Wine, Beer, and Alcohol

The production of alcohol beverages is a process that involves the active participation of microorganisms, most often yeasts.

Humans have been producing alcoholic beverages for thousands of years. The production of alcohol in these drinks is based primarily on yeast fermentation. Yeasts are eukaryotic microorganisms that ferment variety of sugars from different sources into the final products of carbon dioxide and alcohol.

Wine production

Wine is made from grapes or other fruit. The grapes are first cleaned of leaves and stems and the fruit is crushed into must that is ready for fermentation. The yeasts used for the fermentation grow a film on the fruit or in the environment. These wild strains play an important role in the final properties of the drink. However, cultivated strains of Saccharomyces cerevisiae are often added to improve the consistency of the final product. There are hundreds of commercially available yeast strains for wine fermentation.

Wine grapes: The white film that covers the grapes contains wild yeasts.

In the fermentation process, energy that is converted to heat is produced as well. It is important to keep the temperature in the fermentation vessel lower than 40ºC to keep the yeasts alive. To improve yeast growth, additional nutrients, like diammonium phosphate, are sometimes added in the fermentation step.

When making red wine, there is an additional fermentation step after alcoholic fermentation. Malic acid, naturally present in grape juice, can be converted to lactic acid by lactic acid bacteria naturally found in wineries or added artificially.

Beer production

Beer is the most consumed alcoholic beverage in the world. It is made most often of malted barley and malted wheat. Sometimes a mixture of starch sources can be used, such as rice. Unmalted maize can be added to the barley or wheat to lower cost. Potatoes, millet and other foods high in starch are used in different places in the world as the primary carbohydrate source.

The process of making beer is called brewing. It includes breaking the starch in the grains into a sugary liquid, called wort, and fermenting the sugars in the wort into alcohol and carbon dioxide by yeasts. Two main species are used in the fermentation process: Saccharomyces cerevisiae (top-fermenting, since it forms foam on top of the wort) and Saccharomyces uvarum (bottom-fermenting). Top-fermenting yeasts are used to produce ale, while bottom-fermenting produce lagers. The temperature used for top-fermenting (15-24ºC) leads to the production of a lot of esters and flavor products that give beer a fruity taste. Hops are added to introduce a bitter taste and to serve as a preservative.

Brewer’s yeasts are very rich in essential minerals and B vitamins, with the exception of vitamin B12. Beer brewing in modern days is performed by added pure cultures of the desired yeast species to the wort. Additional yeasts species that are used in making beer are Dekkera/Brettanomyces. After the fermentation is finished, the beer is cleared of the yeasts by precipitation or with the use of clearing additives.

Other types of alcohol beverages are made by the fermentation activity of microorganisms as well. A few examples are sake (uses the fungus Aspergillus oryzae to facilitate starch fermentation from rice), brandy, whiskey (both are distilled alcohol), and other alcohol beverages with higher percentage of alcohol compared to wine and beer.

Vinegar

Vinegar is a food product made by acetic acid bacteria that can ferment the alcohol in alcoholic liquids to acetic acid.

Vinegar has been used for cooking and in the household and different industries due to its mildly acidic nature for many centuries. It is one of the foods together with beer, wine, bread and fermented dairy products, that is the result of fermentation by microorganisms and has been around for thousands of years. It is a mixture of acetic acid (most often 5%) and water.

The fermentation is performed usually by acetic acid bacteria, from the genus Acetobacter, from the alcohol in variety of sources (e.g., apple cider, wine, potatoes, fermented grain). Acetobacter bacteria are Gram negative aerobic rods. They are naturally present in environments where alcohol is being produced and can be isolated from damaged fruit, apple cider, etc. In these liquids, the bacteria form a film on the surface, since they are aerobic and need good oxygen supply. This film, called mother of vinegar, can be used as a starter culture of acetic fermentation in fresh alcohol liquids. Mother of vinegar can also be found in unpasteurized store brand vinegar. Acetic acid bacteria are transmitted in nature by vectors like fruit flies and Vinegar eels.

