Asian Journal of Medical Sciences 3(2): 67-70, 2011
ISSN: 2040-8773
© Maxwell Scientific Organization, 2011
Received: January 17, 2011
Accepted: February 15, 2011
Published: April 20, 2011
Choleretic Effect of Curcuma longa and Verapamil in Mice
1
H. Najafzadeh, 2N. Moezzi and 3N. Motamed
Department of Pharmacology and Toxicology,
2
Resident of Veterinary Surgery,
3
Resident of Poultry Sciences, Faculty of Veterinary Medicine,
Shahid Chamran University, Ahvaz-Iran
1
Abstract: Choleretic drugs are used in some conditions e.g., fat maldigestion. Verapamil is a calcium channel
blocker, which inhibits p-glycoprotein (P-gp) and may affect bile accumulation. Curcuma longa is traditionally
used for treatment of maldigestion and as food additive. In this study, the effect of verapamil and Curcuma
longa was compared on bile accumulation in gall bladder of mice. This simple animal model is approved and
used for evaluation and screening pharmacological effect of agents on bile production or secretion. In this
model, the mice arre off-feed for 24 h. The mice were divided in 3 groups. Saline was administrated in group
1, an aqueous suspension of Curcuma longa and verapamil was orally administrated in groups 2 and 3
respectively. One hour after drug administration, the mice were euthanized and gall bladder was immediately
removed. The weight of bile was measured in each mouse and compared between groups. The mean of bile
weight was 4.67±0.615, 14.33±0.792 and 14.17±1.641 mg in groups 1 to 3 respectively. Verapamil as well as
Curcuma longa significantly increased the accumulation of bile in gall bladder in comparison to saline. Thus
verapamil and Curcuma longa may similar action on bile secretion via p-glycoprotein.
Key words: Curcuma longa, choleretic effect, mice, verapamil
INTRODUCTION
mice resembles the disruption of MDR3 in human.
Secondly, mutation in the bile salt exporting pump
(BSEP)/sister of P-glycoprotein (spgp) gene causes
progressive familial intrahepatic cholestasis type 2 in
human. However, in spgp knock-out mice only a mild
persistent cholestasis occurs (Elamiri et al., 2003;
Lam et al., 2005).
Multidrug resistance especially in cancers relates to
P-gp function (Orlowskis and Garrigos, 1999;
Srinivas et al., 2006). Also, P-gp inhibition or induction
can affect pharmacokinetics of drugs and may cause drug
interaction (Verschraagen et al., 1999; Yu, 1999). Class
HI multidrug resistance (MDR) P-glycoproteins (P-gp),
mdr2 in mice and MDR3 in man; mediate the
translocation of phosphatidylcholine across the
canalicular membrane of the hepatocyte. Mice with a
disrupted mdr2 gene completely lack biliary phospholipid
excretion and develop progressive liver disease,
characterized histologically by portal inflammation,
proliferation of the bile duct epithelium, and fibrosis. This
disease phenotype is very similar to a subtype of
progressive familial intrahepatic cholestasis. It was
demonstrate that mutations in the human MDR3 gene lead
to progressive familial intrahepatic cholestasis with high
Bile production and secretion is changed by several
factors including nutrition, diseases and drugs. Choleretic
drugs are used in some conditions such as fat
maldigestion. The Choleretic drugs stimulate production
or secretion the bile. Ultimately, drugs and their
metabolites which present in bile pass into the intestinal
tract during the digestive process. P-Glycoprotein (P-gp)
also is involved in the excretion of endogenous
compounds (Groen et al., 1995; Eytan and Kuchel, 1999;
Leung and Bendayan, 1999; Johnston et al., 2000;
Hardman and Limbird (2001) and this carrier has a
significant impact on systemic and tissue/cellular
bioavailability of drugs (Fromm, 2000; Liu and Hu, 2000;
Hendriks and Vaalburg, 2002).
