Evaluating the Influence of Malolactic Fermentation Inoculation Practices and Ageing on Lees on Biogenic Amine Production in Wine

Biogenic amines are compounds, produced primarily by lactic acid bacteria (LAB) that negatively affect the wholesomeness of wine. Standard winemaking practices can greatly influence the levels of biogenic amines in wine. The aim of this study was to determine the relative contribution of different malolactic fermentation (MLF) practices and ageing of wines on fermentation lees to the final levels of biogenic amines. Wines were made on small scale over two harvest seasons with two red grape cultivars. Treatments included spontaneous MLF, co-inoculated MLF, MLF inoculated after alcoholic fermentation (conventional inoculation) and 4 months of ageing in the presence and absence of fermentation lees of all MLF treatments. Biogenic amine concentrations were measured by high-performance liquid chromatography at key winemaking stages and statistically analysed for the effects of MLF treatment and winemaking stage. Results indicate that the presence of indigenous LAB increased the risk of biogenic amine formation. Inoculation proved to reduce biogenic amine production over time compared to spontaneous MLF and co-inoculation even more than conventional inoculation. The presence of yeast lees during ageing generally led to higher final concentrations of biogenic amines in wines than the absence of lees. This study confirms other works that conclude that spontaneous MLF and uncontrolled ageing on yeast lees are generally unpredictable and pose a risk of biogenic amine contamination in finished wines.     

葡萄酒中的生物胺的生产与工艺控制

生物胺存在于多种发酵食品中，人体吸收过量的生物胺后会引起不良的生理反应，在葡萄酒苹果酸－乳酸发酵（MLF）过程中，有些乳酸菌能够对氨基酸脱羧产生生物胺。利用PCR与DNA探针技术能够快速检测葡萄酒中的组胺产生菌。工艺上采用接种进行MLF，并在MLF完成后对乳酸菌进行有效清除，可以显著降低葡萄酒中生物胺的含量。     

Winemaking biochemistry and microbiology: current knowledge and future trends

The fermentation of grape must and the production of premium quality wines are a complex biochemical process that involves the interactions of enzymes from many different microbial species, but mainly yeasts and lactic acid bacteria. Yeasts are predominant in wine and carry out the alcoholic fermentation, while lactic acid bacteria are responsible for malolactic fermentation. Moreover, several optional winemaking techniques involve the use of technical enzyme preparations. Considerable progress has been made recently in understanding the biochemistry and interactions of enzymes during the winemaking process. In this study, some of these recent contributions in the biochemistry of winemaking are reviewed. This article intends to provide an updated overview (including works published until December, 2003) on the main biochemical and microbiological contributions of the different techniques that can be used in winemaking. As well as considering the transformations that take place in traditional winemaking, the production of special wines, such as sparkling wines, 'sur lie' wines, and biologically aged wines, are also studied.     

Chemometric Analysis for the Detection of Biogenic Amines in Chilean Cabernet Sauvignon Wines: A Comparative Study between Organic and Nonorganic Production

In this work, the presence of biogenic amines (BAs) was correlated with the type of wine grape culture (traditional or organic) and their concentration in the different stages of winemaking (must, alcoholic fermentation [AF] and malolactic fermentation [MLF]). The formation of BA occurred mainly during MLF in which the percentages for putrescine, cadaverine, phenylethylamine, histamine, and tyramine were 100%, 70%, 13%, 61%, and 44% for the wines produced with traditional grapes and 100%, 94%, 25%, 88%, and 13% for the wines produced with organic grapes, respectively. In general, these latter wines exhibited a lower concentration of total amines. The principal component analysis and partial least-square discriminate analysis indicated that the generation of BA has a certain behavioral pattern in the wines analyzed, which is associated with the different stages of wine production and with the type of culture (traditional or organic) used in the wine grapes. Practical Application: Chemometrics tools can be useful as a method of characterization and classification in a global overview of the process variables involved in the development of toxic chemicals in foods, such as the production of BA in wine.     

