The Extraction Technologies Of Polyphenols In Coffee And Evaluation Of Antioxidant Activity

As the largest consumption of drinks in the world, coffee's nutritional value has much more attracted people'eyes. Early in the 19th century, the abroad began to study the function of components of coffee beans. With the advances in technology, innovation in research methods and technology, the nutrient composition of coffee beans and non-nutrient components has been gradually clear, corresponding health function and pharmacological effect has been further confirmed. In particular, a large number of non-nutrient components—polyphenols in the coffee beans has become the focus in international natural medicine field, these substances play an increasingly important role in the leading compound discovery and the innovation of drugs development. Yunnan, Hainan, Guangxi and Guangdong is China's main coffee growing areas, and very rich in resources. Therefore, the establishment of the standard and regular distilling system of coffee polyphenols, in the effective development and rational utilization of the coffee resources, change the export trade mode of raw materials alone, has the very important economic value and practical significance.The key of the exploitation and use of natural products is the advanced extraction and separation technologies, and the accurate, efficient and fast method of analysis. This study established the high performance liquid chromatography (HPLC) analytical methods of the polyphenols in the coffee beans, identified the optimal extraction parameters of the polyphenols from the coffee, and further studyed the optimal conditions that the polyamide resin separated and purified the polyphenol of the coffee, analyzed the specific composition of polyphenols of the coffee by using the HPLC techniques, evaluated the in vitro antioxidant activity of polyphenols in coffee beans, the results were as follows:A method of reversed-phase high performance liquid chromatography coupled with diode array detector (HPLC-DAD) was established for the simultaneous determination of 6 phenolic acids (Caffeic acid,3-O-Caffeoylquinic acid,4-O-Caffeoylquinic acid,5-O-Caffeoylquinic acid, 3,5-O-Dicaffeoylquinic acid,4,5-O-Dicaffeoylquinic acid) in green coffee beans. Separation was achieved using a Kromasil C18 column (4.6 mm×200 mm,5μm), under the gradient elution with the mobile phase of acetonitrile and 0.1%(v/v) formic acid/water. The 6 phenolic acids were well separated within 45 min. The recoveries from 90.76% to 104.73% with the relative standard deviation between 0.7%-3.9%.The method is simple, rapid and highly sensitive, suitable for the simultaneous determination of 6 phenolic acids and quality control of coffee beans. In addition, using the HPLC method the test also compared the content of coffee polyphenol in four areas of Yunnan, the result showed that the highest content of total polyphenols is in Baoshan coffee.The effects on yield of extraction solution, extraction method (temperature), extraction time, and extraction times were researched. The optimization technics parameters of the extraction of coffee polyphenols were as follows:coffee was extracted at 80℃in 70% ethanol twice, with 1:7 as ratio of solid/liquid,extraction 2h every time.7 kinds of macroporous resins were compared for the separation of polyphenol of coffee, Polyamide resin exhibited superior adsorption and regeneration properties. The optimum conditions for separating and purifying polyphenol of coffee were obtained. The results showed that the separation optimization technics parameters with Polyamide resin were as follows:the sample concentration is 4.44mg/ml,the moving speed is 2BV/h, adsorbing twice,and eluting volume is 3BV with 80% ethanol as eluting solvent. The purity of polyphenol of coffee can be improved from 14.43% up to 53.32% and the recovery rate is 78.21%. UV spectrophotometer and HPLC were used to analyze Preliminary purified coffee polyphenols. The former measured the total polyphenol content was 53.32%, the latter measured total content of 6 kinds of the polyphenols was 47.55%, the results indicated that the main Phenolic substances in coffee are caffeyolquinic acid (CQA) and its derivatives.Through five kinds of detection system (scavenging hydrogen peroxide, hydroxyl radical, superoxide anion radical, DPPH radical and reducing power determination), the experiment studied the antioxidant properties of coffee polyphenols. The result showed that coffee polyphenols have certain abilities of scavenging free radieals and reducing Fe3+, and Significant correlation exists between the clearance rate of the free radieals, the abilities of reducing Fe3+ and the Sample concentration.