| The mature Solanum nigrum L. fruits were chosen as material. The extraction; purification; isolation and identification of anthocyanins from Solanum nigrum L. fruits were investigated. The kinetics of degradation of liquid and dried extracts under different pH and temperatures were also studied. The crude extract, purified extract, faction 1 and faction 2 were dried by vacuum freeze dryer and oven at 50℃ and then the antioxidant activity of them were compared.The best solvent to extract anthocyanins from Solanum nigrum L. fruits and the optimal extraction conditions were obtained.14 kinds of solvents were chosen as extraction solvents. The correlation between yields of total phenols; flavonoids; anthocyanins and antioxidant activity were analyzed. Based on the single-factor test analysis, the full factorial design, steepest ascent design and central composite design for response surface analysis were applied. The results were as follows:60% ethanol was the best solvent to extract anthocyanins from Solanum nigrum L. fruits. The content of total phenols; flavonoids and anthocyanins from 60% ethanol extracts were 1.99±0.03mg/g,1.02±0.01mg/g and 0.82±0.02mg/g, respectively. To ABTS+·and ·OH the scavenging activity was up to 88.18%±0.59% and 82.45%±0.59%, respectively. The ferric reducing antioxidant capacity of the extract expressed as 5.45±0.06mmol/L FeSO4 equivalent The polyphenols was strongly positively correlated to antioxidant activity, the correlation indexes were 0.959(FRAP),0.627(·OH), and 0.822(ABTS+·), respectively. The best extraction condition of anthocyanins from Solanum nigrum L. fruits was as follows:pH1.0, temperature 29℃, time 85.5min and liquid-solid ratio 1:25. On this extraction condition, the yield of Solanum nigrum L. anthocyanins was 0.86±0.03mg/g.We selected the best macroporous resin through static adsorption and obtained the optimal purification conditions by the dynamic desorption. The results were as follows:X-5 macroporous resin exhibited the excellent characteristics for purification of Solanum nigrum L. anthocyanins. We found that the time of absorption and desorption equilibrium was 5h and 6h, respectively, the column diameter to length ratio was 1:25, the maximum sample volume was 0.6BV, the concentration of absorption equilibrium was 0.4mg/mL, the eluent volume was 3.5BV, and elution flow rate was 2.0BV/h. We used pH2.060% ethanol solution as eluent. The purity was 19 times of unpurified by combination of those optimal conditions.N-butanol:acetic acid:water=4:1:3 was used as the best developing solvent for Solanum nigrum L. anthocyanins by thin-layer chromatography that compared of different developing solvent systems. A blue fraction and a red fraction were obtained by silica gel column chromatography. Moreover, two kinds of anthocyanins fractions were isolated from Solanum nigrum L. fruits by column chromatography method and HPLC-DAD-ESI-MS/MS analysis and acid hydrolysis were applied for identification. Three anthocyanins were identified for the first time in this fruit, as only delphinidin-3-succinyl-arabinoside was found in faction 1, while both cyanidin-3-galactoside and cyanidin-3-acetyl-galactoside were observed in fraction 2.The degradation kinetics of four anthocyanins extracts from Solanum nigrum L. fruits under different pH and temperature were investigated. The results were as follows:the degradation of anthocyanins from Solanum nigrum L. fruits follows the first-order reaction kinetics. The reaction rate constant of the anthocyanins degrading reaction was increased in exponential pattern as the temperature went up and the half-life was reduced in exponential pattern.60℃ and 70℃, the reaction rate constant of crude and purified extracts degrading reaction were increased in logarithmic pattern as the pH went up and the half-life was reduced in exponential pattern. The reaction rate constant of fraction 1 and fraction 2 degrading reaction were increased in as the exponential pattern pH went up and the half-life was reduced in exponential pattern. However,80℃ and 90℃, the rate constants and half-life were not changed regularly. Four kinds of anthocyanins extracts were stable at 60℃ pH1.0, and crude extract was more stable than others.Vacuum freeze-drying technology was benefit to the stability of anthocyanins. The dried crude extracts were the most unstable than other dried extracts, while the dried fraction 2 showed the best stability. The half-life of fraction 1, which was identified as delphinidin-3-succinyl-arabinoside was 80.6 weeks under the storage condition at -18℃. This is consistent with half-life requirements for commercial anthocyanins.Four extracts (curde, purified, faction 1 and faction 2) from Solanum nigrum L. friuts were evaluated for their antioxidant (DPPH, ABTS, FRAP and liposome peroxidation assays), and use Trolox and Rutin as the positive control. Then we compared the influence on dried methods of the antioxidant of different extracts. The strongest activities were obtained by crude extract for the DPPH, ABTS and FRAP assay (IC50 values of was 33.35±0.23μg/mL, 24.92±0.18μg/mL and 9.25±0.19mM FeSO4, respectively). Fraction 1 showed the best inhibition of lipid peroxidation assay (IC50 value was 0.26±0.02μg/mL). Faction 1 had a stronger activity than faction 2 because of the existence of acylated anthocyanins. Dried method can help improving the antioxidant activity of Solanum nigrum L. fruits anthocyanins. The antioxidant activities of freeze-drying extracts were slightly stronger than low-temperature drying extracts. However, we could chose low-temperature drying method in consideration of the cost of production. |
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