Safety evaluation aims to assess the dose-response relationship to determine a dose/level of exposure for food contaminants below which no deleterious effect is measurable that is 'without appreciable health risk' when consumed daily over a lifetime. These safe levels, such as the acceptable daily intake (ADI) have been derived from animal studies using surrogates for the threshold such as the no-observed-adverse-effect-level (NOAEL). The extrapolation from the NOAEL to the human safe intake uses a 100-fold uncertainty factor, defined as the product of two 10-fold factors allowing for human variability and interspecies differences. The 10-fold factor for human variability has been further subdivided into two factors of 10(0.5) (3.16) to cover toxicokinetics and toxicodynamics and this subdivsion allows for the replacement of an uncertainty factor with a chemical-specific adjustment factor (CSAF) when compound-specific data are available. Recently, an analysis of human variability in pharmacokinetics for phase I metabolism (CYP1A2, CYP2A6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, hydrolysis, alcohol dehydrogenase), phase II metabolism (N-acetyltransferase, glucuronidation, glycine conjugation, sulphation) and renal excretion was used to derive pathway-related uncertainty factors in subgroups of the human population (healthy adults, effects of ethnicity and age). Overall, the pathway-related uncertainty factors (99th centile) were above the toxicokinetic uncertainty factor for healthy adults exposed to xenobiotics handled by polymorphic metabolic pathways (and assuming the parent compound was the proximate toxicant) such as CYP2D6 poor metabolizers (26), CYP2C19 poor metabolizers (52) and NAT-2 slow acetylators (5.2). Neonates were the most susceptible subgroup of the population for pathways with available data [CYP1A2 and glucuronidation (12), CYP3A4 (14), glycine conjugation (28)]. Data for polymorphic pathways were not available in neonates but uncertainty factors of up to 45 and 9 would allow for the variability observed in children for CYP2D6 and CYP2C19 metabolism, respectively. This review presents an overview on the history of uncertainty factors, the main conclusions drawn from the analysis of inter-individual differences in metabolism and pharmacokinetics, the development of pathway-related uncertainty factors and their use in chemical risk assessment.