中国临床药理学杂志

HPLC-MS/MS法同时测定三种抗癫痫药及其活性代谢物的血清浓度

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付胜楠 ¹ , 张家欣 ² , 刘昀 ³ , 林晓纯 ¹ , 裴旭哲 ⁴ , 倪穗琴 ¹

中国临床药理学杂志 | 研究方法 2025,41(17): 2503-2509

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中国临床药理学杂志 |研究方法 2025 , 41 (17) : 2503 -2509

HPLC-MS/MS法同时测定三种抗癫痫药及其活性代谢物的血清浓度

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付胜楠¹, 张家欣², 刘昀³, 林晓纯¹, 裴旭哲⁴, 倪穗琴¹

作者信息

^1.广州市第一人民医院药学部，广东广州 510180

^2.广东药科大学药学院，广东广州 510006

^3.广州市老年医院药学中心，广东广州 510550

^4.上海爱博才思分析仪器贸易有限公司，上海 200050

通讯作者:

倪穗琴，主任药师，硕士生导师 Tel: （020）81048106 E-mail: ywjd510@126.com

作者简介:

付胜楠（1988-），女，副主任药师，主要从事个体化用药方面的工作和研究

Simultaneous determination of three antiepileptic drugs and one active metabolite in human serum by HPLC-MS/MS

Sheng-nan FU¹, Jia-xin ZHANG², Yun LIU³, Xiao-chun LIN¹, Xu-zhe PEI⁴, Sui-qin NI¹

Affiliations

^1.Department of Pharmacy, Guangzhou First People’s Hospital, Guangzhou 510180, Guangdong Province, China

^2.School of Pharmacy, Guangdong Pharmaceutical University, Guangzhou 510006, Guangdong Province, China

^3.Pharmacy Center, Guangzhou Geriatric Hospital, Guangzhou 510550, Guangdong Province, China

^4.Shanghai AB Sciex Analytical Instrument Trading Co., Ltd., Shanghai 200050, China

出版时间: 2025-09-17 doi: 10.13699/j.cnki.1001-6821.2025.17.018

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摘要

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目的

建立高效液相色谱-串联质谱（HPLC-MS/MS）法同时检测拉莫三嗪（LTG）、左乙拉西坦（LEV）、奥卡西平（OXC）及其活性代谢物10,11-二氢-10-羟基卡马西平（MHD）的血清浓度。

方法

该方法用简单的一步蛋白沉淀法对血清样品预处理，以2种抗癫痫药的稳定氘代化合物（OXC-D4、LEV-D6）为内标，用Phenomenex C₁₈（100.0 mm×4.6 mm，3.0 μm）色谱柱分离；以含5 mmol·L^-1乙酸铵的0.1％甲酸水溶液（A）-甲醇（B）为流动相，梯度洗脱6 min，流速0.6 mL·min^-1；柱温40 ℃；用电喷雾离子源，多反应监测模式进行正离子扫描分析。考察该方法的专属性、标准曲线与定量下限、精密度与回收率、基质效应和稳定性。以本院就诊的使用LRG、LEV或OXC治疗的患者纳入为研究对象，于血药浓度达到稳态后采集血样并使用本研究开发的方法对血清浓度进行检测。

结果

该方法专属性好，LTG、LEV、OXC和MHD在（0.20～25.00）、（0.39～50.00）、（0.04～5.00）和（0.31～40.00）μg·mL^-1范围内线性良好，LTG的线性方程为y＝0.99x＋0.02（r＝0.999 2），LEV的线性方程为y＝0.55x＋0.01（r＝0.999 4），OXC的线性方程为y＝4.35x＋0.04（r＝0.998 9），MHD的线性方程为y＝0.56x＋0.01（r＝0.998 7）。定量下限分别为0.20、0.39、0.04和0.31 μg·mL^-1，批内、批间的精密度与准确度良好，相对误差（RE）和相对标准偏差（RSD）均<15.00%，提取回收率为88.56%～112.18%，无明显的基质效应和残留效应。LTG、LEV、OXC和MHD血清样品在－20 ℃放置14 d、反复冻融3次（－20 ℃～室温）、室温3 h和处理后置于进样器（4 ℃）24 h的条件下可保持稳定，RSD均≤10.72%。145例临床样本的分析显示线性范围能够满足临床需求。

结论

本研究成功建立了一种可同时定量分析LTG、LEV、OXC和MHD血药浓度的HPLC-MS/MS法，该方法简单、经济、准确、高效。

关键词

拉莫三嗪 / 左乙拉西坦 / 奥卡西平 / 10-羟基卡马西平 / 抗癫痫药 / 液相色谱-串联质谱法 / 治疗药物监测

Abstract

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Objective

To establish and validate a high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method for simultaneously determining the serum concentration of lamotrigine(LTG), levetiracetam(LEV), oxcarbazepine(OXC) and monohydroxycarbazepine(MHD), which is applied for clinical blood concentration detection.

