树莓干果粗提物的纯化、活性及成分研究

发布时间：2018-04-21 05:36

本文选题：树莓 + 黄酮　；参考：《中北大学》2017年硕士论文

【摘要】：树莓(Rubus idaeus)为多年生小灌木,兼木草两性。其果实营养成分和生物活性成分含量丰富,被美国人称为“生命之果”。本研究利用大孔吸附树脂法对树莓干果粗提物进行纯化,研究了抗生素与提取物的配伍抑菌作用,测定了水溶、醇溶、煮沸和高温高压四种不同条件下样品的总多酚和总黄酮含量,并利用HPLC鉴定了其中的主要黄酮成分和含量变化,最后对高温高压和水溶条件下样品的抗氧化活性和抑菌活性进行分析。结果表明:(1)通过静态吸附和解吸试验,确定XDA-6为纯化树莓粗提物的最佳树脂,吸附率为83.7%,解吸率为83.0%;结合静态与动态吸附解吸试验,确定最佳纯化条件为:原液以6 BV/h的流速上样(大孔树脂的吸附浓度为14.50 mg/mL),60%乙醇以4 BV/h的流速进行洗脱,洗脱液用量约为5 BV。纯化后黄酮含量35.8%较粗提物黄酮含量16.2%提高了1.21倍(以下将纯化前的提取物记为“粗黄酮”,纯化后的记为“纯黄酮”)。对大肠杆菌和金黄色葡萄球菌的最小抑菌浓度(Minimal Inhibitory Concentration,MIC)由粗黄酮的8 mg/m L下降到纯黄酮的2 mg/mL;对小麦赤霉菌和棉花枯萎菌的菌丝抑制率表现为低浓度下提高多,高浓度下提高少的趋势,但是真菌菌丝抑制效率的提高远远低于对细菌的抑制效率的提高幅度。(2)通过研究四种抗生素与黄酮的配伍抑菌效果,得出四种抗生素与黄酮配伍均有协同抑菌效果。氨苄青霉素与黄酮配伍抑制金黄色葡萄球菌和大肠杆菌两种实验菌的协同效果不明显,对沙门氏菌有较好的协同抑菌效果,且青霉素的抑菌浓度降至原浓度的1/10。而红霉素结果与氨苄青霉素正好相反,与黄酮配伍抑制金黄色葡萄球菌和大肠杆菌两种实验菌的协同效果明显,而对沙门氏菌协同抑菌效果不明显。盐酸四环素和硫酸链霉素与黄酮配伍抑制三种菌的协同效果均很好,FICI值均小于0.25,最低可达到0.1。(3)四种条件下样品中总多酚和总黄酮含量的变化趋势为:高温高压沸水浴乙醇水,其中以纯黄酮样品的变化最为明显。高温高压后,粗黄酮和纯黄酮的总多酚含量分别达到331.02 mg/g,395.82 mg/g,相对于水溶条件下的含量分别提高了50.21%,67.07%;总黄酮含量分别增加73.23 mg,166.73 mg,较水溶条件提高了38.72%,86.92%。具体到化学成分,黄酮样品中以维生素C和儿茶素为主,含有少量槲皮素和芦丁,山奈酚含量低于检出线。粗黄酮和纯黄酮高温高压后,维生素C分别减少18.45 mg/g(27.26%),34.24 mg/g(35.20%);儿茶素分别增加9.29 mg/g(12.54%),56.53 mg/g(59.02%);芦丁分别减少0.2 mg/g(57.14%),0.25 mg/g(22.32%),槲皮素和其他未标明的峰变化量极小。(4)高温高压后,样品的抗氧化能力均减弱。清除DPPH自由基实验中,粗黄酮的IC50值由28.36μg/mL升高到38.91μg/m L;纯黄酮的IC50值由22.93μg/mL升高到35.84μg/m L。清除ABTS自由基实验中,粗黄酮的IC50值由14.07μg/m L升高到21.46μg/mL;纯黄酮的IC50值由10.29μg/mL升高到21.3μg/m L。两组实验均说明样品抗氧化能力降低,最高降幅超过50%。高温高压后,样品的抑菌活性显著提高。粗黄酮高温高压后对大肠杆菌的抑菌率提高10%左右;在2 mg/mL时,对沙门氏菌的抑菌率由83.6%上升到100%,完全抑制;对金黄色葡萄球菌的抑菌率由62.3%上升到100%,提高了60.5%。纯黄酮高温高压后对大肠杆菌抑菌率提高了约20%;在1 mg/mL时,对沙门氏菌和金黄色葡萄球菌已完全抑制,达到了杀菌剂量级,分别提高了138%和100%。综上所述,纯化与高温高压条件均可有效提高提取物的总多酚和总黄酮含量,间接提高了其抑菌活性,表明热处理过程不仅有利于树莓提取物的保存还有利于其酚类成分的富集。黄酮与抗生素配伍抑菌时可降低抗生素的用量,为解决抗生素的滥用问题提供新的思路。
[Abstract]:The raspberry (Rubus idaeus) is a perennial shrub and the hermaphrodite of wood grass. The content of its nutritive and bioactive components is rich. It is called "the fruit of life" by the American. This study used macroporous adsorption resin to purify the crude extract of raspberry, and studied the antibacterial effect of antibiotics and extracts. The content of total polyphenols and total flavonoids in four different conditions under boiling and high temperature and high pressure, and the changes of the main flavonoids and content were identified by HPLC. Finally, the antioxidant activity and antibacterial activity of the samples under high temperature and high pressure and water soluble conditions were analyzed. The results showed that: (1) by static adsorption and desorption test, XDA-6 was determined. The best resin to purify raspberry is 83.7% and the desorption rate is 83%. Combining static and dynamic adsorption desorption experiments, the optimum purification conditions are as follows: the original solution is 6 BV/h of the flow rate (the adsorption concentration of macroporous resin is 14.50 mg/mL), 60% ethanol is eluted with 4 BV/h flow speed, and the eluant dosage is about 5 BV. after the purification of Huang Tonghan. The content of the flavonoid content of the crude extract 35.8% was 1.21 times higher than that of the crude extract (below the purified extract as "crude flavonoids", and the purified one was recorded as "pure flavonoids"). The minimum inhibitory concentration (Minimal Inhibitory Concentration, MIC) of Escherichia coli and Staphylococcus aureus (MIC) decreased from 8 mg/m L of crude flavonoids to 2 mg/mL of pure flavonoids; The mycelial inhibition rate of wheat and cotton Fusarium wilt showed a tendency to increase in low concentration and less in high concentration, but the enhancement of fungal mycelium inhibition efficiency was much lower than that of bacteria inhibition efficiency. (2) the compatibility of four antibiotics with flavonoids was studied, and the compatibility of four antibiotics with flavonoids was