of the molecule, thereby overlooking
the other aspects. This should be taken into account in the
design of future research.
Carnosine is a promising therapeutic molecule. A humanspecific problem is the rapid degradation upon oral or intravenous administration by serum carnosinase. The development of carnosinase-resistant carnosine analogs is therefore a major challenge, but a sure hurdle that needs to be
taken to allow future pharmaceutical progress. In addition,
a better understanding of diseases that suffer from reduced
tissue carnosine contents, and of the mechanisms regulating
tissue concentration, may offer therapeutic perspectives as
well.
Over 100 years past the discovery of carnosine, we are still
no where near a good and complete understanding of its
physiological and pathophysiological significance, but we
hope that this review has helped the reader to grasp the
current status on the scientific knowledge of this intriguing
molecule.
ACKNOWLEDGMENTS
During the period of writing and preparing this manuscript,
one of the authors, Alexander Boldyrev, passed away. We
honor Professor Boldyrev with this review paper as a tribute
to the immense contribution he has made to the field of
carnosine research. The assistance of Audrey Baguet in the
preparation of the manuscript is greatly acknowledged.
Address for reprint requests and other correspondence: W.
Derave, Dept. of Movement and Sports Sciences, Ghent
University, Watersportlaan 2, B-9000 Ghent, Belgium (wim.
derave@ugent.be).
B. Future Perspectives
GRANTS
Although the research interest in carnosine is now clearly
gaining momentum (106), we are currently still missing the
critical mass needed to systematically resolve the remaining
missing links in the carnosine system. For this purpose,
This research was supported by Research Foundation-Flanders
Grants FWO G.0046.09, G.0243.11, and G.0352.13N (to W.
Derave). G. Aldini was supported by the Italian Ministry of Uni-
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1835
BOLDYREV, ALDINI, AND DERAVE
versity and Research (Progetti di Ricerca di Interesse Nazionale
2007, 20079SLZMC_001).
18. Aoki A, Shibata Y, Okano S, Maruyama F, Amano A, Nakagawa I, Abiko Y. Transition
metal ions induce carnosinase activity in PepD-homologous protein from Porphyromonas gingivalis. Microb Pathog 52: 17–24, 2012.
19. Arihara K. Strategies for designing novel functional meat products. Meat Sci 74: 219 –
229, 2006.
DISCLOSURES
No conflicts of interest, financial or otherwise, are declared
by the authors.
20. Aristoy MC, Toldra F. Concentration of free amino acids and dipeptides in porcine
skeletal muscles with different oxidative patterns. Meat Sci 50: 327–332, 1998.
21. Aristoy MC, Toldra F. Histidine dipeptides HPLC-based test for the detection of
mammalian origin proteins in feeds for ruminants. Meat Sci 67: 211–217, 2004.
22. Artun BC, Kusku-Kiraz Z, Gulluoglu M, Cevikbas U, Kocak-Toker N, Uysal M. The
effect of carnosine pretreatment on oxidative stress and hepatotoxicity in binge ethanol administered rats. Hum Exp Toxicol 29: 659 – 665, 2010.
REFERENCES
1. Abbasoglu L, Kalaz EB, Soluk-Tekkesin M, Olgac V, Dogru-Abbasoglu S, Uysal M.
Beneficial effects of taurine and carnosine in experimental ischemia/reperfusion injury
in testis. Pediatr Surg Int 28: 1125–1131, 2012.
23. Ashikawa I, Itoh K. Raman-spectra of polypeptides containing L-histidine residues and
tautomerism of imidazole side-chain. Biopolymers 18: 1859 –1876, 1979.
2. Abe H. Distribution of free L-histidine and related dipeptides in the muscle of freshwater fishes. Comp Biochem Physiol B Comp Biochem 76: 35–39, 1983.
24. Attanasio F, Cataldo S, Fisichella S, Nicoletti S, Nicoletti VG, Pignataro B, Savarino A,
Rizzarelli E. Protective effects of L- and D-carnosine on alpha-crystallin amyloid fibril
formation: implications for cataract disease. Biochemistry 48: 6522– 6531, 2009.
3. Abe H. Role of histidine-related compounds as intracellular proton buffering constituents in vertebrate muscle. Biochemistry 65: 757–765, 2000.
4. Abe H, Dobson GP, Hoeger U, Parkhouse WS. Role of histidine-related compounds
to intracellular buffering in fish skeletal muscle. Am J Physiol Regul Integr Comp Physiol
249: R449 –R454, 1985.
25. Attanasio F, Convertino M, Magno A, Caflisch A, Corazza A, Haridas H, Esposito G,
Cataldo S, Pignataro B, Milardi D, Rizzarelli E. Carnosine inhibits A beta 42 aggregation by perturbing the H-bond network in and around the central hydrophobic cluster. Chembiochem 14: 583–592, 2013.
5. Ackermann D, Timpe O, Poller K. Uber das anserin, einen neuen Bestandteil der
Vogelmusculatur. Hoppe-Seil Ztschr Physiol Chem B183: S1–S10, 1929.
26. Aydin AF, Kucukgergin C, Ozdemirler-Erata G, Kocak-Toker N, Uysal M. The effect
of carnosine treatment on prooxidant-antioxidant balance in liver, heart and brain
tissues of male aged rats. Biogerontology 11: 103–109, 2010.
6. Agu R, Cowley E, Shao D, Macdonald C, Kirkpatrick D, Renton K, Massoud E. Protoncoupled oligopeptide transporter (POT) family expression in human nasal epithelium
and their drug transport potential. Mol Pharmacol 8: 664 – 672, 2011.
27. Aydin AF, Kusku-Kiraz Z, Dogru-Abbasoglu S, Gulluoglu M, Uysal M, Kocak-Toker N.
Effect of carnosine against thioacetamide-induced liver cirrhosis in rat. Peptides 31:
67–71, 2010.
7. Ahluwalia TS, Lindholm E, Groop LC. Common variants in CNDP1 and CNDP2, and
risk of nephropathy in type 2 diabetes. Diabetologia 54: 2295–2302, 2011.
28. Babizhayev MA. Antioxidant activity of L-carnosine, a natural histidine-containing dipeptide in crystalline lens. Biochim Biophys Acta 1004: 363–371, 1989.
8. Alaghband-Zadeh J, Mehdizadeh S, Khan NS, O’Farrell A, Bitensky L, Chayen J. The
natural substrate for nitric oxide synthase activity. Cell Biochem Funct 19: 277–280,
2001.
29. Babizhayev MA, Deyev AI, Yermakova VN, Semiletov YA, Davydova NG, Doroshenko VS, Zhukotskii AV, Goldman IM. Efficacy of N-acetylcarnosine in the treatment
of cataracts. Drugs RD 3: 87–103, 2002.
9. Aldini G, Carini M, Beretta G, Bradamante S, Facino RM. Carnosine is a quencher of
4-hydroxy-nonenal: through what mechanism of reaction? Biochem Biophys Res Commun 298: 699 –706, 2002.
30. Babizhayev MA, Deyev AI, Yermakova VN, Semiletov YA, Davydova NG, Kurysheva
NI, Zhukotskii AV, Goldman IM. N-acetylcarnosine, a natural histidine-containing
dipeptide, as a potent ophthalmic drug in treatment of human cataracts. Peptides 22:
979 –994, 2001.
10. Aldini G, Facino RM, Beretta G, Carini M. Carnosine and related dipeptides as quenchers of reactive carbonyl species: from structural studies to therapeutic perspectives.
Biofactors 24: 77– 87, 2005.
11. Aldini G, Gamberoni L, Orioli M, Beretta G, Regazzoni L, Maffei FR, Carini M. Mass
spectrometric characterization of covalent modification of human serum albumin by
4-hydroxy-trans-2-nonenal. J Mass Spectrom 41: 1149 –1161, 2006.
31. Babizhayev MA, Micans P, Guiotto A, Kasus-Jacobi A. N-acetylcarnosine lubricant
eyedrops possess all-in-one universal antioxidant protective effects of L-carnosine in
aqueous and lipid membrane environments, aldehyde scavenging, and transglycation
activities inherent to cataracts: a clinical study of the new vision-saving drug N-acetylcarnosine eyedrop therapy in a database population of over 50,500 patients. Am J Ther
16: 517–533, 2009.
12. Aldini G, Granata P, Carini M. Detoxification of cytotoxic alpha,beta-unsaturated
aldehydes by carnosine: characterization of conjugated adducts by electrospray ionization tandem mass spectrometry and detection by liquid chromatography/mass
spectrometry in rat skeletal muscle. J Mass Spectrom 37: 1219 –1228, 2002.
32. Babizhayev MA, Yermakova VN, Sakina NL, Evstigneeva RP, Rozhkova EA, Zheltukhina GA. N-alpha-acetylcarnosine is a prodrug of L-carnosine in ophthalmic application as antioxidant. Clin Chim Acta 254: 1–21, 1996.
13. Aldini G, Orioli M, Carini M, Maffei FR. Profiling histidine-containing dipeptides in rat
tissues by liquid chromatography/electrospray ionization tandem mass spectrometry.
J Mass Spectrom 39: 1417–1428, 2004.
33. Bae ON, Serfozo K, Baek SH, Lee KY, Dorrance A, Rumbeiha W, Fitzgerald SD,
Farooq MU, Naravelta B, Bhatt A, Majid A. Safety and efficacy evaluation of carnosine,
an endogenous neuroprotective agent for ischemic stroke. Stroke 44: 205–212, 2013.
14. Aldini G, Orioli M, Rossoni G, Savi F, Braidotti P, Vistoli G, Yeum KJ, Negrisoli G,
Carini M. The carbonyl scavenger carnosine ameliorates dyslipidaemia and renal function in Zucker obese rats. J Cell Mol Med 15: 1339 –1354, 2011.
34. Baguet A, Bourgois J, Vanhee L, Achten E, Derave W. Important role of muscle
carnosine in rowing performance. J Appl Physiol 109: 1096 –1101, 2010.
15. Alhamdani MS, Al-Kassir AH, Abbas FK, Jaleel NA, and Al-Taee MF. Antiglycation and
antioxidant effect of carnosine against glucose degradation products in peritoneal
mesothelial cells. Nephron Clin Pract 107: c26 – c34, 2007.
35. Baguet A, Everaert I, Achten E, Thomis M, Derave W. The influence of sex, age and
heritability on human skeletal muscle carnosine content. Amino Acids 43: 13–20, 2012.
16. Allen DG, Lamb GD, Westerblad H. Skeletal muscle fatigue: cellular mechanisms.
Physiol Rev 88: 287–332, 2008.
36. Baguet A, Everaert I, De Naeyer H, Reyngoudt H, Stegen S, Beeckman S, Achten E,
Vanhee L, Volkaert A, Petrovic M, Taes Y, Derave W. Effects of sprint training combined with vegetarian or mixed diet on muscle carnosine content and buffering capacity. Eur J Appl Physiol 111: 2571–2580, 2011.
17. Ansurudeen I, Sunkari VG, Grunler J, Peters V, Schmitt CP, Catrina SB, Brismar K,
Forsberg EA. Carnosine enhances diabetic wound healing in the db/db mouse model
of type 2 diabetes. Amino Acids 43: 127–134, 2012.
37. Baguet A, Everaert I, Hespel P, Petrovic M, Achten E, Derave W. A new method for
non-invasive estimation of human muscle fiber type composition. PLoS One 6: e21956,
2011.
1836
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
38. Baguet A, Koppo K, Pottier A, Derave W. Beta-alanine supplementation reduces
acidosis but not oxygen uptake response during high-intensity cycling exercise. Eur J
Appl Physiol 108: 495–503, 2010.
61. Boldyrev A. Carnosine and Oxidative Stress in Cells and Tissues. New York: Nova
Science, 2007.
39. Baguet A, Reyngoudt H, Pottier A, Everaert I, Callens S, Achten E, Derave W. Carnosine loading and washout in human skeletal muscles. J Appl Physiol 106: 837– 842,
2009.
