Chemistry: Fermentation

Introduction

Fermentation is a biochemical process that is initiated by the actions of naturally occurring microorganisms acting on virtually any type of plant or animal product. It happens anywhere when the environmental conditions are right, with or without man's intervention. If fermentation is carried out under controlled conditions, it enriches the flavor and aroma of foods while adding to their food value and safe storage. It is a relatively easy, efficient, and low energy food enrichment and preservation process. Fermentation as a process was developed using native foods in villages to produce products that were adapted to individual cultures and traditions.

Fermentation of foods as part of cultures and traditions can be found in all parts of the world. It can be traced back to the beginning of recorded history. Bread, beer, wine, and cheese have been found as food staples back thousands of years. Soy sauce, sauerkraut, and yogurts are also fermentation products with a long history linked to different cultures.

Understanding the science behind fermentation did not happen until the microscope was invented, and scientists discovered the world of microorganisms. Fermentation is the decomposition of organic compounds into simpler compounds, which occurs when the right microorganisms are present along with the right conditions for their growth. All living organisms produce organic compounds. Today, organic compounds are also produced in chemical laboratories, some of them through fermentation processes.

The most common compounds associated with fermentation are sugars from fruits; the fermentation products are wines which contain alcohol. Fermented cereal grains are equally common and produce beer, also an alcoholic product. In a different fermentation process, cereal grains are used to produce leavened breads. The breads are leavened, expanded in volume, and made light in texture by carbon dioxide gas that is produced in the fermentation process. Alcohol is not a significant product in making bread, because the fermentation process is so short.

The microorganisms associated with fermentation are all non-green plants classified as bacteria, yeasts, and molds. Yeasts and bacteria are single-celled, with bacteria being smaller than yeasts. There are a great number of different varieties of yeasts, bacteria, and molds; not all of them good are for fermentation. Some microorganisms are actually harmful or produce toxic products.

Fermentation preserves foods, because it produces alcohol and acid products that block the growth of the harmful microorganisms. Bread and alcohol are produced with yeasts. Cheeses are produced with molds and bacteria, and the specific type of microorganism used depends on the type of cheese desired. Soy sauce is produced using molds at the start of a very long process that begins with soybeans.

Historical Background and Scientific Foundations

From the start of recorded history, artifacts have been found that indicate fermentation was being used to produce alcoholic drinks and fermented dairy products. The oldest known ancient wine jar that has been found dates back more than 7,000 years to about 5400 BC. A residue found in the jar was analyzed and identified as having come from wine.

The production of wines and beers was common in ancient countries in the Middle East. In Mesopotamia and Syria as far back as 2600 BC, brewing beer was a home industry. Women were the brewers and vendors. They were the local distillers and alchemists. A woman known as Maria the Jewess, an alchemist in Alexandria, Egypt, about AD 100, is said to have invented a three-armed still.

The distillation of wine and beer was carried out by the early Arabic cultures, but their methods were not

efficient. By 1667 distillation methods had improved, with advances in glass making and distillation equipment. The properties of pure alcohol were observed. The liquid looked like water, but it burned and was a solvent for many things not easily mixed with water. The distillation of alcohol became very important to the alchemist, apothecary, and brewer in Europe during this period in history. Alcoholic distillates were called aqua vitae, or water of life.

The history of fermented milk products such as yogurt has been traced to early civilizations in the Middle East and Asia. The army of Mongolian leader Genghis Khan (c.1160–1227) was reputedly so strong and successful because of their diet, which was largely based on yogurt. In Northern Europe the recipe for Skyr (a fermented, creamy cheese that resembles yogurt) was allegedly brought to Iceland by the Vikings more than 1,100 years ago. It is still a national staple in Iceland.

