Chemistry: Fermentation: A Cultural Chemistry

Introduction

The term fermentation stems from the Latin fervere, “to boil.” It originally referred to a substance whose properties changed through a bubbling or foaming process, like leavening bread or brewing beer. Although fermentation processes have been used by humans for at least 10,000 years, the chemistry behind them was not understood until the nineteenth century, when French chemist and microbiologist Louis Pasteur (1822–1895) proved that fermentation was caused by microorganisms.

Microbes like bacteria and fungi digest glucose, a 6-carbon sugar, and split it into two 3-carbon molecules called pyruvic acid. (This process, called glycolysis, is an anaerobic process, one that takes place without oxygen.) This releases electrons and generates adenosine triphosphate (ATP), an energy storage molecule that powers the cell. Depending on the microorganism involved, the production of pyruvic acid also releases ethyl alcohol, butyl alcohol, and lactic or citric acids. Alcohol is used to brew beer and preserve food; lactic acid is used to make cheese and yogurt. Fermentation also produces carbon dioxide, which makes the foam in beer, the bubbles in some wines, and the air that makes bread rise.

Historical Background and Scientific Foundations

The Ancient World

Fermentation and human history are intricately tied together. Our earliest historical records indicate that humans unknowingly domesticated wild yeasts to ferment food and drink, particularly Saccharomyces cerevisiae for wine production, Saccharomyces carlsbergensis for brewing beer, and “leaven”—most likely a mixture of milk bacteria (lactobacilli) and yeasts—for bread baking. Bakers saved a portion of leaven from each batch of dough as a “starter” for the next one. Each starter has a distinctive flavor, depending on the type and quality of wild yeast in the air. Some strains of yeast, such as those that give sourdough bread its distinctive taste, are cultivated commercially today.

Since grains and fruits ferment naturally, it is likely that the earliest civilizations knew how to make alcoholic beverages. Wild yeasts, for example, collect on the skin of grapes and other fruits, causing fermentation. Ancient pottery shards show the presence of tartrates, a byproduct of fermentation, indicating that the Chinese were brewing a drink made of rice, honey, and fruit as early as 7000 BC. Centuries later, they developed a process to break down complex sugars in rice to simple sugars like glucose, which were then fermented to make rice wine. The first stage in this process is carried out by a mold called Aspergillus oryzae followed by a sake strain of S. cerevisiae.

Wine was also being produced 8,000 years ago in Caucasus Mountain settlements, as well as in ancient Iran. Sumerians in ancient Babylonia (modern Turkey) considered beer a divine drink; they worshipped Ninkasi, the goddess of beer and alcohol, composing a hymn to her 4,000 years ago. In the Gilgamesh epic (c. 3000 BC), Gilgamesh's boon companion, the bestial Enkidu, was “civilized” by learning to eat bread and beer, the two main products of Babylonian fermentation:

They placed food in front of him,

They placed beer in front of him;

Enkidu knew nothing about eating bread for food,

And of drinking beer he had not been taught.

The harlot spoke to Enkidu, saying:

“Eat the food, Enkidu, it is the way one lives.

Drink the beer, as is the custom of the land.”

Enkidu ate the food until he was sated,

He drank the beer—seven jugs!—and became expansive and sang with joy!

He was elated and his face glowed.

He splashed his shaggy body with water,

And rubbed himself with oil, and turned into a human…

The Babylonian ruler Hammurabi (d. 1750 BC), author of the oldest set of written laws, dictated beer rations for each social class, a normal worker receiving two liters per day.

Ancient Egyptian tomb paintings dating from 2400 BC show how they crushed barley and mixed it with water to make dried cakes. When reconstituted with water, an extract of the cakes was fermented in vessels to make a type of beer. Egyptian records from the same period also refer to the use of grapes for viniculture.

Wine Making in Antiquity

Brewing techniques eventually moved east to the Mediterranean, where wild grapes grew in abundance. The ancient Greeks and Etruscans considered wine an essential part of their diet, planting vineyards in their colonies near the Black Sea and in France and Spain. So revered was wine that the Greek religious cult of Bacchus saw drunkenness as a type of connection with the divine. Libations poured on tombs to honor the deceased

involved a mixture of olive oil and wine. Carthage, an ancient Phoenician civilization in North Africa, developed such advanced viniculture that books were written about it; the city became wealthy exporting wine to the nascent Roman republic.

