Study On Preparation And Functional Properties Of Methyl Pyranoanthocyanin

Methyl pyranoanthocyanins are new anthocyanin-derived pigments found inaging fruit wines， which are formed by a series of reactions between berryanthocyanins and secondary metabolites of small molecules． Methylpyranoanthocyanins are one of the important representatives of pyranoanthocyaninfamilies．Methyl pyranoanthocyanins was synthesized by the reaction betweenmalvidin3-glucosides (Mv) from the extracts of red grape skins as raw material andacetone in this paper，and the optimized reaction conditions were studied byorthogonal experiment design．The resulting product pigment was isolated andpurified by hyphenated techniques of polyamide resin，TSK Toyopearl gel columnchromatography and preparative HPLC， and its structure characterized byHPLC-DAD-EI-MS/MS．Methyl pyranoanthocyanins was investigated in terms ofcharacterization of chromatic stability and storage stability including degradationdynamics and CIELab parameters under different conditions， scavenging freeradical and reducing power as well as the protective effect on aged model humanembryonic lung fibroblast cells MRC-5damage by H2O2，and compared withMv． Furthermore， the toxicological safety will also be assessed formethyl-pyranoMv．The main conclusions are as follows：1．HPLC was used to monitor the reaction and the optimal synthesis conditionswere as follows：the ratio of grape anthocyanin pigments (mg) to acetone (mL) was15:1，concentration of grape skin pigments was2.5mg/mL，reaction temperaturewas set at45℃and pH was3.0．The main reaction product was confirmed byHPLC-MS/MS to be methyl pyranomalvidin3-glucoside (pyranoMv) with the yieldof49.4%under these conditions．2．The purity of methyl-pyranoMv was71.16%，91.60%，98.31%after isolatedand purified by hyphenated techniques of polyamide resin，TSK Toyopearl gelcolumn chromatography and preparative HPLC，respectively．And its color valvewas219(citrate buffer, pH3.0)．CIELab parameters showed that methyl-pyranoMvpresents the best yellow hue under pH2.5．Meanwhile，Comparing the smallest△E*values at acidic region pH ranges，it was noticeable that methyl-pyranoMvwas clearly more stable than Mv．In addition，Methyl-pyranoMv was also showeda greater resistance to discoloration by SO2． The ε values of methyl-pyranoMv in different solvents and various pH valueswere higher than Mv，which the order was as follows: in methanol> in ethanol>inwater．Variation of ε values under strong acid conditions suggests no significantdifference as well as under medium acidic conditions．Moreover，ε values ofmethyl-pyranoMv in simulated acidic beverages conditions (water solution，pH4.0)were still much higher as12983mol-1dm3cm-1．3．Methyl-pyranoMv had a obvious stability than Mv in different heatingtemperature and heating time．The retention rate of methyl methyl-pyranoMv was93.2%but Mv was only69.8%after heating1h at90℃．The stability of methyl-pyranoMv under sunlight conditions was worse than Mv，while it keeps a goodstability under dark conditions．Thermal and light degradation kinetics of methyl-pyranoMv fit a first-order reaction model．The half-life of methyl-pyranoMv at50℃and90℃was99d and14d，while Mv was18d and4d．Methyl-pyranoMv hadhigher storage stability than Mv in ethanol solution and the aqueoussystem．However，It had a higher storage stability in ethanol solution than that inwater．Methyl-pyranoMv had a much higher half-life in ethanol (257d) and water(144d) than Mv (131d and71d) at room temperature．4． The hydroxyl radical scavenging ability and reducing power ofmethyl-pyranoMv pigment increased with the increase of concentration． Thehydroxyl radical scavenging ability of methyl-pyranoMv is higher than Mv (P<0.05)and equivalent as ascorbic acid，but the trend was inverted compared with theDPPH assay and the reducing power method(P>0.05)．Moreover，Methyl-pyranoMvpigment could protect the human embryonic lung fibroblast cells MRC-5fromH2O2induced cytotoxicity．5．The acute oral LD50was7945mg/kg in male mice and6816mg/kg in femalemice for methyl-pyranoMv，which belong to actual non-toxic．...