大蒜素与乳清分离蛋白结合物的体外模拟消化特性

大蒜素,即二烯丙基硫代亚磺酸酯,具有广谱抑菌、抑制肿瘤及抗氧化性等生理活性。大蒜素极不稳定,易降解成各种有机硫化合物,使其活性大大降低。研究显示,大蒜素与乳清分离蛋白形成的二硫键结合物可显著增强大蒜素的稳定性。本文在此基础上对大蒜素提取液及二硫键结合物的体外模拟消化特性进行研究,结果表明,大蒜素与乳清分离蛋白的结合物经模拟胃肠消化后,其含硫化合物组成成分与大蒜素提取液的消化产物相同,说明大蒜素与蛋白结合后不会影响大蒜素活性成分的释放。3-乙烯基-1,2-二硫环己-4-烯主要在模拟胃消化阶段释放,而二烯丙基二硫醚主要在模拟肠消化过程中释放。与大蒜素结合不会影响乳清分离蛋白在模拟胃肠消化时的水解度。与蛋白相比,结合物消化产物的小分子质量组分含量增加,说明结合物比蛋白更易被吸收利用。

SDS-PAGE测定大蒜功能性蛋白的相对分子质量

建立用SDS-聚丙烯酰胺凝胶电泳法(SDS-PAGE)测定大蒜功能性蛋白的组成及相对分子质量(M r)。通过电泳考察大蒜冻干粉中的蛋白组分,对标准蛋白进行直线回归分析,确定大蒜冻干粉中功能性蛋白的相对分子质量。此法供试品的处理方便、用量少,分离胶(交联度为10.0%,丙烯酰胺总浓度为3.0%)中能较清晰显示相对分子质量低的蛋白条带,是一种测定相对分子质量低蛋白较好的方法。

大蒜降压肽的制备及超滤分离

为了研究大蒜降压肽的酶解制备与超滤分离工艺,采用响应面法优化制备工艺,酶解后利用超滤技术获得不同分子量范围的组分并进行ACE抑制活性评价。最佳制备工艺为碱性蛋白酶处理、温度50℃、pH11、底物浓度45 g/L、酶底比3%、酶解时间1 h,该条件下ACE抑制率为83.91%±0.13%,半抑制浓度为(157.87±0.44)μg/mL。分子量3 kDa以下部位活性最为显著(P<0.05),优化后的超滤工艺为操作压力0.25 MPa,温度25℃,溶液浓度4%,此时3 kDa超滤膜的膜通量为7.32 L/(m^2·h),ACE的半抑制浓度为(63.27±0.25)μg/mL。氨基酸分析结果表明酸性氨基酸、疏水性氨基酸及天冬氨酸、谷氨酸含量较高,分别为35.78、32.86、21.66、14.12 mg/100 mL,分子量分布显示2000~2500 Da部分最为丰富,占比为49.50%。本研究为大蒜蛋白的开发利用提供了新的思路,具有一定的理论水平和实际应用价值。

Effects of Two Kinds of Heavy Metals Including Cr^(6+) and Pb^(2+) on Garlic Germination

[Objective] This study aimed to revealing the effects of two kinds of heavy metals including Cr6＋ and Pb2＋ on garlic germination.[Method] Root length,enzyme（SOD and CAT）activity and soluble protein content during garlic germination were determined under the stress of two kinds of heavy metals including Cr6＋ and Pb2＋.[Result] During garlic germination,there was an obvious inhibition on buds and seedlings when the concentration of heavy metals including Cr6＋ and Pb2＋ was more than 20 mg/L,while SOD and CAT had certain buffering ability to the heavy metal toxicity with low concentration.With the stress of Cr6＋ in low concentration,SOD activity increased with the increase of metal ion concentration,and then reached maximum when the treatment concentration was 50 mg/L,while CAT activity gradually increased when Cr6＋ concentration was lower than 20 mg/L.With the high concentration of Cr6＋（more than 50 mg/L）or the Pb2＋ stress,SOD and CAT activity continued to decrease with the increase of concentration of heavy metal ions.With various concentrations of heavy metal（Cr6＋ and Pb2＋）solution,soluble protein content showed the decreasing trend with time prolong,and the decreasing trend was obvious with the increase of concentration.Moreover,the tolerance of garlic on Cr6＋ toxicity was much higher than that on Pb2＋.[Conclusion] Two kinds of heavy metals including Cr6＋ and Pb2＋ have impacts on four factors mentioned above during garlic germination,and this study provides a theoretical basis for further discussion of the toxic mechanism of heavy metals on plants.