Characterisation of the main enzymatic activities present in six commercial macerating enzymes and their effects on extracting colour during winemaking of Monastrell grapes

Numerous studies have reported on the use of enzymes in wineries to facilitate the extraction of grape phenolics and to aid colour stability, although no clear conclusions were reached. Information concerning the enzymatic activities of industrial preparations is very difficult to obtain despite its importance. The aim of this work was to characterise and quantify the activities of six macerating enzyme preparations to optimise their use, and to study their effect on the chromatic characteristics of a Monastrell wine. The results indicate that differences exist in the type of enzymatic activities present in the preparations and in their concentration. The chromatic characteristics of the resulting wines, both at the end of alcoholic fermentation and after 12 months of storage in stainless steel tanks, showed differences with the use of the different maceration enzymes when comparing with control wine, although very little differences were observed between the different enzyme treatments. The enzyme-treated wines showed, at the end of alcoholic fermentation, a higher OD₂₈₀ and tannin content, and after 12 months, a higher colour intensity too. The main effect of all the preparations was an increase in the total phenol content, probably by facilitating the release of skin tannins.     

不同皮渣浸渍发酵时间对葡萄酒中缩合单宁构成及感官品质的影响

为明确不同皮渣浸渍时间对葡萄酒中缩合单宁的影响,以赤霞珠和西拉为材料,进行不同浸渍发酵时间处理,对葡萄皮和葡萄籽以及葡萄酒中缩合单宁的含量、组成、缩合单宁以及相关口感指标等进行了分析。结果表明,不同葡萄品种中缩合单宁的初始含量、聚合度及没食子酰化率都有显著差异（P<0.05）。葡萄皮中缩合单宁的平均聚合度显著高于葡萄籽（P<0.05）,而葡萄皮缩合单宁的没食子酰化率则显著低于葡萄籽缩合单宁（P<0.05）。葡萄皮和葡萄籽中缩合单宁组成的最大差异是葡萄籽缩合单宁中不含有（?）-表棓儿茶素亚单元,而且末端单元主要是由（+）-儿茶素、（?）-表儿茶素和（?）-表儿茶素没食子酸酯组成。皮渣浸渍时间对葡萄酒中缩合单宁的组成及含量都有明显影响。随着浸渍时间的延长,葡萄酒中缩合单宁的总含量呈上升趋势,而平均聚合度则呈下降趋势。不同品种的葡萄酒之间缩合单宁含量也存在显著差异（P<0.05）,这与不同葡萄品种中酚类物质的可浸提率有关。葡萄酒中葡萄皮缩合单宁的占比一直显著高于籽粒缩合单宁的占比（P<0.05）,说明葡萄皮中缩合单宁比葡萄籽缩合单宁更容易浸提到葡萄酒中。相对于苦味和酸味,不同浸渍时间处理对葡萄酒的涩味影响更为显著（P<0.05）,综合考虑酒体的丰满度和口感的平衡,皮渣浸渍时间选择14 d左右更好一些。     

Effect of macerating enzyme treatment on the polyphenol and polysaccharide composition of red wines

Effect of macerating enzymes on the polyphenol and polysaccharide composition of Merlot wines after 20 months of ageing was studied over three vintages (2004, 2005, 2006). Pectinase rich enzyme preparations, by degrading grape berry cell walls, led to a modification of the molecular weight distribution of polysaccharides released into the wines. Enzyme-treated wines contained more Rhamnogalacturonan II (RG-II) and less polysaccharides rich in arabinose and galactose (PRAGs) over the three vintages. The enzyme treatment also modified wine polyphenol composition. An increase of colour intensity, of derived pigments resistant to sulphite bleaching and of proanthocyanidins (condensed tannins) was observed, modulated by vintage effect. Principal Component Analysis of all the data indicated both vintage and enzyme effects. Beside the vintage effect separating the wine samples, the impact of enzyme treatment on wines was established for each year.     

不同浸渍工艺对夏黑葡萄醪理化指标及颜色的影响

以热浸渍法、冷浸渍法、常温浸渍法为基础,通过部分添加不同浓度的二氧化硫和果胶酶,探究不同浸渍工艺对葡萄醪中单宁、总酚含量、pH值、色度、色调等指标的影响。结果表明,冷浸渍法（浸渍温度4 ℃、二氧化硫添加量50 mg/L、浸渍时间13 h）浸提时,葡萄醪中单宁含量6.884 mg/L,总酚含量122.332 mg/L,pH值3.85,色调最低,色度最高;常温浸渍法（浸渍温度20 ℃、二氧化硫添加量50 mg/L、浸渍时间13 h）浸提时,葡萄醪中单宁含量7.612 mg/L,总酚含量134.756 mg/L,pH值3.85,色调最低,色度最高;热浸渍法（浸提温度80 ℃,浸渍时间100 min）浸提时,葡萄醪中单宁含量23.451 mg/L,总酚含量264.688 mg/L,pH值3.43,色调最低,色度最高。     

