Design of experiment assisted concurrent enantioseparation of bupropion and hydroxybupropion by high‐performance thin‐layer chromatography

A simple and efficient high‐performance thin‐layer chromatographic method was developed for chiral separation of rac‐bupropion (BUP) and its active metabolite rac‐hydroxybupropion (HBUP). Design of experiment (DoE)‐based optimization was adopted instead of a conventional trial‐and‐error approach. The Box–Behnken design surface response model was used and the operating variables were optimized based on 17 trials design. The optimized method involved impregnation of chiral reagent, L(+)‐tartaric acid, in the stationary phase with simultaneous addition in the mobile phase, which consisted of acetonitrile : methanol : dichloromethane : 0.50% L‐tartaric acid (6.75:1.0:1.0:0.25, v/v/v/v). Under the optimized conditions, the resolution factor between the enantiomers of BUP and HBUP was 6.30 and 9.26, respectively. The limit of detection and limit of quantitation for (R)‐BUP, (S)‐BUP, (R,R)‐HBUP, and (S,S)‐HBUP were 9.23 and 30.78 ng spot−1, 10.32 and 34.40 ng spot−1, 12.19 and 40.65 ng spot−1, and 14.26 and 47.53 ng spot−1, respectively. The interaction of L‐tartaric acid with analytes and their retention behavior was thermodynamically investigated using van't Hoff's plots. The developed method was validated as per the International Conference on Harmonization guidelines. Finally, the method was successfully applied to resolve and quantify the enantiomeric content from marketed tablets as well as spiked plasma samples. A simple and efficient HPTLC method has been developed for chiral separation of rac‐bupropion and its active metabolite rac‐hydroxybupropion using design of experiment. The results obtained using Box‐Behnken design surface response model for the analysis of pharmaceutical preparations and also in human plasma demonstrate good practical utility, especially for resource limited settings. The developed analytical strategy provides an alternative to the existing sophisticated techniques used for the enantioseparation and quantification of rac‐bupropion and rac‐hydroxybupropion.