Aims: To explore the influence of carbohydrates on the production of fengycin by Bacillus amyloliquefaciens fmb-60. Methods and Results: Fermentation and gene expression levels were examined following culture supplementation with fructose, arabinose and sorbitol. Compared with other carbohydrates, fructose most efficiently increased the concentration of the essential component amino acids for fengycin synthesis in the cell. The maximum production of fengycin was found to be 392.87 mg l(-1), increased by 76% in fed-batch fermentation. Furthermore, fructose enhanced the biosynthesis of fengycin by increasing the expression of fengycin synthetase genes (fenA, fenB, fenC, fenD, fenE and fenTE) and regulatory genes (comA, degU, degQ and sigma H). Conclusion: Fructose is important for fengycin production in B. amyloliquefaciens fmb-60, and it could enhance fengycin biosynthesis by raising the expression of the fengycin synthetase genes and upregulating signal factors. Significance and Impact of the Study: The results identify regulatory mechanisms involved in fengycin biosynthesis and provide an effective method to enhance production of fengycin.