牛骨肽钙螯合物制备、表征及其促MC3T3-E1细胞成骨活性

Preparation and Characterization of Bovine Bone Peptide-Calcium Chelate and Its Promoting Effect on Osteogenic Activity of MC3T3-E1 Cells

目的:为制备人体高效吸收利用的补钙剂并探究其对MC3T3-E1细胞成骨活性的影响。方法:对牛骨多肽进行钙螯合处理,以钙螯合能力为考察指标,通过响应面方法确定最佳制备条件;利用红外光谱、X射线衍射和原子吸收等方法探究肽钙螯合物的结构特性及其稳定性;通过细胞实验探究牛骨肽钙螯合物对MC3T3-E1细胞增殖、分化和矿化活性的影响。结果:制备牛骨肽钙螯合物的最佳条件为肽钙质量比2.97∶1、温度62.54℃、pH 9.06。在该条件下,钙螯合能力达到55.65μg/mg。光谱结构分析结果表明氨基氮、羟基氧和羧基氧是钙主要螯合位点,二者之间主要通过配位键结合,且肽螯合钙后分子质量增加;此外,牛骨肽钙螯合物稳定性受温度影响较小,但对pH值较敏感。MC3T3-E1细胞实验结果表明肽钙螯合物具有明显促MC3T3-E1细胞增殖、分化和矿化活性的作用。结论:制备的牛骨肽钙螯合物具有潜在的促MC3T3-E1细胞成骨活性功效。

Purpose:To prepare a calcium supplement for efficient absorption and utilization in the human body and to explore its effect on the osteogenic activity of MC3T3-E1 cells.Methods:Bovine bone peptide was chelated with calcium,and the optimal reaction conditions were determined by response surface methodology(RSM).The structural characteristics and stability of peptide-calcium chelate were studied by infrared spectroscopy,X-ray diffraction and atomic absorption spectrometry.The effects of bovine bone peptide-calcium chelate on the proliferation,differentiation and mineralization of MC3T3-E1 cells were investigated.Results:The optimum conditions for the preparation of bovine bone peptide calcium chelate were peptide/calcium mass ratio 2.97,temperature 62.54℃and pH 9.06.Under these conditions,the calcium chelating capacity was 55.65μg/mg.The results of spectral analysis showed that amino nitrogen,hydroxyl oxygen and carboxyl oxygen were the major calcium-chelating sites.Bovine bone peptide could form coordinate bonds with calcium,and the molecular mass of the peptide increased after chelation with calcium.In addition,the stability of bovine bone peptide-calcium chelate was little affected by temperature,but sensitive to pH.The peptide-calcium chelate could significantly promote the proliferation,differentiation and mineralization of MC3T3-E1 cells.Conclusion:Bovine bone peptide-calcium chelate has the potential to promote the osteogenic activity of MC3T3-E1 cells.