Abstract:

A new flow injection on-line oxidization fluorometry coupled to the technique of dialysis sampling was established for the determination of free thioridazine hydrochloride in serum, and applied to the investigation of binding action of thioridazine hydrochloride with bovine serum albumin(BSA). The thioridazine hydrochloride and BSA were mixed in different molar ratios in 0.050 mol/L phosphate buffer(containing 0.9% NaCl, pH=7.4), and incubated at (37±0.5) ℃ in a water bath. The dialysis sampler was utilized to sample free thioridazine hydrochloride from mixed solution with a relative dialytic efficiency of 8.3%. Then the thioridazine hydrochloride in dialysis solution was injected into carrier and on-line oxidized by lead dioxide solid-phase reactor into fluorescent product with a maximum excitation wavelength of 349 nm and a maximum emission wavelength of 429 nm. The fluorescence intensity measured was linear proportional with the concentration of free thioridazine hydrochloride in mixed solution over the range from 1×10^-5 to 2×10^-4 mol/L with the detection limit of 6×10^-6 mol/L. According to the fluorescence measurement results from mixed solution, the association constant(K) estimated for thioridazine hydrochloride-BSA binding and the number of the binding site(n) with Scatchard analysis were 1.2×10⁴ L/mol and 0.98, respectively. The acting force was suggested to be mainly hydrophobic and the distance between the acceptor and donor was 3.27 nm.

Key words: Flow injection fluorometry, Dialysis sampling, Thioridazine hydrochloride, Bovine serum albumin, Association constant