obtained. The synergistic effect of ampicillin with flavonoids was not obvious, and the synergistic effect of two experimental strains of Staphylococcus aureus and Escherichia coli was not obvious, and Salmonella had better synergistic bacteriostasis effect, and the inhibitory concentration of penicillin decreased to 1/10. of the original concentration, while erythromycin results were opposite to ampicillin and were compatible with flavonoids. The synergistic effect of inhibiting two kinds of experimental bacteria of Staphylococcus aureus and Escherichia coli was obvious, but the synergistic effect of Salmonella was not obvious. The synergistic effect of tetracycline hydrochloride and streptomycin sulfate with flavonoids was good, the FICI value was less than 0.25 and the lowest could reach the total polyphenols and total yellow in the samples of 0.1. (3) and four conditions. The change trend of the content of ketone was as follows: high temperature and high pressure boiling water bath ethanol water, the most obvious change of the YISHION flavone samples. After high temperature and high pressure, the total polyphenols content of the crude flavonoids and pure flavonoids reached 331.02 mg/g, 395.82 mg/g respectively. The content of the total flavonoids increased by 50.21%, 67.07%, respectively, compared with the water soluble conditions, and the total flavonoids content increased by 73.23 mg, 166.7 respectively. 3 mg, higher than water solubility, 38.72%, 86.92%. specific to chemical composition, vitamin C and catechin in flavonoids, containing a small amount of quercetin and rutin, and the content of kaempferol is lower than the detection line. After high temperature and high temperature and high pressure of crude flavonoids and pure flavonoids, vitamin C is reduced by 18.45 mg/g (27.26%), 34.24 mg/g (35.20%), and catechin increases 9.29 mg, respectively. /g (12.54%), 56.53 mg/g (59.02%); rutin decreased by 0.2 mg/g (57.14%), 0.25 mg/g (22.32%), quercetin and other unlabeled peaks. (4) the antioxidant capacity of the samples decreased after high temperature and high pressure. In the DPPH free radical scavenging experiment, the IC50 value of the crude flavonoids increased from 28.36 Mu to 38.91 u g/m L, and the IC50 value of pure flavonoids was from 22.93 mu g. In the 35.84 micron g/m L. scavenging experiment of ABTS free radical, the IC50 value of the crude flavonoids increased from 14.07 mu L to 21.46 mu g/mL, and the IC50 value of the pure flavonoids from 10.29 mu g/mL to 21.3 mu g/m L. two groups showed that the antioxidant capacity of the samples decreased and the highest decrease exceeded the high temperature and high pressure. The antibacterial activity of the samples was significantly increased. The high temperature of the crude flavonoids was high. The bacteriostasis rate of Escherichia coli increased by about 10% after high pressure. At 2 mg/mL, the bacteriostasis rate of Salmonella increased from 83.6% to 100%, completely inhibited, the bacteriostasis rate of Staphylococcus aureus increased from 62.3% to 100%, and increased by high temperature and pressure of 60.5%. pure flavonoids by 20%, and Salmonella and Salmonella at 1 mg/mL. Staphylococcus aureus was completely inhibited and reached the magnitude of bactericide, which increased by 138% and 100%., respectively. Purification and high temperature and high pressure can effectively improve the content of total polyphenols and total flavonoids in the extract, and indirectly improve its antibacterial activity. It shows that the heat treatment process is not only beneficial to the preservation of the extract of raspberry but also to its phenols. The combination of flavonoids and antibiotics can reduce the dosage of antibiotics and provide a new way to solve the problem of abuse of antibiotics.

【学位授予单位】：中北大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TQ914.1

【参考文献】