天然花青苷提取纯化及降血脂作用的研究
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摘要
花青苷是一种水溶性的色素,广泛存在于植物的花、果实和茎等器官中,构成了植物王国中绝大多数品种的蓝色、红色、紫色和黄色等颜色。花青苷是花青素与糖通过糖苷键结合而形成的一类化合物,不但具有很强的着色能力,同时还具有美容养颜、降脂减肥、抗衰抗癌、增加视力等多种重要的生理功能和药用功能,可应用于食品、药品、化妆品等领域。本试验研究了提取花青苷的几种方法,并对这几种方法进行了比较;采用薄层层析和柱层析技术纯化花青苷,并筛选出最佳层析介质和洗脱剂;研究花青苷的降血脂作用。
研究提取花青苷的几种方法:超声波水提法、超声波酸提法、酶解法、超临界—酶解法,其中用超临界一酶解法提取花青苷至今未见报道。通过对几种提取方法的比较,发现超声波水提法的提取率最低,为2.7%;超临界—酶解法的提取率最高,达到6.5%。通过正交试验,确定了酶解法提取花青苷的最佳条件为:温度30℃,pH值7.0,固液比1:6,酶用量为24IU,时间90min,提取率达到6.4%。
采用层析技术对花青苷进行分离纯化,通过正交试验筛选出最佳层析介质和洗脱剂,分别为硅胶和乙酸乙酯-甲酸-盐酸(2mol/L)(74:10:12),层析所得花青苷样品纯度为90.2%。
对大花葵花青苷降血脂作用的研究发现,当大花葵花青苷浓度为0.4mg/mL时,对羟基自由基的清除率可达38.8%,对脂质体氧化的抑制率可达13.4%。通过建立大白鼠的高脂模型,再喂食不同剂量的花青苷,发现喂食0.5g/d花青苷剂量总胆固醇下降22.34%,0.6g/d的剂量甘油三酯下降34.4%。
Anthocyanins, a water-soluble pigment, widely exist in flower, fruit and stem of plants and bring many colour of most plant, such as blue, red, purple and yellow. Its stucture consists of anthocyanidin and saccharide through che mical bond between them. It has strong dyeing ability and many physical functions, such as protecting skin, cutting off blood-fat, dieting, restraining cancer, delaying senescence and improving eyesight. It can be used in many field, such as food, drug and cosmetic. Investigation and research were carried out for several methods of extracting anthocyanins and comparing these methods, purifying anthocyanins through thin layer chromatography and column chromatography and finding out optimum chromatography material and eluting solvent, and ability of reducing blood-fat.Investigation were carried out for several methods of extracting anthocyanin, such as water extracting, ultrasonic extracting, enzyme extracting and super critical fluid-enzyme extract ing. There is no report so far for method of super critical fluid-enzyme extracting. The result showed that extracting rate of water extracting was the lowest 2.7% and that of super critical fluid-enzyme extracting was the highest 6.5% through comparing these extracting methods. Through orthogonal experiment analysis, the optimum condition of enzyme extracting, tempera ture 30℃, pH value 7.0, solid-to-liquid ratio 1:6, enzyme consumption 24IU, time 90min, was found out and its extracting rate was 6.4%.The result of reducing blood-fat showed that cleaning hydroxyl free radical rate of anthocyanin was 38.8% when concentration of anthocyanin was 0.4mg/mL, and restraining lipid dioxition rate of anthocyanin was 13.4% when concentration of anthocyanin was 0.4mg/mL.The high fat model of big white mouse was established. The result showed that cutting cholesterol rate was 22.34% when anthocyanin dosage was 0.5g/d, cutting triglyceride rate was 34.4% when anthocyanin dosage was 0.6g/d.
研究提取花青苷的几种方法:超声波水提法、超声波酸提法、酶解法、超临界—酶解法,其中用超临界一酶解法提取花青苷至今未见报道。通过对几种提取方法的比较,发现超声波水提法的提取率最低,为2.7%;超临界—酶解法的提取率最高,达到6.5%。通过正交试验,确定了酶解法提取花青苷的最佳条件为:温度30℃,pH值7.0,固液比1:6,酶用量为24IU,时间90min,提取率达到6.4%。
采用层析技术对花青苷进行分离纯化,通过正交试验筛选出最佳层析介质和洗脱剂,分别为硅胶和乙酸乙酯-甲酸-盐酸(2mol/L)(74:10:12),层析所得花青苷样品纯度为90.2%。
对大花葵花青苷降血脂作用的研究发现,当大花葵花青苷浓度为0.4mg/mL时,对羟基自由基的清除率可达38.8%,对脂质体氧化的抑制率可达13.4%。通过建立大白鼠的高脂模型,再喂食不同剂量的花青苷,发现喂食0.5g/d花青苷剂量总胆固醇下降22.34%,0.6g/d的剂量甘油三酯下降34.4%。
Anthocyanins, a water-soluble pigment, widely exist in flower, fruit and stem of plants and bring many colour of most plant, such as blue, red, purple and yellow. Its stucture consists of anthocyanidin and saccharide through che mical bond between them. It has strong dyeing ability and many physical functions, such as protecting skin, cutting off blood-fat, dieting, restraining cancer, delaying senescence and improving eyesight. It can be used in many field, such as food, drug and cosmetic. Investigation and research were carried out for several methods of extracting anthocyanins and comparing these methods, purifying anthocyanins through thin layer chromatography and column chromatography and finding out optimum chromatography material and eluting solvent, and ability of reducing blood-fat.Investigation were carried out for several methods of extracting anthocyanin, such as water extracting, ultrasonic extracting, enzyme extracting and super critical fluid-enzyme extract ing. There is no report so far for method of super critical fluid-enzyme extracting. The result showed that extracting rate of water extracting was the lowest 2.7% and that of super critical fluid-enzyme extracting was the highest 6.5% through comparing these extracting methods. Through orthogonal experiment analysis, the optimum condition of enzyme extracting, tempera ture 30℃, pH value 7.0, solid-to-liquid ratio 1:6, enzyme consumption 24IU, time 90min, was found out and its extracting rate was 6.4%.The result of reducing blood-fat showed that cleaning hydroxyl free radical rate of anthocyanin was 38.8% when concentration of anthocyanin was 0.4mg/mL, and restraining lipid dioxition rate of anthocyanin was 13.4% when concentration of anthocyanin was 0.4mg/mL.The high fat model of big white mouse was established. The result showed that cutting cholesterol rate was 22.34% when anthocyanin dosage was 0.5g/d, cutting triglyceride rate was 34.4% when anthocyanin dosage was 0.6g/d.
