High- performance liquid chromatography method applied to investigate mechanism, kinetics, isotherm, and thermodynamics of bile acid adsorption onto bile acid sequestrants

Abstract

The purpose of the study was to develop and validate a high-performance liquid chromatography (HPLC) method which can be further applied to understand the mechanism, kinetics, isotherm, and thermodynamics of bile acid adsorption onto bile acid sequestrants. To investigate these properties a HPLC method was developed using peerless C-8 (150 x 4.6 mm, 5 µm) column with a detection wavelength of 200 nm and run time of about 12.5 min. Bile salts glycocholic (GC), glycochenodeoxycholic (GCDC), and taurodeoxycholic acid (TDC), were used and colesevelam hydrochloride was employed as the bile acid sequestrant. The calibration range was found linear from 10 to 6500 mgL−1 for GC and GCDC and 4to 2400 mg L−1 for TDC. The precision was less than 8.8% and accuracy was found well within the range of 85 to 115%. On treating the data with various established models, it was known that, the adsorption kinetics followed the pseudo second order equation indicating chemisorption mechanism. Equilibrium isotherms revealed that the linear form of Langmuir model was the best fit. The separation factor (RL) calculated revealed that the reaction is favorable and reversible. The positive value of heat of sorption (B) calculated from Temkin model indicated towards the exothermic nature of adsorption. The adsorption energy (E) calculated from Dubinin-Kaganer-Radushkevich model was found to be greater than 8 KJmol−1 conforming chemisorption mechanism. The Gibbs free energy calculated established the affinity of bile salts as TDC > GCDC > GC.