Does the addition of glutathione affect dry white wines?

White wines are clear and transparent, with a refreshing odor, strong aroma and rich nutrition. Wine contains antioxidant components and rich phenolic compounds, which can prevent atherosclerosis and platelet aggregation, protect and maintain the normal physiological functions of the cardiovascular system, protect the heart and prevent strokes. Oxidative spoilage of most white wines has become a well-known problem in the wine industry. White wines are sensitive to oxygen exposure, which leads to the loss of their characteristic aromas, the development of atypical aging characteristics and changes in color[1] . Phenolics, especially o-phenol, are responsible for oxidative browning of wines[2] . Browning may be the result of enzymatic oxidation, which mostly occurs during winemaking, whereas non-enzymatic oxidation occurs mainly during wine aging[3] .

 

SO2 acts as an antioxidant in wine, mainly by effectively cleaving hydrogen peroxide and o-quinone compounds, and it can also form adducts with carbonyl compounds, especially acetaldehyde[4] . Sulphites are considered to be effective agents in controlling oxidative reactions in wine, but they are toxic and allergenic[5] . As a result, wines with lower sulfite levels have been accepted by consumers seeking a healthier diet, and some wine producers have attempted to reduce the use of SO2 in winemaking. Although complete elimination of SO2 is not feasible, substitutes for SO2 can be found to reduce its use [6].

 

Glutathione (GSH) is a tripeptide composed of glutamic acid, cysteine and glycine, which occurs naturally in many plants, animals and microorganisms. The sulfhydryl group of cysteine is the site where its biochemical properties prevent oxidation.7-9 The main functions of GSH can be summarized as antioxidant, immunity booster, and detoxifier.10 GSH has been shown to play an important role in bioreduction in living tissues. In living tissues, GSH plays a key role in bioreduction, anti-oxidative stress, detoxification of xenobiotics and endogenous toxic metabolites, enzyme activity, and sulfur and nitrogen metabolism[11] . Therefore, it is considered to be a powerful, versatile and important self-defense molecule.The addition of GSH allows for the use of lower doses of SO2 in wines and, in contrast to SO2, GSH traps o-quinone during oxidation, limiting the amount of browning pigments[12-13] . In addition, the protective effect of GSH on white wines has been shown to prevent the loss of certain phenols, esters and terpenoids. When GSH is present, quinones formed during oxidation during wine storage and aging react with it, and in the absence of sufficient amounts of GSH, quinones react with other phenolic compounds, and this reaction can lead to the formation of new polymers and larger structural tannins, with consequent alterations in wine organoleptic properties, as these phenols, esters and terpenoids are important contributors to the pleasant bouquet and fruity flavors in white wines [14]. These phenols, esters and terpenoids are important contributors to the pleasant floral and fruity flavors in white wines [14-15]. In this experiment, GSH was added to dry white wines and placed at 50 ℃ for accelerated oxidation, and the physical and chemical indexes, color and monomer phenol content of the wines were measured weekly, compared with those of the wines without GSH, in order to investigate the effect of GSH on dry white wines. This is to investigate the effect of GSH on dry white wines and to provide a theoretical basis for the storage of white wines.

 

1 Materials and Methods

1.1 Materials, reagents and instruments

Dry White Wine: 2017 Longan Dry White Wine, provided by Hebei Huailai Noble Estate. The alcohol content of the wine was 12 %vol, total acid was 7.11 g/L, pH value was 3.63, and reducing sugar was 0.18 g/L. The wine was made with a total alcohol content of 12 %vol, total acid was 7.11 g/L, pH value was 3.63, and reducing sugar was 0.18 g/L.

Reagents and consumables: NaOH, glucose, Na2CO3, foraminol, glacial acetic acid, etc., all analytically pure, Baoding Wanke Reagent Company; acetonitrile, methanol, all chromatographically pure, Shanghai Komeo Company; gallic acid, protocatechuic acid, catechuic acid, vanillic acid, butyric acid, coumalic acid, butyraldehyde, ferulic acid, guaiacol, benzoic acid, salicylic acid, quercetin, U.S.A. Sigma Company.

