Radioimmunoassay for the determination of alosetron in human urine and saliva

Abstract

The development of a radioimmunoassay (RIA) for the sub-ng ml-1 determination of alosetron, a potent and selective 5HT3 receptor antagonist, in human urine and saliva is described. The antiserum was raised in Soay sheep following primary and booster immunizations with an immunogen prepared by conjugating alosetron-p-azobenzoic acid to bovine serum albumin (BSA). The radioligand consisted of alosetron specifically 125-iodinated on the 2-position of the imidazole group. The mean (+/- standard deviation) theoretical sensitivity (minimum detectable dose corresponding to the imprecision of the zero standard) of the RIA is 3.2 +/- 2.6 pg ml-1 (n = 12) of alosetron in assay diluent (0.1% m/v gelatine-0.05% m/v sodium azide in 0.1 mol l-1 phosphate buffer solution, pH 7.4). The working calibration range using 0.1 ml samples of saliva and 20-fold diluted urine is 0.10-6.40 ng ml-1 of alosetron. Urine samples were diluted prior to assay to overcome adverse matrix effects; consequently, the lower limit of quantification for undiluted urine is 2.0 ng ml-1 of alosetron. Inter- and intra-assay bias and imprecision over the working calibration range were generally < +/- 12% and < 13%, respectively, except at the 0.10 ng ml-1 alosetron level, where the corresponding values were < +/- 17.3% and < 20.2%. The antiserum was free from adverse cross-reactivity with either a synthetic precursor of alosetron or with four major metabolites of the drug.(ABSTRACT TRUNCATED AT 250 WORDS)

Application of a UPLC‐MS/MS method for the analysis of alosetron in human plasma to support a bioequivalence study in healthy males and females

“…The inter‐assay precision was 9.5%, and the accuracy was within 1.2% of nominal concentration. A radioimmunoassay method has also been described for determination of ALO in human urine and saliva samples taken from volunteers who had received a 1.0 mg single oral dose of the drug (Wring et al ., ). The working calibration range was established from 0.10–6.40 ng/mL using 0.1 mL samples of saliva and 20‐fold diluted urine samples.…”

Development of a Forced Degradation Profile of Alosetron by Single Mode Reversed-Phase HPLC, LC-MS, and its Validation

“…Wring SA et al . have reported the radioimmunoassay method for the estimation of alosetron in human urine and saliva [ 6 ]. Karthik et al .…”

A novel reverse phase high-performance liquid chromatography (RP-HPLC–UV) method for the estimation of alosetron HCl and its validation

“…It is based on the limit of detection (LOD) and limit of quantification (LOQ) values. ICH guideline [6] specifies three different types of methods to determine the LOD and LOQ values which include visual evaluation, signal to noise ratio, and the standard deviation of response and the slope of the calibration curve. In the current study, the limit of detection and limit of quantification were based on the second approach and were calculated according to S/N 3:1 and 10:1, respectively.…”

“…Thomas L. Lloyd et al have reported estimation of alosetron in human plasma or serum by high-performance liquid chromatography employing robotic sample preparation, cyano propyl stationary phase, and fluorescence detection [5]. S.A. Wring et al have reported the radioimmunoassay (RIA) method for the estimation of alosetron in human urine and saliva [6].…”