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目的建立体外模拟胃肠液消化稳定性试验和热稳定性试验方法,确定试验中的阳性和阴性参考蛋白质。方法在体外模拟胃肠液消化环境和热加工处理环境,通过SDS-PAGE凝胶电泳研究常见食物致敏原(鸡卵清蛋白OVA、牛β-乳球蛋白β-LG)、不常见食物致敏原(牛血清白蛋白BSA)和非致敏原(大豆脂肪水解酶LPE、马铃薯酸性磷酸酶PAP)在其中各观察时间点的降解情况。结果OVA在模拟胃肠液中60min不降解;β-LG在模拟胃液中60min不降解稳定存在,但在模拟肠液中30min内完全降解;BSA在模拟胃液中30min内完全降解,在模拟肠液中60min部分降解;PAP在模拟胃液中15s即完全降解,LPE在30s内完全降解;而在模拟肠液中,PAP60min完全不降解,LPE60min大部分降解。在热稳定性试验中,OVA、β-LG、BSA60min内不降解,PAP60min内完全降解。结论模拟胃液消化稳定性和热稳定性试验中设OVA为常见致敏原对照、BSA为不常见致敏原对照、PAP为非致敏原对照,模拟肠液消化稳定性试验中设OVA为常见致敏原对照、BSA为不常见致敏原对照、LPE为非致敏原对照。并据此初步建立体外消化稳定性和热稳定性试验方法。
Objective To establish an in vitro gastrointestinal juice digestion stability test and thermal stability test method to determine the test positive and negative reference proteins. Methods In vitro simulating gastrointestinal fluid digestion and thermal processing environment, common food allergens (chicken ovalbumin OVA, bovine β-lactoglobulin β-LG) were studied by SDS-PAGE gel electrophoresis, Degradation of allergen (bovine serum albumin BSA) and non-allergen (soybean lipolytic enzyme LPE, potato acid phosphatase PAP) at each observation time point. RESULTS: OVA did not degrade in simulated gastrointestinal fluid for 60min. Β-LG did not degrade in simulated gastric fluid for 60min, but degraded completely in simulated intestinal fluid within 30min. BSA completely degraded within 30min in simulated gastric fluid and in simulated intestinal fluid for 60min Partially degraded. In the simulated gastric juice, PAP completely degraded in 15s and LPE completely degraded in 30s. In simulated intestinal fluid, PAP60min did not degrade completely and most of LPE was degraded in 60min. In the thermal stability test, OVA, β-LG, BSA do not degrade within 60min, completely degraded within 60min. Conclusion Simulated gastric juice digestion stability and thermal stability test with OVA as a common allergen control, BSA is not common allergen control, PAP is non-allergen control, simulated intestinal fluid digestion stability test set OVA common Allergen control, BSA is not common allergen control, LPE non-allergen control. Based on this, a preliminary in vitro digestibility and thermostability test method was established.