Biology
Biology
A lot of the food we eat is alive. Fresh uncooked fruits and vegetables are made of cells that are still living and breathing long after they are picked. But cheese, yogurt, wine, beer, pickles, sauerkraut, olives, and many other foods are produced by the action of living microbes, and in many of the foods we eat, these microbes are still alive, doing their job.
If you think for a moment about how difficult it is to keep something sterile, you will realize that just about everything has something living in or on it. Most bacteria are benign, as are most yeasts, and there may be more beneficial varieties than there are harmful ones.
The yeasts and bacteria in a sourdough starter come from the flour used to make the starter (very little yeast is actually floating in the air). Bees carry yeasts from flower to flower, where they feed on the nectar. When the fruit sets, the yeast grow in the skin of the fruit, where sugars and moisture are available. The powdery coating on grapes is yeast. Crushing the grapes allows the yeast to mix with the juices and ferment them into wine. No added yeast is actually necessary, but is usually added to overwhelm any natural yeasts, and produce a reproducible product.
In most foods that are produced or modified by microorganisms, we adjust the environment to suit the organisms we desire, and to make undesirable organisms have a harder time. In pickles, for example, we add salt to allow lactic acid producing bacteria to outcompete harmful varieties that compete for the sugars in the cucumber. The bacteria then help out further by making the environment too acidic for other life forms. Too much salt, however, allows harmful yeasts to grow, which eat the lactic acid, and thus invite even more undesirable things to grow.
Controlling other factors in the environment, such as temperature, moisture, and oxygen, also help select which of the organisms in the food you are promoting.
Yeast
For thousands of years, people made bread with yeast, not realizing that they were farming microorganisms. That a living thing was responsible for the rising of bread was only recognized after the invention of the microscope.
Under a microscope, we see that yeasts are single cells, like bacteria. Unlike bacteria, however, yeast reproduce by growing smaller yeast cells as buds on the parent cell. In some varieties of yeast, the buds do not detach, and long strings of cells form as the new buds grow to full size and bud new yeast cells themselves.
The yeasts we use in bread, beer, and wine eat sugar. There is little sugar in flour, but uncooked flour has enzymes that break down the starch molecules in the flour into small sugar molecules that the yeast can feed on. The enzymes start to work as soon as water is added to the flour.
In wine, the sugars of the grape feed the yeast. In beer, some of the grain is allowed to sprout, and the sprouting seedling produces large amounts of the enzymes that convert starch into sugars. The sprouts are ground and added to the other grains in the vat, and the yeast feeds on the sugars that are produced by the action of the enzymes.
As the yeast feeds on the sugars, two waste products are excreted. One is alcohol, and the other is carbon dioxide. In beer, both are desirable. In wine, the carbon dioxide is often allowed to escape into the air before the wine is bottled. When a carbonated product is desired, extra sugar is often added just before corking the bottle, so the yeast that remains can produce more of the gas.
In bread, it is the carbon dioxide we are after. Alcohol is still produced, and a measurable amount of alcohol remains in the bread even after an hour of cooking at high temperature. This alcohol may help to dissolve aromatic molecules, and enhance the scent and flavor of the bread, but we generally are only concerned with the amount of gas the yeast produces, to make the dough into a nice light foam.
In modern bread recipes, commercially prepared yeast is generally used. The large amount of yeast that is added to the dough ensures that this one strain of yeast outcompetes all other organisms in the dough, and the flavor of the bread is not affected by uncontrolled bacteria or wild yeast.
Yeast also produces enzymes such as transglutaminase that change the behavior of the gluten in the dough. The dough becomes less extensible as more of the enzyme is produced, and more cross linking happens in the glutenin proteins. This strengthens the dough.
Yeast lives on small sugars like glucose, sucrose, and maltose. The wheat kernel contains enzymes that break down the starch (its stored energy source) into maltose and glucose, which can then be used directly by the growing wheat sprout. Grinding the kernel into flour and wetting the flour allows the enzyme to break down the starch, so the yeast has a food supply.
The yeast has its own enzymes, among them maltase, which breaks down maltose into two glucose molecules. As the yeast feeds on the glucose, it produces not only carbon dioxide and alcohol, but many other molecules (aldehydes, ketones, aromatic heavy alcohols and other metabolic byproducts) that add flavor to bread, beer, and wine.