Although the craft of fermenting food products was well advanced by the 1600s, the science behind the art

was unknown. There were scientific advances in other areas such as those achieved by the Italian scientist Galileo Galilei (1564–1642) who had turned his telescope to the skies. Galileo also made a simple microscope but it was not very clear or powerful. A century later British scientist Robert Hooke (1635–1703) redesigned the microscope and discovered the wonders of seeing details on fleas that were never known before. Hooke published his design for a microscope and illustrations of magnified objects in 1664. His publication, Micrographia, caught the attention of a Dutchman, Antoni van Leeuwenhoek (1632–1723), who then made an even better microscope. Van Leeuwenhoek made a microscope that could clearly magnify over 200 times.

Van Leeuwenhoek was not only skilled in grinding lenses, he was very curious about the world around him as it was seen through his microscope. When he looked at pond water he discovered microorganisms. He wrote so clearly about this wondrous new microscopic world in 1674 that from his descriptions of microorganisms scientists can instantly identify them today.

Although the existence of microorganisms was discovered in the 1600s, no connection was made between microorganisms and fermentation until after French scientist Louis Pasteur (1822–1895) became dean of science at Lille University in 1854. A brewer came to Pasteur with a problem. His beer was turning sour after fermentation. Pasteur conducted a very detailed scientific study of fermentation during which he discovered and proved that living yeast was necessary to produce beer. He also discovered that there are a great many microorganisms and that some of them spoiled foods. Only the correct type of yeast would make good beer, or wine. Fermentation processes entered the scientific era with Pasteur's discoveries.

German chemist Eduard Buchner (1860–1917) was studying the chemistry of fermentation in the 1890s when he discovered the living whole yeast was not necessary for fermentation. He received the Nobel Prize in 1907 for his research that led to the discovery that fermentation only needed something made by the yeast cells; that fermentation could be carried out without the whole yeast cells.

Buchner's discovery was advanced further by the work of British chemist Arthur Harden (1865–1940) and the work of Swedish chemist Hans Karl August Simon von Euler-Chelpin (1873–1964), whose investigations of the fermentation of sugars led to their discovery that the extracted yeast material that makes fermentation possible contains enzymes. It is the enzymes that are needed for fermentation. Enzymes are catalysts, complex organic compounds that make a reaction happen faster.

Shortly before Pasteur discovered the science of fermentation, in 1828 German chemist Friedrich Wöhler (1800–1882) synthesized the organic compound urea, a compound that is found in nature as a waste product of urine. The synthesis of organic compounds was now possible in the laboratory. Until Wöhler accidentally prepared urea, it was thought all organic compounds could only be made in living organisms. Seventeen years after Wöhler synthesized urea, in 1845, his pupil Hermann Kolbe (1818–1884) synthesized acetic acid entirely from constituent elements. Up until this time acetic acid had only been produced in vinegar through fermentation processes.

The synthesis of organic compounds in the laboratory increased greatly over the years, with an estimated 75,000 different compounds being synthesized by 1900. The synthesis of compounds other than alcohol using fermentation was not considered a cost-effective alternative until the 1920s, when efficient fermentation processes began to be developed to produce enzymes, the solvent acetone, another alcohol called butanol, acetic acid, lactic acid, and citric acid on an industrial scale. Along with the industrial production of chemicals came the industrialization of the fermented food products such as wine, beer, cheeses, yogurts, and breads.

Fermentation also played a role in one of the greatest discoveries in medicine during the twentieth century. Although the use of natural materials to treat wounds also goes way back in history, it was not until Scottish biologist Alexander Fleming (1881–1955) observed a common mold had destroyed his culture of a disease bacteria that antibiotics were recognized as powerful tools to fight infections. In 1945 the Nobel Prize was given to Fleming, UK scientist Ernst Chain (1906–1979), and Australian Howard Walter Florey (1898–1968) for their discovery of penicillin and its curative effects.

The name antibiotic was first used by Russian-American microbiologist Selman Waksman (1888–1973) in 1941. Waksman led a research group at Rutgers University that was devoted to the discovery of a treatment for tuberculosis and other critical infectious diseases. Fermentation processes are used in the industrial preparation of antibiotics. Waksman received the Nobel Prize in 1952 for his discovery of streptomycin, the first antibiotic effective against tuberculosis.

