人血浆中甲氨蝶呤的高效液相色谱法测定及其代谢物的质谱定性分析

王漪璇; 毋丹; 曹军宁; 钱隽

doi:10.3969/j.issn.1006-0111.2015.02.013

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人血浆中甲氨蝶呤的高效液相色谱法测定及其代谢物的质谱定性分析

王漪璇, 毋丹, 曹军宁, 钱隽

文章导航 > 药学实践与服务 > 2015 > 33(2): 143-146,158

王漪璇, 毋丹, 曹军宁, 钱隽. 人血浆中甲氨蝶呤的高效液相色谱法测定及其代谢物的质谱定性分析[J]. 药学实践与服务, 2015, 33(2): 143-146,158. doi: 10.3969/j.issn.1006-0111.2015.02.013

引用本文:

WANG Yixuan, WU Dan, CAO Junning, QIAN Jun. Determination of methotrexate in human plasma by HPLC and qualitative analysis of its metabolite by mass spectrometry[J]. Journal of Pharmaceutical Practice and Service, 2015, 33(2): 143-146,158. doi: 10.3969/j.issn.1006-0111.2015.02.013

Citation:

WANG Yixuan, WU Dan, CAO Junning, QIAN Jun. Determination of methotrexate in human plasma by HPLC and qualitative analysis of its metabolite by mass spectrometry[J]. Journal of Pharmaceutical Practice and Service, 2015, 33(2): 143-146,158. doi: 10.3969/j.issn.1006-0111.2015.02.013

人血浆中甲氨蝶呤的高效液相色谱法测定及其代谢物的质谱定性分析

doi: 10.3969/j.issn.1006-0111.2015.02.013

复旦大学附属肿瘤医院肿瘤内科, 复旦大学上海医学院肿瘤学系, 上海 200032

Determination of methotrexate in human plasma by HPLC and qualitative analysis of its metabolite by mass spectrometry

Department of Medical Oncology, Cancer Hospital Affilliated to Fudan University; Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032, China

摘要: 目的建立固相萃取-高效液相色谱法测定人血浆中的甲氨蝶呤浓度,用于临床治疗的监测,并通过质谱对甲氨蝶呤的人体代谢物进行定性分析。方法血浆样本经C₁₈固相萃取小柱净化洗脱后直接进样,以Diamonsil C₁₈分析柱(150 mm×4.6 mm, 5 m)分离,流动相为甲醇和0.5%乙酸溶液(含0.3%三乙胺),采用梯度洗脱;流速为1.0 ml/min;紫外检测波长为306 nm。通过质谱全扫描(QLMS)和多反应监测-信息关联采集-增强子离子扫描(MRM-IDA-EPI)获得甲氨蝶呤代谢物的质谱信息。结果甲氨蝶呤在0.05~100 μmol/L范围内线性关系良好(r=0.999 9),提取回收率>95%,准确度在97%~105%之间,日内与日间精密度(RSD)均<5%。分析甲氨蝶呤代谢物的质谱信息,推断该代谢物为7-羟基甲氨蝶呤。结论本方法灵敏度高、重现性好、操作简便,适用于甲氨蝶呤的血药浓度监测。
- 甲氨蝶呤 /
- 高效液相色谱法 /
- 固相萃取 /
- 代谢物
Abstract: Objective To develop an SPE-HPLC method for the determination of methotrexate (MTX) in human plasma to monitor the clinical drug use of MTX, and to identify the human metabolite of MTX by mass spectrometry. Methods MTX was extracted from human plasma using C₁₈ SPE column and analyzed directly after elution. The separation of MTX was performed on Diamonsil C₁₈ column (150 mm×4.6 mm, 5 m) with a gradient mobile phase of methanol and 0.5% acetic acid solution (containing 0.3% triethylamine) at a flow rate of 1.0 ml/min. The detection wavelength was 306 nm. QLMS and MRM-IDA-EPI scan modes were used to obtain the mass spectrum of MTX metabolite. Results The calibration curve of MTX in plasma was linear over the range of 0.05-100 μmol/L(r=0.999 9). The extraction recovery was above 95% and accuracy was between 97% and 105%. Both intra-and inter-day precision were less than 5%. 7-OH MTX was identified to be the metabolite through the analysis of the mass spectrum. Conclusion The method is sensitive, reproducible, easy to operate and suits for monitoring the concentration of MTX in clinical practice.
- methotrexate /
- HPLC /
- solid phase extraction /
- metabolite

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人血浆中甲氨蝶呤的高效液相色谱法测定及其代谢物的质谱定性分析

doi: 10.3969/j.issn.1006-0111.2015.02.013

复旦大学附属肿瘤医院肿瘤内科, 复旦大学上海医学院肿瘤学系, 上海 200032

收稿日期: 2014-04-03
修回日期: 2014-06-26

关键词:

摘要: 目的建立固相萃取-高效液相色谱法测定人血浆中的甲氨蝶呤浓度,用于临床治疗的监测,并通过质谱对甲氨蝶呤的人体代谢物进行定性分析。方法血浆样本经C₁₈固相萃取小柱净化洗脱后直接进样,以Diamonsil C₁₈分析柱(150 mm×4.6 mm, 5 m)分离,流动相为甲醇和0.5%乙酸溶液(含0.3%三乙胺),采用梯度洗脱;流速为1.0 ml/min;紫外检测波长为306 nm。通过质谱全扫描(QLMS)和多反应监测-信息关联采集-增强子离子扫描(MRM-IDA-EPI)获得甲氨蝶呤代谢物的质谱信息。结果甲氨蝶呤在0.05~100 μmol/L范围内线性关系良好(r=0.999 9),提取回收率>95%,准确度在97%~105%之间,日内与日间精密度(RSD)均<5%。分析甲氨蝶呤代谢物的质谱信息,推断该代谢物为7-羟基甲氨蝶呤。结论本方法灵敏度高、重现性好、操作简便,适用于甲氨蝶呤的血药浓度监测。