Efficient hydrogen production from the lignocellulosic energy crop Miscanthus by the extreme thermophilic bacteria Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana

Abstract

Background: The production of hydrogen from biomass by fermentation is one of the routes that can contribute to a future sustainable hydrogen economy. Lignocellulosic biomass is an attractive feedstock because of its abundance, low production costs and high polysaccharide content.

Results: Batch cultures of Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana produced hydrogen, carbon dioxide and acetic acid as the main products from soluble saccharides in Miscanthus hydrolysate. The presence of fermentation inhibitors, such as furfural and 5-hydroxylmethyl furfural, in this lignocellulosic hydrolysate was avoided by the mild alkaline-pretreatment conditions at a low temperature of 75 degrees C. Both microorganisms simultaneously and completely utilized all pentoses, hexoses and oligomeric saccharides up to a total concentration of 17 g l-1 in pH-controlled batch cultures. T. neapolitana showed a preference for glucose over xylose, which are the main sugars in the hydrolysate. Hydrogen yields of 2.9 to 3.4 mol H2 per mol of hexose, corresponding to 74 to 85% of the theoretical yield, were obtained in these batch fermentations. The yields were higher with cultures of C. saccharolyticus compared to T. neapolitana. In contrast, the rate of substrate consumption and hydrogen production was higher with T. neapolitana. At substrate concentrations exceeding 30 g l-1, sugar consumption was incomplete, and lower hydrogen yields of 2.0 to 2.4 mol per mol of consumed hexose were obtained.

Conclusion: Efficient hydrogen production in combination with simultaneous and complete utilization of all saccharides has been obtained during the growth of thermophilic bacteria on hydrolysate of the lignocellulosic feedstock Miscanthus. The use of thermophilic bacteria will therefore significantly contribute to the energy efficiency of a bioprocess for hydrogen production from biomass.

Figure 1

Enzymatic hydrolysis of alkaline-pretreated Miscanthus on bench scale. (a) The slurry immediately after addition of the enzyme preparation (GC 220 at a final loading rate of 15 IFPU per g dry matter). (b) After 24 h of hydrolysis. Miscanthus was pretreated with 9% NaOH (w w^-1) for 6 h at 75°C.

Figure 2

Hydrogen and organic acid production by cultures of C. saccharolyticus and T. neapolitana grown on Miscanthus hydrolysate. (a) and (c) C. saccharolyticus. (b) and (d) T. neapolitana. (a) and (b) Hydrogen production. (c) and (d) Organic acid (acetic and lactic acid) production. Measurements were done after 16 and 40 h after the start of the fermentation. Hydrolysate concentration is given in g monosaccharides l^-1. In the control without hydrolysate, 10 g l^-1 of a glucose/xylose (7:3, w w^-1) mixture was used as the carbon source. The error bars show the range about the mean of the values from two flasks.

Figure 3

Consumption of carbohydrates by cultures of C. saccharolyticus and T. neapolitana grown on Miscanthus hydrolysate. Monosaccharide consumption (filled symbols) and consumption of di- and/or oligosaccharides (open symbols) by C. saccharolyticus (a) and T. neapolitana (b) in a medium containing Miscanthus hydrolysate. The hydrolysate concentration was at 14 (triangles) and 28 (diamonds) g monosaccharides l^-1.

Figure 4

Simultaneous consumption of monosaccharides by cultures of C. saccharolyticus and T. neapolitana grown on Miscanthus hydrolysate. Consumption of glucose (circle) and xylose (triangle) by C. saccharolyticus (filled symbols) and T. neapolitana (open symbols) in a medium containing Miscanthus hydrolysate. The monosaccharide concentration was 14 g l^-1.

Figure 5

Hydrogen production by cultures of C. saccharolyticus and T. neapolitana grown on pure sugars and Miscanthus hydrolysate. Growth (squares) and hydrogen production (circles) in cultures of C. saccharolyticus (a) and T. neapolitana (b) grown on a glucose/xylose (7:3, w w^-1) mixture (open symbols) or Miscanthus hydrolysate (filled symbols). The monosaccharide concentration was 14 g l^-1.

Figure 6

Effect of Miscanthus hydrolysate concentration on the production of hydrogen, acetate and lactate by cultures of C. saccharolyticus and T. neapolitana. Production of hydrogen (circle), acetate (triangle) and lactate (diamond) in cultures of C. saccharolyticus (filled symbols) and T. neapolitana (open symbols) grown on Miscanthus hydrolysate at concentrations of 10 (a), 14 (b) and 28 (c) g monosaccharide l^-1.