Abstract: Abstract: Fuel ethanol has been recognized as a kind of potential alternative fuel as well as an additive to gasoline because of its oxygenated and high octane characteristics. Syngas fermentation, a novel route for ethanol production, is getting more and more attention. Syngas can be generated from a lot of organic materials including biomass. Gasification technology is used to convert the biomass into a mixture of gases (consisting mainly of CO, CO2 and H2), which is subsequently fermented to ethanol by means of anaerobic microbial catalysts known as homoacetogens. As a homoacetogen, Clostridium autoethanogenum is able to metabolize syngas/CO and synthesize ethanol, but limited work has been accomplished with it and ethanol concentration achieved was low. In order to improve its ethanol production, fermentation process and some factors affecting cell growth or product formation were studied. Although xylose is an easily-used carbon source for C. autoethanogenum, results showed that the primary end product in xylose metabolism was acetate, while ethanol remained in a low level even with high level of xylose (8-10 g/L). C. autoethanogenum grew rapidly in the medium containing xylose and organic nitrogen source, and high cell density was achieved. When nitrogen source was switched to NH4Cl, C. autoethanogenum grew much slowly and the overall cell density diminished. However, nitrogen source didn't have much influence on ethanol production if xylose was used as the carbon source. Two-step fermentation, i.e., growing on xylose (first stage) and then fermenting with CO (second stage), was performed in a 3 L bioreactor to study the batch fermentation of C. autoethanogenum. Results indicated that cell growth and acetate production occurred in the first stage, whereas ethanol was primarily produced in the second stage when xylose was exhausted and CO became the sole carbon source. Fermentation curves showed that pH value and oxidation-reduction potential (ORP) dropped with cell growth and acetate production, while ethanol production was accompanied by the decrease of acetate concentration and the rise of pH value and ORP. CO2 evolution was observed in both growth and fermentation stages, and a small amount of H2 was detected in the outlet gas during the CO fermentation stage. But due to the limitations of bioreactor and operating conditions, gas-liquid mass transfer in the bioreactor was poor, which resulted in the low efficiency of CO utilization, and only 1.71 g/L ethanol was obtained. To eliminate the limitations, the bioreactor was modified and equipped with a specific device which could keep a constant pressure in the headspace. One-step fermentation was carried out in the modified bioreactor using CO as the sole carbon and energy source. In spite of reduced growth rate and cell density, C. autoethanogenum produced more ethanol than it did in two-step fermentation. The maximum ethanol concentration obtained was 7.36 g/L, much greater than that reported in the previous studies. Moreover, during the whole fermentation process, acetate concentration remained lower than 1.1 g/L. Summarily, the results suggest that C. autoethanogenum is a promising strain in ethanol production from CO, and this study presents a reference for the scale-up of CO fermentation to ethanol with C. autoethanogenum.