Quantitative analysis of bucillamine in blood using high‐performance liquid chromatography–mass spectrometry technique

A fast and sensitive method has been developed and validated for the determination of bucillamine in human blood by derivatizing the free sulfhydryl groups with isobutyl acrylate (IA), by APCI‐LC/MS/MS. The collected blood sample was immediately mixed with a mixture of IA and 0.05 m Tris(hydroxymethyl)aminomethane hydrochloride (Tris–HCl) buffer, pH 9.2, to stabilize the sulfyhydryl moieties. The derivatized samples were then extracted by protein precipitation, evaporated, reconstituted and injected using an LC‐APCI/MS/MS instrument. Separation was achieved using a C 18 analytical column and a gradient mobile phase within a chromatographic run time of 5 min. A quadratic (weighted 1/concentration 2 ) relationship was observed during validation over a concentration range of 0.4–40 µg/mL with a correlation value of r ≥ 0.9966. The inter‐batch precision and accuracy at low, medium and high concentrations were 8.1, 8.4 and 7.3%; 113.3, 104.9 and 103.9%, respectively, and the intra‐batch precision and accuracy at low, medium and high concentrations were 7.7, 5.4 and 2.7%; 105.1, 111.9 and 113.2%, respectively. Copyright © 2004 John Wiley & Sons, Ltd.