Sourdough
Stir some water into flour and let it sit for a day. The microbes in the flour will begin to grow. Feed it more flour and water every day, and after a week the ecology in your jar will stabilize, with a mix of yeasts and bacteria that we call sourdough starter.
In practice, feeding a starter involves throwing half of it away when we add more flour and water. There is not very much food in flour that the microbes can actually eat, so throwing away the “used up” part of the mixture keeps it from diluting the new food with what the microbes consider waste matter.
After a week has gone by, and we have a nice stable culture, we no longer waste any starter. Instead, we make bread with it, and feed some of the resulting bread dough back to the starter jar.
Regular baker’s yeast will not survive long in the acid environment of a sourdough starter. The variety of wild yeasts that come to live in harmony with the acid making bacteria in the culture are of a different type, such as Candida milleri, Candida krusei, Pichia saitoi, and Saccharomyces exiguus. Along with these acid tolerant yeasts grow acid producing bacteria such as Lactobacillus sanfranciscensis, a species originally found in sourdough starter from San Francisco, but later found to be in many sourdough starters all over the world.
The bacteria are smaller than the yeasts, and outnumber them 100 to 1 in most sourdough cultures. Candida milleri yeast can’t eat the sugar maltose, but the bacteria can. In this way, they can coexist without competing with one another too much, even though both will eat glucose. The bacteria produce lactic and acetic acids, and another antibiotic called cycloheximide that kill off many of the competing and potentially harmful organisms, but leave the symbiotic yeasts and bacteria unharmed.
The bacteria also excrete glucose as they metabolize maltose, making the glucose available to the yeasts. Maltose is made of two glucose molecules bound together, and when the bacteria break them apart, some of the glucose escapes into the culture medium.
There are many different species and strains (varieties within a species) of bacteria besides Lactobacillus sanfranciscensis. Other Lactobacillus species often found in starter cultures are plantarum, pentosus, rossi, pontis, acidophilus, delbrueckii, homohiochii, hilgardii, viridescens, panis, pastorianus, oris, vaginalis, reuteri, buchneri, fructivorans, salivarius, brevis, fermentum, casei, and paraplantarum. When present in bread, these bacteria act to inhibit mold, and thus serve as preservatives.
The bacteria require more than just sugars to survive, and what they don’t get from the flour, they get from dead yeast cells. So each of the symbionts in the partnership produce something the other needs.
Of course there are many other yeasts and bacteria growing in the culture in smaller numbers. Some grow there despite the non-optimal growing conditions of acidity and antibiotic substances produced by the main colonists in the culture.
A cook might want to control the growth of each of the two types of microorganism. The bacteria provide flavors that the cook might want to make more robust or more subtle, and the yeast provides the main source of carbon dioxide to make the bread lighter in texture. Knowing what growing conditions each type of microbe grows best in allows the cook to trade one off in favor of the other.
Candida milleri (the yeast) is tolerant of a wide range of acidity. The bacteria (Lactobacillus sanfranciscensis) grows best at a pH of 5.5, and grows much less well below 4.5 or above 6.5. Adding vinegar or baking soda to the starter can select against the bacteria, while allowing the yeast to grow (the cook would want to actually measure the pH to make sure it was in the desired range). The bacteria generally grow faster than the yeast at pH levels above 4.5.
The bacteria grow fastest at a temperature of about 90° F (32° C), while the yeast grows fastest at about 80° F (27° C). Since the yeast produces undesirable flavors when growing rapidly, a temperature above or below the optimum for speed of growth is considered optimum for bread flavor. So, if the cook wants to encourage the bacteria, a temperature of 90° F might be worth a try, and if the yeast is to be encouraged instead, a temperature of 70° to 75° F might be sought. It might even be interesting to grow the starter at 90° F to get the flavors from the bacteria, and then proof the actual bread dough at 75° F to grow the yeast for faster rise times. The starter might be left to grow for a day unattended, but the waiting for the bread to rise might be less convenient.
