Biohydrogen production from corncob using natural anaerobic microflora was reported for the first time. The optimum pretreatment condition for the corncob was determined to be 100 degrees C, 30 min, and 1% HGl (w/w). The maximum hydrogen yield of 107.9 ml/g-TVS and hydrogen production rate of 4.20 ml/g-TVS h(-1) was obtained under the condition of 10 g/l substrate concentration and initial pH 8.0. Butyrate and acetate were the dominant metabolic by-products of hydrogen fermentation. Chemical composition analysis, Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) were used to study the mechanism of degrading corncob for hydrogen production. The amorphous domains of cellulose and hemicellulose were hydrolyzed into fermentable saccharides through acid pretreatment and the microorganisms had a devastating effect on the crystallinity of the cellulose. The hydrogen yield from pretreated corncob was much higher than from raw corncob. Therefore, the acid pretreatment played a crucial role on hydrogen production from corncob. (C) 2009 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved.