COMMUNICATION
www.rsc.org/chemcomm | ChemComm
Photo-production of lactate from glyoxylate: how minerals can
facilitate energy storage in a prebiotic world
Marcelo I. Guzman and Scot T. Martin
Received (in Cambridge, UK) 18th November 2009, Accepted 4th January 2010
First published as an Advance Article on the web 21st January 2010
DOI: 10.1039/b924179e
The reaction of glyoxylate with carbon dioxide to produce
lactate is promoted when zinc sulfide is irradiated by ultraviolet
light. These results, representing a model for the action of
colloidal mineral semiconductors on early Earth, complete a
consecutive series that culminates in entry-point molecules of the
reductive tricarboxylic acid cycle.
In a series of chemical steps that fix CO2, many organisms
during the history of Earth have used the reductive tricarboxylic
acid (rTCA) cycle to store energy, ultimately as carbohydrates,
fats, and proteins.1 As such, the rTCA cycle is one of the most
ancient and fundamental of all biochemical pathways, and the
rTCA cycle accordingly has been proposed as a candidate
mechanism for carbon fixation and energy storage at the time
life originated (Scheme 1A).2–5 Moreover, products of the
rTCA cycle are common metabolites that serve as feedstock
for further biosynthesis and assembly. A fundamental challenge
is to provide synthetic access from consecutive reactions
starting from CO2 to the compounds that start the cycle.3,6
Here, we demonstrate for the first time that the C2 compound
glyoxylate (HCOCOO ) reacts with CO2 to produce the C3
compound lactate (CH3–HCOH–COO ) in 15% yield
through a ZnS-photo-promoted reaction.
Photo-generated conduction-band electrons (e CB) and
valence-band holes (h+VB) of semiconductors can facilitate
rapid reactions promoted by radical formation and energy
storage.7 The example of the present study, the mineral sphalerite
(ZnS), is expected to have been plentiful in Earth’s early seas
because of its formation in anoxic conditions from the mix of
zinc and sulfur ejected by extremely active hydrothermal
vents.2,3,8 Formate (HCOO ) and formaldehyde (CH2O) are
the main first-generation C1 products of the photochemistry of
CO2 in the presence of ZnS. Glycolate, oxalate, and glyoxylate
are first-generation C2 coupling products,9,10 promoted by the
CO2 radical formed by electron transfer from the conduction
band of ZnS to CO2.11–13 Once a pool of C2 molecules is present,
second-generation coupling products are produced.2,3,14
Scheme 1 represents reactions that can be promoted by ZnS.
Electrons are excited into the conduction band of ZnS by
absorption of photons having wavelengths shorter than
344 nm (3.6 eV). Their standard reduction potential is 1.04 V
vs. NHE. The resulting overall energy of glyoxylate to lactate
promoted by ZnS is 951 kJ mol 1 based on the thermodynamic data catalogued in ref. 15. From this reaction,
the energy storage per organic carbon atom increases by
School of Engineering and Applied Sciences & Department of Earth
and Planetary Sciences, Harvard University, Cambridge, MA 02138,
USA. E-mail: scot_martin@harvard.edu
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The Royal Society of Chemistry 2010
56.7 kJ mol 1. The highly reducing conduction-band electron
also provides an overpotential that can promote reactions that
are otherwise kinetically sluggish.
These reactions were studied in a series of laboratory
experiments. Colloidal ZnS was prepared by the addition of
Scheme 1 (A) The rTCA cycle in a form highlighting the electrontransfer elements. Blue and orange indicate pathways that provide exit
points from the cycle and that are useful for further synthesis. The
combination of the rTCA cycle and its exit products operates as a
factory for the synthesis of major classes of biologically important
molecules (shown in red). (B) Abiotic anaplerotic-like steps that start
from CO2 and produce pyruvate, a species that serves as an entry point
into the rTCA cycle.