62. Boldyrev A, Fedorova T, Stepanova M, Dobrotvorskaya I, Kozlova E, Boldanova N,
Bagyeva G, Ivanova-Smolenskaya I, Illarioshkin S. Carnosine increases efficiency of
DOPA therapy of Parkinson’s disease: a pilot study. Rejuvenation Res 11: 821– 827,
2008.
40. Bakardjiev A. Carnosine and beta-alanine release is stimulated by glutamatergic receptors in cultured rat oligodendrocytes. Glia 24: 346 –351, 1998.
63. Boldyrev A, Koudinov A, Berezov T, Carpenter DO. Amyloid-beta induced cell death
is independent of free radicals. J Alzheimers Dis 6: 633– 638, 2004.
41. Bando K, Ichihara K, Shimotsuji T, Toyoshima H, Koda K, Hayashi C, Miyai K.
Reduced serum carnosinase activity in hypothyroidism. Ann Clin Biochem 23: 190 –
194, 1986.
64. Boldyrev AA. Carnosine: new concept for the function of an old molecule. Biochemistry 77: 313–326, 2012.
42. Bando K, Ichihara K, Toyoshima H, Shimotuji T, Koda K, Hayashi C, Miyai K. Decreased activity of carnosinase in serum of patients with chronic liver disorders. Clin
Chem 32: 1563–1565, 1986.
43. Bando K, Shimotsuji T, Toyoshima H, Hayashi C, Miyai K. Fluorometric assay of
human serum carnosinase activity in normal children, adults and patients with myopathy. Ann Clin Biochem 21: 510 –514, 1984.
44. Baran EJ. Metal complexes of carnosine. Biochemistry 65: 789 –797, 2000.
45. Baraniuk JN, El-Amin S, Corey R, Rayhan R, Timbol C. Carnosine treatment for gulf
war illness: a randomized controlled trial. Glob J Health Sci 5: 69 – 81, 2013.
46. Baslow MH. A novel key-lock mechanism for inactivating amino acid neurotransmitters during transit across extracellular space. Amino Acids 38: 51–55, 2010.
47. Bate-Smith E. The buffering of muscle in rigour: protein, phosphate and carnosine. J
Physiol 336 –343, 1938.
48. Batrukova MA, Rubtsov AM. Histidine-containing dipeptides as endogenous regulators of the activity of sarcoplasmic reticulum Ca-release channels. Biochim Biophys
Acta 1324: 142–150, 1997.
49. Bauchart C, Savary-Auzeloux I, Patureau MP, Thomas E, Morzel M, Remond D.
Carnosine concentration of ingested meat affects carnosine net release into the portal
vein of minipigs. J Nutr 137: 589 –593, 2007.
65. Boldyrev AA, Dupin AM, Batrukova MA, Bavykina NI, Korshunova GA, Shvachkin Y.
A comparative study of synthetic carnosine analogs as antioxidants. Comp Biochem
Physiol B Comp Biochem 94: 237–240, 1989.
66. Boldyrev AA, Dupin AM, Bunin AY, Babizhaev MA, Severin SE. The antioxidative
properties of carnosine, a natural histidine containing dipeptide. Biochem Int 15: 1105–
1113, 1987.
67. Boldyrev AA, Dupin AM, Pindel EV, Severin SE. Antioxidative properties of histidinecontaining dipeptides from skeletal muscles of vertebrates. Comp Biochem Physiol B
Comp Biochem 89: 245–250, 1988.
68. Boldyrev AA, Gallant SC, Sukhich GT. Carnosine, the protective, anti-aging peptide.
Biosci Rep 19: 581–587, 1999.
69. Boldyrev AA, Kurella EG, Rubtsov AM, Tiulina OV, Shara M, Shentiurts M. [Direct
measurement of the interaction of carnosine and its analogs with free radicals]. Biokhimiia 57: 1360 –1365, 1992.
70. Bonfanti L, Peretto P, De MS, Fasolo A. Carnosine-related dipeptides in the mammalian brain. Prog Neurobiol 59: 333–353, 1999.
71. Bonner AB, Swann ME, Marway JS, Heap LC, Preedy VR. Lysosomal and nonlysosomal protease activities of the brain in response to ethanol feeding. Alcohol 12:
505–509, 1995.
50. Bauer K. Carnosine and homocarnosine, the forgotten, enigmatic peptides of the
brain. Neurochem Res 30: 1339 –1345, 2005.
72. Botka CW, Wittig TW, Graul RC, Nielsen CU, Higaka K, Amidon GL, Sadee W.
Human proton/oligopeptide transporter (POT) genes: identification of putative human genes using bioinformatics. AAPS PharmSci 2: E16, 2000.
51. Bauer K, Hallermayer K, Salnikow J, Kleinkauf H, Hamprecht B. Biosynthesis of carnosine and related peptides by glial cells in primary culture. J Biol Chem 257: 3593–
3597, 1982.
73. Bowden DW, Colicigno CJ, Langefeld CD, Sale MM, Williams A, Anderson PJ, Rich SS,
Freedman BI. A genome scan for diabetic nephropathy in African Americans. Kidney
Int 66: 1517–1526, 2004.
52. Bauer K, Salnikow J, de VF, Tixier-Vidal A, Kleinkauf H. Characterization and biosynthesis of omega-aminoacyl amino acids from rat brain and the C-6 glioma cell line. J
Biol Chem 254: 6402– 6407, 1979.
74. Brown CE. Interactions among carnosine, anserine, ophidine and copper in biochemical adaptation. J Theor Biol 88: 245–256, 1981.
53. Bauer K, Schulz M. Biosynthesis of carnosine and related peptides by skeletal muscle
cells in primary culture. Eur J Biochem 219: 43– 47, 1994.
75. Brown CE, Vidrine DW, Julian RL, Froncisz W. Copper(II) dimers in solution: evidence for motional averaging of coupling tensors without chemical-dissociation. J
Chem Soc Dalton Transactions 2371–2377, 1982.
54. Baumann L, Ingvaldsen T. Concerning histidine and carnosine. The synthesis of carnosine. J Biol Chem 35: 263–276, 1918.
76. Brownson C, Hipkiss AR. Carnosine reacts with a glycated protein. Free Radic Biol Med
28: 1564 –1570, 2000.
55. Baykara B, Cilaker MS, Tugyan K, Tekmen I, Bagriyanik HA, Sonmez U, Sonmez A,
Oktay G, Yener N, Ozbal S. The protective effects of carnosine in alcohol-induced
hepatic injury in rats. Toxicol Ind Health 2012.
77. Calam DH, Waley SG. Amino acids and related compounds in the lens. Biochem J 93:
526 –532, 1964.
56. Bellia F, Vecchio G, Rizzarelli E. Carnosine derivatives: new multifunctional drug-like
molecules. Amino Acids 43: 153–163, 2012.
78. Carini M, Aldini G, Beretta G, Arlandini E, Facino RM. Acrolein-sequestering ability of
endogenous dipeptides: characterization of carnosine and homocarnosine/acrolein
adducts by electrospray ionization tandem mass spectrometry. J Mass Spectrom 38:
996 –1006, 2003.
57. Bharadwaj LA, Davies GF, Xavier IJ, Ovsenek N. L-Carnosine and verapamil inhibit
hypoxia-induced expression of hypoxia inducible factor (HIF-1 alpha) in H9c2 cardiomyoblasts. Pharmacol Res 45: 175–181, 2002.
79. Cartwright SP, Bill RM, Hipkiss AR. L-Carnosine affects the growth of Saccharomyces
cerevisiae in a metabolism-dependent manner. PLoS One 7: e45006, 2012.
58. Bhardwaj RK, Herrera-Ruiz D, Eltoukhy N, Saad M, Knipp GT. The functional evaluation of human peptide/histidine transporter 1 (hPHT1) in transiently transfected
COS-7 cells. Eur J Pharm Sci 27: 533–542, 2006.
59. Biffo S, Grillo M, Margolis FL. Cellular localization of carnosine-like and anserine-like
immunoreactivities in rodent and avian central nervous system. Neuroscience 35:
637– 651, 1990.
60. Boldyrev A. Carnosine as a modulator of endogenous Zn2⫹ effects. Trends Pharmacol
Sci 22: 112–113, 2001.
80. Chan KM, Decker EA. Endogenous skeletal muscle antioxidants. Crit Rev Food Sci Nutr
34: 403– 426, 1994.
81. Chen Y, Getchell TV, Sparks DL, Getchell ML. Cellular localization of carnosinase in
the human nasal mucosa. Acta Otolaryngol 114: 193–198, 1994.
82. Cheng J, Wang F, Yu DF, Wu PF, Chen JG. The cytotoxic mechanism of malondialdehyde and protective effect of carnosine via protein cross-linking/mitochondrial
dysfunction/reactive oxygen species/MAPK pathway in neurons. Eur J Pharmacol 650:
184 –194, 2011.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1837
BOLDYREV, ALDINI, AND DERAVE
83. Chengappa KN, Turkin SR, Desanti S, Bowie CR, Brar JS, Schlicht PJ, Murphy SL,
Hetrick ML, Bilder R, Fleet D. A preliminary, randomized, double-blind, placebocontrolled trial of L-carnosine to improve cognition in schizophrenia. Schizophr Res
142: 145–152, 2012.
105. Derave W, Ozdemir MS, Harris RC, Pottier A, Reyngoudt H, Koppo K, Wise JA,
Achten E. Beta-alanine supplementation augments muscle carnosine content and
attenuates fatigue during repeated isokinetic contraction bouts in trained sprinters. J
Appl Physiol 103: 1736 –1743, 2007.
84. Chez MG, Buchanan CP, Aimonovitch MC, Becker M, Schaefer K, Black C, Komen J.
Double-blind, placebo-controlled study of L-carnosine supplementation in children
with autistic spectrum disorders. J Child Neurol 17: 833– 837, 2002.
106. Derave W, Sale C. Carnosine in exercise and disease: introduction to the International
Congress held at Ghent University, Belgium, July 2011. Amino Acids 43: 1– 4, 2012.
85. Choi SY, Kwon HY, Kwon OB, Kang JH. Hydrogen peroxide-mediated Cu,Zn-superoxide dismutase fragmentation: protection by carnosine, homocarnosine and anserine. Biochim Biophys Acta 1472: 651– 657, 1999.
86. Christman A. Factors affecting anserine and carnosine levels in skeletal muscles of
various animals. Int J Biochem 7: 519 –527, 1976.
87. Clifford WM. The distribution of carnosine in the animal kingdom. Biochem J 15:
725–735, 1921.
88. Cohen M, Hartlage PL, Krawiecki N, Roesel RA, Carter AL, Hommes FA. Serum
carnosinase deficiency: a non-disabling phenotype? J Ment Defic Res 29: 383–389,
1985.
89. Corona C, Frazzini V, Silvestri E, Lattanzio R, La SR, Piantelli M, Canzoniero LM,
Ciavardelli D, Rizzarelli E, Sensi SL. Effects of dietary supplementation of carnosine on
mitochondrial dysfunction, amyloid pathology, and cognitive deficits in 3xTg-AD
mice. PLoS One 6: e17971, 2011.
90. Cross AJ, Major JM, Sinha R. Urinary biomarkers of meat consumption. Cancer Epidemiol Biomarkers Prev 20: 1107–1111, 2011.
91. Crush KG. Carnosine and related substances in animal tissues. Comp Biochem Physiol
34: 3–30, 1970.
92. Cuzzocrea S, Genovese T, Failla M, Vecchio G, Fruciano M, Mazzon E, Di PR, Muia C,
La RC, Crimi N, Rizzarelli E, Vancheri C. Protective effect of orally administered
carnosine on bleomycin-induced lung injury. Am J Physiol Lung Cell Mol Physiol 292:
L1095–L1104, 2007.
93. Daniel H. Molecular and integrative physiology of intestinal peptide transport. Annu
Rev Physiol 66: 361–384, 2004.
94. Daniel H, Kottra G. The proton oligopeptide cotransporter family SLC15 in physiology and pharmacology. Pflügers Arch 447: 610 – 618, 2004.
95. Davey CL. An ion exchange method of determining carnosine, anserine and their
precursors in animal tissue. Nature 179: 209 –210, 1957.
96. Davey CL. The significance of carnosine and anserine in striated skeletal muscle. Arch
Biochem Biophys 89: 303–308, 1960.
