Flow-injection chemiluminescence detection for studying protein binding of terbutaline sulfate with on-line microdialysis sampling

Abstract

The binding of terbutaline sulfate to bovine serum albumin was studied in vitro using the technique of microdialysis sampling combined with flow-injection chemiluminescence analysis (FIA-CL). In the presence of formaldehyde, terbutaline sulfate can be oxidized by KMnO 4 to produce high chemiluminescence emission in sulfate acid media. The concentration of terbutaline sulfate is proportional with the CL intensity in the range of 1×10 −7–2×10 −5 mol l −1 with a detection limit of 3×10 −8 mol l −1. The drug and protein were mixed in different molar ratios in 0.067 mol l −1 phosphate buffer, pH 7.4, and incubated at 37 °C in a water bath. The microdialysis probe was utilized to sample the mixed solution at a perfusion rate of 5 μl min −1 and the dialytic efficiency of terbutaline sulfate under the experimental conditions was 26.3%. The data obtained by proposed microdialysis flow-injection chemiluminescence method was analyzed with Scrathard analysis and Klotz plot. The estimated association constant ( K) and the number of the binding site ( n) on one molecule of BSA by Scrathard analysis were 4.11×10 4 l mol −1 and 1.06, respectively. The proposed system proved that FIA-CL coupled with on-line microdialysis sampling is a simple and reliable technique for the study of drug–protein interaction.