Impact of maceration enzymes on skin softening and relationship with anthocyanin extraction in wine grapes with different anthocyanin profiles

The impact of an enzyme preparation on the extraction of grape skin anthocyanins into a wine-like solution was evaluated during maceration (25 °C, 8 days). The study was performed on Vitis vinifera L. cv. Cabernet Sauvignon and Nebbiolo because of their different anthocyanin profiles, which are mainly composed of tri- and disubstituted forms, respectively. Maceration enzymes enhanced the skin releasing capacity for anthocyanins between 8% and 15% more, depending on the enzyme dose and variety, and shortened the time required to reach the maximum extraction yield by about 40 h, when compared to the control samples. The effect of enzymes on the profile and total content of anthocyanins during maceration was significant only for Cabernet Sauvignon. Therefore, a variety effect was found. In particular, the relative amount of malvidin glucosides increased up to 9% more whereas that of delphinidin, cyanidin and peonidin glucosides decreased up to 5%, 2% and 3% more, respectively, with the addition of the enzyme preparation. The results also showed lower anthocyanin degradation at maceration times longer than 72 h with the use of enzymes. Furthermore, the relationship between this enzyme effect and the instrumental skin hardness was assessed for the first time, as no previous study deals with the variation in the mechanical properties of the berry skin after using maceration enzymes. Significant positive relationships were found between the skin softening and the anthocyanin extraction yield (R > 0.69, p < 0.01), which confirmed that the skin degradation by enzymes facilitates the anthocyanin release.     

葡萄果梗在红葡萄酒酿造中的应用

本文从以下几个方面讨论了葡萄果梗在葡萄酒酿造中的作用：果梗中影响葡萄酒质量的主要物质组成；葡萄浆果各部分酚类化合物的比较；果梗对葡萄酒的影响。葡萄酒酿造过程中，葡萄梗特别是干葡萄梗的添加会增加葡萄酒中酚类物质的含量，并且还会影响到葡萄酒的风味添加经蒸汽处理的葡萄梗，可增加可觉察性黄烷酮类物质的含量。     

Influence of ripening stage and maceration length on the contribution of grape skins,seeds and stems to phenolic composition and astringency in wine-simulated macerations

This paper studies the influence of ripening stage and maceration time (1–23 days) on the contribution of each cluster grape components to phenolic composition and astringency in wine-simulated macerations. In general terms, proanthocyanidin extraction from skins, stems and especially from seeds increased with maceration time. The ripening stage also had a major influence on tannin extraction and astringency. The tannin contribution from skins and stems increased with ripening but the contribution from the seeds decreased. The contribution to astringency from all cluster components was clearly higher when the grapes were unripened. The mean degree of polymerization of the tannins from skin and seed components was unaffected by ripening and maceration length.     

Effect of the prefermentative addition of copigments on the polyphenolic composition of Tempranillo wines after malolactic fermentation

The influence of the prefermentative addition of copigments and different winemaking technologies on the polyphenolic composition of Tempranillo red wines after malolactic fermentation was studied. Six experiments dealing with the prefermentative addition of caffeic acid, rutin, (+) catechin, white grape skin tannin, white grape seed tannin and control wines were realised. Three different winemaking technologies (traditional vinification, prefermentative cold maceration at 6–8 °C and cold soak at 0–2 °C with dry ice) were studied. Prefermentative addition of copigments increases anthocyanin copigmentation reactions and produces wines with a greater colour, a higher anthocyanin concentration, a superior contribution of anthocyanins to the colour of the wine, a superior percentage of tannins polymerised with polysaccharides and less astringency. Cold prefermentative maceration increases the extraction of polyphenols, the anthocyanin copigmentation reactions and the polymerisation reactions between tannins and polysaccharides. The effectiveness of the combination of copigments and prefermentative maceration treatments was demonstrated by the increase of the concentration of the polyphenolic compounds.     