97. Dawson R Jr, Biasetti M, Messina S, Dominy J. The cytoprotective role of taurine in
exercise-induced muscle injury. Amino Acids 22: 309 –324, 2002.
98. De Marchis S, Modena C, Peretto P, Migheli A, Margolis FL, Fasolo A. Carnosinerelated dipeptides in neurons and glia. Biochemistry 65: 824 – 833, 2000.
99. Decker E, Crum A, Calvert J. Differences in the antioxidant mechanism of carnosine
in the presence of copper and iron. J Agricultural Food Chem 40: 758 –759, 1992.
100. Decker EA, Livisay SA, Zhou S. A re-evaluation of the antioxidant activity of purified
carnosine. Biochemistry 65: 766 –770, 2000.
101. Decombaz J, Beaumont M, Vuichoud J, Bouisset F, Stellingwerff T. Effect of slowrelease beta-alanine tablets on absorption kinetics and paresthesia. Amino Acids 43:
67–76, 2012.
102. Del Favero S, Roschel H, Solis MY, Hayashi AP, Artioli GG, Otaduy MC, Benatti FB,
Harris RC, Wise JA, Leite CC, Pereira RM, de Sa-Pinto AL, Lancha-Junior AH, Gualano
B. Beta-alanine (Carnosyn) supplementation in elderly subjects (60 – 80 years): effects
on muscle carnosine content and physical capacity. Amino Acids 2011.
103. Del Rio RM, Orensanz Munoz LM, DeFeudis FV. Contents of beta-alanine and gamma-aminobutyric acid in regions of rat CNS. Exp Brain Res 28: 225–227, 1977.
104. Derave W, Jones G, Hespel P, Harris RC. Creatine supplementation augments skeletal muscle carnosine content in senescence-accelerated mice (SAMP8). Rejuvenation
Res 11: 641– 647, 2008.
1838
107. Deutsch A, Eggleton P. The titration constants of anserine, carnosine and some
related compounds. Biochem J 32: 209 –211, 1938.
108. Dobbie H, Kermack WO. Complex-formation between polypeptides and metals. 2.
The reaction between cupric ions and some dipeptides. Biochem J 59: 246 –257, 1955.
109. Dobrotvorskaya I, Fedorova T, Dobrota D, Berezov T. Characteristics of oxidative
stress in experimental rat brain ischemia aggravated by homocysteic acid. Neurochem
J 5: 42– 46, 2011.
110. Drafi F, Bauerova K, Valachova K, Ponist S, Mihalova D, Juranek I, Boldyrev A,
Hrabarova E, Soltes L. Carnosine inhibits degradation of hyaluronan induced by free
radical processes in vitro and improves the redox imbalance in adjuvant arthritis in
vivo. Neuro Endocrinol Lett 31 Suppl 2: 96 –100, 2010.
111. Dragsted LO. Biomarkers of meat intake and the application of nutrigenomics. Meat
Sci 84: 301–307, 2010.
112. Dringen R, Hamprecht B, Broer S. The peptide transporter PepT2 mediates the
uptake of the glutathione precursor CysGly in astroglia-rich primary cultures. J Neurochem 71: 388 –393, 1998.
113. Drozak J, Chrobok L, Poleszak O, Jagielski AK, Derlacz R. Molecular identification of
carnosine N-methyltransferase as chicken histamine N-methyltransferase-like protein
(HNMT-Like). Plos One 8: e64805, 2013.
114. Drozak J, Veiga-da-Cunha M, Vertommen D, Stroobant V, Van SE. Molecular identification of carnosine synthase as ATP-grasp domain-containing protein 1 (ATPGD1).
J Biol Chem 285: 9346 –9356, 2010.
115. Du Vigneaud V, Hunt M. The synthesis of D-carnosine, the enantiomorph of the
naturally occuring form, and a study of its depressor effect on the blood pressure. J Biol
Chem 93: 1936.
116. Dunnett M, Harris RC. Carnosine and taurine contents of different fibre types in the
middle gluteal muscle of the Thoroughbred horse. Equine Vet J Suppl 18: 214 –217,
1995.
117. Dunnett M, Harris RC. High-performance liquid chromatographic determination of
imidazole dipeptides, histidine, 1-methylhistidine and 3-methylhistidine in equine and
camel muscle and individual muscle fibres. J Chromatogr B Biomed Sci Appl 688: 47–55,
1997.
118. Dunnett M, Harris RC. Influence of oral beta-alanine and L-histidine supplementation
on the carnosine content of the gluteus medius. Equine Vet J Suppl 30: 499 –504, 1999.
119. Dunnett M, Harris RC, Dunnett CE, Harris PA. Plasma carnosine concentration:
diurnal variation and effects of age, exercise and muscle damage. Equine Vet J Suppl
283–287, 2002.
120. Dunnett M, Harris RC, Soliman MZ, Suwar AA. Carnosine, anserine and taurine
contents in individual fibres from the middle gluteal muscle of the camel. Res Vet Sci
62: 213–216, 1997.
121. Dursun N, Taskin E, Ozturk F. Protection against adriamycin-induced cardiomyopathy by carnosine in rats: role of endogenous antioxidants. Biol Trace Elem Res 143:
412– 424, 2011.
122. Dutka TL, Lamb GD. Effect of carnosine on excitation-contraction coupling in mechanically-skinned rat skeletal muscle. J Muscle Res Cell Motil 25: 203–213, 2004.
123. Dutka TL, Lamboley CR, McKenna MJ, Murphy RM, Lamb GD. Effects of carnosine on
contractile apparatus Ca2⫹ sensitivity and sarcoplasmic reticulum Ca2⫹ release in
human skeletal muscle fibers. J Appl Physiol 112: 728 –736, 2012.
124. Egorov SY, Kurella EG, Boldyrev AA, Krasnovsky AA Jr. Quenching of singlet molecular oxygen by carnosine and related antioxidants. Monitoring 1270-nm phosphorescence in aqueous media. Biochem Mol Biol Int 41: 687– 694, 1997.
125. Everaert I, De Naeyer H, Taes Y, Derave W. Gene expression of carnosine-related
enzymes and transporters in skeletal muscle. Eur J Appl Physiol 113: 1169 –1179, 2013.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
126. Everaert I, Mooyaart A, Baguet A, Zutinic A, Baelde H, Achten E, Taes Y, De Heer E,
Derave W. Vegetarianism, female gender and increasing age, but not CNDP1 genotype, are associated with reduced muscle carnosine levels in humans. Amino Acids 40:
1221–1229, 2010.
127. Everaert I, Stegen S, Vanheel B, Taes Y, Derave W. Effect of beta-alanine and carnosine supplementation on muscle contractility in mice. Med Sci Sports Exerc 45: 43–51,
2013.
128. Everaert I, Taes Y, De Heer E, Baelde HJ, Zutinic A, Yard B, Sauerhofer S, Vanhee L,
Delanghe JR, Aldini G, Derave W. Low plasma carnosinase activity promotes carnosinemia following carnosine ingestion in humans. Am J Physiol Renal Physiol 302: F1537–
F1544, 2012.
129. Farkas E, Sovago I, Gergely A. Studies on transition-metal peptide complexes. 8.
Parent and mixed-ligand complexes of histidine-containing dipeptides. J Chem SocDalton Transactions 1545–1551, 1983.
130. Fitzpatrick DW, Fisher H. Carnosine, histidine, and wound healing. Surgery 91: 56 – 60,
1982.
131. Fleisher LD, Rassin DK, Wisniewski K, Salwen HR. Carnosinase deficiency: a new
variant with high residual activity. Pediatr Res 14: 269 –271, 1980.
132. Fleisher-Berkovich S, Abramovitch-Dahan C, Ben-Shabat S, Apte R, Beit-Yannai E.
Inhibitory effect of carnosine and N-acetyl carnosine on LPS-induced microglial oxidative stress and inflammation. Peptides 30: 1306 –1312, 2009.
133. Fontana M, Pinnen F, Lucente G, Pecci L. Prevention of peroxynitrite-dependent
damage by carnosine and related sulphonamido pseudodipeptides. Cell Mol Life Sci 59:
546 –551, 2002.
147. Gjessing LR, Sjaastad O. Letter: homocarnosinosis: a new metabolic disorder associated with spasticity and mental retardation. Lancet 2: 1028, 1974.
148. Goodall MC. Carnosine phosphates as phosphate donor in muscular contraction.
Nature 178: 539 –540, 1956.
149. Gopko AV, Kulibin AY, Semenova ML, Zakhidov ST. Carnosine-induced changes in
the development of spermatogenic epithelial cells in senescence accelerated SAM
mice. Bull Exp Biol Med 140: 235–237, 2005.
150. Gordon AM, Homsher E, Regnier M. Regulation of contraction in striated muscle.
Physiol Rev 80: 853–924, 2000.
151. Greene SM, Margolis FL, Grillo M, Fisher H. Enhanced carnosine (beta-alanyl-L-histidine) breakdown and histamine metabolism following treatment with compound
48/80. Eur J Pharmacol 99: 79 – 84, 1984.
152. Guiotto A, Calderan A, Ruzza P, Borin G. Carnosine and carnosine-related antioxidants: a review. Curr Med Chem 12: 2293–2315, 2005.
153. Gulewitch WS. Zur Kenntnis der Extraktstoffe der Muskeln. Uber die Konstitution
des Carnosine. Hoppe-Seil Ztschr Physiol Chem B73: S434 –S443, 1911.
154. Gulewitch WS, Amiradzibi S. Uber das carnosin, eine neue organische Base des
Fleischextraktes. Ber Deutsch Chem Ges B33: S1902–S1903, 1900.
155. Guney Y, Turkcu UO, Hicsonmez A, Andrieu MN, Guney HZ, Bilgihan A, Kurtman C.
Carnosine may reduce lung injury caused by radiation therapy. Med Hypotheses 66:
957–959, 2006.
156. Gutierrez A, Anderstam B, Alvestrand A. Amino acid concentration in the interstitium
of human skeletal muscle: a microdialysis study. Eur J Clin Invest 29: 947–952, 1999.
134. Fonteh AN, Harrington RJ, Tsai A, Liao P, Harrington MG. Free amino acid and
dipeptide changes in the body fluids from Alzheimer’s disease subjects. Amino Acids
32: 213–224, 2007.
157. Handa O, Yoshida N, Tanaka Y, Ueda M, Ishikawa T, Takagi T, Matsumoto N, Naito
Y, Yoshikawa T. Inhibitory effect of polaprezinc on the inflammatory response to
Helicobacter pylori. Can J Gastroenterol 16: 785–789, 2002.
135. Fouad AA, Morsy MA, Gomaa W. Protective effect of carnosine against cisplatininduced nephrotoxicity in mice. Environ Toxicol Pharmacol 25: 292–297, 2008.
158. Hanson T, Smith EL. Carnosinase: an enzyme of swine kidney. J Biol Chem 179:
789 – 801, 1949.
136. Freedman BI, Hicks PJ, Sale MM, Pierson ED, Langefeld CD, Rich SS, Xu J, McDonough C, Janssen B, Yard BA, van der Woude FJ, Bowden DW. A leucine repeat in
the carnosinase gene CNDP1 is associated with diabetic end-stage renal disease in
European Americans. Nephrol Dial Transplant 22: 1131–1135, 2007.
159. Harding J, Graziadei PP, Monti Graziadei GA, Margolis FL. Denervation in the primary
olfactory pathway of mice. IV. Biochemical and morphological evidence for neuronal
replacement following nerve section. Brain Res 132: 11–28, 1977.
137. Fu Q, Dai H, Hu W, Fan Y, Shen Y, Zhang W, Chen Z. Carnosine protects against
Abeta42-induced neurotoxicity in differentiated rat PC12 cells. Cell Mol Neurobiol 28:
307–316, 2008.
160. Harding J, Margolis FL. Denervation in the primary olfactory pathway of mice. III.
Effect on enzymes of carnosine metabolism. Brain Res 110: 351–360, 1976.
161. Harding JW, O’Fallon JV. The subcellular distribution of carnosine, carnosine synthetase, and carnosinase in mouse olfactory tissues. Brain Res 173: 99 –109, 1979.