The yeast is more tolerant of salt than the bacteria. The yeast will grow in salt up to a concentration of 8%, but the bacteria die out at 4%. Using that much salt in a bread recipe would not be advised, but since the starter will be diluted 5 or 6 to 1, this might be a place where experimentation can be done. Adding small amounts of salt has been shown to stimulate yeast growth in sourdoughs, possibly by eliminating some competition from the bacteria for nutrients they both require.
The yeast grows less as the alcohol content rises. But the bacteria are for the most part unaffected until the alcohol rises to 6% (but the growth falls off sharply to zero in both organisms at 8%).
The yeast is strongly affected by acetic acid (which the bacteria produce from lactic acid in anaerobic conditions), while the bacteria are more tolerant. Both are fairly tolerant of lactic acid.
The reason these two organisms grow well in symbiosis is that they have very similar growth response at temperatures between 68° F and 82° F (20° to 28° C), and pH levels around 4 or 5. When growing a sourdough starter (especially a new one), keeping the conditions in this range will select for a stable symbiotic mix, and select against harmful organisms. Once the starter is well established, short term deviations from these conditions to promote one type over the other can be successful.
When a starter is colonized by bacteria like Enterococcus faecalis, and Pediococcus pentosaceus, some proteolytic (protein breaking) enzymes are produced. These harm the dough by breaking down the gluten that we need to trap the gas bubbles. The Lactobacillus bacteria do this as well, but to a much lesser extent. Nonetheless, selected strains of Lactobacillus have been used to break down the gluten in wheat flours to make it palatable to people with celiac disease, a condition where the immune system is activated by gluten and attacks the lining of the intestine.
The breakup of the gluten protein is not completely harmful to the dough, however. Some of the breakdown products are important to the flavor and aroma of sourdough bread, and as the bacteria break down the protein to free the nitrogen they need for food, they convert some of it into essential amino acids normally missing from wheat flour, such as lysine. They thus improve the nutritional value of the bread to a small extent.
Yogurt
Adding Lactobacillus acidophilus bacteria to warm milk makes yogurt. The bacteria eat the sugar lactose in the milk, and produce lactic acid. The lactic acid denatures the casein proteins in the milk, allowing them to unwind and bind together, forming a gel.
The bacteria compete with harmful bacteria that would otherwise cause the milk to spoil. The acidity of the yogurt prevents some of the harmful organisms from growing well enough to be a problem.
Raw milk can harbor a number of dangerous microbes, the most common of which are Staphylococcus aureus, Campylobacter jejuni, Salmonella, Escherichia coli, Listeria monocytogenes, Mycobacterium tuberculosis, Mycobacterium bovis, Brucella, Coxiella burnetii, and Yersinia enterocolitica.
Staphylococcus aureus grows best at neutral pH (7.0 to 7.5) but can survive in milk as acid as 4.5 pH. The pH of yogurt is just below that level (4.25 to 4.5). Likewise, Salmonella, Campylobacter jejuni, Escherichia coli, Listeria monocytogenes, and Mycobacterium tuberculosis all grow poorly or not at all at pH levels more acid than 4.5. Note, however, that these are not all of the dangerous organisms that can be found in raw milk, and pasteurization is a far better protective mechanism than culturing Lactobacillus.
Sour Cream and Cultured Buttermilk
Lactobacillus is not the only bacterium used to sour dairy products. Both buttermilk and sour cream contain Streptococcus diacetilactis, Streptococcus lactis, Streptococcus cremoris, and Leuconostoc citrovorum along with other less common bacteria.
Both buttermilk and sour cream keep for weeks in the refrigerator, and are easy to make at home. If you have a container of milk that is getting old (but has not already turned sour), you can turn it into buttermilk for baking instead of waiting for it to sour and throwing it away.
The starter culture for buttermilk is some leftover buttermilk, since all of the necessary bacteria are already in the leftovers.
Make sure you have a very clean one quart container. Put a cup of cultured buttermilk that has not passed its “use by” date (since we want the bacteria to be alive) into the container, and fill it up with the milk. If making more or less, keep the 1 cup to 3 cups ratio for best results.
Cover the container tightly, and leave in a warm place for 24 hours. It should have thickened in that time, but if not, you can leave it for up to 36 hours before it no longer tastes good enough to drink (it is still good for baking, however).
