猪小肠黏膜下层脱细胞基质中Ⅰ、Ⅲ型胶原检测方法研究


作者:本站编辑    发布日期:2021-03-10

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

作者:
张扬;陈毅;高建萍;夏磊磊;李勇超;王清石;张贵锋;赵博

中文摘要:
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。

英文摘要:
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.

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

Keywords:
porcine small intestinal submucosa; acellular matrix; type Ⅰ collagen; type Ⅲ collagen; HPLC-MS

生物学杂志,2020年05期

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