Simultaneous Determination of Reserpine, Rescinnamine, and Yohimbine in Human Plasma by Ultraperformance Liquid Chromatography Tandem Mass Spectrometry
A sensitive and selective UPLC-MS/MS method was developed and validated for the determination of three indolic alkaloids (reserpine, rescinnamine, and yohimbine) in human plasma using papaverine as internal standard (IS). After a one step protein precipitation with acetonitrile, separation was carried out using C18 column (50 × 2.1？mm, i.d. 1.7？μm) and mobile phase consisting of acetonitrile？:？water？:？formic acid (60？:？40？:？0.1%, v/v/v) pumped at a flow rate of 0.2？mL/min. The mass spectrometric determination was carried out using an electrospray interface operated in the positive mode with multiple reaction monitoring (MRM) mode. The precursor to product ion transitions of m/z 609.32？>？195.01, m/z 635.34？>？221.03, m/z 355.19？>？144, and m/z 340.15？>？202.02 were selected for the quantification of reserpine, rescinnamine, yohimbine, and IS, respectively. The analytical response was found to be linear in the range of 0.36–400, 0.27–300, and 0.23–250？ng/mL with lower limit of quantification of 0.36, 0.27, and 0.23？ng/mL for reserpine, rescinnamine, and yohimbine, respectively. Validation was made following official guidelines. The proposed method enabled reproducible results and hence could be reliable for pharmacokinetic and toxicological analysis. 1. Introduction Rauwolfia serpentina (family: Apocynaceae) is a medicinally important herb commonly known as Indian snake root. It has been used for centuries in ayurvedic medicines because of their valuable therapeutic actions especially in the treatment of hypertension and psychotic disorders like schizophrenia, anxiety, insomnia, insanity, and so forth [1, 2]. Rauwolfia serpentina has been also used for the treatment of skin cancers, burns, eczema, and snake bite [3, 4]. Rauvolfia verticillata is another traditional Chinese medicinal plant that also belongs to the same Apocynaceae family and also was been used for hypertension and cardiac disorders [5]. Both Rauwolfia serpentine and Rauvolfia verticillata contains pharmacologically active indolic alkaloids, namely, reserpine, rescinnamine, and yohimbine (their chemical structures are given in Figure 1) [5, 6]. Figure 1: Chemical structure of reserpine (a) rescinnamine (b) yohimbine (c) and papaverine (d). Various analytical methods have been reported for the determination of reserpine, rescinnamine, and yohimbine in their plant extracts, pharmaceutical preparation, and biological fluids. Two methods based on high-performance liquid chromatography (HPLC) have been reported for the analysis of Rauwolfia serpentine being one using absorption photometric
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