| Aroma glycosides are the main precursors of aromatic components in grape berries. During fermentation progress, glycosidic aroma precursors are hydrolyzed to release volatile flavor components. As the current methods for grape aroma glycosides have their defectives, glycosidic aroma precursors have not been precisely analyzed.This study aimed to optimize a quick and accurate analysis method of grape glycosidic aroma precursors. After the optimization of key steps of purification, acid hydrolysis and spectral analysis of grape aroma glycosides being performed, the evolution of glycosidic aroma precursors during the winemaking process of enzyme treatments was explored. The main results were as follows:(1) A best column material for grape aroma glycosides and its corresponding solvent elution condition were selected. Through static adsorption- desorption and dynamic adsorption- desorption experiments, XAD-2 was chosen from four column packing materials. After an orthogonal experiment for the elution of aroma glycosides, the best elution condition of the column was built: flow rate 1 ml/min, pentane : dichloromethane = 8:1, ethyl acetate : methanol = 9:1. The recovery of glycosidic aroma precursors of the selected elution condition was above 95 %.(2) After single-factor experiments, orthogonal experiments and response surface method, the optimization design on the determination of aroma glycosides was as below: adding pvpp 20 mg/mL and drying by rotary evaporation or under nitrogen, the extract of aroma glycosides was solubled in a acid solution of 10% ~ 20% of its original volume. The extract was hydrolyzed under 90 ℃ in 1 hour, then neutralized by adding 2 mol/L NaOH in a volume as reaction liquid : alkali = 3:4. HK reaction proportions were 300μL substance, 225μL ATP + NADP+ and 79μL HK, reacting in 30 ℃for 15 min. Repeated experiments showed the detection level of aroma glycosides by optimization design was 3.28 times than the old. Glycosides standard tests showed the R2 were all over 0.97, which proved the optimization design was a high accuracy and well repeatability method.(3) Muscat Blanc, Ecology, Muscat, and Cabernet Sauvignon, those four grape varieties with different colour and aromatic compounds content were selected. The evolution of glycosidic aroma precursors was explored by the optimization design. Results released that the evolution of aromatic glycosides between grape varieties or under different enzyme treatments were clear by the optimized method. As grape varieties or system conditions had a little effects on the measured value, optimization design was suitable to application.(4) The amount of aromatic glycosides was declined in fermentation and risen up in aging. All enzyme treatments during fermentation process made glycosidic aroma precursors reduced, but glycosidase treatments had a greater impact on aroma glycosides than pectinase. Between two selected glycosidase, AR2000 was more sensitive to the frementation environment, glycosidase made from a favor Hanseniaspora- uvarum was insensitive.(5) Enzyme treatments affected the aroma characteristics of all grape varieties. PCA of aroma categories indicated that enzyme treatment enhanced both concentration and complexity of wine aroma. Aroma characteristics showed significant diversities among different enzyme treatments. As pectinase treatment mainly enhanced the aroma of plants, such as asparagus and green beans, glycosidase treatments gave strong and pleasing floral and fruity aroma characteristics to wine. Aroma characteristics of glycosidase purificated from Hanseniaspora –uvarum treatment were quite different from AR2000. Wines of AR2000 treatment exhibited characteristics like pineapple, strawberry, violet, sweat, metal and soil; while the glycosidase from non-Saccharomyces treatment wines tended to enhance sweet fruity and floral characteristics like apple, mango, small fruits and sophora flower. Compared with AR2000, the non-Saccharomyces glycosidase was more practical in flavour enhancement winemaking techology. |
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