title: In vitro metabolism of [14C]-benalaxyl in hepatocytes of rats, dogs and humans.
abstract: The in vitro comparative animal metabolism study is now a data requirement under EU Directive 1107/2009 for registration of plant protection products. This type of study helps determine the extent of metabolism of a chemical in each surrogate species and whether any unique human metabolite(s) are formed. In the present study, metabolism of racemic [14C]-benalaxyl, a fungicide was investigated in cryopreserved rat, dog and human hepatocytes. The metabolites generated were identified/characterized by LC/MS/MS with radiometric detection and comparison with reference standards. [14C]-glucuronide conjugates of benalaxyl metabolites in rat, dog and human hepatocytes were confirmed via additional experiments in which known reference standards were incubated with dog liver microsomes in the presence of UDPGA. After 4 h of incubation, benalaxyl was extensively metabolized in all the species with the following trend: dog (100%) > human (86%) > rat (75%). In all species, the major metabolic pathways consisted of hydroxylation of the methyl group in the xylene moiety to 2-hydroxymethyl-benalaxyl, further oxidation to its carboxylic acid analogue (benalaxyl-2-benzoic acid), and hydrolysis of the methyl ester to yield benalaxyl acid or 2-hydroxymethyl benalaxyl acid. In addition, glucuronidation of phase I metabolites occurred in all species, to a higher extent in dog hepatocytes in which 2-hydroxymethyl-benalaxyl-glucuronide conjugate constituted the most significant metabolite. No major unique metabolite was observed in human hepatocytes. Also, benalaxyl did not undergo stereo-selective metabolism in rat or human hepatocytes.
authors.affiliation: Global Regulatory Science, FMC Agricultural Solutions, 701 PrincetonSouth Corporate Center, Ewing, NJ, 08628, USA. Electronic address: Gopinath.Nallani@fmc.com.
authors.forename: Gopinath C
authors.initials: GC
authors.lastname: Nallani
authors.affiliation: Global Regulatory Science, FMC Agricultural Solutions, 701 PrincetonSouth Corporate Center, Ewing, NJ, 08628, USA.
authors.forename: Shaaban F
authors.initials: SF
authors.lastname: ElNaggar
authors.affiliation: Frontage Laboratories, 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Li
authors.initials: L
authors.lastname: Shen
authors.affiliation: Global Regulatory Science, FMC Agricultural Solutions, 701 PrincetonSouth Corporate Center, Ewing, NJ, 08628, USA.
authors.forename: Appavu
authors.initials: A
authors.lastname: Chandrasekaran
mesh_terms: D000409:Alanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity; D000818:Animals; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D015925:Cryopreservation; D004285:Dogs; D005659:Fungicides, Industrial / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D020719:Glucuronides / Q000378:metabolism; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D008862:Microsomes, Liver / Q000378:metabolism; D015394:Molecular Structure; D010084:Oxidation-Reduction; D051381:Rats; D018570:Risk Assessment; D013045:Species Specificity; D053719:Tandem Mass Spectrometry; D018675:Toxicity Tests
chemical_list: D005659:Fungicides, Industrial; D020719:Glucuronides; C052866:benalaxyl; D000409:Alanine
keywords: Benalaxyl; Comparative in vitro metabolism; Cryopreserved; Hepatocytes