农药对葡萄酒发酵的影响

CHAPTER 2 Contents 44 B. 47 References 60 Abstract Advances in Food and Nutrition Research, Volume 59 # 2010 Elsevier Inc. ISSN 1043-4526, DOI: 10.1016/S1043-4526(10)59002-8 All rights reserved. Department of Toxicology, University of Cagliari, Cagliari, Italy Wine quality strongly depends on the grape quality. To obtain high- quality wines, it is necessary to process healthy grapes at the correct ripeness stage and for this reason the farmer has to be especially careful in the prevention of parasite attacks on the grapevine. B. Effect of malolactic fermentation on pesticide residues 60 A . Pesticide effect on lactic bacteria 59 V. M alo lactic Fermentation 58 p esticide residues 57 C . E ffect of alcoholic fermentation on B . P esticide effect on yeasts 54 A . A lcoholic fermentation 50 IV. F ermentation Process 50 III. P esticides 49 E . V ine and the citrus mealybugs (P. ficus and P. citri) 48 D . G rape moth (L. botrana) 48 C . G Powdery mildew (U. necator) ray mold (B. cinerea) 47 A . Downy mildew (P. viticola) 46 II. G rap evine Pathogens 45 I. In tro duction Pierluigi Caboni and Paolo Cabras Pesticides’ Influence on Wine Fermentation 43 The most common fungal diseases affecting grape quality are I. INTRODUCTION The grapevine req grapes can be cul spheres. These zo 30� to 40� of south The highest co Mediterranean ba ducing nations. In the highest prod most important g nent. Recently, Au grape-producing n million hectares, hectoliters. The E which correspond and Italy compet constantly battled Nevertheless, p ing, most importa 44 Pierluigi Caboni and Paolo Cabras uires particular climatic conditions and, for this reason, tivated only in the temperate zones of the two hemi- nes lie between 50� and 30� of north latitude and from latitude (Fig. 2.1). ncentration of grapevine cultivation is located in the sin where France, Italy, and Spain are the largest pro- the North America, California is the U.S. state with uction of wine, while Chile and Argentina are the rape-producing nations on the South American conti- stralia and New Zealand have emerged as significant ations. Worldwide, the grapevine is cultivated on eight and the wine production reaches about 260 million U possesses 64% of the total grapevine cultivation, s to the 3/4 of the worldwide wine production. France e for the top rank in wine production; they have for first and second place in any given vintage-year. er capita wine consumption is progressively diminish- ntly in those top producing countries such as France, downy and powdery mildew (Plasmopara viticola and Uncinula necator), and gray mold (Botrytis cinerea). On the other hand, the most dangerous insects are the grape moth (Lobesia botrana), vine mealybug (Planococcus ficus), and the citrus mealybug (Planococcus citri). Farmers fight grape diseases and insects applying pesticides that can be found at harvest time on grapes. The persistence of pesticides depends on the chemical characteristic of the active ingredients as well as on photodegradation, thermodegradation, codistillation, and enzymatic degradation. The pesticide residues on grapes can be transferred to the must and this can influence the selection and development of yeast strains. Moreover, yeasts can also influence the levels of the pesticides in the wine by reducing or adsorbing them on lees. During the fermentative process, yeasts can cause the disappearance of pesticide residues by degradation or absorption at the end of the fermentation when yeasts are deposited as lees. In this chapter, we reviewed the effect of commonly used herbicides, insecticides, and fungicides on yeasts. We also studied the effect of alcoholic and malolactic fermentation on pesticide residues. B Pesticides’ Influence on Wine Fermentation 45 Spain, and Italy. For example, the Italian average per capita wine con- sumption decreased from 119.6 to 99.8 L from the beginning of the century through 1960. After a progressive rise in wine consumption reaching the maximum of 116 L per capita in 1968; a constant reduction has been observed from 94.6 L in 1978, to 90.6 L in 1980, to 61.5 in 1990, to 57.6 L in 1995, to 57.2 L in 1996, and to 46.5 in 2005. In the new wine-producing countries, such as New Zealand, individual consump- tion continues to grow quickly (20.3 L per capita in 2005, which is a two- FIGURE 2.1 Worldwide distribution of grape cultivation shown in dark shade. A fold increase in 6 years). Countries with the highest per capita wine consumption are reported in Table 2.1. II. GRAPEVINE PATHOGENS There is no question that the wine quality strongly depends on the grape quality. To obtain high-quality wines, it is necessary to use healthy grapes at the correct ripeness stage and for this reason the farmer has to be especially careful in the prevention of parasite attacks on the grapevine. Many of the grapevine parasites are of animal origin (insects andmites) or from vegetal origin (critograme or parasitic fungi). The most common fungal diseases are downy and powdery mildew (Plasmopara viticola and Uncinula necator), and gray mold (Botrytis cinerea). On the other hand, the most dangerous insects are the grape moth (Lobesia botrana), vine mealy- bug (Planococcus ficus), and the citrus mealybug (Planococcus citri). 