Mother of vinegar: Mother of vinegar is used as a starter culture for vinegar production. It is made of a specific cellulose and acetic acid bacteria

This acetic acid fermentation needs oxygenation. If left at room temperature alcohol containing solution with Acetobacter will be converted to vinegar in months. The industrial process can be completed within hours since air is bubbled and mixed through the solution.

Vinegar can also be an undesired product in wine production. If the temperature in the fermentation vessel is too high, the Acetobacter will outgrow the yeasts and the produced alcohol will be converted to vinegar.

There are bacteria that can convert sugars straight to acetic acid in anaerobic fermentation. Such species include Clostridium and Acetobacterium but they can not tolerate acetic acid of concentrations higher than a few percent. The product made from these bacteria must be concentrated while oxidative fermentation by Acetobacter can produce up to 20% acetic acid.

Vinegar is a food product made all over the world from many different carbohydrate sources where alcohol fermentation has been performed. Some of them are more commonly used, such as apple cider and grapes, while others such as coconut water, dates, kiwifruit are used in specific regions of the world. Vinegar is used not only in food preparation but also as a cleaning agent due to its acidic nature and strong antibacterial properties. It can be used to lower the glycemic index of foods if consumed together with them. It has also been shown to reduce the risk of fatal ischemic heart disease when consumed frequently with oil in salad dressings.

Citric Acid and Other Organic Compounds

Many organic compounds, like citric acid, are produced industrially by microorganisms.

Citric acid (citrate) is an important substance in the Krebs cycle. It is produced from acetyl coenzyme A and oxaloacetate in the presence of the enzyme citrate synthase. The Krebs cycle is key in the oxidation of sugars, proteins and fats to carbon dioxide and water. Many of the cycle compounds are also needed for the synthesis of the cells’ own proteins, carbohydrates, and fats.

Colonies of Aspergillus niger: The black dots covering the colonies are Aspergillus spores. The growth medium is Saboraud’s Dextrose Agar (SDA)

Citrate has been used for centuries in different industries and in the households. It is used as a food additive to give a sour taste to foods or to preserve certain qualities of food products (e.g., prevents separating of the fats in ice cream). It has natural antibacterial properties and is used as a preservative as well. Its buffering property is used in cosmetics and pharmaceuticals to adjust the pH of products.

For centuries, the source of citric acid were citrus fruits. After World War I, the ability of some microorganisms to produce citric acid was further explored and the technology for industrial production was developed. Penicillium mold was the first described organism to produce citric acid but industrially another mold, Aspergillus niger, became the microorganism of choice. The mold is grown in a medium with sucrose or glucose as the main carbon source. The sugar source is usually an inexpensive solution like molasses or corn steep liquor. The microorganism makes more citric acid in the Krebs cycle than needed for the cell’s metabolism and exports it outside the cell. The citric acid is then precipitated out of solution and regenerated.

Microorganisms replaced the industrial chemical production of many different organic compounds, like enzymes and amino acids. Enzymes, such as glucoamylase (used to make high-fructose corn syrup) and pectinase (clearing agent for apple cider and wines) are produced industrially by Aspergillus. The food additivemonosodium glutamate (MSG) is produced in the form of glutamic acid by Corynebacterium glutamicum.

Edible Fungi

Fungi are used as food or as producers of a variety of food products (bread, wine, beer, etc. ) or compounds used in different industries.

Fungi are eukaryotic organisms that are separated taxonomically in the Fungi kingdom. The kingdom includes yeasts and molds (both microorganisms) and mushrooms. These organisms are ubiquitous all over the world. They have been used by people as food or as producers of a huge variety of food products or compounds used in different industries.