The liquid left over from making butter is not the same as cultured buttermilk. It is not thick, and it often has flakes of butter in it. In the U.S. it is referred to as “old fashioned buttermilk”. Some cultured buttermilk has bits of butter added to mimic old fashioned buttermilk. But only true cultured buttermilk will work well as a starter for the buttermilk we need in baking, or for buttermilk pancakes.
Making your own sour cream is as easy as making your own buttermilk. Add a small amount of buttermilk to some cream, and let it sit in a warm place for a day. The higher the fat content of the cream, the thicker the sour cream will be.
Bleu Cheese
When Alexander Fleming discovered penicillin on September 3rd, 1928, it may have been more than just his untidy laboratory that deserves the credit. It may have been his lunch.
He had been studying staphylococci, a disease causing bacteria, and had stacked his cultures in a corner of his lab when he went on vacation for the month of August. On returning September 3rd, he noticed that one culture was contaminated with a fungus, and that the bacteria around the fungus had been killed.
It took another 10 years before chemists at Oxford were able to produce a stable form of the antibiotic, and five more years before industrial methods were developed to produce it in usable quantities.
But Penicillium molds had been used as antibiotics since ancient times, long before bacteria were known to be the cause of many diseases. In 1870, it was noted that cultures with mold in them would not produce bacteria. John Tyndall demonstrated to the Royal Society in 1875 that Penicillium fungus had antibacterial properties. In 1877 Louis Pasteur observed that anthrax was inhibited in cultures that had been contaminated by molds. In 1897 Ernest Duchesne (using a different species of Penicillium than that used to make penicillin) cured guinea pigs of typhoid.
Fleming isolated Penicillium notatum as the species that produced the antibiotic, and later work identified strains of that species that produced even more of the substance. Duchesne used Penicillium glaucum, the species used to make Gorgonzola cheese. That species, along with Penicillium roqueforti, is also used to make bleu, Roquefort, and Stilton cheeses.
Penicillium is a common bread mold, and is the main cause of spoilage in stored grains. Like many of the other organisms we have been discussing, it produces chemicals that inhibit or kill competing organisms. Unlike the alcohol or lactic acids produced by yeasts and bacteria in wine and yogurt, the chemicals produced by the mold is not a waste product that has side benefits, but has evolved because it gives the mold a strong advantage in competing for food.
In cheese, it not only provides the characteristic flavor and aroma of bleu cheese, but it also protects the cheese from spoilage from more harmful bacteria and fungi that would either produce undesirable flavors and odors, or produce toxins that would make the food unfit for consumption.
Alexander Fleming was studying staphylococcus cultures to find something that would kill them, so his rediscovery of the benefits of the mold came when he was well prepared to make use of the information. But it may have been the bread or cheese from his lunch, left in the lab for a month while he was on vacation, that should actually get the credit. We’ll never know.
Roquefort cheese contains the highest levels of glutamates of any naturally produced food. Glutamates are what give savory protein-rich foods their taste, and are found in other fermented foods, such as soy sauce. Purified as monosodium glutamate and used as a flavor enhancer, it is sometimes overused, and people with a sensitivity to it complain. But it is found in almost all foods that contain protein, and our tongues have specialized sensors for it, to help us find nutritious protein rich additions to our diet.
You can make your own cheese fairly easily these days, thanks to the availability of rennet tablets in the supermarket. Adding a little bit of bleu cheese or Roquefort to the curds before pressing the cheese can be a tasty way to experiment with microbial processing and the use of antibiotics to preserve foods.
Wine and beer
Yeast is on the outside of grapes, and on the outside of grains like barley, wheat, and rice. So it is no surprise that grape juice or wet grains eventually ferment. Beer and wine have been around since before agriculture, for more reasons than just to provide a buzz in the head.
As we have seen with other foods made with microorganisms, creating a friendly environment for one organism can make that same environment less friendly to other, potentially harmful organisms. The alcohol that yeast provides helps to preserve the wine and beer, so those calories and nutrients can be saved for a day when food is less available.
As people began to live in more concentrated populations, sanitary drinking water was not always available. Septic systems and sewage treatment plants were not available, and the local water supply was not always fit to drink. But wine and beer, even with low alcohol content, was generally safe to drink. Either the water came from the grape, or it was boiled with the grains, and in either case the germs that cause typhus, diphtheria, dysentery, and other water-borne diseases were not present.