After the Romans conquered Carthage in 241 BC, the Roman Senate decreed that Carthaginian wine treatises be translated into Latin. The writer and politician Cato the Elder (234–149 BC) wrote an agricultural treatise named De Agri Cultura that incorporated Carthaginian techniques. The Romans planted vineyards as far north as Germany in steep river valleys such as the Mosel and Ahr. The valleys protected the grapes from the wind and kept them warm, and the perpendicular sunshine hit the ripening vines with greater heat, producing sweet juice that is better fermented by yeast. Wine was exported to Roman provinces and was particularly popular when sweetened with honey as a beverage called mulsum. Because distillation had not yet been discovered, wine was the drink with the highest alcohol content; it was often diluted with water.

Archaeological ruins at Pompeii and other ancient sites provide evidence of the Roman wine-making process. After the grapes were harvested, they were crushed to produce the first premium pressing of juice. The resulting grape mash was then put through a simple wine press for a second extraction used to make cheaper wine. The final residue from the pressing was mixed with water and given to the farm laborers.

The juice that resulted from pressing, known as must, then fermented either outside or in pitch-covered clay jars that were buried underground. (Modern wineries use stainless steel vats, adding sugar and yeast rather than relying on wild strains from the atmosphere). As Romans liked sweet wines, grapes were often left to ripen on the vine as long as possible until the first frosts of autumn, so the sugar could be concentrated, a technique still used today. Sometimes the must was boiled to concentrate sugars, or spices were added for flavor or to mask a sour vinegary wine. Wines were stored in two-handled tapered clay jugs called amphora, which typically litter most Roman archaeological sites. To extend the wine's life (though most was drunk within the year to prevent spoilage), the Romans also stored it in oak barrels to age the wine and give it added flavor.

The History of Beer Brewing

In colder parts of the Roman Empire where grape cultivation was difficult, beer brewing predominated, usually by making successive extracts from a batch of brown malt.

Malting is the process of immersing barley (or wheat) in water from 55–60°F (12.7–15.5°C) for 40 to 50 hours; its volume increases and the grain begins to sprout, causing its root sheaths, called chits, to break through the husk. As the barley germinates, the seed embryo secretes a hormone called gibberellic acid, which initiates the production of an enzyme called alpha-amylase; this in turn both converts the barley starch into fermentable sugars that the embryo can use as food and produces carbon dioxide. Large amounts of malting barley have to be turned and aerated to dissipate the gases and heat emitted during germination.

The malt is then dried to remove most of the moisture in a process called kilning or roasting, which reduces enzyme activity. Successively wetting and kilning malts imparts different colors and flavors to the finished beer. A high proportion of chocolate malt (a dark-brown roast) mixed with roasted and ungerminated barley is used to make stouts and porters.

Next the malt is mixed with hot water (144–162° F/62–72°C)—and sometimes other grains—in a process called mashing. This completes the conversion of starch into fermentable sugar. The aqueous part of the mixture, called wort, is separated, filtered, and then boiled for 60 to 90 minutes. The excess grains can be used for cattle feed.

By the eighth century brewers had discovered that hops can be added to the wort to impart a tangy, slightly bitter flavor. The female flowers of hop plants contain a chemical called humulone that gives bitters and pilsners their characteristic flavor. Interestingly, recent research has revealed that humulone is an angiogenesis inhibitor—a substance that prevents the growth of new blood vessels. Since cancer tumors can only grow and spread if they make new blood supplies, humulone may be useful in chemotherapy, and there is scientific speculation that beer consumption may help prevent cancer, specifically prostate cancer.

The wort is then filtered again, cooled, and stored in a collecting tank where yeast is added to begin fermentation. Before brewers understood yeast's role in fermentation, a successful brew hinged on the chance of attracting wild yeasts. A bad brew was often blamed on the influence of magic or witches.

In traditional brewing, a first fermentation took a week for ale and more than three weeks to make a lager. A secondary fermentation, in which extra sugar or actively fermenting wort was added, purged the beer of unwanted bitter compounds, a process that could take from a week to three months.

Ancient and medieval brewers waited for the yeast to rise to the top of the brewing vessel, then skimmed it off in a process called top fermentation. The first extract was the strongest, giving the best beer, and the third extract was the poorest, producing what was called small beer. Top-fermented beers, typical of British beers, were known as ales. Another fermenting process, in which the yeast sank to the bottom of the brewing vat, produced lager, a beer that become popular in Germany. As brewing was primarily a winter occupation, the term comes from the German lagern “to store.” Though in the past the yeast that was skimmed off or sank was discarded, modern companies in England and Australia concentrate it into a salty extract called Marmite or Vegemite that is used as a sandwich spread and nutritional supplement, as the yeast provides a high quantity of B vitamins.

After the Roman Empire fell in the fifth century AD, brewing technologies were preserved in Christian monasteries. Monks had originally started making their own sacramental wine and brewed beer as a nutritional alternative to meals. This was important during their many fasts, since drinking liquid was not considered to break the fast. The excess was sold to raise money for the monastery.

