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Metabolism of Benzalkonium Chlorides by Human Hepatic Cytochromes P450
journal contribution
posted on 2019-12-02, 22:29 authored by Ryan P. Seguin, Josi M. Herron, Vanessa A. Lopez, Joseph L. Dempsey, Libin XuBenzalkonium chlorides (BACs) are widely used as disinfectants
in cleaning products, medical products, and the food processing industry.
Despite a wide range of reported toxicities, limited studies have
been conducted on the metabolism of these compounds in animal models
and none in human-derived cells or tissues. In this work, we report
on the metabolism of BACs in human liver microsomes (HLM) and by recombinant
human hepatic cytochrome P450 (CYP) enzymes. BAC metabolism in HLM
was NADPH-dependent and displayed apparent half-lives that increased
with BAC alkyl chain length (C10 < C12 <
C14 < C16), suggesting enhanced metabolic
stability of the more lipophilic, longer chain BACs. Metabolites of d7-benzyl labeled BAC substrates retained all
deuteriums and there was no evidence of N-dealkylation.
Tandem mass spectrometry fragmentation of BAC metabolites confirmed
that oxidation occurs on the alkyl chain region. Major metabolites
of C10-BAC were identified as ω-hydroxy-, (ω-1)-hydroxy-,
(ω, ω-1)-diol-, (ω-1)-ketone-, and ω-carboxylic
acid-C10-BAC by liquid chromatography–mass spectrometry
comparison with synthetic standards. In a screen of hepatic CYP isoforms,
recombinant CYP2D6, CYP4F2, and CYP4F12 consumed substantial quantities
of BAC substrates and produced the major microsomal metabolites. The
use of potent pan-CYP4 inhibitor HET0016, the specific CYP2D6 inhibitor
quinidine, or both confirmed major contributions of CYP4- and CYP2D6-mediated
metabolism in the microsomal disappearance of BACs. Kinetic characterization
of C10-BAC metabolite formation in HLM demonstrated robust
Michaelis–Menten kinetic parameters for ω-hydroxylation
(Vmax = 380 pmol/min/mg, Km = 0.69 μM) and (ω-1)-hydroxylation (Vmax = 126 pmol/min/mg, Km = 0.13 μM) reactions. This work illustrates important
roles for CYP4-mediated ω-hydroxylation and CYP2D6/CYP4-mediated
(ω-1)-hydroxylation during the hepatic elimination of BACs,
an environmental contaminant of emerging concern. Furthermore, we
demonstrate that CYP-mediated oxidation of C10-BAC mitigates
the potent inhibition of cholesterol biosynthesis exhibited by this
short-chain BAC.
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Keywords
food processing industryV maxCYP 4-mediated ω- hydroxylationBAC substratesalkyl chain region0.69 μ MCYP 2D CYP 4FHuman Hepatic Cytochromes P 450 Benzalkonium chlorideshepatic CYP isoformsHLMCYP 2D metabolismC 10BAC metabolite formationtandem mass spectrometry fragmentationHEThepatic cytochrome P 4500.13 μ Mω- carboxylic acid-C 10BAC alkyl chain lengthCYP 2D inhibitor quinidine
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