The Science of the Art of Fermentation

Fermentation is a biochemical process in which complex organic molecules are broken down into smaller molecules. It is called a biochemical reaction because the reaction is catalyzed by enzymes produced by microorganisms. Catalyzed means the reaction speed is increased by the action of a substance that does not become part of the product of the reaction. Enzymes are naturally produced organic catalysts. Organic molecules all contain carbon and are generally large molecules. Until the mid-1800s scientists believed that only living organisms could produce these molecules, thus their name organic compounds.

Catalyzed reactions are described as stepwise reactions because they involve intermediate products. The intermediate products are called activated complexes. In some fermentations several steps are involved between the reacting molecules and the final products when more than one enzyme is involved in the reaction.

The microorganisms used in fermentations are specific to the particular fermentation reaction. For example, certain yeasts are used for the production of ethyl alcohol (commonly just called alcohol) while a specific bacteria is used for the production of the alcohol called butanol and a solvent called acetone that are produced together in one reaction. Not all microorganisms are useful for fermentation. Many microorganisms are involved in breaking down complex organic material into basic constituent molecules—they decompose the material entirely to waste products. In some such decomposition reactions the results are toxic.

The microorganisms used in fermentation are naturally found on plants, in the soil, or from natural fermentations. Pure cultures of useful microorganisms are now available commercially from specialized sources in many countries around the world. For such fermentation products as sourdough bread, a starter culture is continued from one batch of bread to another by the baker.

Fermentation cultures include bacteria, yeasts, and molds. It is not uncommon for more than one type of microorganism to be involved in a fermentation reaction. Fermentation processes produce alcoholic or acidic environments that tend to inhibit the growth of undesirable microorganisms. That is why fermentation is considered a good method to preserve certain foods. Even the microorganisms that produce the enzymes for the specific reaction have a limited tolerance for alcohol, or acid, and so the fermentation reactions are self-limiting. In the case of alcohol production the yeast's limit of tolerance is at about an alcohol concentration of 12 percent.

The growth and activity of microorganisms depend on the temperature, pH, oxygen concentration, moisture content, and nutrients available. The temperature range varies with the microorganism, but it is generally between 41–104°F (5° and 40°C). The pH measures acidity of the solution on a scale of 0–14, with neutral being 7 and the lower the number the more acidic the solution. Above a pH of 7 the solution is alkaline. Bacteria generally do best in near neutral mixtures. Yeasts grow in slightly acidic conditions, and molds tolerate a wide range of pH from acid to alkaline.

Oxygen presence is a significant factor. While oxygen is needed for all metabolic activities, including those of microorganisms, some fermentation reactions have to be conducted without air present and are called anaerobic fermentations; others need air present and are called aerobic fermentations. In anaerobic reactions the oxygen needed for microorganism metabolism is obtained from other compounds present in the reaction mixture. There are also some reactions that grow in reduced amounts of atmospheric oxygen. These microorganisms are described as microaerophilic organisms, meaning they need just a little air.

Yeasts and bacteria are very small unicellular non-green organisms. Being non-green they cannot produce their own food. Yeasts and bacteria are only visible with a microscope as they are on the order of 0.00002 inches (0.005 mm) in diameter. Yeasts are irregularly oval, but bacteria have diverse shapes, including rod shapes. Yeasts were the first microorganisms to be studied as they are used to make wine, beer, and bread. The alcoholic products wine and beer are made in an anaerobic process. Alcohol is fermented in an aerobic process by bacteria to make vinegar. The acid in vinegar is acetic acid. If pure alcohol is used to make acetic acid the flavors that make vinegar tasty are missing.

Bacteria give sourdough bread its special taste. What are known as lactic acid bacteria are used to ferment milks in a microaerophilic fermentation process. The lactic acid bacteria are a diverse group of bacteria that are useful in acid food fermentations. Industrial

production of lactic acid uses sugar from molasses, corn, or milk in a controlled fermentation process. Commercially-produced lactic acid is primarily used in foods, though a small amount is used in the production of other chemicals.