Fast identification of wine related lactic acid bacteria by multiplex PCR

The microflora of must and wine consists of yeasts, acetic acid bacteria and lactic acid bacteria (LAB). The latter group plays an important role for wine quality. The malolactic fermentation carried out by LAB leads to deacidification and stabilisation of wines. Nevertheless, LAB are often associated with wine spoilage. They are mainly responsible for the formation of biogenic amines. Furthermore, some strains produce exopolysaccharide slimes, acetic acid, diacetyl and other off-flavours. In this context a better monitoring of the vinification process is crucial to improve wine quality. Moreover, a lot of biodiversity studies would also profit from a fast and reliable identification method.     

酿酒工艺对葡萄酒中生物胺的影响

探讨了不同酿酒工艺对葡萄酒中生物胺(组胺、苯乙胺、酪胺、色胺、腐胺、尸胺、精胺和亚精胺)含量的影响。结果显示,葡萄酒酿造过程中生物胺主要在酒精发酵和苹果酸-乳酸发酵过程产生。酒精发酵过程生成量较少,主要产生腐胺和精胺,果胶酶的使用、酵母接种量、发酵温度等因素可以调节酒精发酵过程生物胺的生成量;乳酸菌是葡萄酒生物胺最主要来源,在苹果酸-乳酸发酵过程中会产生大量的组胺和色胺。     

龙眼干白葡萄酒发酵过程中生物胺和氨基酸含量的变化

本研究利用高效液相色谱技术分析了龙眼干白葡萄酒酒精发酵和苹乳发酵过程中8种生物胺和22种氨基酸含量的变化。结果表明,在酒精发酵过程中8种生物胺含量均很低,而在苹乳发酵过程中迅速增加。发酵结束时干白葡萄酒中亚精胺和乙醇胺含量最高,其次为组胺和羟色胺。这4种生物胺的前体氨基酸在苹乳发酵阶段含量没有明显变化,而其它大部分氨基酸在酒精发酵过程中含量下降,在苹乳发酵过程中含量呈现不同程度升高。因此,我们认为：葡萄酒发酵过程中氨基酸含量变化与其对应生物胺的变化之间没有相关性,葡萄酒中生物胺的组成比例可能主要取决于乳酸菌中氨基酸脱羧酶的种类和活性。     

葡萄酒中生物胺含量的影响因素研究

研究了葡萄酒生产过程中影响生物胺产生量的主要因素,结果表明,葡萄酒酒精发酵和苹果酸．乳酸发酵过程均有牛物胺的产生,影响葡萄酒中生物胺产生的主要因素是乳酸菌中氨基酸脱羧酶的活性,本研究添加了氨基酸脱羧酶辅酶的样品与对照相比,生物胺含量提高了90．6％。酵母菌和乳酸菌种类、酒精发酵温度,生物胺前体物质也是影响生物胺含量的主要因素,发酵温度越高、生物胺产生量越;在一定范围内,氨基酸的浓度越大产生物胺越多。但酵母菌和乳酸菌的添加量对生物胺浓度的影响较小。     

Formation of biogenic amines throughout the industrial manufacture of red wine

Changes in biogenic amines (histamine, methylamine, ethylamine, tyramine, phenylethylamine, putrescine, and cadaverine) were monitored during the industrial manufacture of 55 batches of red wine. The origin of these amines in relation to must, alcoholic fermentation, malolactic fermentation, sulfur dioxide addition, and wine aging and the interactions between amines and their corresponding amino acids and pH were statistically evaluated in samples from the same batches throughout the elaboration process. Some amines can be produced in the grape or the musts (e.g., putrescine, cadaverine, and phenylethylamine) or can be formed by yeast during alcoholic fermentation (e.g., ethylamine and phenylethylamine), although quantitatively only very low concentrations are reached in these stages (less than 3 mg/liter). Malolactic fermentation was the main mechanism of biogenic amine formation, especially of histamine, tyramine, and putrescine. During this stage, the increase in these amines was accompanied by a significant decline in their amino acid precursors. Significant correlations between biogenic amine formation and the disappearance of their corresponding amino acids were observed, which clearly supports the hypothesis that malolactic bacteria are responsible for accumulation of these amines in wines. No increase in the concentration of biogenic amines was observed after SO2 addition and during wine aging, indicating that sulfur dioxide prevents amine formation in subsequent stages.     

