New Methods for Resolution of Hydroxychloroquine by Forming Diastereomeric Salt and Adding Chiral Mobile Phase Agent on RP-HPLC

Hydroxychloroquine (HCQ), 2-([4-([7-Chloro-4-quinolyl]amino)pentyl]ethylamino)ethanol, exhibited significant biological activity, while its side effects cannot be overlooked. The RP-HPLC enantio-separation was investigated for cost-effective and convenient optical purity analysis of HCQ. The thermodynamic resolution of Rac-HCQ, driven by enthalpy and entropy, was achieved on the C18 column using Carboxymethyl-beta-cyclodextrin (CM-beta-CD) as the chiral mobile phase agent (CMPA). The effects of CCM-beta-CD, pH, and triethylamine (TEA) V% on the enantio-separation process were explored. Under the optimum conditions at 24 degrees C, the retention times for the two enantiomers were tR1=29.39min$$ {t}_{R1}=29.39\ \min $$ and tR2=32.42min$$ {t}_{R2}=32.42\ \min $$, resulting in Rs=1.87$$ {R}_s=1.87 $$. The resolution via diastereomeric salt formation of Rac-HCQ was developed to obtain the active pharmaceutical ingredient of single enantiomer S-HCQ. Di-p-Anisoyl-L-Tartaric Acid (L-DATA) was proved effective as the resolution agent for Rac-HCQ. Surprisingly, it was found that refluxing time was a key fact affecting the resolution efficiency, which meant the kinetic dominate during the process of the resolution. Four factors-solvent volume, refluxing time, filtration temperature, and molar ratio-were optimized using the single-factor method and the response surface method. Two cubic models were established, and the reliability was subsequently verified. Under the optimal conditions, the less soluble salt of 2L-DATA:S-HCQ was obtained with a yield of 96.9% and optical purity of 63.0%. The optical purity of this less soluble salt increases to 99.0% with a yield of 74.2% after three rounds recrystallization. Hydroxychloroquine enantiomers was selectivity separated by di-p-anisoyl-L-tartaric acid through forming diastereomeric salt. Using carboxymethyl-beta-cyclodextrin as the chiral mobile phase agent, the optical purity of hydroxychloroquine enantiomers in the less salt of S-Hydroxychloroquine2di-p-anisoyl-L-tartaric acid was determined conveniently and successfully on the RP-HPLC of OSD-C18 column. image