EFFECTS OF DIFFERENT MACERATION TIMES AND PECTOLYTIC ENZYME ADDITION ON THE ANTHOCYANIN COMPOSITION OF VITIS VINIFERA CV. KALECIK KARASI WINES

Kalecik karasi is an important red grape cultivar for winemaking in Turkey. The effect of three different maceration times (3, 6 and 12 days) and addition of pectolytic enzyme (2 and 4 g/hL) on the anthocyanin and chemical composition of Kalecik karasi wines were studied. High performance liquid chromatography-mass spectrometry coupled with diode array detection was used for analysis. Fourteen anthocyanin compounds were detected in wines. Major anthocyanins in all wines are malvidin-3-glucoside and its acylated esters. The results showed that increasing maceration time, especially with addition of enzymes, gives significant increases in anthocyanin contents. Moreover, the wines treated with enzymes had higher values in total phenolics, tannins, and color intensity than the control wines.

PRACTICAL APPLICATIONS

Anthocyanins are the most important polyphenols in red grapes and red wines with potential health benefits. Therefore, the first analysis of the anthocyanins contents of wine obtained from important turkish cv. Kalecik karasi using liquid-chromatography-mass spectrometry and the influence of different maceration times and addition of pectolytic enzyme on these important phenolic compounds are of interest for scientific literature, the wine industry as well as for the wine consumer.     

Partial purification of a polygalacturonase from a new Aspergillus sojae mutant and its application in grape mash maceration

The use of polygalacturonase (PG) preparations in winemaking promotes the release of phenolic compounds. A PG from a new source, Aspergillus sojae mutant, was semi‐purified and tested for grape mash maceration. Crude extract (CE), a commercial pectinase, and two high PG activity semi‐purified preparations, F_I and F_II, were applied for maceration at PG activity of 3.5 U g^?1 of grape for 46 h. Enzyme‐assisted maceration significantly (P < 0.05) increased the total phenolic content from 255.8 to 916.3 ± 5.2, 5732.9 ± 9.9, 563.4 ± 6.7 and 620.6 ± 18.4 mg L^?1 for CE, commercial pectinase, F_I and F_II, respectively. The content of individual phenolics such as gallic, protocatechuic, chlorogenic and p‐coumaric acids was improved. Principal component and hierarchical clustering analyses suggested that CE has a better performance upon the release of phenols. Semi‐purified preparations acted similar to commercial pectinase. These findings open an opportunity for the potential use of PG from the mutant strain as an alternative macerating enzyme.     

Effects of cold maceration on red wine quality from Tuscan Sangiovese grape

Pre-fermentation cold maceration of must by the direct addition of two cryogens was studied. The experiments were conducted on Sangiovese grape from a typical Chianti vineyard in the year 2001 with two cryogens (solid state carbon dioxide and liquid nitrogen) and at various temperatures (from –5 to +5 °C) in 500-L batches for 48 h. The wines obtained were characterized by chemical analyses and sensory evaluation, and higher quality resulted as seen from the analytical and sensory characteristics with higher extraction of polyphenolic compounds at lower cold maceration temperature. Liquid nitrogen was more effective than solid carbon dioxide at the same temperature. Proportional increase of wine quality with lower cold maceration temperature was observed for solid carbon dioxide. Further investigation is needed to establish the management protocol, a proper engineering of this technique, and to evaluate its economical advantages.     

北冰红山葡萄酒CO2浸渍发酵工艺研究

以北冰红山葡萄为原料酿制山葡萄酒,通过试验确定CO2浸渍时间为10 d,浸渍温度为31 ℃时浸渍效果最佳。在单因素试验的基础上,利用响应面法对北冰红山葡萄酒的发酵工艺条件进行优化。结果表明,最优发酵工艺条件为酵母添加量160 mg/L,发酵温度21 ℃,发酵时间23 d,在此条件下制得北冰红山葡萄酒的平均酒精度为10.40%vol。     

Effect of Maceration Time on Free and Bound Volatiles of Red Wines from cv. Karaoğlan (Vitis vinifera L.) Grapes Grown in Arapgir,Turkey

Skin maceration is an essential winemaking technique that affects sensory properties and aroma of red wines. The aim of this study was to investigate aroma profiles of red wines made from cv. Karao?lan, a grape variety grown in Arapgir (Malatya, Turkey) and the effect of maceration time (5, 10, and 15 d) on free and bound aroma compounds and sensory properties. Three maceration times were applied in durations of 5, 10, and 15 d. Free and glycosidically bound aroma compounds were analyzed using gas chromatography‐mass spectrometry (GC‐MS) in Karao?lan wines, and the effect of different maceration times on aroma composition were evaluated. Results indicate that extended maceration time significantly influenced the aroma composition of wines both qualitatively and quantitatively. A total of 59 free and 22 bound aroma compounds were identified in the final wines. The maximum of free aroma content was achieved with 5 d of maceration, while 15 d of maceration gave the highest amount of glycosidically bound aroma compounds. From sensorial point of view, the wine produced with 10 d of maceration had the best panel score compared to wines with 5 and 15 d of maceration. This is the first study referring to the composition of aroma compounds of red wines made from cv. Karao?lan. It was concluded that this cultivar resulted in a good quality wine in terms of bound and free aroma compounds. This information of vinification practices that influence the chemical properties of wine is useful for grape growers and winemakers.     

