纯化及性质研究

野生毛葡萄皮色素的提取、纯化及性质研究

STUDIES ON THE EXTRACTION,PURIFICATION AND PROPERTIES OF VITIS QUINQUANGULARIS REHD SKIN PIGMENT

作者：薛敏敏　　导师：邓　纪

中南林业科技大学　　食品科学2011届硕士

摘　要

色素是食品中重要的添加剂，随着人类对合成色素毒性作用的认识和天然绿色食品的渴求，天然色素的需求量在增大。我国的野生葡萄资源非常丰富，积极开发这些资源，不论在经济上还是在科学价值上都有重要的意义。本课题对野生毛葡萄皮色素的提取方法、稳定性、纯化和抗氧化性进行了研究，结果如下：

通过显色反应以及可见光谱数据分析，表明该色素属于花色苷类色素。为了对色素提取效果进行研究，采取了传统溶剂提取法和微波—超声波提取法，对两种方法的提取率进行了比较，并采用正交设计优化实验影响因子。结果表明：溶剂浸提的最佳工艺为，以1%盐酸-70%乙醇为提取剂，时间60min，料液比1:5，提取温度50℃。微波—超声波协调提取的最佳工艺为，微波功率60%（540W），微波时间60s，超声波时间60s，料液比1:5。与传统水浴提取法相比，微波—超声波协同提取法提取时间显著缩短，总花色苷含量也提高了41%。

野生毛葡萄皮色素的稳定性试验结果表明：光照、高温、H₂O₂ 、Vc均能引起色素的降解，Ca²⁺对色素稳定性没有显著影响。Zn²⁺、Al³⁺能够增加花色苷的稳定性。Fe²⁺有一定的破坏作用。野生毛葡萄皮色素在低浓度的山梨酸钾、苯甲酸钠中稳定。食盐、蔗糖均有明显的增色作用。

通过考察8种大孔吸附树脂的吸附性及解析性能，确定NKAⅡ为最佳试验树脂。应用Langmuir方程对NKAⅡ树脂的吸附动力学进行了研究。NKAⅡ树脂的吸附速率常数为0.3369，表现出对野生毛葡萄皮色素很好的选择吸附性。NKAⅡ树脂的静态试验最佳条件为：吸附温度35℃，洗脱温度在25℃～35℃范围内选择，pH值为2，吸附平衡时间为5h，洗脱剂为70%乙醇溶液（pH=2）。NKAⅡ树脂的动态吸附最优条件：上样流速为1mL/min，上样液浓度为1.568mg/ml，洗脱速度为1mL/min，洗脱剂用量为9BV，色素液纯化后色价为原来的4.76倍。

通过四种不同的体系研究了野生毛葡萄皮色素的抗氧化性能。发现野生毛葡萄色素有较强的抗氧化性能。野生毛葡萄色素对DPPH、羟自由基OH、O^2-均有清除作用，同等浓度下，Vc的清除效果最好，纯化色素的清除率高于粗色素的清除率，且接近于Vc。野生毛葡萄色素的还原能力随着浓度的增加而增强，纯化色素的还原力高于粗色素的还原力。

关键词　色素　提取　稳定性　纯化　抗氧化

Abstract

Pigment is food additive，and with the people’s understanding to toxicity of synthetic pigments and the desire to the nature green food，the demand for natural pigment is increasing. Our country is very rich in wild grapes，and to develop these resources，whether in economic or scientific value is important. The extraction condition of vitis quinquangularis Rehd pigment，the stabilities of pigment，the purification parameters of pigment，and anoxidation activity of pigment had been studied in this paper. The results were summarized as follows：

The results of experiment of coloration,and analysis of visible spectrum showed that the vitis quinquangularis Rehd pigment was confirmed as a kind of anthocycyanins. In order to research the pigment extraction effect,the two extraction methods: traditional solvent extraction,microwave and ultrasonic extraction were compared in this paper. The factor was optimized by orthogonal design. The optimum extraction parameters of traditional solvent extraction were as following：70% ethanol with 1%HCL(pH=2)as soaking liquid,solvent-to-solid(1:5),extraction 60min at 50℃. The optimum parameters by microwave and ultrasonic extraction were as following：70% ethanol with 1%HCL(pH=2)as soaking liquid,solvent-to-solid(1:5),ultrasonic power(150W),the power of microwave 60%(540W),the treatment time of microwave 60s,the treatment time of ultrasonic 60s,The effect of microwave and ultrasonic extraction　was higher than traditional solvent extraction,The extraction time was clearly reduced， and the total content of anchocyanins improved by 41%.

The result of stability test showed that the sunlight，high temperature，H₂O₂ and Vc accelerated the degradation of the pigment。Ca²⁺ Had little influence on stability of the pigment。Zn²⁺、Al³⁺ Could increase the stability of pigment。Fe²⁺ had some damaging effects. Low concentration of Potassium sorbate and Benzoic Acid had little influence on the stability of the pigment。Salt and sucrose had all copigmentation function,

The purification of vitis quinquangularis Rehd pigment was studied by macroporous resin. It was proved that the NKAⅡ macroporous resin was the most suitable sorbent among the eight kinds of macroporous resin. Adsorption kinetics of NKAⅡ resin by applicating Langmuir equation was studied，and adsorption rate constant the NKAⅡ resin was 0.3369，showing a good choice of adsorption to vitis quinquangularis Rehd pigment。The optimum of static experiment were：adsorption temperature 35℃ ，desorption temperature form 25 to 35℃ ，pH=2,adsorption time 5h,70% ethanol with 1%HCL(pH=2)as enluent. The optimum of dynamic experiment were：the sample velocity 1.0ml/min,the adsorptive concentration 1.568mg/mL,the desorption velocity 1.0mL/min,the eluent consumption 9BV. Under the condition mentioned above,the color value of purified pigment was 4.76 times to that of unpurified.

The antioxidation of samples was studied by four different systems. The results showed that vitis quinquangularis Rehd skin pigment had strong antioxidant properties. Remarkable scavenging effects against DPPH,OH、O^2- were detected,and as the same concentration,the reducing power of Vc was highest,and purified pigment was higher than unpurified pigment. The deoxidative ability of the pigment increased with the concentration,and the deoxidative ability of purified pigment was higher than unpurified pigment.

Key words　Pigment　Extraction　Stability　Purification　Antioxidants