138. Fuchs V, Jaeger KE, Wilhelm S, Rosenau F. The BapF protein from Pseudomonas
aeruginosa is a beta-peptidyl aminopeptidase. World J Microbiol Biotechnol 27: 713–
718, 2011.
162. Harris RC, Dunnett M, Greenhaff PL. Carnosine and taurine contents in individual
fibres of human vastus lateralis muscle. J Sports Sci 16: 639 – 643, 1998.
139. Fujii T, Takaoka M, Muraoka T, Kurata H, Tsuruoka N, Ono H, Kiso Y, Tanaka T,
Matsumura Y. Preventive effect of L-carnosine on ischemia/reperfusion-induced acute
renal failure in rats. Eur J Pharmacol 474: 261–267, 2003.
163. Harris RC, Jones G, Hill C, Kendrick IP, Boobis L, Kim CK, Kim HJ, Dang VH, Edge J,
Wise JA. The carnosine content in V Lateralis of vegetarians and omnivores (Abstract).
FASEB J 6: A944, 2007.
140. Fujii T, Takaoka M, Tsuruoka N, Kiso Y, Tanaka T, Matsumura Y. Dietary supplementation of L-carnosine prevents ischemia/reperfusion-induced renal injury in rats. Biol
Pharm Bull 28: 361–363, 2005.
164. Harris RC, Marlin DJ, Dunnett M, Snow DH, Hultman E. Muscle buffering capacity and
dipeptide content in the thoroughbred horse, greyhound dog and man. Comp Biochem
Physiol A Comp Physiol 97: 249 –251, 1990.
141. Furuta S, Toyama S, Miwa M, Itabashi T, Sano H, Yoneta T. Residence time of
polaprezinc (zinc L-carnosine complex) in the rat stomach and adhesiveness to ulcerous sites. Jpn J Pharmacol 67: 271–278, 1995.
165. Harris RC, Tallon MJ, Dunnett M, Boobis L, Coakley J, Kim HJ, Fallowfield JL, Hill CA,
Sale C, Wise JA. The absorption of orally supplied beta-alanine and its effect on muscle
carnosine synthesis in human vastus lateralis. Amino Acids 30: 279 –289, 2006.
142. Gallant S, Semyonova M, Yuneva M. Carnosine as a potential anti-senescence drug.
Biochemistry 65: 866 – 868, 2000.
166. Harris RC, Wise JA, Price KA, Kim HJ, Kim CK, Sale C. Determinants of muscle
carnosine content. Amino Acids 43: 5–12, 2012.
143. Gardner ML, Illingworth KM, Kelleher J, Wood D. Intestinal absorption of the intact
peptide carnosine in man, and comparison with intestinal permeability to lactulose. J
Physiol 439: 411– 422, 1991.
167. Hartman PE, Hartman Z, Ault KT. Scavenging of singlet molecular oxygen by imidazole compounds: high and sustained activities of carboxy terminal histidine dipeptides
and exceptional activity of imidazole-4-acetic acid. Photochem Photobiol 51: 59 – 66,
1990.
144. Gaunitz F, Hipkiss AR. Carnosine and cancer: a perspective. Amino Acids 43: 135–142,
2012.
145. Geueke B, Kohler HP. Bacterial beta-peptidyl aminopeptidases: on the hydrolytic
degradation of beta-peptides. Appl Microbiol Biotechnol 74: 1197–1204, 2007.
146. Gjessing LR, Lunde HA, Morkrid L, Lenney JF, Sjaastad O. Inborn errors of carnosine
and homocarnosine metabolism. J Neural Transm Suppl 29: 91–106, 1990.
168. Haug A, Rodbotten R, Mydland LT, Christophersen OA. Increased broiler muscle
carnosine and anserine following histidine supplementation of commercial broiler
feed concentrate. Acta Agricult Scand A Anim Sci 58: 71–77, 2008.
169. Herculano B, Tamura M, Ohba A, Shimatani M, Kutsuna N, Hisatsune T. beta-AlanylL-histidine rescues cognitive deficits caused by feeding a high fat diet in a transgenic
mouse model of Alzheimer’s disease. J Alzheimers Dis 33: 983–997, 2013.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1839
BOLDYREV, ALDINI, AND DERAVE
170. Hermanussen M, Gonder U, Stegemann D, Wesolowski M, Ulewicz-Magulska B,
Wartensleben H, Hoffmann GF. How much chicken is food? Questioning the definition of food by analyzing amino acid composition of modern convenience products.
Anthropol Anz 69: 57– 69, 2012.
171. Hill CA, Harris RC, Kim HJ, Harris BD, Sale C, Boobis LH, Kim CK, Wise JA. Influence
of beta-alanine supplementation on skeletal muscle carnosine concentrations and high
intensity cycling capacity. Amino Acids 32: 225–233, 2007.
172. Hill TL, Blikslager AT. Effect of a zinc L-carnosine compound on acid-induced injury in
canine gastric mucosa ex vivo. Am J Vet Res 73: 659 – 663, 2012.
173. Hipkiss AR. Glycation, ageing and carnosine: are carnivorous diets beneficial? Mech
Ageing Dev 126: 1034 –1039, 2005.
174. Hipkiss AR. Carnosine and its possible roles in nutrition and health. Adv Food Nutr Res
57: 87–154, 2009.
175. Hipkiss AR. On the enigma of carnosine’s anti-ageing actions. Exp Gerontol 44: 237–
242, 2009.
176. Hipkiss AR, Brownson C. Carnosine reacts with protein carbonyl groups: another
possible role for the anti-ageing peptide? Biogerontology 1: 217–223, 2000.
177. Hipkiss AR, Brownson C, Bertani MF, Ruiz E, Ferro A. Reaction of carnosine with aged
proteins: another protective process? Ann NY Acad Sci 959: 285–294, 2002.
178. Hipkiss AR, Brownson C, Carrier MJ. Carnosine, the anti-ageing, anti-oxidant dipeptide, may react with protein carbonyl groups. Mech Ageing Dev 122: 1431–1445, 2001.
179. Hipkiss AR, Chana H. Carnosine protects proteins against methylglyoxal-mediated
modifications. Biochem Biophys Res Commun 248: 28 –32, 1998.
180. Hipkiss AR, Michaelis J, Syrris P. Non-enzymatic glycosylation of the dipeptide L-carnosine, a potential anti-protein-cross-linking agent. FEBS Lett 371: 81– 85, 1995.
194. Iovine B, Iannella ML, Nocella F, Pricolo MR, Bevilacqua MA. Carnosine inhibits
KRAS-mediated HCT116 proliferation by affecting ATP and ROS production. Cancer
Lett 315: 122–128, 2012.
195. Ishihara R, Iishi H, Sakai N, Yano H, Uedo N, Narahara H, Iseki K, Mikuni T, Ishiguro
S, Tatsuta M. Polaprezinc attenuates Helicobacter pylori-associated gastritis in Mongolian gerbils. Helicobacter 7: 384 –389, 2002.
196. Jackson MC, Kucera CM, Lenney JF. Purification and properties of human serum
carnosinase. Clin Chim Acta 196: 193–205, 1991.
197. Janssen B, Hohenadel D, Brinkkoetter P, Peters V, Rind N, Fischer C, Rychlik I, Cerna
M, Romzova M, de HE, Baelde H, Bakker SJ, Zirie M, Rondeau E, Mathieson P, Saleem
MA, Meyer J, Koppel H, Sauerhoefer S, Bartram CR, Nawroth P, Hammes HP, Yard
BA, Zschocke J, van der Woude FJ. Carnosine as a protective factor in diabetic
nephropathy: association with a leucine repeat of the carnosinase gene CNDP1.
Diabetes 54: 2320 –2327, 2005.
198. Jappar D, Hu Y, Keep RF, Smith DE. Transport mechanisms of carnosine in SKPT cells:
contribution of apical and basolateral membrane transporters. Pharm Res 26: 172–
181, 2009.
199. Johnson HA, Devaney KO. Thermal instability in the endoplasmic reticulum of the rat
hepatocyte. Virchows Arch B Cell Pathol Incl Mol Pathol 40: 357–364, 1982.
200. Johnson P, Hammer JL. Histidine dipeptide levels in ageing and hypertensive rat
skeletal and cardiac muscles. Comp Biochem Physiol B Comp Biochem 103: 981–984,
1992.
201. Jones G, Smith M, Harris R. Imidazole dipeptide content of dietary sources commonly
consumed within the British diet. Proc Nutr Soc 70: E363, 2011.
202. Jones G, Smith M, Harris R. Imidazole dipeptide content of dietary sources commonly
consumed within the British diet (Abstract). Proc Nutr Soc 70: E363, 2011.
181. Hipkiss AR, Michaelis J, Syrris P, Kumar S, Lam Y. Carnosine protects proteins against
in vitro glycation and cross-linking. Biochem Soc Trans 22: 399S, 1994.
203. Jones NR. The free amino acids of fish; 1-methylhistidine and beta-alanine liberation by skeletal muscle anserinase of codling (Gadus callarias). Biochem J 60:
81– 87, 1955.
182. Hipkiss AR, Preston JE, Himsworth DT, Worthington VC, Keown M, Michaelis J,
Lawrence J, Mateen A, Allende L, Eagles PA, Abbott NJ. Pluripotent protective effects
of carnosine, a naturally occurring dipeptide. Ann NY Acad Sci 854: 37–53, 1998.
204. Junge W, McLaughlin S. The role of fixed and mobile buffers in the kinetics of proton
movement. Biochim Biophys Acta 890: 1–5, 1987.
183. Hipkiss AR, Preston JE, Himswoth DT, Worthington VC, Abbot NJ. Protective effects
of carnosine against malondialdehyde-induced toxicity towards cultured rat brain
endothelial cells. Neurosci Lett 238: 135–138, 1997.
184. Hipkiss AR, Worthington VC, Himsworth DT, Herwig W. Protective effects of carnosine against protein modification mediated by malondialdehyde and hypochlorite.
Biochim Biophys Acta 1380: 46 –54, 1998.
185. Hobson RM, Saunders B, Ball G, Harris RC, Sale C. Effects of beta-alanine supplementation on exercise performance: a meta-analysis. Amino Acids 2012.
186. Hoffmann AM, Bakardjiev A, Bauer K. Carnosine-synthesis in cultures of rat glial cells
is restricted to oligodendrocytes and carnosine uptake to astrocytes. Neurosci Lett
215: 29 –32, 1996.
187. Holliday R, McFarland GA. Inhibition of the growth of transformed and neoplastic cells
by the dipeptide carnosine. Br J Cancer 73: 966 –971, 1996.
188. Horii Y, Shen J, Fujisaki Y, Yoshida K, Nagai K. Effects of L-carnosine on splenic
sympathetic nerve activity and tumor proliferation. Neurosci Lett 510: 1–5, 2012.
189. Horinishi H, Grillo M, Margolis FL. Purification and characterization of carnosine
synthetase from mouse olfactory bulbs. J Neurochem 31: 909 –919, 1978.
190. Horning MS, Blakemore LJ, Trombley PQ. Endogenous mechanisms of neuroprotection: role of zinc, copper, and carnosine. Brain Res 852: 56 – 61, 2000.
191. Hultman E, Sahlin K. Acid-base balance during exercise. Exercise Sports Sci Rev 8:
41–128, 1980.
192. Ibrahim W, Tatumi V, Yeh CC, Hong CB, Chow CK. Effects of dietary carnosine and
vitamin E on antioxidant and oxidative status of rats. Int J Vitam Nutr Res 78: 230 –237,
2008.
193. Imanari M, Higuchi M, Shiba N, Watanabe A. Accurate analysis of taurine, anserine,
carnosine and free amino acids in a cattle muscle biopsy sample. Anim Sci J 81: 369 –
376, 2010.
1840
205. Kalyankar GD, Meister A. Enzymatic synthesis of carnosine and related beta-alanyl
and gamma-aminobutyryl peptides. J Biol Chem 234: 3210 –3218, 1959.