Some religious houses, in fact, made brewing a big business: eleven out of twelve monastic houses in medieval Yorkshire, for example, had brew houses on their premises. It is not only likely that monks discovered the advantages of adding hops to beer, but medieval monasteries such as those in St. Gall in Switzerland also added secret proprietary mixtures of herbs to their brews to widen their public appeal. The beer industry became so important in Germany that the first food purity laws, the Reinheitsgebot of 1516, regulated beer purity, requiring that it only contained water, malted barley, malted wheat, and hops.

Other Domestic Fermentation

Outside of the monasteries, brewing, like baking, was primarily a female occupation. Because of this, taverns and alehouses were often run by female alewives; the labor shortage that followed the Black Plague (1347) only increased this trend. Women in the medieval era also used other fermentation techniques to pickle food and to make cheese and yogurt.

In these processes, salting to control the growth of the microbes is combined with fermentation to pickle food. When cucumbers are pickled, for example, microbes turn carbohydrates into acid, and their color changes from bright to olive green. To stop microbial activity, pickles are stored in a brine solution of 8–10% for the first week, with the salt concentration increased by 1% a week until it stands at 16%. The pickles, then cured, have the excess salt leached from them by soaking them in warm water for 10–14 hours.

Cheese-making dates back to ancient Babylonia, and it is likely its production was accidental, as milk was transported in the stomachs of young animals slaughtered for food. Lactobacilli in the milk combined with the sun's warmth permitted the milk sugars to ferment. The acid in the stomach lining, called rennet, caused milk to curdle. Swaying the stomach bag broke up the solid curds, producing the liquid whey. As the curds set in a soft gel, the cheese was drained, salted, and packaged to make soft cheese. For hard cheese, the cheese would be baked to further drain the whey and make it more solid. Molds were used to drive out water and shape the final product. As cheese ages, microbes (usually lactobacilli) typically break down the milk protein (called casein) into amino acids and reduce milk fats into simpler fatty acids; both molecules impart flavor and texture.

Yogurt, or fermented milk, was made simply by allowing milk to heat in the sun. The milk's sugar was fermented by natural lactobacillus to produce lactic acid, which reacted with the milk protein to thicken and solidify the milk.

Though in ancient Egypt the secret of cheese-making was kept by temple priests, in ancient Greece and Rome cheese was part of the daily diet, as it was a way to store otherwise perishable milk year-round. In the Middle Ages, monks in North Yorkshire abbeys made Wensleydale cheese from ewe's milk; cheddar cheese was invented near England's Cheddar Gorge in 1500.

Louis Pasteur Discovers the Fermentation Process

Though fermentation was used throughout history to make a variety of foodstuffs, the biological and chemical mechanisms behind it were not revealed until the nineteenth century. No one knew, for example, why beer and wine would suddenly sour during production. And, as the Industrial Revolution commercialized beer and wine production on a vast scale, a “bad batch” could mean great financial losses.

Louis Pasteur, dean of sciences at the University of Lille, was asked for help by an industrialist who was producing beer from beet sugar. The batches continually were going sour. By examining the brewer's samples under the microscope, Pasteur realized that correctly aged beer contained spherical yeast, but sour batches contained yeasts that were elongated. Pasteur also noted that the desirable globe-shaped yeasts produced alcohol, but the rod-shaped yeasts produced lactic acid, making the batches go sour. These findings made him realize 1) yeasts were responsible for fermentation, 2) different types of yeasts made different byproducts, and 3) yeasts did not need oxygen to metabolize. Anaerobic respiration (without oxygen) to this day is sometimes referred to as the “Pasteur effect.”

Pasteur advised the industrialist that after the “good” yeasts did their work and fermented to produce alcohol, the liquid be heated gently to 122° F (50° C) to kill the “bad yeast” and prevent souring as the beer aged. This process, now called pasteurization, is used today in milk, cheese, yogurt, beer, and wine production. In 1876 Pasteur published the results of his findings, the Études sur la Bière (Studies on beer). His work led to a scientific understanding of the brewing and winemaking processes. Pasteur modestly wrote in the preface of his book:

I need not hazard any prediction concerning the advantages likely to accrue to the brewing industry from the adoption of such a process of brewing as my study of the subject has enabled me to devise, and from an application of the novel facts upon which this process is founded. Time is the best appraiser of scientific work, and I am not unaware that an industrial discovery rarely produces all its fruits in the hands of its first inventor.

From his research, Pasteur discovered that sour batches could also be caused by microbes other than yeast, such as molds or bacteria. Realizing that each type of fermentation was the result of a specific type of microbe, he applied this conclusion to disease, suggesting diseases were also microbe or “germ”—specific. This insight is known as the germ theory of disease, and Pasteur went on to develop vaccines for anthrax, chicken cholera, and rabies based on this principle.

