The Study Of The Determination Of Four Polyphenols In Different Parts Of Pomegranate Fruit

The study of pomegranate fruit was attached importance by scientist all over the world day by day for plentiful biological activity compounds in it. So far, the biological activity compounds are pomegranate polyphenols, such as gallic acid, punicalin, punicalagins and ellagic acid are chief compounds which was widely accepted. But all determination methods of polyphenols were for single composition, the method of several polyphenols were determined in different parts of pomegranate fruit hadn’t been developed. In this study, the HPLC method was developed for determination of gallic acid, punicalin, punicalagins and ellagic acid in pomegranate peel, pomegranate juice, pomegranate seed, pomegranate leaf and pomegranate flesh. The pretreatment method of samples for several parts (pomegranate peel, pomegranate juice, pomegranate seed, pomegranate leaf and pomegranate flesh) and the chromatogram condition for simultaneous determination of gallic acid, punicalin, punicalagins and ellagic acid were studied in this study. The results suggested:the pomegranate peel, pomegranate seed, pomegranate leaf and pomegranate flesh were distilled by mixture of methanol, ethanol, acetone and water, the samples gained after centrifugating could be used to determine punicalin, punicalagins and ellagic acid accurately, and the samples after extracting by ethyl acetate could be used to determine gallic acid accurately; freeze-drying and ultrasonic-extract by methanol was carried out on pomegranate juice, the samples gained after centrifugating could be used to determine punicalin, punicalagins and ellagic acid accurately, pomegranate juice was extracted by ethyl acetate could be used to determine gallic acid accurately.The RP-HPLC was carried out on Arcus EP-C18(250mm×4.6mm,5μm). The gradient elution was performed by a mobile phase consisted a mixture of methanol (A) and solvent0.1%TFA (B), was used under binary linear gradient conditions as follows:0-10min,2%-13%solvent A in solvent B;10-20min,13%solvent A in solvent B;20-20.5min,13%-40%solvent A in solvent B;20.5-42min,40%solvent A in solvent B; at a flow rate of1.0mL-min-1, and the detection wavelength of gallic acid was at254nm; punicalin, punicalagins and ellagic acid were at377nm.The characteristic peak were right and the linear equation were y=2353.15x-17496, R=0.9997for gallic acid at0.020-0.320μg; y=880.44x-23405.09, R=0.9971for punicalin at0.038-0.608μg; y=403.18x-11971.98, R=0.9968and y=413.95x-12127.42, R=0.9978for punicalagins at0.074-1.184μg; y=1112.91x-18658.51, R=0.9994for ellagic acid0.039-0.624μg. The HPLC method which had good precision, stability, repeatability, and average recoveries of samples could be used to determine gallic acid, punicalin, punicalagins and ellagic acid in pomegranate peel, pomegranate juice, pomegranate seed, pomegranate leaf and pomegranate flesh. The five parts of pomegranate fruit, the contents of the four polyphenols are maximum in pomegranate peel, second in pomegranate flesh and least in pomegranate juice. In pomegranate peel, the contents of punicalagins is maximum, punicalin and ellagic acid are second, gallic acid is least; in pomegranate juice, the contents of gallic acid, punicalin and ellagic acid are considerable and punicalagins is maximum; in pomegranate seed, the contents of gallic acid and ellagic acid are considerable, punicalin and punicalagins were not determined. The contents of ellagic acid in pomegranate leaf is maximum in the five parts of pomegranate fruit; punicalagins in pomegranate flesh is of high content which is behind of pomegranate peel.The method is accurate for quality control of the four polyphenols in the five parts of pomegranate. The gallic acid, punicalin, punicalagins and ellagic acid were not only determined in different parts of one pomegranate fruit but also in different kinds of pomegranate fruit by the HPLC method. So, the distribution of four important polyphenols in different parts of pomegranate fruit and in different kinds of pomegranate fruit. Then, pomegranate fruit can be distinguished between good and bad to a certain degree. Scientific basis was provided for deep study and the quality control of related products about pomegranate fruit.