Spectrophotometric determination of choline in pharmaceutical formulations via host-guest complexation with a biomimetic calixarene receptor

Choline is a quaternary ammonium bioactive compound that is used as a dietary supplement. Direct determination of choline in pharmaceutical formulations is a challenging analytical task. The strong UV absorbance of p-sulfonatocalix[4]arene (SC4) and its outstanding complexation properties towards quaternary ammonium compounds have been utilized to develop a method to detect and quantify choline in aqueous solution. This method has been applied to a commercial pharmaceutical choline formulation showing results with no statistically significant difference from those obtained by the official U.S. Pharmacopeial method. The method was validated according to ICH guidelines and the limit of detection for choline was calculated to be 7.2 x 10(-6) mol L-1 (1.30 mu g mL(-1)). The host-guest complexation between SC4 and choline were investigated using UV and H-1 NMR spectroscopy and DFT calculations. The peak amplitude of the host-guest complexes formed were found to be linearly proportional to the concentration of the choline and L-carnitine analytes in the ranges of 1.0 x 10(-5) to 1.6 x 10(-4) mol L-1 and 1.0 x 10(-5) to 1.8 x 10(-4) mol L-1 respectively. The reaction stoichiometry between either choline or L-carnitine with SC4 in the formed complexes were determined to be 1:1 while the stability constants for the complexes formed were estimated to be 8.3 x 10(4) M-1 and 2.8 x 10(4) M-1 in the case of choline and L-carnitine, respectively. Both H-1 NMR spectroscopy and DFT calculations suggest the complexation between choline and SC4 involves the insertion of the trimethylammonium group of choline into the cavity of SC4 in agreement with previously published X-ray data. The spectrophotometric method presented here should not be limited to the determination of choline. The linear relationship between the peak amplitude of the host-guest complex formed between SC4 and L-carnitine presented here suggests the applicability of the method for the detection of other small molecular species possessing a quaternary ammonium group.