Modern Cultural Connections

After Pasteur made his discoveries, production of the industrial solvents butyl and ethyl alcohol became the most important industrial fermentations. In the 1960s, however, chemical engineers devised nonbiological ways to produce alcohol and acetone that relied on synthetic starting materials derived from then-cheap and plentiful petroleum. Growing concerns in the 1980s about petroleum conservation led to renewed interest in producing alcohols from microbial fermentation, using cellulose and starches from agricultural wastes that would otherwise go to a landfill. Fermentations using microbes also do not produce toxic byproducts, as do chemical processes.

Ethanol, fuel for automobiles produced by microbial fermentation of cornstarch, has also become a topic of current scientific and political interest. In 1999 Brazil mandated that by 2003 all new cars in the country had to run on E85, a fuel that was 85% ethyl alcohol and 15% gasoline. American, Japanese, and European manufacturers began to produce cars that were E85 compatible as well.

Molecular biologists, in turn, have genetically engineered bacteria, yeasts, and even some mammalian cells to produce ethyl alcohol more efficiently via fermentation in giant vessels called bioreactors. The bioreactors provide appropriate nutrients, stirrers, temperature, and pH (level of acidity or alkalinity). Genetically modified yeasts are also being used in modern brewing to shorten the time it takes to ferment the wort. Transgenic barleys have been developed with genes that encode for certain desired enzymes like alpha-amylase.

Industrial fermentation with fungi such as yeasts is also used to produce a large variety of other commercial products as well. Other products of industrial mycology include modern antibiotics like penicillin or streptomycin; food coatings, such as pullulan, a complex sugar produced from glucose by yeast; and vitamin B₂ from brewer's yeast. Citric acid, used in foods, detergent, and pharamaceutical applications, is produced industrially from Aspergillus niger. From bread and beer to fuel, humans have depended on microbial fermentation to create a wide variety of products necessary to their well-being.

See Also Chemistry: Biochemistry: The Chemistry of Life; Chemistry: Fermentation;

bibliography

Books

Bennett, Judith M. Ale, Beer, and Brewsters in England: Women's Work in a Changing World, 1300–1600. New York: Oxford University Press, 1996.

Bigelis, Ramunas. “Fungal Fermentation: Industrial.” In Encyclopedia of Life Sciences. New York: John Wiley and Sons, Ltd., 2001.

Epic of Gilgamesh. Translated by Maureen Gallery Kovacs. Stanford: Stanford University Press, 1990.

Johnson, Hugh. Vintage: The Story of Wine. New York: Simon & Schuster, 1989.

Pasteur, Louis. Louis Pasteur's Studies on Fermentation: The Diseases of Beer, Their Causes, and the Means of Preventing Them. MacMillan & Co., 1879. Reprint Edition, BeerBooks.com, 2005.

Sandler, Merton, and Roger Pinder. Wine: A Scientific Exploration. London: Taylor & Francis, 2002.

Stansbury, P.F., A. Whitaker, and S.J. Hall. Principles of Fermentation Technology. New York: Butterworth-Heinemann, 1997.

Periodicals

Brown, William. “Description of the Buildings of Twelve Small Yorkshire Priories at the Reformation.” Yorkshire Archaeological Journal ix (1886), 197–215.

Linko, Matti, Auli Haikara, Anneli Ritala, and Merja Penttil. “Recent Advances in the Malting and Brewing Industry.” Journal of Biotechnology 65, nos. 2–3 (October 27, 1998): 85–98.

Pausch, Mark H., Donald R. Kirsch, and Sanford J. Silverman. “Saccharomyces cerevisiae: Applications.” In Encyclopedia of Life Sciences. New York: John Wiley and Sons, Ltd. (2001).

Rossiter, J.J. “Wine and Oil Processing at Roman Farms in Italy.” Phoenix 35, no. 4. (Winter 1981): 345–361.

Shimamura M., et al. “Inhibition of Angiogenesis by Humulone, a Bitter Acid from Beer Hop.” Biochemical and Biophysical Research Communications 289, no. 1 (November 23, 2001): 220–224.

Web Sites

Encyclopedia Romana. “Wine and Rome.” http://penelope.uchicago.edu/̃grout/encyclopaedia_romana/wine/wine.html (accessed October 22, 2007).

National Health Museum: Access Excellence. “Microbial Fermentations: Changed the Course of Human History.” http://www.accessexcellence.org/LC/SS/ferm_background.html (accessed October 22, 2007).

New Scientist.com. “World's Earliest Tipple Discovered in China.” http://www.newscientist.com/article/dn6759.html accessed October 22, 2007).

Anna Marie Eleanor Roos