Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia.
National Organization for Drug Control and Research, Cairo P.O. Box 29, Egypt.
Molecules. 2021 Apr 6;26(7):2091. doi: 10.3390/molecules26072091.
A novel, fast and sensitive enantioselective HPLC assay with a new core-shell isopropyl carbamate cyclofructan 6 (superficially porous particle, SPP) chiral column (LarihcShell-P, LSP) was developed and validated for the enantiomeric separation and quantification of verapamil (VER) in rat plasma. The polar organic mobile phase composed of acetonitrile/methanol/trifluoroacetic acid/triethylamine (98:2:0.05: 0.025, ) and a flow rate of 0.5 mL/min was applied. Fluorescence detection set at excitation/emission wavelengths 280/313 nm was used and the whole analysis process was within 3.5 min, which is 10-fold lower than the previous reported HPLC methods in the literature. Propranolol was selected as the internal standard. The S-(-)- and R-(+)-VER enantiomers with the IS were extracted from rat plasma by utilizing Waters Oasis HLB C18 solid phase extraction cartridges without interference from endogenous compounds. The developed assay was validated following the US-FDA guidelines over the concentration range of 1-450 ng/mL (r ≥ 0.997) for each enantiomer (plasma) and the lower limit of quantification was 1 ng/mL for both isomers. The intra- and inter-day precisions were not more than 11.6% and the recoveries of S-(-)- and R-(+)-VER at all quality control levels ranged from 92.3% to 98.2%. The developed approach was successfully applied to the stereoselective pharmacokinetic study of VER enantiomers after oral administration of 10 mg/kg racemic VER to Wistar rats. It was found that S-(-)-VER established higher C and area under the concentration-time curve (AUC) values than the R-(+)-enantiomer. The newly developed approach is the first chiral HPLC for the enantiomeric separation and quantification of verapamil utilizing a core-shell isopropyl carbamate cyclofructan 6 chiral column in rat plasma within 3.5 min after solid phase extraction (SPE).
建立了一种新型的、快速的、灵敏的手性高效液相色谱分析方法，使用新型核壳型异丁基氨基甲酸酯环果糖 6（表面多孔颗粒，SPP）手性柱（LarihcShell-P，LSP），对大鼠血浆中维拉帕米（VER）的对映异构体进行分离和定量分析。流动相为乙腈/甲醇/三氟乙酸/三乙胺（98:2:0.05:0.025），流速为 0.5mL/min。采用荧光检测，激发/发射波长为 280/313nm，整个分析过程在 3.5min 内完成，比文献中以前报道的 HPLC 方法快 10 倍。选择普萘洛尔作为内标。S-（-）-和 R-（+）-VER 对映异构体与 IS 经 Waters Oasis HLB C18 固相萃取小柱提取，不受内源性化合物干扰。根据美国 FDA 指南，在每个对映体（血浆）的 1-450ng/mL 浓度范围内（r≥0.997）验证了该方法的线性关系，两种异构体的定量下限均为 1ng/mL。日内和日间精密度均不超过 11.6%，S-（-）-和 R-（+）-VER 在所有质控水平的回收率均在 92.3%-98.2%之间。该方法成功应用于口服 10mg/kg 消旋 VER 后 Wistar 大鼠 VER 对映体的立体选择性药代动力学研究。结果发现，S-（-）-VER 的 C 和浓度-时间曲线下面积（AUC）值均高于 R-（+）-对映体。该方法是在手相萃取后 3.5min 内，利用核壳型异丁基氨基甲酸酯环果糖 6 手性柱首次实现大鼠血浆中维拉帕米对映体分离和定量的手性高效液相色谱分析方法。
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