中国组织工程研究

中国组织工程研究 ›› 2017, Vol. 21 ›› Issue (2): 291-295.doi: 10.3969/j.issn.2095-4344.2017.02.023

• 生物材料基础实验 basic experiments of biomaterials • 上一篇    下一篇

液质谱联用法同时检测体外生殖培养液中的3种能量成分

黄元礼，柯林楠，王春仁，杨昭鹏
  

  1. 中国食品药品检定研究院医疗器械所，北京市  100050
  • 收稿日期:2016-11-23 出版日期:2017-01-18 发布日期:2017-02-27
  • 作者简介:黄元礼，男，1985年生，北京市人，汉族，2015年北京协和医学院毕业，硕士，助理研究员，主要从事医疗器械化学性能评价研究、组织工程、生物材料的性能评价研究。
  • 基金资助:

    国家重点研发计划(2016YFC1101202)

Simultaneous determination of three main bioactive constituents in the in vitro fertilization medium by UHPLC-MSMS

Huang Yuan-li, Ke Lin-nan, Wang Chun-ren, Yang Zhao-peng
  

  1. Institute of Medical Equipment, National Institute for Food and Drug Control, Beijing 100050, China
  • Received:2016-11-23 Online:2017-01-18 Published:2017-02-27
  • About author:Huang Yuan-li, Master, Research assistant, Institute of Medical Equipment, National Institute for Food and Drug Control, Beijing 100050, China
  • Supported by:

    the National Research and Development Plan of China, No. 2016YFC1101202

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摘要：

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文题释义：
人类辅助生殖技术：指采用医疗辅助手段使不育夫妇妊娠的技术，包括人工授精和体外受精-胚胎移植及其衍生技术两大类。
体外生殖培养液：是含有能量成分、渗透压调节成分、酸碱度调节成分、氨基酸、生长因子等，保证生殖细胞在体外可以存活并成长的一种培养液。不同的体外生殖细胞使用不同的培养液，在不同的生长阶段，也会使用不同的培养液，因此体外生殖培养液的成分对最终体外生殖手术的成功率有很大影响。

背景：目前，对于体外生殖培养液的质量控制文件或技术标准中，对成分含量的指标及检测方法未做明确规定，为保证该类产品的安全有效性，应及早建立质量标准。
目的：建立采用液相质谱联用法检测体外生殖培养液中葡萄糖、乳酸钠、丙酮酸钠3种指标成分的方法，分析不同用途培养液中3种成分的含量。
方法：采用超高效液相色谱三重四级杆质谱联用法，使用SUPELCO Discovery HS F5-3色谱柱(15 cm×2.1 mm，3 μm)，流动相为0.1%甲酸水溶液-0.1%甲酸乙腈溶液，梯度洗脱，流速0.35 mL•min-1，柱温40 ℃；质谱采用电喷雾离子源，负离子模式扫面，扫描方式为多反应监测。
结果与讨论：葡萄糖、丙酮酸钠、乳酸钠分别在0.1-10 mg/L(r=0.999 8)、0.05-5 mg/L(r=0.999 4)、0.1-       10 mg/L(r=0.999 4)范围线性关系练好；平均加样回收率在96.4%-98.1%之间，相对标准偏差均小于2.8%。说明采用液相质谱联用法检测体外生殖培养液灵敏、快速、准确、专属性强，可为体外生殖培养液的质量标准研究提供依据。

关键词: 生物材料, 材料相容性, 超高效液相, 三重四级杆质谱仪, 体外生殖培养液, 葡萄糖, 乳酸钠, 丙酮酸钠, 含量, 质量控制

Abstract:

BACKGROUND: In the present quality control file or technique standards of in vitro fertilization medium, the indicators of the component contents and detection methods have not been clearly defined. To ensure the safety and effectiveness of these products, we should establish the quality standards as early as possible.
OBJECTIVE: To establish a method for determining the three main bioactive constituents of in vitro fertilization
medium including glucose, lactic acid sodium salt, pyruvic acid sodium salt by ultra-high performance liquid chromatography tandem mass spectrometric method (UHPLC-MSMS), and to analyze the content of each constituent.
METHODS: The UHPLC-MSMS was used, and UHPLC separation was performed on a SUPELCO Discovery HS F5-3 column (15 cm × 2.1 mm, 3 μm) in a gradient elute mode with acetonitrile and water (both containing 0.1% formic acid) as the mobile phase at a flow rate of 0.35 mL/min. The column temperature was 40℃. Mass spectrometry detection was performed with multi-reaction monitoring mode using negative electro spray ionization.
RESULTS AND CONCLUSION: The linearity was achieved in the range of 0.1-10 μg/mL (r=0.999 8) for glucose, 0.05-5 μg/mL (r=0.999 4) for lactic acid sodium salt, and 0.1-10 μg/mL (r=0.999 4) for pyruvic acid sodium salt. The recoveries were 96.4%-98.1% with relative standard deviation less than 2.8%. To conclude, the UHPLC-MSMS method is sensitive, rapid, accurate and specific, thus providing a basis for the quality standard study of in vitro fertilization medium.

Key words: Glucose, Sodium Lactate, Quality Control, Tissue Engineering

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