Instruments: digital refractometer, ATAGO, Japan; colorimeter (CR-400), Konica Minolta; high-performance liquid chromatograph (2489 UV detector, autosampler, CLASS-VP workstation), Waters, USA; ultrasonic degasser, Ningbo Xinzhi Bio-Tech Co. Shanghai Yarong Biochemical Instrument Factory.

 

1.2 Experimental Methods

1.2.1 Handling of wine samples

Accurately weigh 30 mg of GSH standard into 1.5 L of white wine samples, divided into 3 bottles of 500 mL each, and take 3 bottles of 500 mL of wine samples without GSH as control. The 6 bottles of wine samples were stored in a thermostat at 50 ℃, and the samples were taken once every 7 days for the determination of various indexes.

 

1.2.2 Determination of basic physical and chemical indicators

The pH value was determined by pH meter; soluble solids were determined by refractometer; total acid was determined by NaOH titration; reducing sugar was determined by DNS method.

 

1.2.3 Determination of color difference

The L*, a* and b* values of the wine samples were determined using a colorimeter. The L* value represents the brightness (L*=0 means black, L*=100 means white); a* value is the red-green parameter (+ is red, - is green); and b* value is the yellow-blue parameter (+ is yellow, - is blue)[16-17] . The experiment was repeated three times and the samples were measured under natural light.

 

1.2.4 Determination of phenolics

1.2.4.1 Determination of total phenols

The forintol colorimetric method was used.

 

1.2.4.2 Determination of monomeric phenols

10 mL of wine sample was extracted with 10 mL of ethyl acetate for three times, and then the organic phases were combined, concentrated to dryness by rotary evaporation, and the residue was dissolved in 3 mL of methanol for chromatography, and stored at -20 ℃, protected from light, and used for liquid chromatographic analysis. The liquid chromatographic analysis was based on the method of Hou Lijuan[18-19] .

 

1.3 Statistical analysis

All the experimental data were averaged from the results of three replications and the results were expressed as X ± SD. Origin 8.6 or SPSS 17.0 software was used for data statistics and ANOVA, and ANOVA and Duncan's multiple analysis of variance (P < 0.05) were used.

 

2 Results and analysis

2.1 Changes in basic physical and chemical indicators

The changes in the physicochemical parameters of the wines with increasing storage time are shown in Table 1. The soluble solids content decreased slightly with time, but there was no significant difference between the samples without and with GSH. Reducing sugars decreased with time in both non-GSH-added and GSH-added wines, indicating that the effect of GSH on reducing sugars was very small. The pH values of the wines with and without GSH did not change significantly with storage time. The total acid content increased slightly with time, but decreased significantly after the addition of GSH, probably due to the reaction between GSH and organic acids in the wine samples, which consumed part of the acids or inhibited the oxidation of alcohols and aldehydes into acids. The relationship between pH value and total acid in wine samples is as follows: the acid substances in wine are mainly composed of organic acids, i.e. tartaric, malic and citric acids from grapes, and succinic, lactic and acetic acids from processes, most of which are present in the free state, and a few are present in the form of salts. The total acidity of wine refers to the total amount of free acid in wine, i.e. the titratable acid, which does not directly indicate the acidity of a certain acid in wine, but only the state in which it exists. The pH value is the negative logarithm of the concentration of hydrogen ions and indicates the actual concentration of hydrogen ions. The organic acids in wine are all weak organic acids, and their ability to dissociate hydrogen ions varies, so the pH value of wine depends on the nature of the organic acids, their relative content, and the condition of the wine [20].

