Several bacteria from different genera are able to produce hydrogen. Major group constitute stringent anaerobic microorganisms, such as clostridia, paunch bacteria or hyperthermophilic archae bacteria. Additionally, facultative anaerobe, e.g. enterobacter, e.coli or citrobacter can produce hydrogen. Alcaligenes and bacillus (aerobic microorganisms) are able to produce hydrogen. Facultative anaerobe show less hydrogen yields in fermentations, compared to stringent anaerobic organisms. Highest hydrogen yields (more than 3 mol hydrogen per mol glucose) can be achieved with thermophilic microorganisms. Therefore, these organisms are definitively better for hydrogen production than mesophilic bacteria (Nath et al., 2004).
Major thermophilic producers of hydrogen belong to eubacteria, including species such as clostridium, caldicellulosiruptor, fervidobacterium, thermoanaerobacter, thermotoga and thermococcus (Goorissen et al., 2006).
The majority of hydrogen producing bacteria belong to the species clostridium. These are spore forming bacteria, which grow at temperatures up to 60 °C. Products are variable - ethanol, lactate, acetate, butyrate, propionate and hydrogen can be produced.
The genus caldicellulosiruptor has four closely related species, which form no spores and grow at temperatures up to 80 °C. Advantage of these microorganisms is the closer product palette.
The order of thermotogales includes the genera thermotoga and fervidobacterium. Thermotoga (hyperthermophilic) are mostly isolated from marine hydrothermal sources and can grow at temperatures up to 90 °C. Some of the thermotoga species are halophilic and therefore need complex nutrient sources, being disadvantageous for these bacteria. The species of fervidobacterium are not halophilic and need less nutrients for growth. Thermoanaerobacter include various species, which can grow in acidic surroundings and produce elemental sulfur from thiosulfate. They grow at temperatures of 85 °C and mostly form spores.
Thermococci belong to archaebacteria, are halophilic and can grow on different protein and sugar-containing substrates.
Caldicellulosiruptor derives from latin words "caldus", "cellulosum" and "ruptor" and stands for "Bacterium, which cleaves hot cellulose". The organisms of caldicellulosiruptor-species are gram-positive straight rods with stringent anaerobic living. They grow in temperature range of 45-80 °C with an optimum of 70 °C. These are extremely thermophilic bacteria (figure 1).
Saccharolyticus derives from the Greek: "Sakchar" stands for sugar and "lytikos" for the ability to dissolve. Caldicellulosiruptor saccharolyticus produces 0,4-0,6 ∙ 3-4 µm straight rods, which exist separately or pairwisely. The pH-range, in which the organism can live, is between 5.5 and 8, but a pH-value of 7 constitutes an optimum. This organism has been isolated for the first time in the 1980s from driftwood in a hot spring in Taupo, New Zealand (Rainey et al., 1994).
Figure 1: Microscopy of Caldicellulosiruptor saccharolyticus (source: genome.jgi-psf.org)
Caldicellulosiruptor is taxonomically classified as follows (NCBI Taxonomy Browser):
Experiments with C13-labeling have indicated, that 99 % of the produced acetate from Caldicellulosiruptor saccharolyticus is synthesized during Embden-Meyerhof metabolic pathway. They have not shown any indication, that the Entner-Doudoroff or the pentose phosphat pathway are participated (De Vrije et al., 2007).
Entire sequencing of the organism´s genome (Van de Werken et al., 2008) indicated the components of a complete Embden-Meyerhof-pathway. Genes for oxidative part of the pentose phosphate or Entner-Doudoroff-pathway have not been detected. However, the not-oxidative part of the pentose phosphate pathway could be detected, where xylose is converted to glycolysis intermediate products. Additionally, genome sequences for polysaccharide-decomposing enzymes were found.
These sequences give to Caldicellulosiruptor saccharolyticus the ability to produce hydrogen not only from α-bound polymeranes, but also from complex ß-bound glycanes, such as cellulose or hemicellulose. The pyruvat, produced during conversion is decarboxylated to acetyl-CoA through the enzyme PFOR. Acetyl-CoA is converted to acetate and ATP or is used for biosynthetical functions. As fermentation end products, the reduction equivalents NAD and ferredoxin are produced. The species saccharolyticus is able to produce hydrogen from ferredoxin or directly from NADH, as two hydrogenase clusters exist in the genome. Genome sequences of ferredoxin-dependent Ni-Fe hydrogenase and of membrane-bound, NAD-dependent Fe-hydrogenase were found. At too high hydrogen partial pressures, production of hydrogen of NADH is disadvantageous thermodynamically. Then, NADH is oxygenated via production of lactate or ethanol. This change of metabolic pathway leads to reduced hydrogen yield.