Study On HPLC Determination Methods And Content Changes Of Vitamins In Japanese Persimmon

Japanese persimmon(Diospyros kaki Thunb.) belongs to the genus Diospyros(Ebeuaceae), which is utilized as the representative species of fruit trees and is also one of the important fruit-tree species in China. The fruit of Japanese persimmon is sweet, juicy and colorful. It has unique flavor and rich nutrition, with high edible and medicinal value. Vitamins are necessary for the maintenance of human and animal physiological functions and play important roles in material metabolism but natural foods with less vitamin content. Vitamins are a well-known group of compounds with complex structure and have a lot of extraction and analysis mehod. High performance liquid chromatography(HPLC) with high separation efficiency and analysis advantages of fast speed, high sensitivity, is applicated for determination of vitamins more and more extensively.There are less studies on the main nutritional components of persimmon, research on persimmon vitamin is also more concentrated in leaves and vitamin C and carotenoids, researches on the main nutrients such as vitamins in persimmon are limited and lack of systematic.In this experiment, we established the Japanese persimmon vitamins extraction and determination method by HPLC with the non-astringent persimmon-Yang Feng and astringent persimmon-Mopan as the test materials. On the basis of that,we have studied the vitamin C,vitamin B1,B2,B3,B6,β-carotene(prime of vitamin A)and α-tocopherol(prime of vitamin E)content at different developmental stages in different parts of the(persimmon flesh, persimmon peel, persimmon leaf, persimmon calyx),and we have also studied a variety of vitamins content the leaves and fruits of 46 mature persimmon main cultivars.In order to develop new products,improve the commodity performance and added value and provide the basis for promoting the healthy development of the Japanese persimmon industry.The main results are as follows:1. A simple and rapid HPLC method for the determination of ascorbic acid in Japanese persimmon was developed and validated.Chromatographic conditions: liquid chromatograph: Aglient 1200 system, chromatographic column:Elite Hypersil BDS- C18 column(250 x 4.6 mm,5μm); Mobile phase: 0.02 M phosphate buffer(p H5.8); Flow rate: 1 ml/min; Column temperature 25℃; detection wavelength: 260 nm; Sample quantity: 25?L.The method provided low limits of detection(LOD) and quantification(LOQ), good linearity, accuracy and precision.2. A simple, fast and accurate liquid chromatography method for simultaneous separation and determination of four vitamin compounds, including vitamin B1(thiamine), B3(nicotinic acid), B6(pyridoxine) and B2(riboflavin) in Japanese persimmon, was developed and validated. For chromatographic analysis, the column used for chromatographic separation was Elite Hypersil BDS- C18 column(250 x 4.6 mm,5μm). The mobile phase adopted was acetonitrile(A) and 0.1% phosphoric acid aqueous solution(B)(v/v) using a linear gradient elution. The flow-rate was kept at 0.8 m L/min all the times. The column was operated at 25℃. The injection volume was 20μL.The eluent was monitored at 270 nm for all analytes. The method provided low limits of detection(LOD) and quantification(LOQ), good linearity, accuracy and precision.3. A simple and rapid HPLC method for the extraction and determination of beta-carotene and alpha-tocopherol in Japanese persimmon was developed and validated. The chromatographic separations were performed on a Agilent ZORBAX ODS C18 column(250 mm x 4.6 mm, 5μm). A solution of acetonitrile and methylene chloride(7:3) : methanol = 60:40 was used as the mobile phase with a flow-rate of 1 m L/min. The column was operated at 25℃. The detection wavelength was monitored at 450 nm for beta-carotene and 300 nm for alpha-tocopherol.The injection volume was 25μL. The method provided high accuracy and precision, high sensitivity, good effective and feasible.4. At different developmental stages of the Japanese persimmon pulp, peel, leaf and calyx,the general trend of vitamin C,vitamin B1,vitamin B3,vitamin B6 content during annual variation is high to low, vitamin B2, beta-carotene and alpha-tocopherol content changes with time from low to high, but different varieties and different parts of fruit changes different.5. At the same Japanese persimmon developmental stage in different organs(parts), beta-carotene and alpha-tocopherol content level is leaf > calyx > peel >pulp. Vitamin B1,B2,B3,B6 content level is leaf > calyx > pulp >peel. Vitamin C content level is calyx > leaf > peel > pulp,but the content of different vitamins changes slightly during different period.6. The contents of vitamin in different organs(parts) of different varieties were different. In the leaves of all the varieties tested, vitamin C, vitamin B content is highest in Jiro and Xiaoguotianshi; vitamin A content is highest in : Hanguopanshi, Taianmantanghong; vitamin E content is highest in Dahebaimu,Lichengdamianhu. In the fruit of all the varieties tested, vitamin C content is highest in Xihong and fuyou; vitamin B content is highest in Xihong and Changanfujianding;vitamin A content is highest in Changanfujianding and Boaibayuehuang; vitamin E content is highest in Changanfujianding and Taianmantanghong.