As a result of increasing price of transportation fuel, we have intensified our research on butanol production from agricultural residues using Clostridium beijerinckii. Butanol has superior fuel properties compared to ethanol. In this paper, wheat straw was evaluated as a feedstock for butanol production. Wheat straw was pretreated with dilute sulfuric acid and hydrolyzed to simple sugars using commercial carbohydrases. Hydrolysis, fermentation, and product recovery were combined in a single step using a 2.5 L bioreactor. Pretreated wheat straw was successfully hydrolyzed to produce glucose, xylose, arabinose, galactose, and mannose and these sugars were fermented by C. beijerinckii. The fermentation performance was enhanced by simultaneously recovering products [Acetone-butanol (AB)] from the fermentation broth by gas stripping, thereby, avoiding end product inhibition. The reactor was operated in a fed-batch mode and the fermentation lasted for >500 h. These studies demonstrated that production of AB from wheat straw in a single reactor is possible when hydrolytic enzymes are used and product (AB) is simultaneously produced and recovered.