46 Pierluigi Caboni and Paolo Cabras A. Downy mildew (P. viticola) The downy mildew was introduced in France from North America in 1878. It moved into Italy the following year and subsequently into the other countries of the Mediterranean basin. Furthermore, it was intro- duced into Australia in 1919 and into New Zealand in 1926. Today downy TABLE 2.1 World per capita wine consumption Countries Per capita wine consumption (L) France 55.4 Luxembourg 54.6 Portugal 46.7 Italy 46.5 Slovenia 44.7 Croatia 40.8 Switzerland 39.3 Hungary 34.7 Greece 32.2 Spain 31.8 Austria 29.3 Denmark 28.7 Year 2005, source O.I.V. mildew is present in all vine cultivation areas. The downy mildew is the most prevalent form of mildew and is usually spread by rainfall. It attacks leaves, shoots, and berries and can quickly defoliate the vine leading to loss of the entire crop. Optimum conditions for primary infection take place at 10:10:10, which corresponds to at least 10 mm of rain at a temperature 10 �C or more, over 10 h. The fungus survives in the form of spores for 3–5 years in old, infected leaf material that is remaining in the soil and, with rain, is splashed onto the foliage. If the spores remain wet long enough, the disease begins to develop. This shows up as ‘‘oil spots’’ on leaves. Spores form under the oil spot and show up as a ‘‘white down.’’ If conditions are right, second- ary infection occurs from these spores and the spread of the disease becomes quite rapid. Downy mildew can be controlled by the spray application of various chemicals either as preinfection or postinfection treatments. There are two groups of spray chemicals, those with single site activity which act on only one site within the fungus organism or those with multisite activity, which act on more than one site within the fungus. The most used multisite chemicals for the preventive control of downy mildew are Pesticides’ Influence on Wine Fermentation 47 copper salts such as copper oxychloride. In the past several years, the overuse of chemicals has lead to small mutational changes within the fungus, which in turn can lead to the fungus being resistant. At present, many of the nonsystemic active ingredients against downy mildew such as metiram, mancozeb, folpet, tolylfluanide are commonly used. On the other hand, the systemic fungicides in current use are cymoxanyl, dimethomorf, famoxadone, fenamidone, zoxamide, meta- laxil-m, iprovalicarb, and strobirulines (azoxystrobin and pyraclostrobin). B. Powdery mildew (U. necator) The powdery mildew is a pathogen that was brought into England from North America in 1845. Subsequently, powdery mildew was introduced into France in 1847, Belgium in 1848, and finally Italy in 1849. By 1853, it was discovered, in France, that vine treatments with sulfur were able to control this pathogen. Powdery mildew attacks leaves, shoots, and bunches. It is evidenced by an ash gray to white powdery growth on both the upper and lower surfaces of the leaves. Moreover, the disease attacks the bunches with the same ash gray/white powder showing up on the berries and stalks. Other than crop losses, the most negative aspect is that the disease causes off flavors in wine production. Powdery mildew spores hide in the buds of dormant vines. Mild cloudy weather and low light in the canopy encourage development of this disease. There are no approved fungicides for postinfection treatments that make the application of a protective spray from budburst necessary. There are multiple chemicals from both the singlesite and multisite groups. In Italy, the relatively safe and multisite active wettable sulfur is utilized. Other than sulfur, many active ingredients such as dinocap, fungi- cides, QoI-STAR derived from strobilurins (azoxystrobin, kresoxy-methyl, trifloxystrobin), quinoxyfen and IBS (inhibitors of sterol biosynthesis) such as fenarimol, triadimenol, penconazole, myclobutanil, fenbuconazole, hexaconazole, fusilazole, tetraconazole, and tebuconazole are used. Other active ingredients used are proquinazid and spiroxamine. C. Gray mold (B. cinerea) Gray mold is a common bunch rot in regions with warm, wet conditions. In addition to the fruit, it can also attack shoots and leaves. It causes large crop losses while infected grapes can cause off flavors in the wine. It should bementioned here that not all botrytis infections are unfavorable. Under specific conditions, the fungus takes hold and dehydrates the bunches increasing the sugar content without causing rot. This enables very sweet dessert wines with their traditional marmalade favor caused There are virtually no curative sprays, and