Saccharomyces cerevisiae: Single cells of baker’s yeasts under a light microscope. Total magnification is 1,500 x; gradation marks are 1 µm apart

Yeasts

The yeast species Saccharomyces cerevisiae has been used as leavening agentfor the production of bread since ancient times. The yeasts ferment the carbohydrates in the dough and produce carbon dioxide that causes the dough to rise and the bread to be softer after baking. Different sources provided the starter cultures. Dough could be left exposed to the air before cooking. Beer foam or grape juice paste were alos used as yeasts sources. Nowadays, the common used starters are pure cultures of Saccharomyces cerevisiae produced and sold as baker’s yeasts, although some artisan bakers maintain their own starter cultures. Other yeasts and some bacteria can be used as leavening agents too. For example, sourdough is made with Saccharomyces exiguus and Lactobacillus cultures that give it its sour taste.

Molds

Molds are fungi which cells grow in long chains of filamentous hyphae. The first antibiotic used in modern medicine, penicillin, was isolated form Penicillium mold. Different species of the mold Penicillium are added to milk or curd when making soft cheese to produce blue cheese. The mold adds specific smell and flavor to the cheese. Some bacteria, such as Brevibacterium linens, are also used to give blue cheese its characteristic odor.

Boletus edulis: An edible wild mushroom widely distributed in the Northern Hemisphere and artificially introduced to the Southern Hemisphere

Each mold species is usually found in the environment of the local region where the production of specific brand started. To enhance the mold growth in cheese, different techniques are applied to improve the access of air. The cheese is ripen for weeks to months in dark cold places. Even before the discovery of penicillin, people used blue cheese to prevent gangrene in wounds.

Mushrooms

Edible mushrooms are macrofungi since they are visible with a naked eye. Mushrooms have fleshy fruit body with certain aroma and flavors as well as good nutritional properties and are used mostly as food. A few species of mushrooms have been cultivated but wild mushrooms are harvested as well. However, some mushrooms produce toxic compounds that can be life-threatening. Proper identification is key since quite often poisonous mushrooms mimic edible ones in appearance. Even if not dangerous, mushrooms in general are great absorbants of chemicals from the environment and sometimes they can make them toxic, e.g., pesticides, insecticides, heavy metals. Certain mushrooms have been used for their medicinal properties in some cultures.

Edible Algae

Edible algae have been used as food for centuries in many coastal regions all over the world.

Algae are a very diverse group of generally simple unicellular or multicellular eukaryotic organisms. Most of them are autotrophic which means that they can harvest carbon dioxide from the atmosphere and convert it to organic matter. They inherited their photosynthetic apparatus from cyanobacteria. Cyanobacteria are sometimes called blue-green algae but they are prokaryotic organisms and are not true algae. Some cyanobacterial species are used as food as well.

Seaweeds are edible algae that have been used for centuries as food in many coastal regions all over the world. They may belong to one of three groups of multicellular algae: red, green or brown. In countries such as China, Japan, Korea and to some extent Iceland, Ireland, Chile and New Zealand algae are part of people’s regular diet. They are usually of marine origin since freshwater algae are often poisonous.

Sea grapes: Sea grapes (Caulerpa lentillifera) is a type of seaweed consumed raw as a salad or snack

Algae are of excellent nutritional value since they contain complete protein (in contrast to plant food harvested on land), fiber, and sometimes high levels of omega-3 fatty acids. In fact, the omega-3 acids in fish comes from the microalgae consumed at the bottom of the food pyramide and gradually passed up to the fish at the top. Algae are also rich in many vitamins, such as A, C, B₁, B₂, B₃ and B₆, as well as minerals, such as iodine, calcium, potassium, magnesium and iron. They are consumed both cooked, dried and raw.

Cultivated microalgae and cyanobacteria such as Spirulina and Chlorella are sold as nutritional supplements. Hydrocolloids such as agar, alginate and carrageenan are isolated from wild and cultivated algae and used as additives in the food industry for their emulsifying and thickening properties. Some of the complex polysaccharides found in algae may be digested by bacteria in the gut since the needed enzymes for digestion are abundantly present in Japanese people but absent in people from North America.

Microbes and Dairy Products

Fermented dairy products have been prepared and consumed by people for centuries due to their high nutritional values.