The inhibition of canalicular glycoprotein
transporters has been suggested as the cause of familial
intrahepatic cholestasis. Mutations in multidrug resistance
3-glycoprotein (MDR3) gene induce progressive familial
intrahepatic cholestasis type 3 (PFIC3) (Belinsky et al.,
2005; Elamiri et al., 2003). PFIC2 patients secrete less
than 1% of bile salts compared with normal infants
(Wang et al., 2001). Phenotypically, mutation in mdr2 in
Corresponding Author: H. Najafzadeh, Department of Pharmacology and Toxicology, Faculty of Veterinary Medicine, Shahid
Chamran University, Ahvaz-Iran. Ph: +986113330073/+989166182496
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Asian J. Med. Sci., 3(2): 67-70, 2011
serum (-GT. The histopathological picture in these
patients is very similar to that in the corresponding mdr2(/-) mouse, in which mdr2 P-gp deficiency induces
complete absence of phospholipid in bile (De
Vree, 1998).
Curcuma longa L., which belongs to the
Zingiberaceae family, is a perennial herb that measures up
to 1 m high with a short stem, distributed throughout
tropical and subtropical regions of the world, being
widely cultivated in Asiatic countries, mainly in India and
China. Its rhizomes, a powder, called turmeric, it has
been in continuous use for its flavoring, as a spice in both
vegetarian and non-vegetarian food preparations and it
also has digestive properties. Current traditional Indian
medicine claims the use of its powder against biliary
disorders, anorexia, coryza, cough, diabetic wounds,
hepatic disorder, rheumatism and sinusitis (Ammon and
Wahl, 1991; Araújo and Leon, 2001; Chainani-Wu, 2003;
Zeng et al., 2007).
The extract from C. longa specifically inhibits gastric
acid secretion by blocking H2 histamine receptors in a
competitive manner. C. longa has specifically
antidepressant effects in vivo. The activity of C. longa in
antidepression may mediated in part through MAO A
inhibition in mouse brain (Yu et al., 2002;
Kim et al., 2005). The main constituents of the extract
were identified as curcumin, demethoxycurcumin,
bisdemethoxycurcumin, and ar-turmerone. Both
curcuminoids and sesquiterpenoids in turmeric exhibit
hypoglycemic
effects
(Kuroda
et al., 2005;
Nishiyama et al., 2005).
Verapamil, a calcium channel blocker, is specific
inhibitor of the P-glycoprotein (mdr1) in the hepatocyte
canalicular membrane and other tissue. This drug is used
as
antihypertensive
and
antiarrhytmic
agent
(Mickisch et al., 1991; Barth et al., 2006).
Thus in this study, the effect of verapamil and
Curcuma longa was compared on bile accumulation in
gall bladder of mice for evaluation their potency and
possible mechanism.
volume of suspension for each mouse was nearly 0.5 mL.
Suspension of verapamil was orally administrated in 0.24
mg/10 g body weight of mice in group 3. One hour later,
the mice were euthanized by diethyl ether and
immediately the gall bladder was carefully removed. The
weight of gall bladder (with bile) for each mouse was
measured with analytical scale. Then, the bile is extracted
from gall bladder and again it was weighted. The
difference of weight of full and empty gall bladder was
concerned as amount of bile.
The mean of amount of bile in groups was compared
by SPSS program and ANOVA test. The p-values less
than 0.05 were concerned as significant.
RESULTS AND DISCUSSION
The mean of bile in gall bladder was 4.67±0.615 mg
in group 1 as control. This amount was 14.33±0.792 in
group 2 that received Curcuma longa. The mean of bile in
gall bladder was 14.17±1.641 mg in group 3 that received
verapamil.
The bile accumulation in gall bladder was
significantly increased by Curcuma longa and verapamil
in comparison to saline (p = 0.001).
Final effect of verapamil and Curcuma longa was
similar on the bile accumulation probably by same
mechanism on P-glycoprotein. P-glycoprotein (Mdr1) is
capable of transporting bile acids (Lam et al., 2005).
Verapamil was studied for its effects on secretive function
of liver in rats. In the animals with low initial level of bile
secretion, infusion of verapamil resulted in increase of the
bile flow. In the animals with high initial level of the liver
secretive function, verapamil decreased the bile flow. The
changes of bile flow and biliary secretion of bile acids and
lipids in two groups of animals suggest that verapamil
could be influenced in regulation of bile secretion
depending on its initial level
(Tukaev et al., 2002).
The similar effects of Verapamil and Cyclosporin A,
on bile acid transport and metabolism, can be explained
by mdr1 mediated disturbances of cellular ATP transport
rather than by inhibition of individual bile acid
transporters (Barth et al., 2006).
