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  • For human health risk assessment of chemicals a default unce

    2021-03-02

    For human health risk assessment of chemicals, a default uncertainty factor (UF) of 3.16 is used to capture inter individual variation in toxicokinetics (Dorne and Renwick, 2005). However, the scientific background for this default UF remains unsatisfactory., Several studies have reported that a factor of 3.16 may not be enough to reflect the inter individual differences in toxicokinetics arising due to enzymatic polymorphism (CYP2D6, 2C9 and 2C19) and a factor of minimum 4.5 has been suggested to capture this inter individual variability in toxicokinetics (Falk-Filipsson et al., 2007). For example, owing to the poorly metabolizing CYP2C9 isoform, the uncertainty factor of 3.16 has been reported to be inadequate to account for the observed variability since poor metabolizers would be at increased risk (Dorne et al., 2005). Similarly, the uncertainty factor of 3.16 has been found to be inadequate to include CYP3A4 mediated differences in the metabolism of environmental contaminants, and hence the risk factor associated with it, in neonates (Dorne et al., 2003). In general, metabolism is considered as a most important factor responsible for inter individual variability and interspecies difference in toxicokinetics. An insightful knowledge about the olopatadine hcl involved in the metabolism would help to better understand the impact of genetic polymorphism on inter individual toxicokinetics and xenobiotic interaction potential of BNZ. Thus, it is of utmost importance to understand the metabolic pathways involved in the clearance of xenobiotics. To identify the CYP isoforms involved in its metabolism, BNZ was screened using those recombinant human CYPs that are most commonly involved in xenobiotic metabolism. All of the rCYPs (3A4, 2C9, 2E1, 1A2, 2D6 and 2C19) tested were found to significantly olopatadine hcl metabolize BNZ. The relative contribution of each of the tested CYPs was determined using both relative activity factor and relative abundance approach. To confirm the results of the recombinant enzyme assay, CYP phenotyping of BNZ was also performed using human liver microsomes. Using CYP isoform specific inhibitors, BNZ was found to be a substrate of CYP3A4, 2C9, 2E1, 2C19, 2D6 and 1A2. Qualitatively and quantitatively similar results were obtained using both the methodologies. None of the chemical inhibitors used could considerably inhibit the overall metabolism of BNZ indicating that the major contributing isoform could not be ascertained and all the tested isoforms contributed to its metabolism. Since the multiple enzymes are involved in BNZ metabolism and none of the inhibitors could considerably inhibit overall metabolism suggest that single enzyme genetic polymorphism (e.g. CYP3A4, 2C9, 2E1, 2C19, 2D6 and 1A2) is unlikely to have profound effect on the toxicokinetics of BNZ and the traditional uncertainity factor of 3.16 might be sufficient to capture the intraspecies toxicokinetic variability. Further, the involvement of multiple CYP rules out the probability of CYP-mediated interactions of BNZ that could have caused unexpected BNZ toxicity due to alterations in its metabolic clearance. The plasma clearance is the most crucial parameter determining the internal dose/systemic availability of any compound governing the biophase and other local tissue concentrations, responsible for exerting any effect or toxicity. Thus, the in vitro data generated was used to predict the in vivo clearance in rat and human. Using the degradation constant value obtained from log percent remaining versus time profile, the intrinsic clearance was calculated which was found to be 383.64 and 296.86 μL/min/mg protein in rat and human liver microsomes. The in vivo hepatic clearance predicted using the well-stirred model approach, 63.55 and 18.91 mL/min/kg in rat and human, respectively, was found to be close to the hepatic blood flow rate in both rat and human, suggesting it to be a high clearance compound (Davies and Morris, 1993). The in vivo studies conducted in rat in our lab also suggest that BNZ is a high clearance compound (data unpublished) as is indicated by the predicted in vivo hepatic clearance in this study. The extraction ratio was more than 0.9 in both the species. In general, the clearance of high extraction compounds is blood flow limited and thus, would not be affected by the phenotypic differences. Being blood flow limited, the clearance of BNZ could vary with changes in hepatic blood flow. Factors which decrease hepatic blood flow like aging, exercise and differences in posture could result in higher concentrations reaching systemic circulation as the clearance decreases. However, this inter-individual variability (30–40%) in hepatic blood flow would be accounted for in the default factor of 3.16 for individual variability in toxicokinetics to predict risk assessment in humans with confidence. Also, the high predicted in vivo hepatic clearance indicates that liver could be a major contributor towards the first pass metabolism of BNZ leading to its low oral bioavailability (internal dose) (Davies and Morris, 1993). This would be favorable in limiting the higher systemic concentration of BNZ when exposed through food and water.