引文
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[2] 崔介君,孙培龙,马新.原花青素的研究进展.食品科技.2003,(2):92～95
[3] 钟瑞敏.花色苷结构与稳定性的关系及其应用研究.韶关学院学报.2001,22(12):79～83
[4] G. Mazza. Recent Development in the stabilization of Anthocyanins in Food Products. Food Chemistry, 1987, (88): 207～225
[5] S. Renand, M. Zorgelil. Wine, alcohol, platelets and the French paradox for coronary heart disease. Lancet, 1992, 339: 1523～1526
[6] G. Pifferi, R. Cultrera. Enzymatic degradation of anthoyanins: The role of sweet cherry polyphe noloxidase. Food sci.,1974, 39,786
[7] G. Daravingas, RF Gain. Changes in the anthocyanin of raspberries during processing and storage. Food Sci., 1965, 30(3): 400～405
[8] N. Palamindis, P. Matkakis. Stability of grape anthocyanin in a carbonated beverage. Food Sci., 1975, 40: 1047～1049
[9] 刘小芬,刘剑秋,陈炳华等.轮叶蒲桃果实红色素提取与基本性质分析.亚热带植物科学.2003,32(4):15～18
[10] J. G. Swweeny. Effect of flavonoid sulfonates on the photobleaching of anthocyanins in acid solution. Food Chem.,1981. 29: 563～567
[11] R. E. Wrolstad, J. A. Erlandson. Effect of metal ions on the color of strawberry paree. Food Sci, 1973, 38:460
[12] Sitrunk and Cash. The effect of certain chemicals on the color and polysaccharides of strawberry puree. Food Techn., 1970, 24: 473
[13] G. Mazza, R. Brouillard. Recent development in the stabilization of anthocyanins in food products. Food Chem.,1987, 25(3):207～225
[14] 陈健初,苏平,叶兴乾.杨梅花色苷及色泽稳定性研究.浙江大学学报.1994,20(2):178～182
[15] 李进,彭宇,彭子模.鸡冠花红色素理化性质及其稳定性研究.生物技术.2004,14(1):21～24
[16] 励建荣,岑沛霖,Joyce D.C.杨梅汁内花色苷热降解动力学研究.科技通报.2002,18(1):1～5
[17] R. E. Wrolstad, M. M. Giusti. Anthocyanins from Radishes and Red-fleshed Potatoes. Chemistry and Physiology of Selected Food Colorants, J M Ames and T F Hofrnann (eds). ACS Symposium Series No. 775, Washingfton, DC: American Chemical Society., 2001: 66～89
[18] 赵新淮,刘宏芳.黑加仑果渣红色素的稳定性及热降解动力学.食品工业科技.1998,19(3):8～9
[19] 陈勇.不同PH下紫苏色素热动力学研究.青岛大学学报.1996,11(3):25～28
[20] F. M. Clydesdale, J. H. Main, J Francis. Roselle (Hibiscus sabdariffa L.) Anthocyanins as Colorants for Beverages and Gelatin Desserts. Food Prot.,1979,42: 204～207
[21] M. M. Giusti, H Ghanadan, R. E. Wrolstad. Elucidation of the Structure and Conformation of Red Radish (Raphanus sativus) Anthocyanins Using One- and Two-dimensional Nuclear Magnetic Resonance Techniques. Food Chem., 1998. 46: 4858～4863
[22] M. Monica Giusti. Applications of acylated anthocyanins as natutre food colourants. Business Briefing: Innovative. Food Ingredients, 2002: 1～3
[23] O. Dangles, N. Saito, R. Brouillard. Anthocyanin intramolecular copigment effect. Phytoche mistry, 1993.34:119～124
[24] N. Saito, F. Tatsuzawa, K. Yoda, et al. Acylated cyaniding glycosides in the violet-blue flow ers of Ipomoea purpurea. Phytochemistry, 1995.40(4): 1283～1289
[25] K. Yoshida, T. Kondo, T. Goto. Unusually stable monoacylated anthocyanin from purple yam Dioscorea alata. Tetrahedron Lett., 1991. 32(40): 5579～5580
[26] C. A. Winlliams, J. Greenham, J.B.Harbome. Acylatted anthocyanins and flavonls from purple flowers of Dedrobium cv.Pompadour.Biochemical Systematics and Ecology,2002.30:667～675
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