The fermentation of starch-containing grains using a particular bacteria strain to produce butyl alcohol and acetone was developed and patented by Russian-British chemist Chaim Weizmann (1874–1952). Acetone is a solvent used in the chemical industry and also in the manufacture of cordite. Weizmann's process became particularly important during World War I (1914–1918) because cordite is used as a propellant for cartridges and shells in wartime.

Yeasts and molds are both classified as fungi. Although molds are multi-cellular plants, they are microorganisms visible individually only with a microscope. They are also non-green but they may have other colors such as blue or black. Since they are non-green, they are also dependent on other organisms for food. Yeasts multiply by budding, but bacteria multiply by dividing. Molds grow by spreading filaments and reproduce by shedding spores into the air.

Antibiotics are mass produced in aerobic fermentation processes, using specifically cultured strains of microorganisms for maximum yield. Some molds are useful in fermentation processes to produce antibiotics and add flavor to cheeses. Others are helpful in industrial processes to produce citric and other specific organic acids. But the vast majority of molds are spoilers of foods and, when they are in the air, can cause allergic reactions for some people.

Influence on Science and Society

The impact of Pasteur's discovery of the part microorganisms play in the fermentation process in the 1800s brought a centuries-old debate about spontaneous generation of organisms in certain matter to a public conclusion. Spontaneous generation seemed to explain the sudden appearance of organisms where there were no obvious parents for them. Maggots in decaying matter were an example of organisms that did not appear to descend from any living organisms of the same kind. Although the relation between flies and maggots was determined before Pasteur's time, the discovery of microorganisms by van Leeuwenhoek opened up a new mystery.

French naturalist Félix A. Pouchet (1800–1872) was a leader of the proponents of spontaneous generation at the time Pasteur was describing to colleagues his meticulous experiments. Pasteur was certain that he had proved the concept of spontaneous generation was wrong. The debate on whether spontaneous generation explained the presence of microorganisms in fermentation mixtures, or whether the organisms only appeared if there were invisible organisms present in the environment surrounding a culture medium, was very heated.

Pouchet challenged Pasteur to prove in a public demonstration that no organisms would grow inside a heated, then sealed, flask containing a culture medium—as Pasteur insisted would be the case. If no organisms were found when the flask was opened, Pouchet said he would admit he was wrong. Pouchet had tried a similar experiment on his own and found there were microorganisms growing.

Pasteur took his experiment to the appointed meeting with the Academy of Sciences, but Pouchet did not appear. However, the witnesses to Pasteur's experiment all saw that he was right. There were no microorganisms in the flasks when they were opened. The scientists who witnessed Pasteur's success attested to his results, and the debate of spontaneous generation was finally settled. It was recognized that Pouchet's experiment had been flawed because he did not completely sterilize his flask.

In current times, the influence of the development of the science of fermentation processes has vastly improved the yield and quality of products. Some traditional fermentation processes are steeped in local culture but are disappearing from common use. The Food and Agricultural Organization (FAO) of the United Nations has publicized the need for governments to create a supportive environment to preserve the heritage and culture of traditional fermentation processes around the world.

FAO points out that the introduction of western-style foods is displacing traditional fermented foods that have been staples for thousands of years. The nutritional value of fermented foods has been very important, especially in regions where the diet would otherwise be lacking in essential vitamins or other nutrients. Lack of proper nourishment has historically been a problem in underdeveloped countries. In places where fermented foods have been traditional staples, the disease rate from poor nutrition is lower. FAO promotes improving native fermentation practices for traditional foods to be even more effective as diet supplements. At the same time, FAO is encouraging protection of the culture and traditions that make each region unique.

Advances in the efficiency of fermentation processes have had a positive influence on the industrial level preparation of chemicals. The use of fermentation is becoming more cost-effective and it can be a green process for manufacturing some chemicals. Greener means the process is more environmentally friendly, a very important consideration.