Effects of pre-fermentation enzyme maceration on extraction and colour stability in Pinot Noir wine

Effects of pre-fermentation enzyme maceration on anthocyanins, phenolics, colour and colour parameters were investigated in Pinot Noir ( Vitis vinifera L.) wines made from musts that were macerated for 12 hours at 20°C with a commercial enzyme preparation, Rohapect VR-C (Rohm, Darmstadt, Germany), with and without additions of SO₂. Anthocyanins and phenolics were determined by spectrophotometric and HPLC methods. Standard triplicate microvinifications were conducted in glass vessels. Extraction of anthocyanins was not increased by enzyme treatment. However, wines produced by enzyme maceration had increased visible colour intensity, colour density and polymeric pigment formation. By the end of fermentation, enzyme-treated wines had more polymeric pigment, and less monomeric anthocyanins than non-enzyme-treated wines. The wines were bottled, stored at 12°C and sampled at three, six, eight and 18 months after bottling. Over 18 months, differences between the treatments were maintained. The promotion of the formation of polymeric pigments in enzyme-treated wines continued after the wines were bottled, and the greater content of polymeric pigment led to greater colour density in the enzyme-treated wines. The promotion of polymeric pigment formation in wines by macerating enzyme treatment of the must may be due to the breakdown of protective polysaccharide-protein colloids by hemicellulase and protease activities in the enzyme preparation.     

Effects of pulsed electric fields on the extraction of phenolic compounds during the fermentation of must of Tempranillo grapes

The effects of a pre-treatment of the grape skin with electric field treatments (PEF) at 5 and 10 kV/cm on the evolution of colour, anthocyanin and index of total polyphenols during the Tempranillo grape vinification and on the wine characteristics after fermentation have been investigated.Results showed that the permeabilization of the grape skin by application of a PEF treatment at room temperature caused an increment of the colour intensity, anthocyanin content and of total polyphenolic index with respect to the control during all the vinification process. Generally along the maceration process the values of these three parameters increased when the electric field strength raised from 5 to 10 kV/cm.Colour intensity and anthocyanin content of the wine significantly increased when the PEF treatment was applied at 10 kV/cm. However, the total phenolic index increased considerably with the application of a PEF treatment at 5 kV/cm. A further increment on the electric field strength did not appreciably augment this attribute. PEF did not affect the ratio between the components of the red wine colour (tint and yellow, red and blue components) and other wine characteristics such as alcohol content, total acidity, pH, reducing sugar concentration and volatile acidity.These results indicate that the PEF treatment previous to the maceration step in the winemaking process of the red wine can contribute to reduce the duration of the maceration during vinification or to increase the quantity of these attributes in the final wine.

Industrial relevance

This study investigates the effect of the application of a PEF pre-treatment of the grape skin on the evolution of colour, anthocyanin and index of total polyphenols during the Tempranillo grape vinification. Results obtained indicate that the application of a PEF treatment preceding the maceration step in the winemaking process of red wine could contribute to reduce the duration of the maceration during vinification or to increase the quantity of these parameters in the final wine. Latter effect could be of interest in grapes poor in polyphenols.     

Changes in skin cell wall composition during the maturation of four premium wine grape varieties

BACKGROUND: Grape skin cell walls form a barrier against the diffusion of colour. The colour of red wines is mainly due to anthocyanins, although their concentration in wine is not always correlated with the anthocyanin content of grape skins. The cell wall composition changes during fruit ripening, so it was thought that a study of the ripening behaviour of four premium varieties might provide information on how the composition changes during ripening and explain the technological differences. RESULTS: The largest quantities of skin cell wall material (CWM) were isolated from Monastrell grapes, probably owing to the greater number of cell layers they contain and the thicker cell walls. During ripening, a loss of skin CWM (e.g. 120–60 and 65–45 mg CWM g^?1 fresh skin for Monastrell J and Cabernet Sauvignon respectively) and galactose (e.g. 36–20 and 55–30 mg g^?1 CWM for Cabernet Sauvignon and Monastrell J respectively) was observed, together with a decrease in the degree of pectin methylation and acetylation, except in Syrah, in which pectin methylation remained unchanged. These changes were accompanied by the accumulation of glucose, while other neutral sugars showed no significant variations. A correlation was found between the degree of ripening and those constituents changing in the grape skin cell wall. CONCLUSION: The cell wall composition differs sufficiently between grape varieties to allow discrimination between them. Ripening was associated with a decrease in the quantity of CWM and galactose in the cell walls. Monastrell showed the largest quantities of both throughout ripening. The differences observed between Monastrell and the other varieties would explain the technological differences between them. Copyright © 2007 Society of Chemical Industry     

