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Toxicokinetics of 11 Gelsemium Alkaloids in Rats by UPLC-MS/MS

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Toxicokinetics of 11 Gelsemium Alkaloids in Rats by UPLC-MS/MS Hindawi BioMed Research International Volume 2020, Article ID 8247270, 13 pages https://doi.org/10.1155/2020/8247270 Research Article Toxicokinetics of 11 Gelsemium Alkaloids in Rats by UPLC- MS/MS Xiuwei Shen ,1 Jianshe Ma ,2 Xianqin Wang ,3 Congcong Wen ,4 and Meiling Zhang 3 1Ruian People’s Hospital, The Third Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, China 2School of Basic Medicine, Wenzhou Medical University, Wenzhou 325035, China 3Analytical and Testing Center, School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou 325035, China 4Laboratory Animal Center of Wenzhou Medical University, Wenzhou 325035, China Correspondence should be addressed to Meiling Zhang; [email protected] Received 22 April 2020; Revised 1 June 2020; Accepted 16 June 2020; Published 15 July 2020 Academic Editor: Damião Pergentino de Sousa Copyright © 2020 Xiuwei Shen et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Gelsemium elegans (Gardn. & Champ.) Benth. is a plant belonging to the genus Gelsemium (family Gelsemiaceae), and its main components are alkaloids. It is a Chinese traditional medicinal plant and notoriously known as a highly toxic medicine. However, a method has not yet been found for the simultaneous detection of 11 Gelsemium alkaloids in rat plasma, and the toxicokinetics of 11 Gelsemium alkaloids after intravenous administration has not been reported. In this work, we have developed a sensitive and rapid method of ultraperformance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) for the detection of 11 Gelsemium alkaloids in rat plasma. The toxicokinetic behavior was also investigated, so as to provide a reference of the scientific properties of Gelsemium elegans and improve the efficacy and safety of drugs. Sixty-six Sprague-Dawley rats were randomly divided into 11 groups, six rats in each group. Each group was intravenously given one alkaloid (0.1 mg/kg), respectively. A Waters UPLC BEH C18 column (50 mm × 2:1mm,1.7μm) was used for chromatographic separation. Methanol and water (containing 0.1% formic acid) were used for the mobile phase with gradient elution. Multiple reactions were monitored, and positive electrospray ionization was used for quantitative analysis. The precision was less than 16%, and the accuracy was between 86.9% and 113.2%. The extraction efficiency was better than 75.8%, and the matrix effects ranged from 88.5% to 107.8%. The calibration curves were in the range of 0.1–200 ng/mL, with a correlation coefficient (R2) greater than 0.995. The UPLC-MS/MS method was successfully applied to the toxicokinetics of 11 Gelsemium alkaloids in rats after intravenous administration (0.1 mg/kg for each alkaloid). The results of the toxicokinetics provide a basis for the pharmacology and toxicology of Gelsemium alkaloids and scientific evidence for the clinical use of Gelsemium alkaloids. 1. Introduction tension. These officinal values have attracted great attention from the international medical and pharmaceutical circles About 12,000 kinds of alkaloids have been found, most of [4]. Gelsemium elegans (Gardn. & Champ.) Benth. is a plant which have relatively strong physiological activity. Many belonging to the genus Gelsemium (family Gelsemiaceae), alkaloids have been widely used in the field of medicine [1]. whose main components are alkaloids. It is warm, pungent, For example, morphine plays an analgesic role in opium bitter, and poisonous. It is mainly distributed in the Fujian [2]. Codeine, also an alkaloid, can relieve a cough, and ephed- and Zhejiang provinces in China. It is a Chinese traditional rine plays an antiasthmatic role in ephedra [3]. Chinese medicinal plant and also a world-famous highly toxic medi- herbal medicines often contain poisonous alkaloids, which cine [5]. In China, it is mainly used for external application have the functions of antitumor, antivirus, antiplatelet aggre- because of its high toxicity. It can dispel wind and blood sta- gation, anti-inflammatory, antiarrhythmia, and antihyper- sis, reduce swelling, relieve pain, and kill insects. In recent 2 BioMed Research International years, pharmacological studies have shown that Gelsemium spectrometer was obtained from Waters Corp. (Milford, elegans has many functions, including analgesic effects, MA, USA). sedation, anti-inflammation, mydriatic effects, antitumor, Eleven Gelsemium alkaloids (humantenirine, humante- reduced heart rate, heart contraction inhibition, increasing nine, akuammidine, gelsevirine, rankinidine, n-methoxyanhy- blood pressure, and improving immunity [6, 7]. The antitu- drovobasinediol, gelsenicine, gelsemine, koumine, koumidine, mor