The effects of verapamil on the dopamine-induced
pancreatic exocrine secretion were investigated in the
isolated and blood-perfused canine pancreas. Dose-related
increases in the volume of pancreatic secretion induced by
dopamine (1-10 micrograms) were reduced by infusion of
50 and 100 :g/min of verapamil (Iwatsuki et al., 1986).
The effects of various P-glycoprotein transporters is
evaluate on bile formation and the canalicular transport of
taurocholic acid in an attempt to understand the combined
role of these transporters in the pathogenesis of familial
intrahepatic cholestasis. Total bile acid and cholic acid
secretion rate were decreased in the mdr2 knock-out mice.
MATERIALS AND METHODS
This study was conducted in Department of
pharmacology and toxicology Shahid Chamran
University, Ahvaz, Iran during 2008-2009.
Mice were purchased from lab animals' research
center, Jundishapour university of Ahvaz. Mice (NMRI
strain) with 20±3 grams weight were divided 3 groups (6
mice in each group). The mice were kept without feed but
water ad libitum in 24 hours prior study.
In day of study, in group 1, saline was administrated
(as control group). Suspension of Curcuma longa was
orally administrated in 20 mg/10 g body weight of mice
in group 2. This suspension was prepared by 600 mg
powder of Curcuma longa and 10 mL distilled water. The
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Asian J. Med. Sci., 3(2): 67-70, 2011
However, bile flow and the secretion of muricholic acids
were increased (Elamiri et al., 2003).
The pathogenesis of manganese-bilirubin (Mn-BR)
induced cholestasis has only been studied in rats and is
associated with alteration in the hepatic homeostasis of
cholesterol and phospholipids. Multidrug resistance-2
(mdr2) transporter, which mediates excretion of these
lipids, is suggested to be involved in this phenomenon.
Akoume et al. (2004) examined the role of mdr2 in its
pathogenesis, using mice with disrupted mdr2 gene (mdr2
(-/-)). Results showed that Mn-BR combination decreased
bile flow in mice. These data indicate that Mn-BR
induced cholestasis is reproducible in mice
(Akoume et al., 2004).
The knockout of spgp gene in mice results in
intrahepatic cholestasis, but with significantly less
severity than PFIC2 in humans. Notably, although the
secretion of cholic acid in mutant mice is greatly reduced
(6% of wild-type), total bile salt output in mutant mice is
about 30% of wild-type. Also, secretion of an
unexpectedly large amount of tetra-hydroxylated bile
acids (not detected in wild-type) is observed. These
results suggest that hydroxylation and an alternative
canalicular transport mechanism for bile acids
compensate for the absence of Spgp function and protect
the mutant mice from severe cholestatic damage. In
addition, the spgp-/- mice display a significant increase in
the secretion of cholesterol and phospholipids into the
bile. This latter observation in spgp-/- mice suggests that
intrahepatic, rather than intracanalicular, bile salts are the
major driving force for the biliary lipid secretion. The
spgp-/- mice thus provide a unique model for gaining new
insights into therapeutic intervention for intrahepatic
cholestasis and understanding mechanisms associated
with lipid homeostasis (Wang et al., 2001).
Former studies have shown that curcumin, which can
be extracted from different Curcuma species, is able to
stimulate
bile
flow
in
rats,
whereas
bisdemethoxycurcumin, which is mainly found in
rhizomes of Curcuma longa, is believed to inhibit bile
flow. The influence of both curcuminoids on bile flow,
bile acid concentration and excretion over a time period
of 180 min in the bile fistula model in rats. Furthermore,
it was tested the ability of both curcuminoids to reduce
cyclosporin-induced cholestasis. The choleretic effect of
bisdemethoxycurcumin lasted longer than that of
curcumin. However, only bisdemethoxycurcumin
statistically significantly attenuated cyclosporin-induced
reduction of bile acid excretion (Deters et al., 1999).
In conclusion, from present study it is concluded that
Verapamil has choleretic effect similar to Curcuma longa
in mice as experimental model. The important finding of
this study is the choleretic effect of Curcuma longa may
be carried by p-glycoprotein (P-gp). But more study is
needed for finding the exact its mechanism and this effect
of Curcuma longa can be evaluated in human.
ACKNOWLEDGMENT
The authors wish to express their gratitude to the
research council of Shahid Chamran University for their
financial supports.
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