Fermented milk or dairy products have been part of human diet since ancient times. Various fermented products are made by different strains. Lactic acid fermentation is performed most often by lactic acid bacteria. Due to their abundance in nature, including mucosal surfaces of the human body, and their use in fermented foods they are labeled as GRAS (generally recognized as safe). The main genera that belong to the lactic acid bacteria group are: Lactobacillus, Leuconostoc, Lactococcus, Pediococcus and Streptococcus. These bacteria ferment the carbohydrates in milk, the major one being lactose, to lactic acid and some other products. The acid precipitates the proteins in the milk and that is why fermented products are usually of thicker consistency than milk. The high acidity and low pH hinders the growth of other bacteria, including pathogens. Some lactic acid bacteria can produce agents with antimicrobial properties. Since milk is rich in many nutrients such as protein, calcium, phosphorus, and B vitamins dairy products are an excellent food.

Some of the most popular and widespread cultured dairy products are yogurt and cheese.

Records of yogurt preparation as food date back to centuries BCE. Classic yogurt is the result of the fermentation of two main bacterial species: Lactobacillus bulgaricus and Streptococcus thermophilus. Sometimes other lactic acid bacteria are added as well. Yogurt is most often made of cow’s milk although milk from sheep, goat, water buffalo, camels and yaks is used as well depending on the region of cultivation.

To make yogurt, the milk is first heated to 80ºC or boiled to kill any pathogenic bacteria and to denature the milk proteins to prevent the formation of curds. After it is cooled down to about 45ºC, the starter culture of the two species is mixed well with the milk and incubated at the same temperature for a few hours. In many countries, the traditional food is yogurt without any sweeteners which could be consumed plain or used to prepare a variety of dishes usually with vegetables. Yogurt has been traditionally consumed in Eastern cultures as a cold drink after mixing with water (e.g., lassi, ayran, doogh). After the industrialization of yogurt production in the twentieth century, yogurt with added sweetener and fruit or fruit jam has become popular in the Western world.

Turkish cacik: A yogurt dish made of plain yogurt and cucumbers.

Cheese is another popular and ancient dairy product. It consists of milk proteins and fat together with lactic acid bacteria. It has longer shelf life than uncultured milk. Currently there are a few hundred varieties of cheese produced all over the world. Making cheese is similar to yogurt but after acidification usually with lactic acid bacteria (Lactococci, Lactobacilli, Streptococci), the solids are separated from the whey by coagulation with rennet and processed further to yield the final product. Depending on the type of cheese, the solids could go straight to packaging or other bacteria or mold could be added (e.g., Penicillium mold for blue cheese) for additional fermentation.

Other fermented and widely consumed cultured dairy products include kefir (lactic acid bacteria and yeasts are used for the fermentation), sour cream (fermented cream), cultured buttermilk (fermented cow’s milk with Streptococcus lactis or Lactobacillus bulgaricus only).

Lactic acid bacteria have been researched for medicinal health benefits. In the early twentieth century, the Nobel laureate in medicine, Elie Metchnikoff, believed that the longevity of peasants in Bulgaria and the Russian steppes was due to their high consumption of milk-fermented products. He hypothesized that the lactic acid bacteria would inhabit the gut after consumption, create and acidic environment as they grow and multiply, and hence prevent the growth of proteolytic. After it was discovered that Lactobacillus bulgaricus can not live in the human gut, the idea was abandoned. Years later, strains of Lactobacillus acidophilus were found to thrive in the gut after implantation and the research started again. The term “probiotics” was introduced and defined as live microorganisms that provide beneficial effects for their host when administered in adequate concentration. Most of the researched species were isolated from different fermented dairy products. The research has been focused on curing or preventing a number of diseases like diarrhea, intestinal inflammations, urogenital infections, allergies, etc. Some species have been prepared and sold as nutritious supplements. However, so far there has not been enough evidence to establish a definite cause and effect relationship about any of the marketed products.