色谱

色谱

• 研究论文 • 上一篇    下一篇

高效液相色谱-质谱联用测定胶原蛋白中的羟脯氨酸

夏金根,陈波,姚守拙   

  1. Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research of Ministry of Education, Hunan Normal University, Changsha 410081, China
  • 收稿日期:2008-10-27 修回日期:2008-10-27 出版日期:2008-10-27 发布日期:1983-09-25
  • 通讯作者: 陈波
  • 基金资助:

Determination of hydroxyproline in collagenous proteins by high performance liquid chromatography-mass spectrometry

XIA Jingen, CHEN Bo, YAO Shouzhuo   

  1. Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research of Ministry of Education, Hunan Normal University, Changsha 410081, China
  • Received:2008-10-27 Revised:2008-10-27 Online:2008-10-27 Published:1983-09-25
  • Contact: CHEN Bo

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1054

被引次数

99

摘要： 建立了一种简单、快速地测定胶原蛋白中羟脯氨酸(HYP)的高效液相色谱-电喷雾质谱(HPLC-ESI/MS)方法。胶原蛋白经酸水解后,以乙腈-0.05%的三氟乙酸水溶液(体积比为5∶95)为流动相,以1.0 mL/min的流速在C8反相柱上进行分离。以茶氨酸为内标,利用质谱定性定量测定HYP。在电喷雾正离子模式下,对m/z 132和m/z 175离子进行选择离子监测。在11.7～117 mg/L范围内,HYP与内标物茶氨酸的峰面积比和HYP的质量浓度呈良好的线性关系,相关系数为0.9993。含量测定的相对标准偏差为1.87%,回收率为97.85%～101.76%。此方法流动相简单,分析时间短且无需衍生处理,抗干扰能力强,能准确快速地测定胶原蛋白中HYP的含量。

关键词: 高效液相色谱-质谱, 胶原蛋白 , 羟脯氨酸

Abstract: A simple, rapid and accurate method for the determination of hydroxyproline in collagenous proteins by high performance liquid chromatography-electrospray mass spectrometry (HPLC-ESI/MS) was developed. After hydrolysis of collagenous proteins with hydrochloric acid, the hydroxyproline was separated on a Spherigel C8 column using acetonitrile-0.05% trifluoroacetic acid aqueous solution (5∶95, v/v) at a flow rate of 1.0 mL/min. Theanine was used as the internal standard for the quantification. Hydroxyproline was identified and quantified by electrospray ionization mass spectrometry (ESI-MS), which was operated in positive ion mode while the m/z 132 and m/z 175 ions were recorded in the selective ionization monitoring (SIM) mode. The peak area ratio of hydroxyproline to theanine versus the hydroxyproline concentration was linear over a concentration range of 11.7-117 mg/L for hydroxyproline. The correlation coefficient was 0.9993. The reproducibility and average recovery of the method were 1.87% and 98.85% respectively. The method has the advantages of easy operation, without derivatization, less analysis time, good precision and accuracy.

Key words: collagenous proteins , hydroxyproline, high performance liquid chromatography-mass spectrometry (HPLC-MS)