Purification, Composition Analysis and Anti-oxidation Activity of Anthocyanidins in Blueberries (Vaccinium Uliginosum) of China
|School||Ocean University of China|
|Keywords||proanthocyanidins of Vaccinium uliginosum extraction and purification composition analysis anti-oxidation free radicals removal|
Blueberry is a fruit with a high economic value and belongs to Ericaceae ,Vaccinium sp.Wild Blueberry of China is mainly located in the northern Da Hinggan (Heilongjiang, Inner Mongolia), Jilin Changbai Mountain. Blueberry fruit contains a large number of nutrients.There are a lot of phenolic compounds found in the Blueberry. The major constituent of phenolic compound is proanthocyanidins (PC) which has a highly hydroxyl structure. It can act as an anti-oxidation,remove free radicals, and reduce risk of chronic diseases such as cardiovascular disease and cancer. It has a very high value of development and utilization.Proanthocyanidins, flavanols mainly by the monomer (+) - Catechin [(+)- catechin, C ],(-) a epicatechin [(-)- epicatechin, EC] and (-) a table catechin gallate [(-)- epicatechin-3-O-gallate, ECG] polymerization by the highest average degree of polymerization 15 Tatsu polymers. Usually, 2-4 polymers are known as low - polymer (procyanidolic oligomers. referred to as OPC or PCO), and more than pentamer are known as high polymer (procyanidolic polymers, referred to as PPC). PC connects between the modules mainly by the C4-C6 and C4-C8. As the differences of single body conformation or bonding location, there are a variety of isomers, and the polarity between the polymer and polyisomers are very similar.In this study, wild blueberry fruits were washed, homogenate broken Marc by centrifugal separation as the raw material, taking the Marc to experiment. and the PC was extracted from Marc with precooling acetone .The best technical marameters were 4℃as extracting temperature, 75%(v/v) acetone content,Marc and acetone the ratio of 1:3(v/v), and 1 hour of extracting time. 4000rpm centrifuged for 30min, the collection of the supernatant filter, vacuum concentration of acetone to all steamed out, recycling reusable acetone, concentrated liquid volume ratio of 4:1 by adding a mixture of non-polar solvents, such as the volume of extraction of 2 or 3 times.The adsorption yield of PC in HP-2MG was 90.8%.However the desorption yield was 96.0%. macroporous resin column was loaded with wet, after pretreatment standby. Concentrated extract in the column before the pole with access to water away deoxidization nitrogen 1~2 size, to exclude the column oxygen, preventing procyanidins from oxidation, and will be used xibozhi parcel post, to the effect of light . Tai-chu, the macroporous resin adsorption saturation, and then pass into the nitrogen deoxidization water to rinse fluid no obvious color. Then 10% -50% -75% -95% ethanol for washing, the concentration of ethanol pH of 3 ~ 6, and has access to nitrogen in order to maintain system stability. 0.5 ~ 4 column volume / hour over the speed of column after column to collect the liquid. Outflow of liquid the color of light, stop on the column, record the size of the eluate. Collecting the eluate from the 10%, 50%, 75%, 95% ethanol were concentrated by Rotary Evaporator,concentrated solution was freeze-dried.The procedure is:After the macroporous resin chromatography, the yield of anthocyanidins came up to 0.72%. The total content of flavanol was measured up to 95% by vanillin method.A RP-HPLC method and ESI-MS were established to separate and analyze the component of anthocyanidins from blueberries of Vaccinium uliginosum of China. The method was first preparing the crude extract of anthocyanidins from blueberries. Then the RP-HPLC was performed on Alltima C18 column (250 mm×4.6 mm,5μm), with the mobile phase of solution A 2% formic acid in water and solution B 2% formic acid in methanol, to elute the component of anthocyanidins by two gradient.The main elution peak was collected and then analyzed using ESI-MS in positive ion mode. According to the value of m/z, the degree of polymerization was detected. The result showed that the extract of anthocyanidins in blueberries of Vaccinium uliginosum was separated into mainly eight elution peaks, the total content up to 85% measured by peak areas.According to their mass spectra, the peaks 4、8 and 12 are monomers, the peaks 6 and 10 are dimers, the peak 7 is trimer and the peaks 13 and 14 are tetramers. It is showed that blueberries of Vaccinium uliginosum of China contain a wealth of anthocyanidins and the oligomer is the main component. This paper reported the composition of anthocyanidins in blueberries of Vaccinium uliginosum of Da Xing An Ling of China for the first time, which has some references value in the subsequent study of the structure, the function, the quality inspection, the production research and development.To further study the nature of proanthocyanidins, catechin and epicatechin mixture (recorded as PCM group), different proanthocyanidin dimers (recorded as PCD group), degree of polymerization> 2, a mixture of blueberry proanthocyanidins (credited for the PCT group) were used to study the blueberry proanthocyanid oins to restore the ability to make the determination. The experimental results show that proanthocyanidins with the relative average degree of polymerization of lower molecular weight and narrow molecular weight distribution, that is, its ability to restore the PCM <PCT <PCD. At the same time, linoleic acid system,superoxide anion radical of O2 - ? ,the removal of hydroxyl radical scavenging effect of HO ? experimental were used study antioxidant activity of blueberry proanthocyanidins. The results showed that Blueberry proanthocyanidins were more powerful than Vc in anti-oxidation capacity in the linoleic acid system , and with the concentration increasing ,its antioxidant capacity similar to the synthetic antioxidant BHT. Blueberry proanthocyanidins were more powerful than grape seed proanthocyanidins and Vc in antioxidation capacity and free radicals removal.