Serum Institute of India Research Foundation, Hadapsar, Pune 411 028, India.
J Pharm Biomed Anal. 2013 Jun;80:203-12. doi: 10.1016/j.jpba.2013.03.001. Epub 2013 Mar 26.
Withania somnifera (WS) is one of the popular botanical medicines widely used in Ayurveda. Withanolides such as withaferin A (WA) and withanolide A (WLD) are its bioactive constituents reported as promising drug candidates in cancer and neurological disorders respectively. A new, selective and rapid high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method has been developed and validated for simultaneous determination of WA and WLD in mice plasma. Simple liquid-liquid extraction procedure was followed using ter-butyl methyl ether (TBME) for plasma sample pretreatment. Analytes were separated on Hypurity C18 column using methanol and ammonium acetate (95:5, v/v) as a mobile phase and detected by electrospray ionization in the multiple reaction monitoring (MRM) mode. The mass transition ion-pair was followed as m/z 471.3→281.2 for WA; m/z 437.2→292.2 for tianeptine (IS) and m/z 488.3→263.1 for WLD; m/z 315.9→270 for clonazepam (IS). The method showed excellent linearity (r(2)>0.997) over the concentration range of 0.484-117.880ng/ml for WA and from 0.476-116.050ng/ml for WLD. The lower limits of quantification (LLOQs) were found to be 0.484ng/ml and 0.476ng/ml for WA and WLD respectively. Precision (% CV) and accuracy (% bias) were found in the range of 3.7-14.3% and -14.4-4.0%, respectively. The validated method was successfully applied to a pharmacokinetic (PK) study for estimation of WA and WLD in mice plasma following oral administration of W. somnifera root aqueous extract (WSE). The PK study suggested rapid oral absorption of these withanolides. The PK study revealed that withaferin A has one and half times more relative bioavailability as compared to withanolide A.
睡茄（Withania somnifera）是一种广受欢迎的植物药，在阿育吠陀医学中被广泛应用。其生物活性成分包括醉茄内酯 A（WA）和醉茄内酯 A（WLD），分别被报道为癌症和神经紊乱的有前途的候选药物。本研究建立并验证了一种用于同时测定小鼠血浆中 WA 和 WLD 的新型、选择性和快速高效液相色谱-串联质谱（HPLC-MS/MS）方法。采用叔丁基甲醚（TBME）进行简单的液-液萃取，对血浆样品进行预处理。分析物在 Hypurity C18 柱上进行分离，以甲醇和乙酸铵（95:5，v/v）作为流动相，通过电喷雾电离在多重反应监测（MRM）模式下进行检测。质量过渡离子对分别为 m/z 471.3→281.2（WA）；m/z 437.2→292.2（替奈普汀，IS）和 m/z 488.3→263.1（WLD）；m/z 315.9→270（氯硝西泮，IS）。该方法在 WA 的浓度范围为 0.484-117.880ng/ml 和 WLD 的浓度范围为 0.476-116.050ng/ml 时表现出优异的线性（r(2)>0.997）。WA 和 WLD 的定量下限（LLOQ）分别为 0.484ng/ml 和 0.476ng/ml。精密度（%CV）和准确度（%偏差）分别在 3.7-14.3%和-14.4-4.0%的范围内。该方法已成功应用于口服睡茄根水提物（WSE）后 WA 和 WLD 在小鼠血浆中的药代动力学（PK）研究。PK 研究表明这些醉茄内酯吸收迅速。PK 研究表明，与醉茄内酯 A 相比，醉茄内酯 A 的相对生物利用度高 1.5 倍。
