2.1. Isolation of hydrogen-producing strains
The hydrogen-producing strains were isolated from the
anaerobic sewage sludge of the municipal wastewater plant
(Xuhui District, Shanghai, PR China). The sludge was firstly
diluted with sterile distilled water, spread onto the sterile agar
plates and incubated at 37 C in an anaerobic incubator
flushed with N2. Then separated colonies were transferred to
inoculate in the cuvette with a Durham tube in it, and cultured
in the anaerobic incubator. Evolved gas in the Durham tube
would be observed immediately. The bacteria which could
yield gas were finally cultured in a syringe. The volume and
composite of evolved gas inside the syringe was measured by
the syringe and a gas chromatography, respectively. And the
strains with higher hydrogen production yield or rate were
selected for the further analysis.