Chem. Commun., 2010, 46, 2265–2267 | 2265
O2-free Na2S (52 mM) to ZnSO4 (52 mM) with stirring under
continuous argon bubbling. The size distribution and the
structure of the colloidal particles prepared by this method
were described previously.8 The 500 mL colloid was placed
in a photochemical reactor and then augmented first with
additional Na2S (1.8 mL; 2.22 M) and second with NaOOCHCO
(18.2 mL; 48 mM). Carbon dioxide was bubbled continuously
through the solution to decrease the pH from 12.3 to 7.5. The
colloidal ZnS had a mass concentration of 2.3 g L 1, and the
initial concentrations prior to irradiation were 1730 mM
glyoxylate and 1760 mM sulfide available as HS . After sample
preparation and under continuous CO2 bubbling, the mixture
was irradiated at 15 1C under anoxic conditions for 2 h by an
immersion medium-pressure photochemical lamp (Hanovia
7825). An intensity of 7.4 10 6 Einstein s 1 was determined
by actinometry with KFe(C2O4)2.16
The reaction products as well as the remaining reactants
were withdrawn and passed through a syringe filter (25 mm
syringe filters, 0.2 mm pore size) to prepare for ion chromatography analysis (Dionex ICS-3000, equipped with an IonPac
AS11-HC analytical column, suppression system, and a
conductivity detector).3 The flow was set to 1.5 mL min 1
with a NaOH gradient, in which a mobile phase of 1.0 mM
NaOH was used for 2 min followed by a linear increase of
2.16 mM min 1 up to 55.0 mM NaOH. The final concentration
was held for 1 min. For some analyses, sulfate was removed by
a 1 : 5 dilution of the supernatant (centrifugation for 5 min,
5000 rpm) in a 1 : 2 dilution with saturated barium hydroxide
solution (i.e., totalling a 1 : 10 dilution). Lactate (1), glycolate
(2), formate (3), glyoxylate (4), bicarbonate (5), sulfite (6),
sulfate (7), oxalate (8), and thiosulfate (9) were quantified
(Fig. 1). Although acetate (10) was separated from lactate
using this method, it eluted simultaneously with glycolate.
Thus, a new round of analyses using 1 mM NaOH with 10%
methanol isocratic mobile phase at 1.5 mL min 1 was used to
confirm that the reduction of glyoxylate stopped at glycolate
and did not continue further to produce acetate (Fig. 2).
In addition to ion chromatography, sulfide and pH were
measured using ion-selective electrodes.
A summary of results is given in Table 1 for the experiments
and controls that demonstrate the heterogeneous photoproduction of the organic products. Measurements were carried
out in the presence of one or more of glyoxylic acid (‘‘GA’’),
colloid (‘‘ZnS’’), ultraviolet irradiation (‘‘hn’’; 200 to 400 nm),
carbon dioxide (‘‘CO2’’), and sulfur-based hole scavenger (ST).
The results summarized in Table 1 imply that the conditions
Fig. 1 Chromatograms (A) before and (B) after irradiation. The
number at the top of each peak corresponds to the labeling in the text.
The inset shows an expanded region of chromatogram B.
2266 | Chem. Commun., 2010, 46, 2265–2267
Fig. 2 Chromatograms showing (A) glycolate and formate formed as
products, (B) these products spiked with additional glycolate, and (C)
these products spiked with acetate. The number at the top of each peak
corresponds to the labeling in the text.
Table 1 Experiments and controls to demonstrate that heterogeneous photoelectrochemistry is the production mechanism of
lactate, glycolate, formate, and oxalate starting with glyoxylatea
Productsc
Conditions
Experiment
Control A
Control B
Control C
Control D
Control E
GA
ZnS
hn
CO2
ST
LA
Glyc
FA
OA
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
b
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
a
GA = glyoxylate, LA = lactate, Glyc = glycolate, FA = formate,
and OA = oxalate. Table entries of ‘‘+’’ and ‘‘ ’’ indicate the
presence or absence of a species, respectively. b [Na2S]0 o 10 3 mM.
c
Major products identified.
for the formation of lactate (15% yield) require a mechanism
that is driven by heterogeneous photoelectrochemistry.
For the conditions of the full experiment (cf. Table 1), after
2 h of irradiation the initial glyoxylate concentration of
1730 mM decreased by 25% to 1290 mM. In addition to lactate,
formate (1100 mM), oxalate (48 mM), and glycolate (18 mM)
were also formed, as expected. Oxalate and glycolate were
formed both as first-generation C2 coupling products of CO2
as well as by the oxidation and the reduction of glyoxylate,
respectively.
Further investigation to detect the best conditions for the
formation of lactate are underway. Among other variables, the
light intensity, the concentration of reactants, the pH value,
and the temperature may affect the observed yield. For
example, one experiment showed that the effect of dropping
from 1760 to 570 mM initial sulfide was to decrease the yield of
lactate from 15% to 4%. The explanation may be that
stoichiometrically insufficient hole scavenger was present at
the lower sulfide loading so that conduction-band electrons
underwent recombination photophysics within the ZnS
instead of interfacial electron transfer to form CO2 . As of
yet, the full range of experimental conditions has not been
investigated to determine the optimized yield, but the presented
conditions of the experiment for 15% yield were designed as a
scenario to resemble as closely as is presently known what
were the conditions on early Earth.