Modern Cultural Connections

The development of the science of fermentation has had a profound impact on society. One of the greatest societal impacts started with Weizmann, who discovered a fermentation process to develop acetone from maize (corn) in 1916. The availability of acetone was critical to the British government's efforts in World War I, as acetone was needed to supply the troops with ammunition.

During the war the First Lord of the Admiralty asked Winston Churchill (1874–1965), Prime Minister of the United Kingdom from 1940–1945, to supply thirty thousand tons of acetone. Weizmann succeeded in scaling up his small laboratory production process to an industrial scale, a feat that later gave him the unofficial title of father of industrial fermentation.

The Second World War (1939–1945) provided another story of the impact of the science of fermentation. The recently discovered life-saving antibiotic penicillin was needed in large quantities. The task to mass-produce such a drug was an enormous challenge. This was met with a deep-tank fermentation process that was developed at a major drug manufacturing company to produce the quantities of the antibiotic needed. The antibiotic penicillin saved many lives that would have been lost to infections from war-related injuries. The commercial scale development of many antibiotics can be traced to that discovery.

One of the most ancient of fermentation products, alcohol, has become the center of efforts to replace fossil fuels with renewable energy sources. Considerable research is being devoted to the development of enzymes that will improve the fermentation process to produce ethanol (pure alcohol) for fuel from a variety of biomass sources. Corn, as well as corn stalks and other waste vegetative materials, is being used to produce ethanol. Although all sources for fermentation are not yet economical, considerable research is being conducted to make the fermentation of waste plant products practical. There is even an effort by a New Zealand firm to make ethanol from carbon monoxide using a bacterial fermentation process. The carbon monoxide is a waste gas from a steel mill process. Carbon monoxide is a very simple carbon compound, but it is a source of carbon, and carbon is the key to the production of ethanol.

Fermentation processes use microorganisms to reduce large organic molecules to smaller molecules. The basic process is primarily associated with the production of a desirable product, but it can also be used to reduce large undesirable organic molecules to small harmless ones. Microorganisms have long been known to reduce all kinds of waste materials naturally. Bacterial strains now have been developed that can digest oil to clean up oil spills.

In 2001, scientists at a U.S. National Laboratory developed microorganisms that can clean up coal to make it cleaner burning as a fuel. The bacteria they use have been adapted from geothermal locations in the South Pacific and North America. Development through altering the traits of microorganisms can include genetic engineering.

Genetic engineering of organisms is opening up new frontiers in the science of fermentation. Genetic engineering means altering the inherited traits of an organism to make it even more desirable for a specific trait. Not only microorganisms are genetically engineered today; certain plant crops used in fermentation are being genetically engineered for greater productivity, disease and pest resistance, or other desirable traits—though not without controversy.

See Also Biomedicine and Health: Antibiotics and Antiseptics; Biomedicine and Health: Bacteriology; Biotechnology and the manipulation of Genes; Chemistry: Chemical Bonds; Chemistry: Fermentation: A Cultural Chemistry; Chemistry: Chemical Reactions and the Conservation of Mass and Energy; Chemistry: States of Matter: Solids, Liquids, Gases, and Plasma.

bibliography

Books

De Kruif, Paul. Microbe Hunters. New York: Harcourt, Brace & World, Inc., 1954.

Moore, F.J. A History of Chemistry. New York and London: McGraw-Hill Book Company, Inc., 1939.

Salzberg, Hugh W. From Caveman to Chemist. Washington, D.C., American Chemical Society, 1991.

Shreve, R. Norris. Chemical Process Industries. New York and London: McGraw-Hill Book Company, 1967.

Taylor, F. Sherwood. An Illustrated History of Science. London: William Heinemann, 1956.

Web Sites

Chemical Heritage Foundation. http://www.chemheritge.org (accessed August 17, 2007).

Food and Agricultural Organization of the United Nations. http://www.fao.org (accessed August 17, 2007).

Nobel Prize Organization. http://nobelprize.org (accessed August 17, 2007).

Miriam C. Nagel