effect of Gelsemium elegans provides an attractive pros- and sempervirine) and IS were separated using a Waters pect for research and development. Indole alkaloids are the UPLC® BEH C18 column (50 mm × 2:1mm,1.7μm) main- main chemical components in Gelsemium elegans and the tained at 40°C. The mobile phase consisted of methanol and main source of its toxicity. However, as the toxic dose is close water (containing 0.1% formic acid). The gradient elution to the effective dose, poisoning events of the Gelsemium was as follows: from 0 to 0.2 min, the methanol was kept at elegans are common in clinical practice. Seventeen kinds of 10%; from 0.2 to 2.0 min, methanol increased from 10% to poisonous alkaloids have been isolated from Gelsemium 80%; from 2.0 to 2.5 min, the methanol was kept at 80%; from elegans, among which koumine is the most abundant, 2.5 to 2.8 min, methanol was changed from 80% to 10%; and at followed by gelsemine. As the alkaloid types and contents last, methanol was kept at 10% for 2.2 min. The flow rate was from different places and different plant parts of Gelsemium set at 0.4 mL/min. elegans are slightly different, there are currently not many Nitrogen was used for the cone gas (50 L/h) and desolva- reports regarding the toxicokinetics of Gelsemium alkaloids tion gas (900 L/h). The capillary voltage was 2.5 kV, the in vivo [8–10]. It has been a popular research topic to analyze source temperature was 150°C, and the desolvation tem- alkaloids from different sources in complex systems quickly, perature was 450°C. The employed collision gas for fragmen- sensitively, and reliably. UPLC-MS/MS, because of its flexi- tation in the multiple reaction monitoring (MRM) mode was bility, exhibits advantages of good sensitivity, excellent sepa- argon. The MRM mode (Table 1) was used for quantitative ration ability, wide application range, and strong specificity, analysis in the electrospray ionization (ESI) positive interface which is why it is widely used in alkaloid analysis [11–14]. (Figure 2). There have been methods reported for the determination of Gelsemium alkaloids, including high performance liquid 2.3. Stock Solutions. Stock solutions of 100 μg/mL each of chromatography-tandem (HPLC) [15, 16], ultraperformance fl humantenirine, humantenine, akuammidine, gelsevirine, ran- liquid chromatography-quadrupole-time of ight mass kinidine, n-methoxyanhydrovobasinediol, gelsenicine, gelse- spectrometry (UPLC-Q-TOF/MS) [17, 18], liquid chromato- – mine, koumine, koumidine, sempervirine, and IS were graphy/tandem mass spectrometry (LC/MS/MS) [19 23], prepared in methanol. The working standard solutions of each and ultraperformance liquid chromatography/tandem mass – of the 11 Gelsemium alkaloids were prepared by dilution of the spectrometry (UPLC-MS/MS) [24 28] in vivo. However, a stock solution with methanol. The stock solutions and work- method has not been found for the simultaneous deter- ing standard solutions were stored at 4°C. mination of 11 Gelsemium alkaloids in rat plasma, and the The calibration standards were prepared by spiking blank toxicokinetics of 11 Gelsemium alkaloids (humantenirine, rat plasma with appropriate amounts of the 11 Gelsemium humantenine, akuammidine, gelsevirine, rankinidine, n-meth- alkaloid working standard solutions. Calibration plots of oxyanhydrovobasinediol, gelsenicine, gelsemine, koumine, each of the 11 Gelsemium alkaloids were constructed in the koumidine, and sempervirine) after intravenous administra- 0.1-200 ng/mL range for plasma (0.1, 0.5, 2, 10, 20, 50, 100, tion has not been reported. In this work, a sensitive and rapid and 200 ng/mL). Quality-control (QC) samples (0.4, 18, and method of UPLC-MS/MS was developed for the determination 180 ng/mL) were prepared in the same manner as the calibra- of 11 Gelsemium alkaloids in rat plasma, and the toxicokinetic tion standards. behavior was investigated, so as to provide a scientificunder- standing of Gelsemium elegans and improve the efficacy and safety of drugs. 2.4. Sample Preparation. In a 1.5 mL centrifuge tube, an aliquot of 50 μL plasma was added; then, 150 μL acetonitrile (containing IS 20 ng/mL) was added, and the vortex was 2. Materials and Methods mixed for 1.0 min, then centrifuged (14900 g) for 10 min, μ 2.1. Materials and Reagents. Humantenirine, humantenine, and then, the supernatant (2 L) was injected into the akuammidine, gelsevirine, rankinidine, n-methoxyanhydro- UPLC-MS/MS system for analysis. vobasinediol, gelsenicine, gelsemine, koumine, koumidine, sempervirine (all >98%, Figure 1), and the internal standard 2.5. Method Validation. Calibration curves were established strychnine (IS, all >98%) were purchased from Chengdu by analyzing calibration samples on three different days. Mansite
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