生物学杂志

生物学杂志 ›› 2020, Vol. 37 ›› Issue (5): 108-.doi: 10.3969/j.issn.2095-1736.2020.05.108

• 技术方法 • 上一篇    下一篇

猪小肠黏膜下层脱细胞基质中I、Ⅲ型胶原检测方法研究

  

  1. 1. 北京博辉瑞进生物科技有限公司， 北京 100026； 2. 中国科学院过程工程研究所生化工程国家重点实验室， 北京 100190； 3. 河南科技学院 生命科技学院， 新乡 453003
  • 出版日期:2020-10-18 发布日期:2020-10-14
  • 通讯作者: 张贵锋，博士，研究员，研究方向为蛋白分析检验，E-mail：gfzhang@ipe.ac.cn；赵 博，博士，副主任医师，研究方向为消化道恶性肿瘤基础与临床研究及外科修复医用生物材料，E-mail：zhaobo@biosishealing.com
  • 作者简介:张扬，硕士，研究方向为蛋白质分离纯化，生物材料制备与表征，E-mail:zyang94@foxmail.com
  • 基金资助:
    中关村科技园区管理委员会“重大前沿原创技术成果转化和产业化”；北京市科技新星计划（XX2018008）；国家重点研发计划（2018YFA0108200）；国家重点研发计划（2018YFC1106400）；广东省自然科学基金项目(2014A030312013)；海口市海洋经济创新发展示范城市产业链协调创新类项目（HHCL201804）

Study on the detection method of type I/III collagen in porcinesmall intestinal submucosa extracellular matrix

  1. 1. Beijing Biosis Healing Biological Technology Co., Ltd., Beijing 100026; 2. State Key Laboratory of BiochemicalEngineering, Institute of Process Engineering, CAS, Beijing 100190; 3. School of Life Science andTechnology, Henan Institute of Science and Technology, Xinxiang 453003, China
  • Online:2020-10-18 Published:2020-10-14

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摘要： 猪小肠黏膜下层脱细胞基质（SIS，Small intestinal submucosa）的主要成分为I、III型胶原，拟建立一种基于液相质谱联用（HPLC-MS）的胶原类型识别和定量检测方法。首先利用胰蛋白酶将胶原标准品和样品酶解，利用离子阱质谱（Ion trap mass spectrometry, LCQ）识别出I、III型胶原的特征多肽分别为GETGPAGPAGPVGPVGAR、GPPGAVGPSGPR，再利用三重四极杆质谱（Triple quadrupole mass spectrometry, TSQ）对其进行定量；然后通过胶原标准品浓度和峰面积的线性关系计算I、III型胶原的含量。结果表明VIDASISTM和Biodesign中I、III型胶原的含量总和与高效液相色谱法检测结果一致，通过HPLC-MS检测猪跟腱和真皮中I、III型胶原含量的结果符合其在组织中的分布规律；方法学实验表明HPLC-MS的精密度（RSD=2.38%）较高、稳定性良好。

关键词: 猪小肠黏膜下层, 脱细胞基质, Ⅰ型胶原, Ⅲ型胶原, 液质联用

Abstract: Type I and III collagen are the major components of porcine small intestine submucosal extracellular matrix (SIS). This study intends to establish a collagen type identification and quantitative detection method based on liquid chromatography-mass spectrometry. Firstly, the ion trap mass spectrometry (LCQ) was used to detect the digestion of the collagen standard and SIS products that had been marketing to identify the type of collagen and its marker peptides. The marker peptide of type I collagen was GETGPAGPAGPVGPVGAR, and type III was GPPGAVGPSGPR. The triple quadrupole mass spectrometry (TSQ) was used to detect the marker peptides, and then the contents of type I and III collagen in SIS were calculated by the linear relationship between the concentration and the peak area. The result showed that the contents of I and III in VIDASISTM and Biodesign were close to that of the total collagen content. the contents of I and III collagen in porcine Achilles tendon and dermis were detected by HPLC-MS, and the results accorded with the distribution of type I and III collagen in tissues.

Key words: porcine small intestinal submucosa, acellular matrix, type Ⅰ collagen, type III collagen, HPLC-MS

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