Caenorhabditis elegans is becoming an important model in pharmacology and toxicology (Leung et al., 2008), but there is little biochemical information regarding its xenobiotic metabolic capacity. The current study investigated the in vivo activity of C. elegans with two human cytochrome P450 (CYP) substrates, phenacetin and testosterone, which are metabolized by human CYP1A2 and CYP3A enzymes, respectively, into acetamidophenol and 6β-hydroxytestosterone via deethylation and hydroxylation reactions.
Each experiment included (a) 50 ml liquid cultures with 30 µM substrate, a mixed-stage N2 (Bristol) population, and OP50 bacteria in S medium; (b) N2 cultures without OP50; (c) OP50 cultures without N2;, and (d) substrate-only controls. All cultures were done in triplicate and incubated on a shaker at 20 °C for 24 hr. OP50 and C. elegans were isolated afterward using centrifugation and sucrose flotation, respectively. The resultant extract was homogenized using sonication and resuspended in 1.5 ml 0.1 N NaCl solution. The culture medium and homogenate were analyzed for substrates and metabolites using HPLC-MS/MS (Micromass Quattro Ultima, Beverly, MA) and published methods (Walsky and Obach, 2004).
C. elegans generated the human CYP3A diagnostic metabolite 6β-hydroxytestosterone in liquid culture with or without OP50 (Table 1). The medium concentration of testosterone in the N2 cultures was nearly eliminated after 24 hr, but was unchanged in both substrate controls and OP50 cultures. This indicated that C. elegans either metabolized or bioconcentrated testosterone. 6β-hydroxytestosterone was detected in the N2 homogenate, demonstrating human CYP3A-like metabolism.
The results of the phenacetin experiment were less clear-cut. The medium concentration of phenacetin in the N2 cultures but not the OP50-only cultures was reduced after 24 hr incubation, but acetamidophenol was never detected at levels exceeding 0.04 µM in any culture medium or homogenate (Table 1). This is likely to result from further metabolism to chemical forms that we did not measure, but testing this hypothesis will require additional experiments.
The current findings suggest that C. elegans possessed human CYP3A-like activities, consistent with previous evidence for a functional role of human CYP3 homologs in C. elegans genome (Leung et al., 2010). However, the finding of modest CYP1A2-like activity was surprising based on previous work (Leung et al., 2010). Clearly further experiments are needed to explore how C. elegans metabolizes pharmaceuticals and toxicants (Kosel et al., 2011; Shäfer et al., 2009).
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References
Kosel M, Wild W, Bell A, Rothe M, Lindschau C, Steinberg CE, Schunck WH, Menzel R. (2011) Eicosanoid formation by a cytochrome P450 isoform expressed in the pharynx of Caenorhabditis elegans. Biochem J 435, 689-700. 
Leung MC, Goldstone JV, Boyd WA, Freedman JH, Meyer JN. (2010) Caenorhabditis elegans generates biologically relevant levels of genotoxic metabolites from aflatoxin B1 but not benzo[a]pyrene in vivo. Toxicol Sci 118, 444-53.
Leung MC, Williams PL, Benedetto A, Au C, Helmcke KJ, Aschner M, Meyer JN. (2008) Caenorhabditis elegans: an emerging model in biomedical and environmental toxicology. Toxicol Sci 106, 5-28.
Shäfer P, Müller M, Krüger A, Steinberg CE, Menzel R. (2009) Cytochrome P450-dependent metabolism of PCB52 in the nematode Caenorhabitis elegans. Arch Biochem Biophys 488, 60-68.
Walsky RL, Obach RS. (2004) Validated assays for human cytochrome P450 activities. Drug Metab Dispos 32, 647-660.
