【作者】 王惠；

【导师】 赵国群； 冯凤琴；

【作者基本信息】 河北科技大学 ， 发酵工程， 2020， 硕士

【摘要】 树莓（Rubus corchorifolius L.）为蔷薇科悬钩子属灌木型植物。树莓果实甘爽多汁,含有丰富的鞣花酸、黄酮等生物活性成分。树莓果实成熟后极易腐烂,不耐贮运,非常有必要开展树莓果实的深加工。果蔬汁乳酸菌发酵,不仅能够增加果蔬汁有机酸、香气物质的种类,改善风味,而且还能够提高其降血糖、抗氧化等保健功效。为了开发功能性树莓汁,本研究首先筛选适于树莓汁发酵的优良功能性乳酸菌,使用该乳酸菌进行树莓汁发酵,研究了工艺条件对树莓汁乳酸菌发酵的影响,并对发酵树莓汁的抗氧化、抗肿瘤性能进行了评价。主要研究结论如下:（1）优良发酵树莓汁乳酸菌的筛选。从模拟胃肠道耐受性能、抑菌性能、粘附性能、抗氧化性能、抑制肿瘤细胞活性性能等5个方面对12株乳酸菌进行筛选,得到5株性能良好的乳酸菌,分别为:副干酪乳杆菌（Lactobacillus paracasei）YJ1、YJ10,鼠李糖乳杆菌（Lactobacillus rhamnosus）SL、植物乳杆菌（Lactobacillus plantarum）T34、Lactobacillus acidipiscis YJ5。用这5株乳酸菌发酵树莓汁,其中副干酪乳杆菌YJ1在树莓汁中的生长能力最强。因此,副干酪乳杆菌YJ1为适宜的树莓汁发酵菌株。（2）树莓汁发酵工艺研究及成分分析。以活菌数为指标,研究了发酵温度、发酵时间、接种量、加糖量、初始pH对树莓汁发酵工艺的影响,并进行了工艺优化,其最佳发酵工艺为:接种量3%、加糖量6%、初始pH为4.5、发酵温度35℃、发酵时间65.6 h、,最终活菌数达到9.71 log CFU/mL。发酵后树莓汁中总酚含量增加了18.58%,黄酮含量增加了23.00%,总酸含量显著增加,有机酸、酚酸种类增加。乳酸菌发酵后的树莓汁口感良好。（3）发酵树莓汁的体外抗氧化功能评价。与未发酵的树莓汁相比,发酵树莓汁清除1,1-二苯基-2-三硝基苯肼（DPPH）和羟基自由基能力分别提高了35.13%和25.08%。用2,2’-联氨-双-3-乙基苯并噻唑啉-6-磺酸（ABTS）和铁离子还原/抗氧化能力（FRAP）法测定的总抗氧化能力分别增加了36.31%、39.48%。树莓汁中总酚、总黄酮含量与DPPH、ABTS自由基清除能力及FRAP总抗氧化性能呈显著正相关。总之,乳酸菌发酵后树莓汁抗氧化能力显著提高。（4）发酵树莓汁的体外抗肿瘤性能评价。采用Cell Counting Kit（cck-8）法测定细胞活性,判断树莓汁对人结肠癌细胞HCT116的作用,并从乳酸脱氢酶（LDH）释放率、细胞周期、细胞凋亡等三方面进一步分析原因。研究发现,发酵树莓汁能够呈剂量依赖性抑制人结肠癌细胞HCT116的细胞活性,其IC₅₀为17.047μL/mL;能够呈剂量依赖性提高乳酸脱氢酶（LDH）释放率;能够在G0/G1期阻滞细胞周期;能够促进细胞凋亡,浓度为16μL/mL时,凋亡细胞达66.37%。与未发酵树莓汁相比,发酵树莓汁抑制肿瘤细胞活性能力、促进乳酸酸脱氢酶释放能力、促进细胞凋亡能力显著提高。更多还原

