摘要:
In this study,the hydrolysis conditions for preparing egg yolk protein hydrolysate (EYPH) from defatted egg yolk were optimized based on the viability of MC3T3-E1 cells when cultured in the presence of EYPH.Additionally,we also evaluated the promoting effect of EYPH on proliferation and differentiation of MC3T3-E1 cells.The results showed that the hydrolysis with trypsin alone yielded a product with stronger cell proliferation promoting activity than when it was used in combination with flavourzyme.The optimum hydrolysis conditions for trypsin were as follows:hydrolysis time,6 h;temperature,37 °C;enzyme-to-substrate ratio,1∶50;and pH,8.0.EYPH prepared under the optimized conditions increased the viability of MC3T3-E1 cells by 9.1% and the proportion of S-phase cells by 4.69% indicating that EYPH significantly increased the proliferation of MC3T3-E1 cells.EYPH significantly affected differentiation and mineralization of MC3T3-E1 cells as indicated by a 9.1% increase in alkaline phosphatase activity,an increase in osteocalcin secretion of 2 mg/mL,and the formation of more mineralized nodules of MC3T3-E1.Furthermore,EYPH also could stimulate the expression level of RUNX2 gene by 6.12 times.These finding showed that EYPH could significantly promote proliferation and differentiation in osteoblastic MC3T3-E1 cells.Therefore,EYPH has the potential to be used as a functional food ingredient for its anti-osteoporosis activity.%鸡蛋黄经脱脂得到蛋黄蛋白质,直接以MC3T3-E1细胞增殖活力为指标,对水解条件进行优化,从而制备蛋黄蛋白质水解产物(egg yolk proteins hydrolysate,EYPH)并对其促MC3T3-E1细胞增殖分化的活性进行研究.结果表明:在对比胰蛋白酶与风味蛋白酶水解过程中,发现单一胰蛋白酶反应得到产物活性最好,胰蛋白酶反应的最佳水解条件为水解时间6h、水解温度37°C、酶与底物质量比1∶50、pH 8.0.EYPH使MC3T3-E1细胞增殖活力得到提高,为129.89%,细胞周期分析得到S期细胞增加了4.69%,这2个方面证实EYPH对MC3T3-E1细胞的增殖活力有显著的提高.EYPH对MC3T3-E1细胞分化矿化有显著影响,碱性磷酸酶活力提高了9.1%;刺激骨钙素分泌,其含量上升2 mg/mL;形成更多的矿化结节.此外,EYPH还能刺激RUNX2特异性转录因子的基因表达,表达量提高了6.12倍.结果说明EYPH对成骨细胞MC3T3-E1增殖分化有显著的促进作用,因此,EYPH对于抗骨质疏松方面有较好的应用潜力,为开发其功能性食品提供一定的数据参考.