206. Kamal MA, Jiang H, Hu Y, Keep RF, Smith DE. Influence of genetic knockout of Pept2
on the in vivo disposition of endogenous and exogenous carnosine in wild-type and
Pept2 null mice. Am J Physiol Regul Integr Comp Physiol 296: R986 –R991, 2009.
207. Kamei J, Ohsawa M, Miyata S, Tanaka S. Preventive effect of L-carnosine on changes
in the thermal nociceptive threshold in streptozotocin-induced diabetic mice. Eur J
Pharmacol 600: 83– 86, 2008.
208. Kang JH, Kim KS. Enhanced oligomerization of the alpha-synuclein mutant by the
Cu,Zn-superoxide dismutase and hydrogen peroxide system. Mol Cells 15: 87–93,
2003.
209. Kang JH, Kim KS, Choi SY, Kwon HY, Won MH, Kang TC. Protective effects of
carnosine, homocarnosine and anserine against peroxyl radical-mediated Cu,Zn-superoxide dismutase modification. Biochim Biophys Acta 1570: 89 –96, 2002.
210. Karton A, O’Reilly RJ, Pattison DI, Davies MJ, Radom L. Computational design of
effective, bioinspired HOCl antioxidants: the role of intramolecular Cl⫹ and H⫹
shifts. J Am Chem Soc 134: 19240 –19245, 2012.
211. Kato S, Tanaka A, Ogawa Y, Kanatsu K, Seto K, Yoneda T, Takeuchi K. Effect of
polaprezinc on impaired healing of chronic gastric ulcers in adjuvant-induced
arthritic rats–role of insulin-like growth factors (IGF)-1. Med Sci Monit 7: 20 –25,
2001.
212. Kendrick IP, Harris RC, Kim HJ, Kim CK, Dang VH, Lam TQ, Bui TT, Smith M, Wise
JA. The effects of 10 weeks of resistance training combined with beta-alanine supplementation on whole body strength, force production, muscular endurance and body
composition. Amino Acids 547–554, 2008.
213. Kendrick IP, Kim HJ, Harris RC, Kim CK, Dang VH, Lam TQ, Bui TT, Wise JA. The
effect of 4 weeks beta-alanine supplementation and isokinetic training on carnosine
concentrations in type I and II human skeletal muscle fibres. Eur J Appl Physiol 106:
131–138, 2009.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
214. Kim KS, Choi SY, Kwon HY, Won MH, Kang TC, Kang JH. The ceruloplasmin and
hydrogen peroxide system induces alpha-synuclein aggregation in vitro. Biochimie 84:
625– 631, 2002.
234. Liu H, Liu Y, Zhou Y, Peng S. Comparison between carnosine and pirenoxine in
prevention and treatment of rat with diabetic cataract. Int J Opthalmol 8: 1566 –1567,
2008.
215. Kim MY, Kim EJ, Kim YN, Choi C, Lee BH. Effects of alpha-lipoic acid and L-carnosine
supplementation on antioxidant activities and lipid profiles in rats. Nutr Res Pract 5:
421– 428, 2011.
235. Liu P, Ge X, Ding H, Jiang H, Christensen BM, Li J. Role of glutamate decarboxylaselike protein 1 (GADL1) in taurine biosynthesis. J Biol Chem 287: 40898 – 40906, 2012.
216. Kish SJ, Perry TL, Hansen S. Regional distribution of homocarnosine, homocarnosinecarnosine synthetase and homocarnosinase in human brain. J Neurochem 32: 1629 –
1636, 1979.
217. Klebanov GI, Teselkin Y, Babenkova IV, Popov IN, Levin G, Tyulina OV, Boldyrev AA,
Vladimirov Y. Evidence for a direct interaction of superoxide anion radical with carnosine. Biochem Mol Biol Int 43: 99 –106, 1997.
218. Ko JK, Leung CC. Ginger extract and polaprezinc exert gastroprotective actions by
anti-oxidant and growth factor modulating effects in rats. J Gastroenterol Hepatol 25:
1861–1868, 2010.
219. Kohen R, Yamamoto Y, Cundy KC, Ames BN. Antioxidant activity of carnosine,
homocarnosine, and anserine present in muscle and brain. Proc Natl Acad Sci USA 85:
3175–3179, 1988.
220. Kopec W, Jamroz D, Wiliczkiewicz A, Biazik E, Hikawczuk T, Skiba T, Pudlo A, Orda
J. Antioxidation status and histidine dipeptides content in broiler blood and muscles
depending on protein sources in feed. J Anim Physiol Anim Nutr 2012.
221. Koppel H, Riedl E, Braunagel M, Sauerhoefer S, Ehnert S, Godoy P, Sternik P, Dooley
S, Yard BA. L-Carnosine inhibits high-glucose-mediated matrix accumulation in human
mesangial cells by interfering with TGF-beta production and signalling. Nephrol Dial
Transplant 26: 3852–3858, 2011.
222. Korolkiewicz RP, Fujita A, Seto K, Suzuki K, Takeuchi K. Polaprezinc exerts a salutary
effect on impaired healing of acute gastric lesions in diabetic rats. Dig Dis Sci 45:
1200 –1209, 2000.
223. Kotamraju S, Matalon S, Matsunaga T, Shang T, Hickman-Davis JM, Kalyanaraman B.
Upregulation of immunoproteasomes by nitric oxide: potential antioxidative mechanism in endothelial cells. Free Radic Biol Med 40: 1034 –1044, 2006.
224. Kubomura D, Matahira Y, Nagai K, Niijima A. Effect of anserine ingestion on hyperglycemia and the autonomic nerves in rats and humans. Nutr Neurosci 13: 183–188,
2010.
236. Liu Q, Sikand P, Ma C, Tang Z, Han L, Li Z, Sun S, Lamotte RH, Dong X. Mechanisms
of itch evoked by beta-alanine. J Neurosci 32: 14532–14537, 2012.
237. Liu WH, Liu TC, Yin MC. Beneficial effects of histidine and carnosine on ethanolinduced chronic liver injury. Food Chem Toxicol 46: 1503–1509, 2008.
238. Liu Y, Xu G, Sayre LM. Carnosine inhibits (E)-4-hydroxy-2-nonenal-induced protein
cross-linking: structural characterization of carnosine-HNE adducts. Chem Res Toxicol
16: 1589 –1597, 2003.
239. Liu YF, Liu HW, Peng SL. [Effects of L-carnosine in preventing and treating rat cataract
induced by sodium selenite]. Zhonghua Yan Ke Za Zhi 45: 533–536, 2009.
240. Lunde HA, Gjessing LR, Sjaastad O. Homocarnosinosis: influence of dietary restriction
of histidine. Neurochem Res 11: 825– 838, 1986.
241. Ma J, Xiong JY, Hou WW, Yan HJ, Sun Y, Huang SW, Jin L, Wang Y, Hu WW, Chen Z.
Protective effect of carnosine on subcortical ischemic vascular dementia in mice. CNS
Neurosci Ther 18: 745–753, 2012.
242. Ma XY, Jiang ZY, Lin YC, Zheng CT, Zhou GL. Dietary supplementation with carnosine improves antioxidant capacity and meat quality of finishing pigs. J Anim Physiol Anim
Nutr 94: e286 – e295, 2010.
243. Mannion AF, Jakeman PM, Dunnett M, Harris RC, Willan PL. Carnosine and anserine
concentrations in the quadriceps femoris muscle of healthy humans. Eur J Appl Physiol
Occup Physiol 64: 47–50, 1992.
244. Mannion AF, Jakeman PM, Willan PL. Effects of isokinetic training of the knee extensors on high-intensity exercise performance and skeletal muscle buffering. Eur J Appl
Physiol Occup Physiol 68: 356 –361, 1994.
245. Mannion AF, Jakeman PM, Willan PL. Skeletal muscle buffer value, fibre type distribution and high intensity exercise performance in man. Exp Physiol 80: 89 –101, 1995.
246. Margolis FL. Carnosine in the primary olfactory pathway. Science 184: 909 –911,
1974.
225. Kurashige M, Imamura M, Araki SI, Suzuki D, Babazono T, Uzu T, Umezono T,
Toyoda M, Kawai K, Imanishi M, Hanaoka K, Maegawa H, Uchigata Y, Hosoya T,
Maeda S. The influence of a single nucleotide polymorphism within CNDP1 on susceptibility to diabetic nephropathy in Japanese women with Type 2 diabetes. Plos One
8: 2013.
247. Margolis FL, Grillo M. Axoplasmic-transport of carnosine (beta-alanyl-L-histidine) in
mouse olfactory pathway. Neurochem Res 2: 507–519, 1977.
226. Kurata H, Fujii T, Tsutsui H, Katayama T, Ohkita M, Takaoka M, Tsuruoka N, Kiso Y,
Ohno Y, Fujisawa Y, Shokoji T, Nishiyama A, Abe Y, Matsumura Y. Renoprotective
effects of L-carnosine on ischemia/reperfusion-induced renal injury in rats. J Pharmacol
Exp Ther 319: 640 – 647, 2006.
249. Marusic N, Aristoy MC, Toldra F. Nutritional pork meat compounds as affected by
ham dry-curing. Meat Sci 93: 53– 60, 2013.
227. Lamont C, Miller DJ. Calcium sensitizing action of carnosine and other endogenous
imidazoles in chemically skinned striated muscle. J Physiol 454: 421– 434, 1992.
228. Lee YT, Hsu CC, Lin MH, Liu KS, Yin MC. Histidine and carnosine delay diabetic
deterioration in mice and protect human low density lipoprotein against oxidation and
glycation. Eur J Pharmacol 513: 145–150, 2005.
229. Lenney JF. Separation and characterization of two carnosine-splitting cytosolic dipeptidases from hog kidney (carnosinase and non-specific dipeptidase). Biol Chem Hoppe
Seyler 371: 433– 440, 1990.
230. Lenney JF, George RP, Weiss AM, Kucera CM, Chan PW, Rinzler GS. Human serum
carnosinase: characterization, distinction from cellular carnosinase, and activation by
cadmium. Clin Chim Acta 123: 221–231, 1982.
231. Lenney JF, Peppers SC, Kucera CM, Sjaastad O. Homocarnosinosis: lack of serum
carnosinase is the defect probably responsible for elevated brain and CSF homocarnosine. Clin Chim Acta 132: 157–165, 1983.
248. Margolis FL, Grillo M, Hempstead J, Morgan JI. Monoclonal antibodies to mammalian
carnosine synthetase. J Neurochem 48: 593– 600, 1987.
250. Matsukura T, Tanaka H. Applicability of zinc complex of L-carnosine for medical use.
Biochemistry 65: 817– 823, 2000.
251. McDonough CW, Hicks PJ, Lu L, Langefeld CD, Freedman BI, Bowden DW. The
influence of carnosinase gene polymorphisms on diabetic nephropathy risk in AfricanAmericans. Hum Genet 126: 265–275, 2009.
252. McFarland GA, Holliday R. Retardation of the senescence of cultured human diploid
fibroblasts by carnosine. Exp Cell Res 212: 167–175, 1994.
253. McFarland GA, Holliday R. Further evidence for the rejuvenating effects of the dipeptide L-carnosine on cultured human diploid fibroblasts. Exp Gerontol 34: 35– 45, 1999.
254. McManus IR. Enzymatic synthesis of anserine in skeletal muscle by N-methylation of
carnosine. J Biol Chem 237: 1207–1211, 1962.
255. Miller DJ, O’Dowd A. Vascular smooth muscle actions of carnosine as its zinc complex
are mediated by histamine H(1) and H(2) receptors. Biochemistry 65: 798 – 806, 2000.
232. Li YF, He RR, Tsoi B, Kurihara H. Bioactivities of chicken essence. J Food Sci 77:
R105–R110, 2012.
256. Mineo P, Vitalini D, La MD, Rizzarelli E, Scamporrino E, Vecchio G. Electrospray mass
spectrometric studies of L-carnosine (beta-alanyl-L-histidine) complexes with copper(II) or zinc ions in aqueous solution. Rapid Commun Mass Spectrom 16: 722–729,
2002.
233. Lin H, King N. Demonstration of functional dipeptide transport with expression of
PEPT2 in guinea pig cardiomyocytes. Pflügers Arch 453: 915–922, 2007.