【Abstract】 Rubus corchorifolius L.is a shrub plant of the genus Rubus of Rosaceae.Raspberry fruit is sweet and juicy,which is rich in ellagic acid,flavonoids and other bioactive components.Raspberry fruit is very perishable after ripening,so it is very necessary to carry out deep processing of raspberry fruit.Fruit and vegetable juice with fermentation of lactic acid bacteria can not only increase the types of organic acids and aroma substances,but also improve its hypoglycemic,antioxidant and other health effects.In order to develop functional raspberry juice,the excellent functional lactic acid bacteria suitable for raspberry juice fermentation were screened and used to ferment raspberry juice in this study.The effects of technological conditions on raspberry juice lactic acid bacteria fermentation were studied.The antioxidant and anti-tumor properties of fermented raspberry juice were evaluated.The main conclusions are as follows:(1)A suitable strain of lactic acid bacteria was screened out for raspberry juice fermentation.Twelve strains of lactic acid bacteria were screened from five aspects of tolerance to simulated gastrointestinal environment,antibacterial activity,adhesion,antioxidation and inhibition of tumor cell activity.Five strains of lactic acid bacteria with good properties were obtained: Lactobacillus paracasei YJ1,YJ10,Lactobacillus rhamnosus SL,Lactobacillus plantarum T34,Lactobacillus acidipiscis YJ5.These five strains of lactic acid bacteria were used to ferment raspberry juice,among which Lactobacillus paracasei YJ1 had the strongest growth ability in raspberry juice.Therefore,Lactobacillus paracasei YJ1 is a suitable strain for raspberry juice fermentation.(2)The fermentation technology of raspberry juice was studied and its composition was analyzed by HPLC.Taking the viable count as the response value,the effects of fermentation temperature,fermentation time,the amount of inoculation,amount of sugar and initial pH on the fermentation process of raspberry juice were studied,and the process was optimized.The optimum fermentation process were shown as following: inoculation3%,sugar 6%,temperature 35 ℃,time 65.6 h,initial pH 4.5,and the final viable count reached 9.71 log CFU/mL.After fermentation,the concentration of total phenols in raspberry juice increased by 18.58%,the concentration of flavonoids increased by 23.00%,the concentration of total acids increased significantly,and the types of organic acids and phenolic acids increased.The raspberry juice fermented by lactic acid bacteria tastes good.(3)Antioxidant function of fermented raspberry juice was evaluated in vitro.Compared with the raspberry juice without fermentation of lactic acid bacteria,the antioxidant activity of fermented red raspberry juice was significantly improved(P<0.05),and its 1,1-diphenyl-2-trinitrophenylhydrazine(DPPH)and hydroxyl free radical scavenging capacity increased by 35.13% and 25.08%,respectively.The total antioxidant capacities of fermented raspberry juice were determined respectively by the methods of 2,2’-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid(ABTS)and Ferric ion reducing antioxidant power(FRAP),and its total antioxidant capacity of ABTS and FRAP increased by 36.31% and 39.48%,respectively.The contents of total phenols and total flavonoids in raspberry juice were positively correlated with the free radical scavenging ability of DPPH and ABTS and the total antioxidant activity of FRAP.In a word,compared with that of pre-fermentation,the antioxidant capacity of raspberry juice was significantly improved after fermentation.(4)Antitumor activity of fermented raspberry juice was evaluated in vitro.The cell activity was determined by Cell Counting Kit(cck-8)method to determine the effect of raspberry juice on human colon cancer cell HCT116,and the causes were further analyzed from three aspects: lactate dehydrogenase(LDH)release rate,cell cycle and apoptosis.It was found that raspberry juice with fermentation of lactic acid bacteria could inhibit the cell activity of human colon cancer cell HCT116 in a dose-dependent manner,and its IC50 of 17.047 μL/mL.It could increase the release rate of lactate dehydrogenase(LDH)in a dose-dependent manner.It could block the cell cycle in G0/G1 phase and promote apoptosis.When the concentration was 16 μL/mL,the apoptotic cells reached 66.37%.Compared with unfermented raspberry juice,fermented raspberry juice significantly improved the ability of inhibiting tumor cell activity,promoting lactate dehydrogenase release and promoting cell apoptosis.更多还原