title: High-performance liquid chromatographic determination of N-[2(S)-(mercaptomethyl)-3-(2-methylphenyl)-1-oxopropyl]-L-methionine, the active plasma metabolite of a prodrug atriopeptidase inhibitor (SCH 42495), using a thiol selective (Au/Hg) amperometric detector.
abstract: A high-performance liquid chromatographic assay for the determination of N-[2(S)-(mercaptomethyl)-3-(2-methylphenyl)-1-oxopropyl]-L-methionine (SCH 42354; II), the active metabolite of the atriopeptidase inhibitor prodrug, N-[2(S)-(acetylthiomethyl)-3-(2-methylphenyl)-1-oxopropyl]-L-methi onine ethyl ester (SCH 42495; I), in human plasma was validated for use in clinical pharmacokinetic studies. Plasma (200 microliters) was processed by protein precipitation with acetone containing the internal standard, N-[2(S)-(mercaptomethyl)-3-(2-methylphenyl)-1-oxopropyl]-L- ethionine (III). Compound II was recovered (ca. 90%) in the supernatant after centrifugation and prepared for injection by the addition of 0.15 M monochloroacetic acid containing 0.2 mM EDTA. Separation of II and III was accomplished on commercially available reversed-phase C8 columns designed for the separation of basic compounds. Both compounds were detected using amperometric detection (+0.125 V versus Ag/AgCl) on a thin-layer Au/Hg amalgam electrode. The lower limit of quantitation was 10 ng/ml, where the inter-assay precision (coefficient of variation) was +/- 11.4% and the inter-assay accuracy (bias) was +1.0%. No endogenous interferences were observed in the extracts obtained from drug-free plasma. The detector response (using either peak area or height ratios of II to III) was linear from 0.01 to 1.0 micrograms/ml. Compound II was stable in plasma supplemented with EDTA and sodium hydrogensulfite for at least 3 months when stored frozen at -78 degrees C; no significant decomposition of II was observed following three freeze-thaw cycles. The feasibility of this liquid chromatographic assay with electrochemical detection was demonstrated with plasma samples from hypertensive subjects administered 100 mg of compound I.
authors.affiliation: Department of Drug Metabolism and Pharmacokinetics, Schering-Plough Research Institute, Bloomfield, NJ 07003.
authors.forename: K B
authors.initials: KB
authors.lastname: Alton
authors.forename: A
authors.initials: A
authors.lastname: Hernandez
authors.forename: N
authors.initials: N
authors.lastname: Alvarez
authors.forename: J E
authors.initials: JE
authors.lastname: Patrick
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006046:Gold; D006801:Humans; D006973:Hypertension / Q000097:blood; D008628:Mercury; D008715:Methionine / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism; D015260:Neprilysin / Q000037:antagonists & inhibitors* / Q000097:blood / Q000378:metabolism; D013438:Sulfhydryl Compounds
chemical_list: D013438:Sulfhydryl Compounds; C069018:SCH 42495; C086374:CGS 26129; D006046:Gold; D008715:Methionine; D015260:Neprilysin; D008628:Mercury