257. Ming X, Stein TP, Barnes V, Rhodes N, Guo L. Metabolic perturbance in autism
spectrum disorders: a metabolomics study. J Proteome Res 11: 5856 –5862, 2012.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1841
BOLDYREV, ALDINI, AND DERAVE
258. Mishima T, Yamada T, Sakamoto M, Sugiyama M, Matsunaga S, Maemura H, Shimizu
M, Takahata Y, Morimatsu F, Wada M. Chicken breast attenuates high-intensityexercise-induced decrease in rat sarcoplasmic reticulum Ca2⫹ handling. Int J Sport
Nutr Exerc Metab 18: 399 – 411, 2008.
259. Mooyaart A. Diabetic Nephropathy: Pathology, Genetics and Carnosine Metabolism
(PhD thesis). Leiden, The Netherlands: Leiden University Medical Center, 2011.
260. Mooyaart AL, Zutinic A, Bakker SJ, Grootendorst DC, Kleefstra N, van Valkengoed
IG, Bohringer S, Bilo HJ, Dekker FW, Bruijn JA, Navis G, Janssen B, Baelde HJ, de HE.
Association between CNDP1 genotype and diabetic nephropathy is sex specific.
Diabetes 59: 1555–1559, 2010.
261. Mora L, Sentandreu MA, Toldra F. Contents of creatine, creatinine and carnosine in
porcine muscles of different metabolic types. Meat Sci 79: 709 –715, 2008.
262. Nagai K, Niijima A, Yamano T, Otani H, Okumra N, Tsuruoka N, Nakai M, Kiso Y.
Possible role of L-carnosine in the regulation of blood glucose through controlling
autonomic nerves. Exp Biol Med 228: 1138 –1145, 2003.
263. Nagai K, Suda T. [Antineoplastic effects of carnosine and beta-alanine–physiological
considerations of its antineoplastic effects]. Nihon Seirigaku Zasshi 48: 741–747, 1986.
264. Nagai K, Suda T, Kawasaki K, Mathuura S. Action of carnosine and beta-alanine on
wound healing. Surgery 100: 815– 821, 1986.
265. Nagai K, Tanida M, Niijima A, Tsuruoka N, Kiso Y, Horii Y, Shen J, Okumura N. Role
of L-carnosine in the control of blood glucose, blood pressure, thermogenesis, and
lipolysis by autonomic nerves in rats: involvement of the circadian clock and histamine.
Amino Acids 43: 97–109, 2012.
266. Nagai R, Murray DB, Metz TO, Baynes JW. Chelation: a fundamental mechanism of
action of AGE inhibitors, AGE breakers, and other inhibitors of diabetes complications. Diabetes 61: 549 –559, 2012.
267. Nagasawa T, Yonekura T, Nishizawa N, Kitts DD. In vitro and in vivo inhibition of
muscle lipid and protein oxidation by carnosine. Mol Cell Biochem 225: 29 –34, 2001.
268. Negre-Salvayre A, Coatrieux C, Ingueneau C, Salvayre R. Advanced lipid peroxidation
end products in oxidative damage to proteins. Potential role in diseases and therapeutic prospects for the inhibitors. Br J Pharmacol 153: 6 –20, 2008.
269. Ng RH, Marshall FD. Regional and subcellular distribution of homocarnosine-carnosine synthetase in the central nervous system of rats. J Neurochem 30: I87–I90, 1978.
270. Ng RH, Marshall FD, Henn FA, Sellstrom A. Metabolism of carnosine and homocarnosine in subcellular fractions and neuronal and glial cell-enriched fractions of rabbit
brain. J Neurochem 28: 449 – 452, 1977.
271. Nicoletti VG, Santoro AM, Grasso G, Vagliasindi LI, Giuffrida ML, Cuppari C, Purrello
VS, Stella AM, Rizzarelli E. Carnosine interaction with nitric oxide and astroglial cell
protection. J Neurosci Res 85: 2239 –2245, 2007.
272. Niijima A, Okui T, Matsumura Y, Yamano T, Tsuruoka N, Kiso Y, Nagai K. Effects of
L-carnosine on renal sympathetic nerve activity and DOCA-salt hypertension in rats.
Auton Neurosci 97: 99 –102, 2002.
273. Nishiwaki H, Kato S, Sugamoto S, Umeda M, Morita H, Yoneta T, Takeuchi K.
Ulcerogenic and healing impairing actions of monochloramine in rat stomachs: effects
of zinc L-carnosine, polaprezinc. J Physiol Pharmacol 50: 183–195, 1999.
274. Noori S, Mahboob T. Antioxidant effect of carnosine pretreatment on cisplatin-induced renal oxidative stress in rats. Indian J Clin Biochem 25: 86 –91, 2010.
275. Nordsborg N, Mohr M, Pedersen LD, Nielsen JJ, Langberg H, Bangsbo J. Muscle
interstitial potassium kinetics during intense exhaustive exercise: effect of previous
arm exercise. Am J Physiol Regul Integr Comp Physiol 285: R143–R148, 2003.
276. Numata Y, Terui T, Okuyama R, Hirasawa N, Sugiura Y, Miyoshi I, Watanabe T,
Kuramasu A, Tagami H, Ohtsu H. The accelerating effect of histamine on the cutaneous wound-healing process through the action of basic fibroblast growth factor. J
Invest Dermatol 126: 1403–1409, 2006.
279. O’Dowd A, O’Dowd JJ, O’Dowd JJ, MacFarlane N, Abe H, Miller DJ. Analysis of novel
imidazoles from isolated perfused rabbit heart by two high-performance liquid chromatographic methods. J Chromatogr 577: 347–353, 1992.
280. O’Dowd JJ, Robins DJ, Miller DJ. Detection, characterisation, and quantification of
carnosine and other histidyl derivatives in cardiac and skeletal muscle. Biochim Biophys
Acta 967: 241–249, 1988.
281. Okuma E, Abe H. Major buffering constituents in animal muscle. Comp Biochem Physiol
B Comp Physiol 102: 37– 41, 1992.
282. Orioli M, Aldini G, Benfatto MC, Facino RM, Carini M. HNE Michael adducts to
histidine and histidine-containing peptides as biomarkers of lipid-derived carbonyl
stress in urines: LC-MS/MS profiling in Zucker obese rats. Anal Chem 79: 9174 –9184,
2007.
283. Orioli M, Aldini G, Beretta G, Facino RM, Carini M. LC-ESI-MS/MS determination of
4-hydroxy-trans-2-nonenal Michael adducts with cysteine and histidine-containing
peptides as early markers of oxidative stress in excitable tissues. J Chromatogr B Analyt
Technol Biomed Life Sci 827: 109 –118, 2005.
284. Ozel TU, Bilgihan A, Biberoglu G, Mertoglu CO. Carnosine supplementation protects
rat brain tissue against ethanol-induced oxidative stress. Mol Cell Biochem 339: 55– 61,
2010.
285. Pamplona R. Advanced lipoxidation end-products. Chem Biol Interact 192: 14 –20,
2011.
286. Pan JW, Hamm JR, Rothman DL, Shulman RG. Intracellular pH in human skeletal
muscle by 1H NMR. Proc Natl Acad Sci USA 85: 7836 –7839, 1988.
287. Pandya V, Ekka MK, Dutta RK, Kumaran S. Mass spectrometry assay for studying
kinetic properties of dipeptidases: characterization of human and yeast dipeptidases.
Anal Biochem 418: 134 –142, 2011.
288. Panzanelli P, Cantino D, Sassoe-Pognetto M. Co-localization of carnosine and glutamate in photoreceptors and bipolar cells of the frog retina. Brain Res 758: 143–152,
1997.
289. Parkhouse WS, McKenzie DC, Hochachka PW, Ovalle WK. Buffering capacity of
deproteinized human vastus lateralis muscle. J Appl Physiol 58: 14 –17, 1985.
290. Pattison DI, Davies MJ. Evidence for rapid inter- and intramolecular chlorine transfer
reactions of histamine and carnosine chloramines: implications for the prevention of
hypochlorous-acid-mediated damage. Biochemistry 45: 8152– 8162, 2006.
291. Pavlov AR, Revina AA, Dupin AM, Boldyrev AA, Yaropolov AI. The mechanism of
interaction of carnosine with superoxide radicals in water solutions. Biochim Biophys
Acta 1157: 304 –312, 1993.
292. Pedersen TH, Nielsen OB, Lamb GD, Stephenson DG. Intracellular acidosis enhances
the excitability of working muscle. Science 305: 1144 –1147, 2004.
293. Pegova A, Abe H, Boldyrev A. Hydrolysis of carnosine and related compounds by
mammalian carnosinases. Comp Biochem Physiol B Biochem Mol Biol 127: 443– 446,
2000.
294. Penafiel R, Ruzafa C, Monserrat F, Cremades A. Gender-related differences in carnosine, anserine and lysine content of murine skeletal muscle. Amino Acids 26: 53–58,
2004.
295. Pepper ED, Farrell MJ, Nord G, Finkel SE. Antiglycation effects of carnosine and other
compounds on the long-term survival of Escherichia coli. Appl Environ Microbiol 76:
7925–7930, 2010.
296. Perel’man MI, Kornilova ZK, Paukov VS, Boikov AK, Priimak AA. [The effect of
carnosine on the healing of a lung wound]. Biull Eksp Biol Med 108: 352–356, 1989.
297. Perry TL, Hansen S, Love DL. Serum-carnosinase deficiency in carnosinaemia. Lancet
1: 1229 –1230, 1968.
277. O’Dowd A, Miller DJ. Analysis of an H1 receptor-mediated, zinc-potentiated vasoconstrictor action of the histidyl dipeptide carnosine in rabbit saphenous vein. Br J
Pharmacol 125: 1272–1280, 1998.
298. Perry TL, Hansen S, Tischler B, Bunting R, Berry Carnosinemia K. A new metabolic
disorder associated with neurologic disease and mental defect. N Engl J Med 277:
1219 –1227, 1967.
278. O’Dowd A, O’Dowd JJ, Miller DJ. The dipeptide carnosine constricts rabbit saphenous vein as a zinc complex apparently via a serotonergic receptor. J Physiol 495:
535–543, 1996.
299. Perry TL, Kish SJ, Sjaastad O, Gjessing LR, Nesbakken R, Schrader H, Loken AC.
Homocarnosinosis: increased content of homocarnosine and deficiency of homocarnosinase in brain. J Neurochem 32: 1637–1640, 1979.
1842
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
300. Peters V, Jansen EE, Jakobs C, Riedl E, Janssen B, Yard BA, Wedel J, Hoffmann GF,
Zschocke J, Gotthardt D, Fischer C, Koppel H. Anserine inhibits carnosine degradation but in human serum carnosinase (CN1) is not correlated with histidine dipeptide
concentration. Clin Chim Acta 412: 263–267, 2011.
301. Peters V, Kebbewar M, Jansen EW, Jakobs C, Riedl E, Koeppel H, Frey D, Adelmann
K, Klingbeil K, Mack M, Hoffmann GF, Janssen B, Zschocke J, Yard BA. Relevance of
allosteric conformations and homocarnosine concentration on carnosinase activity.
Amino Acids 38: 1607–1615, 2010.
302. Peterszegi G, Molinari J, Ravelojaona V, Robert L. Effect of advanced glycation endproducts on cell proliferation and cell death. Pathol Biol 54: 396 – 404, 2006.
303. Petroff OA, Mattson RH, Behar KL, Hyder F, Rothman DL. Vigabatrin increases
human brain homocarnosine and improves seizure control. Ann Neurol 44: 948 –952,
1998.
304. Pfister F, Riedl E, Wang Q, vom HF, Deinzer M, Harmsen MC, Molema G, Yard B,
Feng Y, Hammes HP. Oral carnosine supplementation prevents vascular damage in
experimental diabetic retinopathy. Cell Physiol Biochem 28: 125–136, 2011.
305. Pietkiewicz J, Bronowicka-Szydelko A, Dzierzba K, Danielewicz R, Gamian A. Glycation of the muscle-specific enolase by reactive carbonyls: effect of temperature and
the protection role of carnosine, pyridoxamine and phosphatidylserine. Protein J 30:
149 –158, 2011.