 

2.2 Changes in Color Difference

The color of dry white wines is an important organoleptic indicator that affects consumer acceptance. Many factors affect the color of dry white wines. When dry white wines are exposed to oxygen (air) in warm conditions, they are oxidized, resulting in a darker color. Once a wine has been oxidized, the damage is irreparable, as redox reactions are irreversible. The color of the wine samples gradually changed from light yellow to yellowish-brown with the increase of storage time, as shown by the decrease of the color difference L* and the increase of a* and b*, as shown in Figs. 1 to 3. All the values of the samples with GSH were smaller than those of the samples without GSH, and the final color was lighter. It can be seen that GSH can reduce the rate of color change of wine samples and have a certain protective effect on the color of dry white wines.

 

2.3 Changes in phenolics (Figure 3)

2.3.1 Changes in total phenols

Phenolics in wine are compounds that contain phenolic groups in their molecular structure. Phenolics are important components of wine, which not only contribute to the organoleptic properties of wine, such as color, flavor, and astringency, but also have important antioxidant properties, including scavenging of free radicals and chelating with metals[21-22] . The changes in total phenols are shown in Fig. 4. The total phenol content of the wine samples decreased with time, and even more so in the wine samples without GSH. Phenols are easily oxidized, and the addition of GSH protects them from oxidation.

 

2.3.2 Changes in monomeric phenols

There is a wide variety of monomer phenols in wine, including flavanols, flavonols, hydroxycinnamic acids, etc. They have various biological activities and are one of the most important natural products. In this study, only several different types of monomer phenols with high content in wine were analyzed.

 

2.3.2.1 Flavanols

Flavanols are the most abundant phenols in wine, imparting bitterness and structure to the wine, and are introduced into the wine during vinification through maceration of grape seeds and pomace[23] . Catechins are typical flavanol phenols[24] . The variation of catechins in wine samples is shown in Figure 5. The catechin content decreased with time, and the rate of decrease was faster in the samples without GSH than in the samples with GSH.

 

2.3.2.2 Flavonols

The anthocyanins in wine combine with flavonols to give blue-purple and orange-yellow hues to wine. Quercetin is one of the flavonols, and it has been studied that quercetin has a better complementary color effect on wine than other flavonols[25] . The variation of quercetin in wine samples is shown in Figure 6. The quercetin content decreased with time in both samples with and without the addition of GSH. The decrease in quercetin content may also be a reason for the darkening of the wine samples.

 

2.3.2.3 Hydroxycinnamic acids

Between 20% and 25% of the phenolic acids in grape berries exist in the free form, with hydroxycinnamic acid derivatives being the most abundant, and they play an important role in the oxidative discoloration of wine. Some basic anthocyanosides are acetylated, caffeoylated and coumaroylated to form more anthocyanosides[26] , and through a series of reactions, more complex structures of pyranosides and polymeric pigments are formed, which lead to the change of wine color[27] . The changes of coumaric acid in wine samples are shown in Figure 7. The amount of coumaric acid slowly increased and then decreased with time. In wine samples without GSH, the coumaric acid content decreased first, and the magnitude of the decrease was larger. The increase in coumaric acid may be the result of the decomposition of complex macromolecules, which are then oxidized, and the level of coumaric acid then decreases. GSH is oxidized first, so the coumaric acid content is higher in samples to which GSH has been added.

 

3 Conclusion

In this study, the effects of GSH on dry white wines were investigated by adding GSH to dry white wines. Under the condition of high temperature accelerating the oxidation of dry white wine, the content of soluble solids and reducing sugars decreased, the pH value did not change significantly, and the total acid content increased. The addition of GSH to wine had little effect on the basic physicochemical indexes of the wine, but had a significant effect on the total acid content; GSH could protect the color of dry white wine and slow down the tendency of darkening of the color of the dry white wine; it had a large effect on the total phenol content of the wine. GSH has a strong effect on the total phenolic content of the wine, and although the chemical reaction with flavanols, flavonols and hydroxycinnamic acid phenols varies, it is effective in slowing down the reduction of phenolics and prolonging the storage time of dry white wines.

 

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