VALIDATION STUDIES ON AN INVITRO SEMICONTINUOUS CULTURE SYSTEM DESIGNED TO SIMULATE A BACTERIAL ECOSYSTEM OF THE HUMAN INTESTINE

A semicontinuous culture system (SCCS) inoculated with human intestinal bacteria was used to determine if the metabolic activity in the human intestine could be maintained in vitro. After 43 days, a diverse bacterial population was still present with counts above 4 x 10(9) bacteria/ml. Bacteroides spp. accounted for approximately 50% of the colonies isolated. Other isolates included Eubacterium, Fusobacterium, Peptostreptococcus, Lactobacillus, Peptococcus, Clostridium, Streptococcus, and Acidaminococcus spp. Acetic, propionic, and butyric acid made up over 90% of the volatile fatty acids. Beta-glucosidase, beta-glucuronidase, nitroreductase, and azoreductase, which were present in the inoculum, were also present after 43 days. Carbon dioxide was the predominant gas (17.81 mmol/l). Hydrogen and methane were detected at 0.66 mmol/l and 3.67 mmol/l, respectively. After 17 days, the azo dye Direct blue 15, was added to the culture and over 90% was metabolized in the first 24 h. Two metabolites, 3,3'-dimethoxybenzidine and 4-amino-3,3'-dimethoxybiphenyl, were isolated and identified by high pressure liquid chromatography (HPLC), gas chromatography and mass spectrometry. The results indicate that diverse, metabolically active cultures can be maintained for prolonged periods and used for studies of xenobiotic metabolism.