306. Preston JE, Hipkiss AR, Himsworth DT, Romero IA, Abbott JN. Toxic effects of
beta-amyloid(25–35) on immortalised rat brain endothelial cell: protection by carnosine, homocarnosine and beta-alanine. Neurosci Lett 242: 105–108, 1998.
307. Purchas RW, Busboom JR. The effect of production system and age on levels of iron,
taurine, carnosine, coenzyme Q10, and creatine in beef muscles and liver. Meat Sci 70:
589 –596, 2005.
308. Purchas RW, Rutherfurd SM, Pearce PD, Vather R, Wilkinson BH. Cooking temperature effects on the forms of iron and levels of several other compounds in beef
semitendinosus muscle. Meat Sci 68: 201–207, 2004.
309. Purchas RW, Triumf EC, Egelandsdal B. Quality characteristics and composition of the
longissimus muscle in the short-loin from male and female farmed red deer in New
Zealand. Meat Sci 86: 505–510, 2010.
310. Raghavan M, Lindberg U, Schutt C. The use of alternative substrates in the characterization of actin-methylating and carnosine-methylating enzymes. Eur J Biochem 210:
311–318, 1992.
319. Roberts PR, Zaloga GP. Cardiovascular effects of carnosine. Biochemistry 65: 856 –
861, 2000.
320. Rubio-Aliaga I, Frey I, Boll M, Groneberg DA, Eichinger HM, Balling R, Daniel H.
Targeted disruption of the peptide transporter Pept2 gene in mice defines its physiological role in the kidney. Mol Cell Biol 23: 3247–3252, 2003.
321. Sakata K, Yamashita T, Maeda M, Moriyama Y, Shimada S, Tohyama M. Cloning of a
lymphatic peptide/histidine transporter. Biochem J 356: 53– 60, 2001.
322. Sale C, Saunders B, Harris RC. Effect of beta-alanine supplementation on muscle
carnosine concentrations and exercise performance. Amino Acids 39: 321–333,
2010.
323. Sanchez-Hernandez L, Marina ML, Crego AL. A capillary electrophoresis-tandem
mass spectrometry methodology for the determination of non-protein amino acids in
vegetable oils as novel markers for the detection of adulterations in olive oils. J
Chromatogr A 1218: 4944 – 4951, 2011.
324. Sassoe-Pognetto M, Cantino D, Panzanelli P, Verdun di CL, Giustetto M, Margolis FL,
De BS, Fasolo A. Presynaptic co-localization of carnosine and glutamate in olfactory
neurones. Neuroreport 5: 7–10, 1993.
325. Sato M, Karasawa N, Shimizu M, Morimatsu F, Yamada R. Safety evaluation of chicken
breast extract containing carnosine and anserine. Food Chem Toxicol 46: 480 – 489,
2008.
326. Sauerhofer S, Yuan G, Braun GS, Deinzer M, Neumaier M, Gretz N, Floege J, Kriz W,
van der WF, Moeller MJ. L-Carnosine, a substrate of carnosinase-1, influences glucose
metabolism. Diabetes 56: 2425–2432, 2007.
327. Schiaffino S, Reggiani C. Fiber types in mammalian skeletal muscles. Physiol Rev 91:
1447–1531, 2011.
328. Schonherr J. Analysis of products of animal origin in feeds by determination of carnosine and related dipeptides by high-performance liquid chromatography. J Agric
Food Chem 50: 1945–1950, 2002.
329. Schroder L, Schmitz CH, Bachert P. Carnosine as molecular probe for sensitive
detection of Cu(II) ions using localized (1)H NMR spectroscopy. J Inorg Biochem 102:
174 –183, 2008.
330. Seidler N, Shokry S, Nauth J. Properties of a glycation product derived from carnosine. J Biochem Mol Biol Biophys 5: 153–162, 2001.
331. Seidler NW. Carnosine prevents the glycation-induced changes in electrophoretic
mobility of aspartate aminotransferase. J Biochem Mol Toxicol 14: 215–220, 2000.
311. Rajanikant GK, Zemke D, Senut MC, Frenkel MB, Chen AF, Gupta R, Majid A. Carnosine is neuroprotective against permanent focal cerebral ischemia in mice. Stroke
38: 3023–3031, 2007.
332. Seidler NW, Yeargans GS, Morgan TG. Carnosine disaggregates glycated alpha-crystallin: an in vitro study. Arch Biochem Biophys 427: 110 –115, 2004.
312. Rashid I, van Reyk DM, Davies MJ. Carnosine and its constituents inhibit glycation of
low-density lipoproteins that promotes foam cell formation in vitro. FEBS Lett 581:
1067–1070, 2007.
333. Seto K, Yoneta T, Suda H, Tamaki H. Effect of polaprezinc (N-(3-aminopropionyl)-Lhistidinato zinc), a novel antiulcer agent containing zinc, on cellular proliferation: role
of insulin-like growth factor I. Biochem Pharmacol 58: 245–250, 1999.
313. Razenkov I, Derwies G, Severin S. Zur frage nach Carnosin-wirkung auf die magensaftsekretion. Ztschrift Phys Chem 162: 95–99, 1926.
334. Severin SE, Kirzon MV, Kaftanova TM. Effect of carnosine and anserine on action of
isolated frog muscles (article in Russian). Dokl Akad Nauk SSSR 91: 691– 694, 1953.
314. Renner C, Zemitzsch N, Fuchs B, Geiger KD, Hermes M, Hengstler J, Gebhardt R,
Meixensberger J, Gaunitz F. Carnosine retards tumor growth in vivo in an NIH3T3HER2/neu mouse model. Mol Cancer 9: 2, 2010.
335. Severina IS, Bussygina OG, Pyatakova NV. Carnosine as a regulator of soluble guanylate cyclase. Biochemistry 65: 783–788, 2000.
315. Riedl E, Koeppel H, Brinkkoetter P, Sternik P, Steinbeisser H, Sauerhoefer S, Janssen
B, van der Woude FJ, Yard BA. A CTG polymorphism in the CNDP1 gene determines
the secretion of serum carnosinase in Cos-7 transfected cells. Diabetes 56: 2410 –
2413, 2007.
316. Riedl E, Koeppel H, Pfister F, Peters V, Sauerhoefer S, Sternik P, Brinkkoetter P,
Zentgraf H, Navis G, Henning RH, van den Born J, Bakker SJ, Janssen B, van der
Woude FJ, Yard BA. N-glycosylation of carnosinase influences protein secretion and
enzyme activity: implications for hyperglycemia. Diabetes 59: 1984 –1990, 2010.
336. Sewell DA, Harris RC, Dunnett M. Carnosine accounts for most of the variation in
physico-chemical buffering in equine muscle. Equine Exercise Physiol 3: 276 –280,
1991.
337. Sewell DA, Harris RC, Marlin DJ, Dunnett M. Estimation of the carnosine content of
different fibre types in the middle gluteal muscle of the thoroughbred horse. J Physiol
455: 447– 453, 1992.
338. Shen Y, He P, Fan YY, Zhang JX, Yan HJ, Hu WW, Ohtsu H, Chen Z. Carnosine
protects against permanent cerebral ischemia in histidine decarboxylase knockout
mice by reducing glutamate excitotoxicity. Free Radic Biol Med 48: 727–735, 2010.
317. Riedl E, Pfister F, Braunagel M, Brinkkotter P, Sternik P, Deinzer M, Bakker SJ,
Henning RH, van den Born J, Kramer BK, Navis G, Hammes HP, Yard B, Koeppel H.
Carnosine prevents apoptosis of glomerular cells and podocyte loss in STZ diabetic
rats. Cell Physiol Biochem 28: 279 –288, 2011.
340. Skulachev VP. Membrane Bioenergetics. Berlin: Springer, 1988.
318. Ririe DG, Roberts PR, Shouse MN, Zaloga GP. Vasodilatory actions of the dietary
peptide carnosine. Nutrition 16: 168 –172, 2000.
341. Soliman K, Mohamed A, Metwally N. Attenuation of some metabolic deteriorations
induced by diabetes mellitus using carnosine. J Appl Sci 7: 2252–2260, 2007.
339. Skaper SD, Das S, Marshall FD. Some properties of a homocarnosine-carnosine synthetase isolated from rat brain. J Neurochem 21: 1429 –1445, 1973.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1843
BOLDYREV, ALDINI, AND DERAVE
342. Stegen S, Blancquaert L, Everaert I, Bex T, Taes Y, Calders P, Achten E, Derave W.
Meal and beta-alanine co-ingestion enhances muscle carnosine loading. Med Sci Sports
Exercise. 45: 1478 –1485, 2013.
363. Thomas S, Kotamraju S, Zielonka J, Harder DR, Kalyanaraman B. Hydrogen peroxide
induces nitric oxide and proteosome activity in endothelial cells: a bell-shaped signaling response. Free Radic Biol Med 42: 1049 –1061, 2007.
343. Stellingwerff T, Anwander H, Egger A, Buehler T, Kreis R, Decombaz J, Boesch C.
Effect of two beta-alanine dosing protocols on muscle carnosine synthesis and washout. Amino Acids 2011.
364. Tiedje KE, Stevens K, Barnes S, Weaver DF. Beta-alanine as a small molecule neurotransmitter. Neurochem Int 57: 177–188, 2010.
344. Stellingwerff T, Decombaz J, Harris RC, Boesch C. Optimizing human in vivo dosing
and delivery of beta-alanine supplements for muscle carnosine synthesis. Amino Acids
43: 57– 65, 2012.
345. Stenesh JJ, Winnick T. Carnosine-anserine synthetase of muscle. 4. Partial purification
of the enzyme and further studies of beta-alanyl peptide synthesis. Biochem J 77:
575–581, 1960.
346. Stuerenburg HJ, Kunze K. Concentrations of free carnosine (a putative membraneprotective antioxidant) in human muscle biopsies and rat muscles. Arch Gerontol Geriatr 29: 107–113, 1999.
365. Tinbergen BJ, Slump P. The detection of chicken meat in meat products by means of
the anserine/carnosine ratio. Z Lebensm Unters Forsch 161: 7–11, 1976.
366. Tolkatschewskaya NF. Zurr Kenntnis der Extraktstoffe der Muskeln. Uber des Extraktivstoffe des Hunherfleisherm. Hoppe-Seil Ztschr Physiol Chem B185: S32, 1929.
367. Tomonaga S, Hayakawa T, Yamane H, Maemura H, Sato M, Takahata Y, Morimatsu F,
Furuse M. Oral administration of chicken breast extract increases brain carnosine and
anserine concentrations in rats. Nutr Neurosci 10: 181–186, 2007.
368. Tomonaga S, Tachibana T, Takahashi H, Sato M, Denbow DM, Furuse M. Nitric oxide
involves in carnosine-induced hyperactivity in chicks. Eur J Pharmacol 524: 84 – 88,
2005.
347. Stvolinskii SL, Fedorova TN, Yuneva MO, Boldyrev AA. Protective effect of carnosine
on Cu,Zn-superoxide dismutase during impaired oxidative metabolism in the brain in
vivo. Bull Exp Biol Med 135: 130 –132, 2003.
369. Torreggiani A, Tamba M. Free radical scavenging and metal chelating activity of some
therapeutic heterocyclic agents. Trends Heterocyclic Chem 10: 115–137, 2005.
348. Suyama M, Maruyama M. Identification of methylated beta-alanylhistidine in the muscles of snake and dolphin. J Biochem 66: 405– 407, 1969.
370. Torreggiani A, Bonora S, Fini G. Raman and IR spectroscopic investigation of zinc(II)carnosine complexes. Biopolymers 57: 352–364, 2000.
349. Suzuki Y, Ito O, Mukai N, Takahashi H, Takamatsu K. High level of skeletal muscle
carnosine contributes to the latter half of exercise performance during 30-s maximal
cycle ergometer sprinting. Jpn J Physiol 52: 199 –205, 2002.
371. Torreggiani A, Tamba M, Fini G. Binding of copper(II) to carnosine: raman and IR
spectroscopic study. Biopolymers 57: 149 –159, 2000.
