Akira Yamauchi

An enzyme immunoassay for plasma betamethasone

A sensitive enzyme immunoassay for plasma betamethasone was developed using betamethasone-3-(O-carboxymethyl)oxime-beta-D-galactosidase conjugate as a labelled antigen and 4-methylumbelliferyl-beta-D-galactoside as a fluorescence substrate. The performances of the enzyme immunoassay were compared with that of a radioimmunoassay using 3H-betamethasone and the same antiserum. The minimal detectable level for the enzyme immunoassay was 0.15 pg/tube or 0.15 ng/ml of plasma, which was remarkably more sensitive than the radioimmunoassay level of 10 pg/tube or 2 ng/ml of plasma. The specificity was sufficient, in particular, the cross reactivity of cortisol as 0.008%. However, the precision of the enzyme immunoassay was inferior to that of the radioimmunoassay.

Radioimmunoassay and gas chromatography/mass spectrometry for a novel antiglaucoma medication of a prostaglandin derivative, S-1033, in plasma

A radioimmunoassay (RIA) and a gas chromatographic/mass spectrometric (GC/MS) method for a new antiglaucoma medicament, the prostaglandin derivative sodium (5Z, 9 alpha, 11 alpha, 13E)-9,11-dihydroxyprosta-5, 13-dienoate (S-1033), in human and rabbit plasma were investigated. For a competitive RIA, antisera from rabbit and radioiodine-labeled S-1033 were prepared by immunizing a conjugate of S-1033 with bovine serum albumin and by the Bolton and Hunter method, respectively. Pretreatment by C18 solid-phase extraction (SPE) for rabbit plasma sample and further purification by high-performance liquid chromatography (HPLC) for human plasma samples followed by the RIA (SPE/RIA and HPLC/RIA, respectively) were developed. The assay recoveries of SPE/RIA and HPLC/RIA were both excellent and the limits of quantitation were 320 and 10 pg ml-1, respectively. GC/MS for plasma samples after solid-phase extraction and thin-layer chromatographic purification was also developed using deuterium-labeled S-1033 as internal standard. The limit of quantitation was 100 pg ml-1 in human or rabbit plasma. Rabbit plasma samples after administration of this drug were measured by SPE/RIA and GC/MS and the assay results from both methods agreed well. The SPE/RIA, HPLC/RIA and GC/MS assay methods were suitable for measuring samples from preclinical studies, clinical studies and cross-validation, respectively.