A radioimmunoassay to measure piretanide in human serum and urine.

This paper describes a specific radioimmunological method for determining the diuretic agent piretanide (4 phenoxy-3-(1-pyrrolidinyl)-5-sulfamoyl benzoic acid) in human serum, plasma and urine. The antiserum was raised in rabbits against an immunogen of piretanide coupled to bovine serum albumin. The iodinated hydroxy derivative of piretanide was used as tracer. The separation of free and antibody-bound piretanide was performed by precipitating the antibody-tracer complex by polyethylene glycol. The limit of detection was 4 ng/ml. Studies on specificity showed less than 0.5% cross-reactivity of an identified metabolite. Interassay reproducibility showed an average coefficient of variation of 7.3%, the interassay variation was 5%. A recovery experiment yielded 100.9% recovery. There is good agreement between parallel determinations of piretanide by RIA and HPLC in both human serum and urine.

A radioimmunoassay for determination of glibenclamide and other sulfonylureas.

An antiserum was prepared for the determination of glibenclamide and for the estimation of other commercially available sulfonylureas. Rabbits were immunized with a glibenclamide-BSA conjugate. Tritiated glibenclamide was used as the tracer. The assay was performed in the presence of 8-anilinonaphthalenesulfonic acid to displace glibenclamide bound to serum protein, and free and antibody bound tracer were separated by dextran-coated charcoal. For glibenclamide determination in serum and plasma the limit of detection was 3 ng/ml. Sensitivity calculated for the whole determination range was 102 cpm for a 10 % concentration difference. Specificity studies showed a cross-reaction of less than 0.1 % for glibenclamide metabolite M1 and 9 % for metabolite M2. Other sulfonylurea drugs display cross-reactivities from 0.1% (chlorpropamide) to 190% (gliquidone). Both intra-assay and inter-assay imprecision were below 10 %. Accuracy was established by comparison of the present method with HPLC. The assay was applied to the specific determination of glibenclamide in clinical trials and for diagnosing factitious hypoglycemia caused by sulfonylureas.