Malolactic fermentation of Tempranillo wine: contribution of the lactic acid bacteria inoculation to sensory quality and chemical composition

Malolactic fermentation (MLF) of Tempranillo Rioja wines (Spain) inoculated with two lactic acid bacteria (LAB) strains were studied and compared with spontaneous MLF. Inoculation with selected Oenococcus oeni lyophila shortened MLF duration up to 19 days and lead to wines with more fresh and fruity characters, especially when implantation was 100%. We demonstrated modifications in the concentration of volatile and nitrogenous compounds and a good correlation between analytical and sensory attributes was also noted. In addition, the low initial amino acid concentration and the consumption of these compounds by the inoculated yeast strain during alcoholic fermentation resulted in wines with very low biogenic amines levels (under 3.75 mg L^?1) after MLF and 3 month storage period in all cases. The results showed the significance of choose the most suitable starter to elaborate quality wines and suggest the control of amino acid content in must and wine to prevent the formation of biogenic amines.     

Occurrence of biogenic amines in wine: The role of grapes

The evolution of biogenic amines from must to wine has been studied in seven different grape cultivars before and after malolactic fermentation. Alcoholic and malolactic fermentations have been carried out using selected yeasts and bacteria that, in a previous study, were unable to produce biogenic amines. The study has been performed under aseptic conditions to exclude possible interferences due to uncontrolled contaminating microorganisms present in grapes and/or in the environment. The goal of this work was to investigate the influence of grape on biogenic amines content of the wine. The results obtained showed that grape variety is related to the presence of some biogenic amines in wines and that, climatic conditions also affect the accumulation of these compounds in grapes.     

Biogenic amines in wines: Influence of oenological factors

Biogenic amines formation results from the decarboxylation of the corresponding amino acids by action of microorganisms. The presence of these compounds is considered by some authors a fundamental parameter for detriment of alcoholic beverages. The aim of this work was to assay the effect of some oenological factors (viticulture region, grape variety, anti-fungi treatment of grapes, fermentation activators, malolactic starters and storage on lees) from the point of view of their influence on the biogenic amines content of wines. According to our results, it was possible to show that the viticulture region affects the amounts of amines, since wines of some regions present higher contents of amines than wines from other regions. Grape varieties appear to influence the wine amines content. Commercial malolactic starters, after careful selection, should be added to the vinification process in order to decrease the formation of biogenic amines, since in our assays the wines that were inoculated with starters present lower amounts of biogenic amines. The wine storage on lees contributes for a biogenic amines increase.     

葡萄酒中生物胺的产生与工艺控制

生物胺存在于多种发酵食品中，人体吸收过量的生物胺后会引起不良的生理反应。在葡萄酒苹果酸—乳酸发酵(MLF)过程中，有些乳酸菌能够对氨基波脱投产生生物胺。利用PCR与DNA探针技术能够快速检测葡萄酒中的组胺产生菌。工艺上采用接种法进行MLF，并在MLF完成后对乳酸菌进行有效清除、可以显著降低葡萄酒中生物胺的含量。     

苹果酒酿造中的苹果酸-乳酸发酵

在苹果酒酿造中进行苹果酸-乳酸发酵,乳酸菌通过分解苹果酸,产生乳酸,引起其他有机酸的变化,使苹果酒的口感、质量得以改善。pH、温度、SO_2、酒度通过影响乳酸菌的活动而影响苹果酸-乳酸发酵的进行。保证苹果酒苹果酸-乳酸发酵进行的条件为:温度16～18℃,总SO_2含量<70mg/L,pH<3.70,酒精体积分数低于13％。     