Impact of enzyme on quality of blackcurrant and plum juices

The aim of work was optimization of technology of cloudy blackcurrant and plum juices production. The major concern was the increase of product cloudiness and its stability. Red fruit were processed with commercial pectinolytic preparations (Pectinex BE Colour, Pectinex BE XXL, and mixture of Rohapect PTE with Rohament PL). The effect of enzyme dose, maceration time and addition of ascorbic acid (in case of plum) on juices quality was investigated. Using single enzyme, either polygalacturonase or pectin lyase did not allow obtaining juices with high enough turbidity, however appropriate enzyme mixture allowed to obtain cloudy juice with the turbidity of at least 230 NTU for blackcurrant and above 500 NTU for plum juice. The best pressing-yield for blackcurrant was achieved with polygalacturonase and pectin lyase, 65 g/100 g after 1 h and 74 g/100 g after 4 h of pectinolysis. The macerating mixture gave about 58÷59 g/100 g yield, irrespectively of enzymation time. Pressing-yield of plum juices was in the range of 94÷97 g/100 g due to the fact that practically only skins were retained on pressing cloth. Addition of ascorbic acid (AA) during plum juice processing in quantity of 500 mg/kg had protective effect on anthocyanins. In control plum juices range of anthocyanins contents were 12.1÷16.5 mg/100 ml while in juice with AA addition even 21.4÷24.5 mg/100 ml.     

Colour and phenolic compounds in sweet red wines from Merlot and Tempranillo grapes chamber-dried under controlled conditions

Chamber drying under controlled temperature and humidity conditions of the red grape varieties Merlot and Tempranillo grown in Andalusia (Spain), and the fortification and maceration of the musts in the presence of skin from both types of grapes, to obtain sweet red wines, has been studied. Changes in colour and in monomeric and polymeric phenols during the vinification process were examined. Chamber drying increased the sugar content to about 31.4 °Brix within 48 h in Merlot grapes and 72 h in Tempranillo grapes. This drying process also causes skin rupture, facilitating the access of phenolic compounds to the pulp. The resulting musts exhibited slight browning and increased red hues, due to a high concentration of anthocyanins; maceration in the presence of grape skins for 24 h provided the best results. The end-product contained highly acceptable tannin, colour and phenolic compounds for marketing as a high quality sweet red wine, in comparison with other commercial sweet red wines.     

The maceration process during winemaking extraction of anthocyanins from grape skins into wine

The rate of anthocyanin extraction from skins of Monastrell grapes into wine during the maceration process has been studied by following anthocyanin accumulation in wine and its decrease in crushed skins. The most important changes occurred during the first 7 days of maceration; the following 7 days saw very little variation in the levels of the anthocyanins found in the wine or the crushed skins. The anthocyanin profile of the wines was quite different from that of the grape skins, whereas the crushed grape skins had a profile very similar to the profile of the grape skins at the time of harvest. Any differences in the proportions of the different anthocyanins found in Monastrell grapes and wines cannot therefore be attributed to different rates of extraction for each of the anthocyanins during winemaking. Other explanations, such as degradation or yeast cell wall adsorption, must be sought.     

Influence of low temperature prefermentative techniques on chromatic and phenolic characteristics of Syrah and Cabernet Sauvignon wines

Low temperature prefermentative techniques (cold maceration, superficially frozen grapes and dry-ice frozen must) were studied as an enological alternative for obtaining highly colored wines and compared with other more common practices, such as the use of maceration enzymes during the maceration step. The experiments were conducted with two grape varieties (Syrah and Cabernet Sauvignon) during the 2007 season. Significant differences were found between the treatments at the three moments analyzed (at the end of alcoholic and malolactic fermentation, and at the moment of bottling). The cold maceration technique produced the highest concentration of anthocyanins in Cabernet Sauvignon wines. As regards chromatic parameters, the prefermentative low temperature techniques led to wines with a better color than the control wine although similar to the color obtained when a commercial maceration enzyme was used. Taking into account the results in both Cabernet Sauvignon and Syrah wines, freezing the must with dry-ice and the use of maceration enzyme were the most interesting techniques.