Extraction And Purification Of ?-glucosidase And Its Improvement On The Flavor Of Red-orange Juice

Lianjiang red-orange had big fruit and brilliant flesh color,and very popular at domestic and foreign country.But Lianjiang red-orange as raw material processing products in the market were rarely reported,so the Lianjiang red-orange for processing products was a hot spot of research.With the rapid development of the domestic juice industry,fruit juice became another sales channels of fruit.?-glucosidase could promote the release of aroma compoments,it played an important role in the processing of fruit juice,fruit wine.After years of research,the domestic and foreign researchers had extracted and purifired various ?-glucosidase from microorganisms,animals and plants,the research had relatively mature.But the demand for ?-glucosidase was still far greater the supply,so improving the efficiency of producing ?-glucosidase was also one of the key point of research.In order to improve the efficiency of microbial production ?-glucosidase and its effect on Lianjiang red-orange,this research improved the efficiency of ?-glucosidase of Aspergillus niger CICC2475 from four factors: fermentation time,broth content,initial pH and inoculation amount through the single factor experiment and neural network optimization.The optimum process conditions of enzyme production was the inoculation amount 11.0%,the initial pH 5.6,fermentation time 130 hours and broth content 70.0 mL.The enzyme activity of Aspergillus niger fermentation ?-glucosidase reached 118.73 U/mL,compared with the previous optimization,the enzyme activity increased by 48.61%.The crude enzyme was precipitated with ammonium sulfate,purified with DEAE-52 anion exchange chromatography and sephacryl S-300 High resolution.The SDS-PAGE test its purity reached the level of electrophoresis,and the enzyme was finally purified to 1.78-fold with a specificity activity from 481.15 U/mg to 854.57 U/mg protein and a total yield of 27.94%.The bioinformatics analysis of the purified ?-glucosidase by LC/MS/MS,and got part of the amino acid sequence of peptides,and the homology of ?-glucosidase was from Aspergillus niger,the molecular weight and isoelectric point was 93626 and 4.6.The purification ?-glucosidase had been put in Lianjiang red-orange juice.By sensory evaluation,the results showed that the red-orange juice actioned by ?-glucosidase was significantly better than that of non enzyme on the sensory,the sweetness and aroma increased,the bitter decreased,the taste and the flavor was improved.It was found that the reducing sugar content in red-orange juice increased by 18.98% after ?-glucosidase,and ?-glucosidase could change the species and content of volatile components in red-orange juice determined by GC-MS.39 kinds of volatile substances were detected in red-orange juice before enzymatic hydrolysis and 48 kinds of volatile substances were detected in red-orange juice after enzymatic hydrolysis.The contents of alcohols,acids and aldehydes increased in both the quantity and the total content in the red-orange juice actioned by ?-glucosidase,the esters was no change in the quantity,but the total content increased.On the basis of the unique flavor of Lianjiang red-orange,?-glucosidase could increased the flavor components such as 3-carene,benzaldehyde,cyclohexanol,et al,thus increasing the aroma of fruit juice.In summary,?-glucosidase increased the type and content of flavors,improved the flavor of red-orange juice,and played an important role in release of aroma components.