Biogenic amine production by Oenococcus oeni during malolactic fermentation of wines obtained using different strains of Saccharomyces cerevisiae

Twenty-six wild Oenococcus oeni strains were investigated for their ability to form biogenic amines during malolactic fermentation in synthetic medium and in wine. Eight strains produced histamine and tyramine in screening broth at concentrations of 2.6-5.6 mg/L and 1.2-5.3 mg/L, respectively. Based on their ability to form biogenic amines, five strains were selected to inoculate three wines obtained by the fermentation of three different Saccharomyces cerevisiae strains (A, B, and C). All bacterial strains could perform malolactic fermentation for short periods in wine C, whereas only one strain performed complete malolactic fermentation in wines A and B. Two O. oeni strains (261 and 351) produced histamine and tyramine in wine C. Time-course analysis of these compounds showed that for both strains, histamine and tyramine production began at day 10 and finished on day 25, after the end of malolactic fermentation. These results indicate that the ability of O. oeni to produce histamine and tyramine is dependent on the bacterial strain and on the wine composition, which in turn depends on the yeast strain used for fermentation, and on the length of bacteria-yeast contact time after the completion of malolactic fermentation.     

Malolactic Fermentation and Secondary Metabolite Production by Oenoccocus oeni Strains in Low pH Wines

Abstract: Different wine varieties, including some with low pH, were studied to determine the ability to grow and produce secondary metabolites of a previously selected autochthonous Oenococcus oeni strain (C22L9), compared with a commercial strain. Monitoring of malolactic fermentation (MLF) was carried out by microbiological and chemical analysis of wines. The concentration of some major volatile compounds and biogenic amines in wines before and after malolactic fermentation was also determined. The results showed major differences in MLF duration both between wines and strains, although the differences between strains were slight for most of the analyzed compounds. Statistically significant differences in citric acid degradation were found in all wine varieties and it was confirmed that O. oeni C22L9 is a poor degrader of citric acid; this means that MLF can be prolonged without the risk of producing high concentrations of diacetyl and acetoin. Sensory analysis of wines after MLF showed similar characteristics in wines from both strains. This study thus shows that O. oeni C22L9 possesses even better sensory and fermentation properties than the commercial strain and can be used in wines with different characteristics, which makes it highly valuable for industrial use. Practical Application: The increasingly use of grape varieties of low pH in winemaking and the higher alcohol content of wines, as a consequence of the climatic change, make interesting the study of the behavior during MLF of O. oeni strains in order to determine their ability to grow, when growth conditions are not optimal, and to produce secondary metabolites. A comparative study was conducted using an autochthonous O. oeni strain (C22L9) and a commercial O. oeni strain and 4 wine varieties.     

Amino acids and biogenic amines in red varietal wines: the role of grape variety, malolactic fermentation and vintage

This work studies for the first time the amino acid and biogenic amine composition of Rioja red wines made with the red minority varieties Vitis vinífera cv. Monastel and Maturana Tinta de Navarrete, using Tempranillo as a reference variety. The role of malolactic fermentation and vintage on these compounds was also analysed, and discriminate analyses were applied to achieve a possible differentiation of the wines. Amino acid composition allowed a differentiation of wines according to grape variety. Monastel was characterised by the highest value in β-alanine and Maturana Tinta de Navarrete by its highest value in OH-proline. However, biogenic amines were no able to classify varietal wines. The malolactic fermentation had significant changes on the amino acid and biogenic amine content, and allowed distinguishing wines that underwent this process from wines without malolactic fermentation. No correlation was found between total amino acids and total biogenic amines after malolactic fermentation, suggesting that a higher initial concentration of amino acids in the medium did no affect the concentration of biogenic amines after malolactic fermentation. Vintage influenced the amino acid and biogenic amine pattern, obtaining a clear separation of wines by vintages. Monastel and Maturana Tinta de Navarrete wines showed a minor varietal character and were more influenced by the climatic conditions of each vintage than Tempranillo wines. All the wines showed histamine levels below the human physiological threshold and implemented regulations.     

