Homofermentative production of reduced products requires additional reducing power output (NADH) from glucose catabolism. Anaerobic expression of the pyruvate dehydrogenase complex (PDH, encoded by aceEF-lpd, a normal aerobic operon) is able to provide the additional NADH required for production of reduced products in Escherichia coli fermentation. The multiple promoters (pflBp(1-7)) of pyruvate formate lyase (pflB) were evaluated for anaerobic expression of the aceEF-lpd operon. Four chromosomal constructs, pflBp(1-7)-aceEF-lpd, pflBp(1-6)-aceEF-lpd, pflBp(6,7)-aceEF-lpd, and PflBp6-aceEF-lpd efficiently expressed the PDH complex in anaerobically grown cells. Doubling the reducing power output was achieved when glucose was oxidized to acetyl-CoA through glycolysis and pyruvate oxidation by the anaerobically expressed PDH complex (glucose -> 2 acetyl-CoA + 4 NADH). This additional reducing power output can be used for production of reduced products in anaerobic E. coli fermentation. (C) 2009 American Institute of Chemical Engineers Biotechnol. Prog., 26: 45-51, 2010