Ultraviolet spectrophotometric, solvent extraction, high-performance liquid chromatographic and polarographic study of the H+/K+ adenosine triphosphatase inhibitor (SK&F 95601) and its acid-catalysed degradation products

Abstract

pK_a values of 3.3, 5.35, 9.0 and 12.0 have been established for the H⁺/K⁺ adenosine triphosphatase inhibitor SK&F 95601 (1) with optimum solvent extractability into ethyl acetate in the pH range 7.5–8.5. Following solvent or solid-phase extraction of 1 × 10^–6 mol l^–11 from urine, a recovery of 92.3% was calculated using quantification by high-performance liquid chromatography (HPLC) with ultraviolet detection at 300 nm. The complex degradation of 1 at pH 2 can be followed by HPLC and differential-pulse polarography, which results in the detection of D₁(the sulfenamide) and D₂(the sulfide) in a 10 min time scale. Differential-pulse polarography was also used in situ to follow the reaction between a thiol (2-mercaptoethanol) and 1 in 1 × 10^–2 mol l^–1 HCl as a simulation of its believed reaction in vivo.