不同接种方式下小片球菌C30对‘赤霞珠’葡萄酒品质的影响

以甘肃河西走廊葡萄酒产区小片球菌C30为苹果酸乳酸发酵(malolactic fermentation,MLF)启动菌,比较以不同接种方式进行MLF后对葡萄酒品质的影响,探讨小片球菌C30用于MLF的可行性及其优势。将小片球菌C30在酒精发酵(alcohol fermentation,AF)的早、中、后期进行接种,以商品乳酸菌为对照进行酿造实验。在研究小片球菌C30的MLF动力学与酿造葡萄酒安全性的基础上,分析其对葡萄酒基本理化指标、香气成分和感官品质等的影响。结果表明,MLF后期接种小片球菌C30,完成发酵时间最短(27 d),酒样中多糖、生物胺等物质的含量和残糖、酒精度等理化指标符合国家标准,酒样的香气物质总含量显著高于商品菌株。小片球菌C30可完成MLF,且具有安全性,在AF后期接种进行MLF,性能优良,对提升赤霞珠葡萄酒品质有积极影响。     

黄酒前酵中生物胺生成规律的研究

本研究对黄酒前酵工序的生物胺生成规律及影响因素进行了分析探讨。采用反相高效液相色谱技术,改进了生物胺定量检测法,该检测体系准确可靠,样品峰型对称分离度好且缩短了检测时间。分析了前酵工序中主要微生物,氨基酸,发酵醪酸度、糖度、酒精度、p H对生物胺生成的影响。发现前酵工序对成品酒生物胺的影响度为77.67%,第一次开耙阶段生物胺总量增幅最大,达到7.63 mg/L。研究发现主要酿造微生物中乳酸菌总数与生物胺总量成正相关,乳酸菌在搭窝期间生长速率最大,为7.13×106CFU/(m L·h)。在前酵工序中生物胺总量与发酵醪酸度、酒精度、p H呈显著正相关,与糖度呈负相关;前酵过程中主要生物胺与前体氨基酸也呈明显正相关。本研究分析了黄酒前酵中生物胺生成规律,有助于建立降低黄酒生物胺含量的更安全、科学的工艺。     

Kinetics of sugars, organic acids and acetaldehyde during simultaneous yeast-bacterial fermentations of white wine at different pH values

Successfully inducing malolactic fermentation in the production of grape wines can be challenging, especially in wines with multiple inhibitors such as low pH values and high ethanol concentrations. In the present study, the kinetics of several chemicals of enological relevance was studied in Chardonnay vinified by traditional, consecutive alcoholic (AF) and malolactic fermentations (MLF), and simultaneous AF/MLF, where bacteria were co-inoculated with yeast. The Chardonnay must was adjusted to four pH values (3.2, 3.35, 3.5 or 3.65), and the concentrations of sugars, organic acids as well as acetaldehyde were followed throughout the fermentations. The degradation of glucose and fructose was slower at the lowest must pH value (3.2) but independent from the time point of bacterial inoculation. In all cases, malolactic conversion was faster after yeast-bacterial co-inoculation and was completed in simultaneous treatments at pH values of 3.35-3.65, and consecutive treatments at pH 3.5 and 3.65. No statistically significant difference was observed among the final acetic acid concentrations among all inoculation and pH treatments, but there was a trend towards higher acetic acid residues in wines produced by co-inoculation, especially at high pH values. Overall, simultaneous AF/MLF allowed for greatly reduced fermentation times, but the must pH remained a strong factor for fermentation success and determined the final concentration of various wine components. The time point of inoculation influenced formation and degradation kinetics of organic acids and acetaldehyde considerably, and these are of relevance for vinification decisions.