Among the reasons suggested for the discrepancy between N balance and tracer-derived indispensable amino acid (IAA) requirement estimates is the possibility that the metabolic requirement is met not only by the diet but also by IAA synthesized de novo by the gastrointestinal microflora, which are then absorbed. It is therefore crucial to better understand and quantify the microbial biosynthesis of amino acids in the human gastrointestinal tract and its potential role in providing IAA to meet human amino acid requirement. Here, the available evidence on the contribution of microbial amino acids to the host's amino acid homeostasis, applying the (15)N labeling paradigm, is summarized. Between 1 and 20% of circulating plasma lysine, urinary lysine and body protein lysine of the host, respectively, is derived from intestinal microbial sources and corresponds to a gross microbial lysine contribution of 11-68 mg. kg(-1). d(-1) in adult humans with an adequate protein intake when fecal or ileal microbial lysine enrichment is used as precursor. Factors affecting estimates of net microbial IAA contribution are discussed. It appears that the small intestine is responsible for a large part of microbial lysine uptake, although some absorption from the large intestine cannot be excluded. Nonoxidative lysine losses from the human gastrointestinal tract, which were found to be between 3.9 to 8.5 mg. kg(-1). d(-1), are necessary to estimate the net contribution of microbial IAA. It is reasonable to assume that microbial amino acid synthesis in the human gastrointestinal tract utilizes a mixture of various nitrogen sources, i.e., endogenous amino acids, urea and ammonia. Microbes in the small intestine may rely more on endogenous amino acids. Deprivation of nutrients, the intake of certain dietary nonstarch oligosaccharides, lipids, as well as protein intake level and source and level of consumption of certain amino acids can affect the composition and metabolic activity of the intestinal microflora and thus its fermentation products potentially available to the host. In conclusion, with the use of the (15)N labeling paradigm, a significant contribution of microbial lysine to the host lysine homeostasis is found. However, to assess the net contribution of microbial IAA and its importance in defining the adult IAA requirement, this is not the ultimately successful experimental strategy because the interpretation of results is complicated by the nitrogen recycling in the gut, the uncertainty of the precursor pool of absorption and the limited data on nonoxidative IAA losses from the human gastrointestinal tract.