These new results can be combined with some already
known facts, namely that (1) CO2 reacts in the presence of
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tetramethylammonium chloride salt to produce C2 compounds
including glyoxylate and (2) that lactate can be oxidized to
pyruvate at high yield using ZnS photochemistry. Pyruvate is
one entry point into the rTCA cycle (Scheme 1). Our present
results that describe a pathway for the production of lactate
from glyoxylate, taken together with what is already known,
constitute a plausible production scheme on early Earth for
the first metabolite of the rTCA cycle (i.e., through a series of
cascading reactions starting from CO2). Our earlier work has
examined the continuation from pyruvate to energy storage in
other species of the rTCA cycle by the pathway of ZnSphoto-promoted reduction steps (Scheme 1).2,3,14 A direct
mechanism thus emerges for the synthesis of useful energy-rich
molecules through abiotic and nonenzymatic anaplerotic-like
reactions in a prebiotic world.
These results can be related to how life might have emerged
on an early Earth composed only of simple chemical
compounds.17 The scenario envisages a primary role for the
mineral-based catalysis of processes. The minerals photocatalyze the primary steps of energy storage in small organic
molecules. When immersed in sterile water with a suitable
electron donor like HS and exposed to sunlight, sphalerite
and other similarly reactive minerals conduct a train of
reactions starting from CO2 to produce consecutively
two-carbon, three-carbon, and longer chain compounds
(Scheme 1). These steps have been demonstrated individually,
and the next step in research would be a demonstration in toto.
Also marked in Scheme 1 are more complex conversions
observed for lactate and pyruvate, such as the one-step
production of succinate (12% yield), a-ketoglutarate (50%),
and isocitrate (11%). Complex conversions for other
compounds include the reduction of oxaloacetate to malate
(75%) and of fumarate to succinate (95%). The carboxylation
of a-ketoglutarate to oxalosuccinate (2.5%) has also been
demonstrated.2,3,14
These transformations combine to a generalized postulated
model for carbon fixation promoted by photo-transformations
on minerals. This hypothesized cycle was running prior to the
appearance of enzymes and is evolutionarily linked in this way
to present-day metabolism. This model, if supported in future
studies by in toto operation of the cycle, would become an
important achievement in linking the prebiotic and the living
worlds.
The science of the origins of life is divided into advocates of
a replicator-first school and those who support a metabolismfirst vision.18 Both descriptions, however, require a supply of
small molecules, either as building blocks for the replicator or
as components for a primordial energy-driven self-sustaining
metabolic cycle. Possible sources for this organic feedstock
have been suggested, including atmospheric lighting-driven
fixation in the Miller experiment,19 extraterrestrial infall,20
and mineral-catalyzed CO and CO2 fixation in hot springs
and volcanos.21,22 The ZnS photochemical studies add an
additional pathway to this group, one for which evidence
continues to accumulate.2,3,8,14 The ZnS pathways can be
applied as described to the rTCA cycle or alternatively in
strong support of other proposed pathways, such as the
glyoxylate scenario23 or the sugar model.24 All pathways
may have been simultaneously operative on the early Earth.
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The Royal Society of Chemistry 2010
Our research is summarized in Scheme 1 and apparent
therein is that the formation of two rTCA cycle intermediates,
cis-aconitate and citrate, have not been demonstrated by ZnS
photoelectrochemistry. We have not yet investigated these
pathways. Finding simple mechanisms for their production
would advance the case for the chemical foundations of the
rTCA cycle as being manufactured easily and routinely
in early Earth’s oceans. Other pathways described in the
literature have led to compounds of the rTCA cycle but only
in low yield (o0.1%) and only for high temperatures
(>500 K) and pressures (>50 MPa).21 For comparison, the
hypothesized photo-driven mineral cycle integrates the use of
ultraviolet energy, captured and transmitted through semiconductor minerals, to produce molecules that are otherwise
inaccessible. The prospect that sphalerite can catalyze versatile
and complex reactions all by itself is promising and possibly
brings us much closer to understanding the chemical origins of
life, supporting a view that life is bound to emerge because
simple molecules central to metabolism can result and be
driven in an energy-storing direction by carbon dioxide, light,
and minerals.
Support from the Harvard Origins of Life Initiative and
NASA Grant NNX07AU97G issued through the Office of
Space Science is gratefully acknowledged.
Notes and references
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