Quantitative determination of zwitterionic detergents using salt-induced phase separation of Triton X-100

Zwitterionic detergents interfere with the salt-induced phase separation for nonionic detergents in a concentration-dependent manner by shifting the normal cloud point of nonionic detergents to a higher ionic strength at room temperature. This phenomenon was used to determine the concentration of the zwitterionic detergents CHAPS, CHAPSO, and sulfobetaine SB-12 in solution by titration with ammonium sulfate in the presence of Triton X-100. Among the ionic detergents tested, the method was only applicable to sodium cholate. The assay can be used to control the removal of zwitterionic detergents during the reconstitution of membrane proteins in liposomes. However, it cannot be used to determine the specific binding of zwitterionic detergents to highly diluted, pure membrane proteins because of the limited sensitivity. Neither proteins nor phospholipids interfered with this method at concentrations up to 20 mg/ml of test solution (human serum albumin) or 10 mg/ml (phospholipids), respectively. Since the assay is based on the competition between salts and nonionic detergents for water molecules, it is important to equalize the ionic strength of samples and calibration standards.