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

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.

A novel analytical approach towards in-vitro bile acid binding studies to Colesevelam Hydrochloride tablets: An ultra-high performance liquid chromatography tandem mass spectrometric method.

Colesevelam hydrochloride is a bile acid sequestrant used as a low density lipoprotein (LDL) reducing agent in hyperlipidemia with an additional advantage to improve glycemic control in type 2 diabetes patients. The objective of the study was to develop and validate a liquid chromatography tandem mass spectroscopic method for the simultaneous in-vitro estimation of bile acid salts of Glycocholic acid (GC), Glycochenodeoxycholic acid (GCDC) and Taurodeoxycholic acid (TDC) and its application in performing in-vitro binding study with Colesevelam Hydrochloride tablets. The method was developed using C-18 (50 x 4.6 mm, 3 µm) column with detection on negative ion mode and acquisition time of 3.5 min. The calibration range was linear from 0.0002 mM to 0.0065 mM for GC, 0.0002 mM to 0.0065 mM for GCDC and 0.0001 mM to 0.0021 mM for TDC. The precision was less than 3.0% and accuracy was found well within the range of 85 to 115%. The validated method was further applied to conduct in-vitro equilibrium binding study. The data was subjected to Langmuir isotherm and affinity constant (k1) and capacity constant (k2) were calculated.