利用枯草芽孢杆菌制备大豆发酵液(soybean fermentation broth,S-FB)并研究了其抗氧化活性。通过分析抗氧化活性物含量得到S-FB中总酚和总肽的质量浓度比大豆发酵前提取液(soybean extract,SE)分别提高了187%和229%。抗氧化测试结果显示,1% S-FB对2,2′-联氨双-(3-乙基苯并噻唑啉-6-磺酸)[2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS]阳离子自由基的清除效果接近10% SE。因此,发酵提高了S-FB中的活性物含量并增强了其抗氧化活性。试验进一步研究了S-FB在人永生化角质形成细胞(HaCaT)内的抗氧化作用。结果表明,S-FB (5%、10%和20%)对活性氧的诱导作用随其体积分数的提高而增强。H2O2+S-FB (5%、10%和20%)提高的活性氧水平均低于H2O2或5% S-FB。因此,当活性氧水平提高,S-FB对活性氧的清除效果增强,诱导作用减弱。5% S-FB处理后,核因子E2相关因子2上调,并且超氧化歧化酶活力在36和48 h分别提高了32.5%和32.1%,对应的过氧化氢酶活力分别提高了19.6%和17.5%,同时该细胞被紫外线B损伤后的存活率是常规HaCaT细胞的1.3倍。因此,S-FB能够通过降低氧化应激反应和激活抗氧化通路来保护HaCaT细胞。
Soybean fermentation broth (S-FB) was prepared with Bacillus subtilis and its antioxidant activity was studied. Analysis of the content of active compounds revealed that the mass concentration of total polyphenols and total peptides in S-FB increased by 187% and 229%, respectively, compared with the extract of unfermented soybean (SE). The antioxidant test result suggested that the scavenging effect of 1% φ(S-FB) on ABTS (2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid), ABTS) cation radical was close to 10% φ(SE). Thus, fermentation increased the active compounds in S-FB and enhanced its antioxidant activity. Further research was conducted to reveal its antioxidant activity in human immortalized keratinocytes (HaCaT), which showed that φ(S-FB) (5%, 10% and 20%) induced more reactive oxygen species with the increase of its volume fraction. Moreover, H2O2+φ(S-FB) (5%, 10% and 20%) all led to a smaller increase of reactive oxygen species than H2O2 or 5%φ(S-FB) did. Hence, in high reactive oxygen species (ROS)-level cells, the scavenging effect of S-FB on reactive oxygen species increased, followed by its inducting effect declined. After treatment with 5% φ(S-FB), the intracellular nuclear factor erythroid-2-related factor 2 was upregulated, and after 36 and 48 h, the superoxide dismutase activity increased by 32.5% and 32.1%, respectively, while catalase activity increased by 19.6% and 17.5%, respectively. Meanwhile, their cell viability in damage test by ultraviolet radiation B was 1.3 times higher than normal HaCaT cells. Therefore, S-FB could protect HaCaT cells by reducing the oxidative stress and activating the antioxidant pathway.
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