Methods

A simple one-step precipitation method was used for the serum sample pretreatment, and stable isotopes of two antiepileptic drugs (OXC-D4, LEV-D6) as the internal standard. The mobile phases were aqueous solution of 0.1% formic acid and 5 mM ammonium acetate(A) and methyl alcohol(B), and the gradient elution was performed at the speed of 0.6 mL·min^-1 for 6 min on Phenomenex C₁₈(100.0 mm×4.6 mm, 3.0 μm). The column temperature was 40 ℃. Positive ion scan analysis was carried out in multi-reaction monitoring mode, using electrospray ion source. The specificity, standard curve and lower limit of quantification (LLOQ), accuracy and precision, extraction recovery, matrix effect and stability were evaluated. The patients treated with LRG, LEV or OXC in hospital were included as the research object. Blood samples were collected after the blood concentration reached a steady state and the serum concentration was detected by the method developed in this study.

Results

The method demonstrated good specificity. For LTG, LEV, OXC and MHD, outstanding linearity was observed in the ranges of (0.20-25.00), (0.39-50.00), (0.04-5.00), (0.31-40.00) μg·mL^-1, respectively. The linear equation of LTG was y=0.99x+0.02 (r=0.999 2), LEV was y=0.55x+0.01 (r=0.999 4), OXC was y=4.35x+0.04 (r=0.998 9), and MHD was y=0.56x+0.01 (r=0.998 7). The LLOQs were 0.20, 0.39, 0.04, 0.31 μg·mL^-1, respectively. Acceptable accuracy as well as intra-batch and inter-batch precision were achieved; relative error (RE) and relative standard deviation (RSD) were all <15.00%. The extraction recoveries reached the standard (88.56%~112.18%, RSD<15.00%), without significant matrix effects and carryover effects. The serum samples of LTG, LEV, OXC, and MHD exhibited stability under the following conditions: storage at -20 ℃ for 14 days, three freeze-thaw cycles (-20 ℃ to room temperature), room temperature exposure for 3 hours, and post-treatment storage in the autosampler (4 ℃) for 24 hours, with all RSDs ≤10.72%. The analysis of 145 clinical samples showed that the linear range could meet the clinical needs.

Conclusion

This study successfully established a HPLC-MS/MS method to simultaneously quantify the serum concentration of LTG, LEV, OXC and MHD. The method is simple, economical accurate and efficient, which has promotional value.

Key words

lamotrigine / levetiracetam / oxcarbazepine / 10-hydroxycarbamazepine / antiepileptic drugs / high performance liquid chromatography tandemmass spectrometry / therapeutic drug monitoring

引用本文

付胜楠, 张家欣, 刘昀, 林晓纯, 裴旭哲, 倪穗琴. HPLC-MS/MS法同时测定三种抗癫痫药及其活性代谢物的血清浓度. 中国临床药理学杂志, 2025 , 41 (17) : 2503 -2509 . DOI: 10.13699/j.cnki.1001-6821.2025.17.018

Sheng-nan FU, Jia-xin ZHANG, Yun LIU, Xiao-chun LIN, Xu-zhe PEI, Sui-qin NI. Simultaneous determination of three antiepileptic drugs and one active metabolite in human serum by HPLC-MS/MS[J]. Chinese Journal of Clinical Pharmacology, 2025 , 41 (17) : 2503 -2509 . DOI: 10.13699/j.cnki.1001-6821.2025.17.018

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2025年第41卷第17期

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文章信息

doi: 10.13699/j.cnki.1001-6821.2025.17.018

接收时间：2024-11-04
首发时间：2026-08-05
出版时间：2025-09-17

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收稿日期：2024-11-04

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作者信息

^1.广州市第一人民医院药学部，广东广州 510180

^2.广东药科大学药学院，广东广州 510006

^3.广州市老年医院药学中心，广东广州 510550

^4.上海爱博才思分析仪器贸易有限公司，上海 200050

通讯作者:

倪穗琴，主任药师，硕士生导师 Tel: （020）81048106 E-mail: ywjd510@126.com

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https://castjournals.cast.org.cn/joweb/zglcylxzz/CN/10.13699/j.cnki.1001-6821.2025.17.018

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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