it is essential that a protective almost ripe fruits and various molds, in particular Botrytis, develop very 48 Pierluigi Caboni and Paolo Cabras rapidly on thewounds; the attacked fruits turn brown at the place of attack and begin to rot. The presence of larvae and rotten fruits lowers the quality of the crop; molds render wine making difficult and may require the crop to be harvested prematurely. The following pesticides are commonly used to control the grape moth: pyrethroids (cypermethrin and deltamethrin), organophosphorus (chlorpyrifos, chlorpyrifos-methyl), nicotinoids (imi- dacloprid), oxadiazine insecticides (indoxacarb), chitin synthesis inhibitor insecticides (flufenoxuron, lufenuron), and moulting hormone agonists (tebufenozide). E. Vine and the citrus mealybugs (P. ficus and P. citri) The two insects, morphologically similar, are the vine mealybug (P. ficus), and the citrus mealybug (P. citri). They are currently the most economi- cally important pseudococcids in vineyards in Italy. All life stages of vine mealybug are found throughout the vine, including on the roots, under spray is applied at very definite times of bunch development. Applications are commonly at 80% capfall (toward the end of flowering) and again just before bunch closure (just before the berries have stopped growing and become ‘‘squished’’ together in the bunch). Chlorothalonil is commonly used for this purpose. This chemical is also a protectant against downy mildew so it can replace the copper. Pesticides used to control botrytis are the following: dicarboxymides (iprodione, procymidone, and vinclozonil), new generation products such as pyrimethanil, mepanypirim, fenhexamid, ciprodinil þ fludioxonil, and fluazinam. D. Grape moth (L. botrana) The life cycle of L. botrana can allow 3–4 generations depending on geo- graphical and environmental variability and whether the summer has been hot. The moths first appear at the end of April when the vine has 3 or 4 leaves and they emerge at intervals and the flights spread over 2–3weeks. The caterpillar finishes its development at the time of flowering and then it pupates. The second flight takes place toward the end of June and into July; then the caterpillars pupate again and the third flight occurs between mid-August and the end of September. The caterpillars gnaw the by the action of fungal enzymes (e.g., Sauterne in France or in Australia). The disease in this case is known as noble rot. The disease hides in decaying plant debris such as dead canes and mummified fruit. Spores are spread by wind and find a place in the developing bunch flowers. If the ‘‘closed’’ bunch coincides with wet weather and high humidity, the disease spreads rapidly. sap as it dries. In addition, a fungus called ‘‘sooty mold’’ grows on the honeydew. This black fungus covers the grape leaves interfering with Pesticides’ Influence on Wine Fermentation 49 photosynthesis and fouling the grape bunches. The vine mealybug is known to transmit leaf roll virus in grapes. This same behavior is exhib- ited in P. citri. Generally, the chemical control is done in the spring time to coincide with the emerging of nymphs from winter sites using mineral oil or calcium polysulfur. Chlorpyrifos, chlorpyrifos-methyl, imidacloprid, methomyl, buprofezin, and dimethoate can be used as alternatives. III. PESTICIDES Before entering the market, pesticides need to be registered. Starting in 2008, pesticide registration has been done by the EU and not by individual countries. The registration process for each pesticide set requires the authorized culture, the dose, preharvest interval, and the maximum resi- due limit (MRL). The legal limit of the residue does not coincide with the toxicological limit and for this reason still if the legal limit is exceeded it will not pose a serious risk to human health. The legal limit is determined from toxicological data establishing a lack of risk to human health (NOEL ¼ no observed effect level) commonly corrected by a safety factor of 100. Field residues of pesticides are affected by the environmental conditions (temperature, wind, rain, solar irradiance, etc.). Field residues, if below the corrected acceptable daily intake, are used to set the legal limit of the pesticide residue. Residues limits can vary between countries because of the different climatic conditions, leading to EU trade difficulties. the bark on the trunk and cordons, on canes, and leaves. There is no overwintering stage, rather all life stages can be found throughout the year. There are usually 3–7 generations per year. During the winter months, eggs, nymphs, and adults can be found under the bark, within developing buds, and on the roots as well. As temperatures warm in the spring, the density of vine mealybug increases, and the mealybugs move out to the cordons and aerial parts of the vine. Vine mealybug can be found on all parts of the vine including leaves and clusters by late