350. Suzuki Y, Ito O, Takahashi H, Takamatsu K. The effect of sprint training on skeletal
muscle carnosine in humans. Int J Sport Health Sci 2: 105–110, 2004.
351. Suzuki Y, Nakao T, Maemura H, Sato M, Kamahara K, Morimatsu F, Takamatsu K.
Carnosine and anserine ingestion enhances contribution of nonbicarbonate buffering.
Med Sci Sports Exercise 38: 334 –338, 2006.
352. Swietach P, Youm JB, Saegusa N, Leem CH, Spitzer KW, Vaughan-Jones RD. Coupled
Ca2⫹/H⫹ transport by cytoplasmic buffers regulates local Ca2⫹ and H⫹ ion signaling.
Proc Natl Acad Sci USA 110: E2064 –E2073, 2013.
353. Szwergold BS. Carnosine and anserine act as effective transglycating agents in decomposition of aldose-derived Schiff bases. Biochem Biophys Res Commun 336: 36 – 41,
2005.
354. Takahashi S, Nakashima Y, Toda K. Carnosine facilitates nitric oxide production in
endothelial f-2 cells. Biol Pharm Bull 32: 1836 –1839, 2009.
355. Tallon MJ, Harris RC, Boobis LH, Fallowfield JL, Wise JA. The carnosine content of
vastus lateralis is elevated in resistance-trained bodybuilders. J Strength Cond Res 19:
725–729, 2005.
356. Tallon MJ, Harris RC, Maffulli N, Tarnopolsky MA. Carnosine, taurine and enzyme
activities of human skeletal muscle fibres from elderly subjects with osteoarthritis and
young moderately active subjects. Biogerontology 8: 129 –137, 2007.
357. Tamaki N, Tsunemori F, Wakabayashi M, Hama T. Effect of histidine-free and -excess
diets on anserine and carnosine contents in rat gastrocnemius muscle. J Nutr Sci
Vitaminol 23: 331–340, 1977.
358. Tamba M, Torreggiani A. A pulse radiolysis study of carnosine in aqueous solution. Int
J Radiat Biol 74: 333–340, 1998.
359. Tamba M, Torreggiani A. Hydroxyl radical scavenging by carnosine and Cu(II)-carnosine complexes: a pulse-radiolysis and spectroscopic study. Int J Radiat Biol 75:
1177–1188, 1999.
360. Teufel M, Saudek V, Ledig JP, Bernhardt A, Boularand S, Carreau A, Cairns NJ, Carter
C, Cowley DJ, Duverger D, Ganzhorn AJ, Guenet C, Heintzelmann B, Laucher V,
Sauvage C, Smirnova T. Sequence identification and characterization of human carnosinase and a closely related non-specific dipeptidase. J Biol Chem 278: 6521– 6531,
2003.
372. Torreggiani A, Trinchero A, Tamba M, Fini G. Vibrational characterisation and biological activity of carnosine and its metal complexes. Ital J Biochem 52: 87–97, 2003.
373. Trombley PQ, Horning MS, Blakemore LJ. Carnosine modulates zinc and copper
effects on amino acid receptors and synaptic transmission. Neuroreport 9: 3503–3507,
1998.
374. Tsai SJ, Kuo WW, Liu WH, Yin MC. Antioxidative and anti-inflammatory protection
from carnosine in the striatum of MPTP-treated mice. J Agric Food Chem 2010.
375. Tsubone S, Yoshikawa N, Okada S, Abe H. Purification and characterization of a novel
imidazole dipeptide synthase from the muscle of the Japanese eel Anguilla japonica.
Comp Biochem Physiol B Biochem Mol Biol 146: 560 –567, 2007.
376. Unno H, Yamashita T, Ujita S, Okumura N, Otani H, Okumura A, Nagai K, Kusunoki
M. Structural basis for substrate recognition and hydrolysis by mouse carnosinase
CN2. J Biol Chem 283: 27289 –27299, 2008.
377. Van Heeswijk PJ, Trijbels JM, Schretlen ED, van Munster PJ, Monnens LA. A patient
with a deficiency of serum-carnosinase activity. Acta Paediatr Scand 58: 584 –592,
1969.
378. Van Thienen R, Van Proeyen K, Vanden Eynde B, Puype J, Lefere T, Hespel P.
Beta-alanine improves sprint performance in endurance cycling. Med Sci Sports Exerc
41: 898 –903, 2009.
379. Vardarli I, Baier LJ, Hanson RL, Akkoyun I, Fischer C, Rohmeiss P, Basci A, Bartram
CR, van der Woude FJ, Janssen B. Gene for susceptibility to diabetic nephropathy in
type 2 diabetes maps to 18q22.3–23. Kidney Int 62: 2176 –2183, 2002.
380. Vaughan-Jones RD, Spitzer KW, Swietach P. Spatial aspects of intracellular pH regulation in heart muscle. Prog Biophys Mol Biol 90: 207–224, 2006.
381. Vaughan-Jones RD, Spitzer KW, Swietach P. Intracellular pH regulation in heart. J Mol
Cell Cardiol 46: 318 –331, 2009.
382. Velez S, Nair NG, Reddy VP. Transition metal ion binding studies of carnosine and
histidine: biologically relevant antioxidants. Colloids Surf B Biointerfaces 66: 291–294,
2008.
383. Vistoli G, Carini M, Aldini G. Transforming dietary peptides in promising lead compounds: the case of bioavailable carnosine analogs. Amino Acids 43: 111–126, 2012.
361. Teuscher NS, Keep RF, Smith DE. PEPT2-mediated uptake of neuropeptides in rat
choroid plexus. Pharm Res 18: 807– 813, 2001.
384. Vistoli G, Straniero V, Pedretti A, Fumagalli L, Bolchi C, Pallavicini M, Valoti E, Testa
B. Predicting the physicochemical profile of diastereoisomeric histidine-containing
dipeptides by property space analysis. Chirality 24: 566 –576, 2012.
362. Teuscher NS, Shen H, Shu C, Xiang J, Keep RF, Smith DE. Carnosine uptake in rat
choroid plexus primary cell cultures and choroid plexus whole tissue from PEPT2 null
mice. J Neurochem 89: 375–382, 2004.
385. Vongerichten KF, Klein JR, Matern H, Plapp R. Cloning and nucleotide sequence
analysis of pepV, a carnosinase gene from Lactobacillus delbrueckii subsp. lactis DSM
7290, and partial characterization of the enzyme. Microbiology 140: 2591–2600, 1994.
1844
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
PHYSIOLOGY AND PATHOPHYSIOLOGY OF CARNOSINE
386. Wang JP, Yang ZT, Liu C, He YH, Zhao SS. L-Carnosine inhibits neuronal cell apoptosis
through signal transducer and activator of transcription 3 signaling pathway after acute
focal cerebral ischemia. Brain Res 1507: 125–133, 2013.
387. Wanic K, Placha G, Dunn J, Smiles A, Warram JH, Krolewski AS. Exclusion of polymorphisms in carnosinase genes (CNDP1 and CNDP2) as a cause of diabetic nephropathy in type 1 diabetes: results of large case-control and follow-up studies.
Diabetes 57: 2547–2551, 2008.
388. Watanabe S, Wang XE, Hirose M, Kivilioto T, Osada T, Miwa H, Oide H, Kitamura T,
Yoneta T, Seto K, Sato N. Insulin-like growth factor I plays a role in gastric wound
healing: evidence using a zinc derivative, polaprezinc, and an in vitro rabbit wound
repair model. Aliment Pharmacol Ther 12: 1131–1138, 1998.
389. Willi SM, Zhang Y, Hill JB, Phelan MC, Michaelis RC, Holden KR. A deletion in the long
arm of chromosome 18 in a child with serum carnosinase deficiency. Pediatr Res 41:
210 –213, 1997.
390. Williams HM, Krehl WA. The microbiological determination of carnosine and its
formation by rat liver slices. J Biol Chem 196: 443– 448, 1952.
391. Winnick T, Winnick RE. Pathways and the physiological site of anserine formation.
Nature 183: 1466 –1468, 1959.
392. Wisniewski K, Fleisher L, Rassin D, Lassmann H. Neurological disease in a child with
carnosinase deficiency. Neuropediatrics 12: 143–151, 1981.
393. Xiang J, Hu Y, Smith DE, Keep RF. PEPT2-mediated transport of 5-aminolevulinic acid
and carnosine in astrocytes. Brain Res 1122: 18 –23, 2006.
394. Yamada S, Tanaka Y, Ando S. Purification and sequence identification of anserinase.
FEBS J 272: 6001– 6013, 2005.
395. Yamashita T, Shimada S, Guo W, Sato K, Kohmura E, Hayakawa T, Takagi T, Tohyama
M. Cloning and functional expression of a brain peptide/histidine transporter. J Biol
Chem 272: 10205–10211, 1997.
396. Yan H, Guo Y, Zhang J, Ding Z, Ha W, Harding JJ. Effect of carnosine, aminoguanidine,
and aspirin drops on the prevention of cataracts in diabetic rats. Mol Vis 14: 2282–
2291, 2008.
397. Yan H, Harding JJ. Carnosine protects against the inactivation of esterase induced by
glycation and a steroid. Biochim Biophys Acta 1741: 120 –126, 2005.
398. Yan SL, Wu ST, Yin MC, Chen HT, Chen HC. Protective effects from carnosine and
histidine on acetaminophen-induced liver injury. J Food Sci 74: H259 –H265, 2009.
399. Yeargans GS, Seidler NW. Carnosine promotes the heat denaturation of glycated
protein. Biochem Biophys Res Commun 300: 75– 80, 2003.
400. Yeum KJ, Orioli M, Regazzoni L, Carini M, Rasmussen H, Russell RM, Aldini G. Profiling histidine dipeptides in plasma and urine after ingesting beef, chicken or chicken
broth in humans. Amino Acids 38: 847– 858, 2010.
401. Yuneva AO, Kramarenko GG, Vetreshchak TV, Gallant S, Boldyrev AA. Effect of
carnosine on Drosophila melanogaster lifespan. Bull Exp Biol Med 133: 559 –561, 2002.
402. Yuneva M, Bulygina E, Gallant S, Kramarenko G, Stvolinsky S, Semyonova M, Boldyrev
A. Effect of carnosine on age-induced changes in senescence-accelerated mice. J
Anti-aging Med 2: 337–342, 1999.
403. Zaloga GP, Roberts PR, Black KW, Lin M, Zapata-Sudo G, Sudo RT, Nelson TE.
Carnosine is a novel peptide modulator of intracellular calcium and contractility in
cardiac cells. Am J Physiol Heart Circ Physiol 272: H462–H468, 1997.
404. ZapataSudo G, Sudo RT, Lin M, Nelson TE. Calcium-sensitizing function for the
dipeptide carnosine in skeletal muscle contractility. Cell Physiol Biochem 7: 81–92,
1997.
405. Zapp J, Wilson D. Quantitative studies of carnosine and anserine in mammalian muscle. J Biol Chem 19 –27, 1938.
406. Zhang X, Song L, Cheng X, Yang Y, Luan B, Jia L, Xu F, Zhang Z. Carnosine pretreatment protects against hypoxia-ischemia brain damage in the neonatal rat model. Eur J
Pharmacol 667: 202–207, 2011.
407. Zhou S, Decker EA. Ability of carnosine and other skeletal muscle components to
quench unsaturated aldehydic lipid oxidation products. J Agric Food Chem 47: 51–55,
1999.
408. Zhou S, Dickinson LC, Yang L, Decker EA. Identification of hydrazine in commercial
preparations of carnosine and its influence on carnosine’s antioxidative properties.
Anal Biochem 261: 79 – 86, 1998.
409. Zschocke J, Nebel A, Wicks K, Peters V, El Mokhtari NE, Krawczak M, van der WF,
Janssen B, Schreiber S. Allelic variation in the CNDP1 gene and its lack of association
with longevity and coronary heart disease. Mech Ageing Dev 127: 817– 820, 2006.
Physiol Rev • VOL 93 • OCTOBER 2013 • www.prv.org
1845