Analysis of Styrene and its Metabolites in Blood and Urine of Workers Exposed to both Styrene and Acetone

Prieto, M.J.; Marhuenda, D.; Cardona, Antonio

doi:10.1093/jat/26.1.23

Cited by 16 publications

(10 citation statements)

References 21 publications

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“…The allosteric mechanism for in vitro styrene metabolism may occur under in vivo conditions, according to biomonitoring studies for industrial workers (Löf et al, 1986;Somorovská et al, 1999;Prieto et al, 2002). Typical styrene levels in blood of low to medium exposed workers, e.g., maintenance workers and sprayers, ranged from 24 to 188 ng/ml (0.24 -1.9 M).…”

Section: Discussionmentioning

confidence: 99%

CYP2E1 Metabolism of Styrene Involves Allostery

2012

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ABSTRACT:We are the first to report allosterism during styrene oxidation by recombinant CYP2E1 and human liver microsomes. At low styrene concentrations, oxidation is inefficient because of weak binding to CYP2E1 (K s ‫؍‬ 830 M). A second styrene molecule then binds CYP2E1 with higher affinity (K ss ‫؍‬ 110 M) and significantly improves oxidation to achieve a k cat of 6.3 nmol ⅐ min ؊1 ⅐ nmol CYP2E1 ؊1. The transition between these metabolic cycles coincides with reported styrene concentrations in blood from exposed workers; thus, this CYP2E1 mechanism may be relevant in vivo. Scaled modeling of the in vitro-positive allosteric mechanism for styrene metabolism to its in vivo clearance led to significant deviations from the traditional model based on Michaelis-Menten kinetics. Low styrene levels were notably much less toxic than generally assumed. We interrogated the allosteric mechanism using the CYP2E1-specific inhibitor and drug 4-methylpyrazole, which we have shown binds two CYP2E1 sites. From the current studies, styrene was a positive allosteric effector on 4-methylpyrazole binding, based on a 10-fold increase in 4-methylpyrazole binding affinity from K i 0.51 to K si 0.043 M. The inhibitor was a negative allosteric effector on styrene oxidation, because k cat decreased 6-fold to 0.98 nmol ⅐ min ؊1 ⅐ nmol CYP2E1 ؊1. Consequently, mixtures of styrene and other molecules can induce allosteric effects on binding and metabolism by CYP2E1 and thus mitigate the efficiency of their metabolism and corresponding effects on human health. Taken together, our elucidation of mechanisms for these allosteric reactions provides a powerful tool for further investigating the complexities of CYP2E1 metabolism of drugs and pollutants.

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Section: Discussionmentioning

confidence: 99%

CYP2E1 Metabolism of Styrene Involves Allostery

2012

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“…The common emphasis on erCYP2E1 metabolism would then underestimate styrene activation to the epoxide metabolite under these conditions. In fact, individuals, such as workers in the polymer industry, were previously exposed to higher levels of styrene and report plasma levels of styrene up to 22 μM (Löf 1986; Prieto 2002; Somorovska 1999); however, more recent studies of industrial workers suggest exposures are lower and result in styrene plasma levels reaching only 0.06 μM (Bonanni et al 2015). The difference between assumed and actual metabolic activation of styrene would be even larger for more common environmental exposures of less than 10 μM.…”

Section: Discussionmentioning

confidence: 99%

Subcellular localization of rat CYP2E1 impacts metabolic efficiency toward common substrates

et al. 2015

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Cytochrome P450 2E1 (CYP2E1) detoxifies or bioactivates many low molecular-weight compounds. Most knowledge about CYP2E1 activity relies on studies of the enzyme localized to endoplasmic reticulum (erCYP2E1); however, CYP2E1 undergoes transport to mitochondria (mtCYP2E1) and becomes metabolically active. We report the first comparison of in vitro steady-state kinetic profiles for erCYP2E1 and mtCYP2E1 oxidation of probe substrate 4-nitrophenol and pollutants styrene and aniline using subcellular fractions from rat liver. For all substrates, metabolic efficiency changed with substrate concentration for erCYP2E1 reflected in non-hyperbolic kinetic profiles but not for mtCYP2E1. Hyperbolic kinetic profiles for the mitochondrial enzyme were consistent with Michaelis-Menten mechanism in which metabolic efficiency was constant. By contrast, erCYP2E1 metabolism of 4-nitrophenol led to a loss of enzyme efficiency at high substrate concentrations when substrate inhibited the reaction. Similarly, aniline metabolism by erCYP2E1 demonstrated negative cooperativity as metabolic efficiency decreased with increasing substrate concentration. The opposite was observed for erCYP2E1 oxidation of styrene; the sigmoidal kinetic profile indicated increased efficiency at higher substrate concentrations. These mechanisms and CYP2E1 levels in mitochondria and endoplasmic reticulum were used to estimate the impact of CYP2E1 subcellular localization on metabolic flux of pollutants. Those models showed that erCYP2E1 mainly carries out aniline metabolism at all aniline concentrations. Conversely, mtCYP2E1 dominates styrene oxidation at low styrene concentrations and erCYP2E1 at higher concentrations. Taken together, subcellular localization of CYP2E1 results in distinctly different enzyme activities that could impact overall metabolic clearance and/or activation of substrates and thus impact the interpretation and prediction of toxicological outcomes.

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“…P&T sampling, in combination with chromatographic techniques, has also been successfully applied to the analysis of other sample matrices {e.g., APIs [47], urine and blood [71], fecal samples [72]} (Table 2). …”

Section: Other Samplesmentioning

confidence: 99%