spring and summer. Shortly after harvest, the density of vine mealybug declines. This generalized biology fits most vine mealybug populations; however, it varies slightly with location and cultivar. At high densities, the vine mealybugs can reduce plant vigor by removing large amounts of sap, which carries the nutrients to the grape roots and growing tissues including the grape bunches. The vine mealy- bugs excrete large amounts of fluids that have high concentrations of sugars. This ‘‘honeydew’’ can foul the grapevine with a layer of sticky over, yeasts can also influence the levels of the pesticides in the wine by fermentation step (malolactic fermentation), which corresponds to the transformation of L-malic acid to L-lactic acid. 50 Pierluigi Caboni and Paolo Cabras A. Alcoholic fermentation In winemaking, the fermentative process may take place due to ambient yeasts that are naturally present in wine cellars, vineyards and on the grapes themselves (sometimes known as a grape’s ‘‘bloom’’). Otherwise, it can be conducted using cultured yeast which are specifically isolated and inoculated for use in winemaking. Yeasts responsible for alcoholic fermentation belong to the genus Saccharomyces spp. However, other yeasts, especially non-Saccharomyces yeasts are present in the initial stages of the fermentation process and may have an influence on the final organoleptic properties of the wine (Pretorius et al., 1999). These genera include Candida, Klo¨ckera/Hanseniaspora, Pichia, and Zygosaccharomyces. These yeasts grow to about 106, 107 cfu/mL but, by midfermentation begin to decline and die off. At this time, Saccharomyces cerevisiae becomes predominant (107, 108 cfu/mL) and continues the fermentation until its completion. Evidence exists that non-Saccharomyces yeasts may influence the unique oenological characteristics of each wine-producing zone, while reducing or adsorbing them on lees (Cabras et al., 1987). IV. FERMENTATION PROCESS In the fermentative process, the first step is due to yeasts which transform sugars to alcohol (alcoholic fermentation). This is followed by a second Currently the EU is working for the harmonization of the MRLs of pesticides. In Italy, pesticides currently registered for use on grapes are listed in Table 2.2. Italy is one of the few countries with legal limits also set on wine (Table 2.3). In other countries, where there is a lack of a legal limit for processed foods, the amount of the raw food corresponding to pro- cessed food unit (e.g., 1.5 kg of grapes for 1 L of wine), and the incidence of technological process should be taken into account. Since each active ingredient has its particular behavior, residue changes during the trans- formation process should be determined. In the absence of these data, the unique and safe reference is the MRL of the primary food. Different levels of pesticides can be found at harvest on grapes depending on the chemical characteristic of the active ingredients. More- over, the persistence of pesticides can depend on photodegradation, thermodegradation, codistillation, and enzymatic degradation. The pesticide residues on grapes can be transferred to the must and this can influence the selection and development of yeast strains. More- TABLE 2.2 Pesticides registered on grapes in Italy Pesticide MRL (mg/kg) Pesticide MRL (mg/kg) Pesticide MRL (mg/kg) Abamectin 0.01 Esfenvalerate 0.1 Methoxyfenozide 1 Acrinathrin 0.1 Ethephon 0.1 Metiram 2 Alcalines solphites 10 Etofenprox 0.05 Myclobutanil 1 Alphamethrin 0.3 Etoxazole 1 Oxadiazon 0.05 Azadirachtin 0.5 Famoxadone 0.02 Oxyfluorfen 0.05 Azinphos-methyl 1 Fenamidone 2 Paraquat 0.05 Azociclotin 0.3 Fenamiphos 0.5 Penconazole 0.2 Azoxystrobin 2 Fenarimol 0.02 Phosalone 1 Benalaxyl 0.2 Fenazaquin 0.3 Phosetyl-al 2 Benfuracarb 0.05 Fenbuconazole 0.2 Piperonyl butoxide 3 Bifenthrin 0.2 Fenbutatin oxide 0.2 Pirimicarb 0.2 Bifentrin 0.2 Fenhexamid 2 Pirimiphos-methyl 2 Bromopropylate 2 Fenoxycarb 0.5 Procymidone 5 Bromuconazole 0.5 Fenpropidin 0.2 Propargite 2 Buprofezin 1 Fenpropimorph 2 Propiconazole 0.5 Calcium polysulfide 50 Fenpyroximate 0.05 Propineb 2 Captan 10 Flazasulfuron 0.3 Propyzamide 0.02 Carbaryl 3 Fluazifop-p-butyl 0.01 Pyraclostrobin 2 Carbendazim 2 Fluazinam 0.1 Pyrethrins 1 Chloropicrin 0.05 Fludioxonil 1 Pyridaben 0.1 Chlorothalonil 3 Fludioxonil 2 Pyrimethanil 3 Chlorpropham 0.05 Flufenoxuron 2 Quinoxyfen 0.5 (continued) TABLE 2.2 (continued ) Pesticide MRL (mg/kg) Pesticide MRL (mg/kg) Pesticide MRL (mg/kg) Chlorpyrifos 0.5 Flusilazole 0.1 Rotenone 0.05 Chlorpyrifos-methyl 0.2 Fluvalinate 0.01 Spinosad 0.2 Clofentezine 1 Folpet 0.5 Spiroxamine 1 Cyanamide 0.05 Glufosinate ammonium 10 Sulfur 50 Cyazofamid 1 Glyphosate 0.1 Tebuconazole 1 Cycloxidim 0.1 Glyphosate trimesium 0.1 Tebufenozide 0.5 Cyfluthrin 0.3 Hexaconazole 0.1 Tebufenpyrad 0.3 Cyhexatin 0.3 Hexythiazox 0.5 Teflubenzuron 1 Cymoxanil 0.1 Indoxacarb 0.5 Tetraconazole 0.5 Cypermethrin 0.5 Ip