Biofuels in Perspective

Biological Hydrogen Production by Anaerobic Microorganisms 209
cellulolytic capacity, Ca. saccharolyticus was selected for further research on biohydrogen
production. 5,70 Moreover, Ca. saccharolyticus was selected by the DOE for genome
sequencing, which is expected to be finished early 2007. Ca. saccharolyticus efficiently
converts a wide range of biomass components viz. cellulose (Avicel, amorph), hemicellulose,
pectin, α-glucans (starch, glycogen, pullulan), β-glucans (lichenan, laminarin), guar
gum and gum locust bean resulting in the formation of hydrogen. 67
Batch incubations (pH-controlled) resulted in hydrogen/glucose ratios of 3.3. 5,71 Relatively
high P(H2) were found to initiate a shift to lactate production (∼2 × 10 4 Pa)
or to completely inhibit growth (∼6 × 10 4 Pa). 70 However, continuous culture cultivation
on glucose or xylose combined with hydrogen removal by nitrogen purging, resulted
in complete fermentation to acetate, without formation of lactic acid or ethanol. Under
these conditions the hydrogen acetate ratio comes close to the theoretical maximum of
4 (H. Goorissen et al., unpublished results). Moreover, preliminary fermentation data
showed that glucose and xylose can be utilized simultaneously, and that catabolite repression
by glucose does not occur. 72 From the viewpoint of hydrogen formation from
biomass this is an important phenomenon, which enables efficient conversion of both
cellulose and hemicellulose. Ca. saccharolyticus has been tested for hydrogen formation
on various biomass hydrolysates (Paper sludge, Miscanthus, potato steam peels, Sweet
Sorghum). 73 Ca. saccharolyticus displayed efficient behavior, producing 3-4 hydrogen per
glucose equivalent. Ca. saccharolyticus has a preference for xylose if grown in mixtures
with glucose (Figure 11.3a and 11.3b), although this xylose repression is concentration
dependent. At a high initial xylose concentration xylose acted as a repressor. At a low initial
xylose concentration, glucose repressed xylose utilization. Similar results were obtained
for other pentose/hexose mixtures (results not shown), although some small differences in
lactate production were observed.
The available genome sequence will enable detailed analysis of the catabolic and redoxmediating
pathways by functional genomics . As soon as a genetic system can be developed
for this organism, metabolic engineering may be used to improve the hydrogen generating
system.
Ca. saccharolyticus uses the EM pathway for its glucose catabolism which suggests
that reducing equivalents are produced as NADH and reduced ferredoxin. 71 Hydrogen
formation may proceed by an NADH-dependent cytoplasmic Fe-only hydrogenase and a
Concentration (mM)
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40 (a) 40 (b)
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Time (h)
Time (h)
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Figure 11.3 Growth of Ca. saccharolyticus at high (a) and low (b) xylose concentrations in mixtures of
glucose and xylose. Symbols: * hydrogen; ♦, glucose; �, xylose; �, acetate; �, lactate. (72)
Concentration (mM)
Hydrogen (%)