There are between 700 and 900 calories in a liter of wine, and 360 or so in a liter of beer. So besides being safer to drink than the local water, they provided energy and some amount of nutrition.
Beer has been called “liquid bread”, and weak beers with low alcohol content could be consumed without compromising mental faculties or losing bodily fluids from the diuretic effects of alcohol.
Making beer and wine is simple. Making good beer or wine is an art. Like most of the foods made with microorganisms, much of the science is involved with making sure we encourage the organisms we want, and discourage those we don’t want.
Sterilization helps, so that there are no harmful organisms to start out. Controlling the temperature also helps to make a consistent brew. Paying attention to the nutrient requirements of the desired organism, and limiting the nutrition of the undesired ones will help to keep the ecology of the brew in balance. Knowing or controlling the mineral content of the water used (in the case of beer) affects both the taste of the resulting beer, and the health and vigor of the yeasts that make it. Making sure the brew gets enough oxygen helps to keep the yeast healthy and to prevent harmful anaerobic bacteria from growing. Brewers carefully control when aeration occurs. The yeast multiply when there is enough oxygen, and produce alcohol and carbon dioxide when there is less. Aeration when the temperature is too high causes bad flavors to develop.
Controlling the temperature helps to promote yeast growth and inhibit bacterial growth. The wort (the liquid that becomes beer after fermentation) is boiled, but then quickly cooled, since undesired bacteria grow well at temperatures between 130°F and 90°F (54°C to 32°C). Aeration is begun when the temperature is below 80°F (26°C) to prevent oxidation while still providing oxygen for the yeast to multiply. Rapid cooling also prevents sulfur containing compounds from re-dissolving in the wort, causing a cooked cabbage taste.
In beer making, there are two main types of yeast – top fermenting, and bottom fermenting. Top fermenting yeasts (called ale yeasts) prefer warmer temperatures, and go dormant below about 55°F (12°C). Bottom fermenting yeasts (called lager yeasts) stay active down to about 40°F (4°C). While lagers are usually brewed at lower temperatures than ales, steam beer uses lager yeasts at the higher temperatures, to get a different flavor.
To keep foreign organisms out of the beer as it ferments, the gas is allowed to escape through a tube to bubble up through water. This is called an air-lock.
As with other foods produced by microorganisms, it helps to start with a large number of the desired organisms, to overwhelm any of the undesirable ones. In the case of beer and wine, yeast is often added rather than relying on the natural yeast on the grapes or on the grain.
Brewers use a device called a hydrometer to measure the density of the liquid. This indicates how much sugar is dissolved in the wort, and thus how the fermentation is progressing (as the yeast consumes the sugar and produces less dense alcohol).
To make sparkling wine, or control the carbonation in beer, sugar is sometimes added just before bottling. This allows the small amount of remaining yeast to produce carbon dioxide, which cannot escape the bottle. The pressure rises, and the gas dissolves in the water.
Preserving
Many of the processed foods we enjoy were invented as ways of preserving fresh foods for consumption later. Dried fruits, cheese, salted meats, fermented beverages, yogurt, pickles – all are ways of extending the useful storage time of fresh fruits, grains, meats, and dairy products.
Salt and drying
Life needs water to survive and grow. Eliminating water prevents harmful organisms from consuming the food and spoiling it. Many fresh fruits and meats can be simply dried in the sun to preserve them. Fresh cut grass is dried to store as livestock feed.
Salting food to preserve it is another way of preventing harmful organisms from the water they need to survive and thrive. If the outside environment is saltier than the inside of an organism, the water diffuses out to dilute the salt. The organism dries out, even if there is lots of salty water outside.
We salt meat and fish more than fruits because the sugar in the fruit has the same osmotic effect on spoilage organisms as salt does, once the fruit has dried enough to concentrate the sugar a bit. Fish and meats spoil quickly in the sun without some salt to prevent bacteria, molds and yeasts from getting a foothold before the drying has progressed enough to preserve the food.
Heat sterilization and smoking
Another way to keep organisms from getting started is to keep the temperature high enough to kill them or prevent growth. This can be done while drying the food, or in the case of canning, it can be done while the food is sealed to prevent organisms from getting into the packaging after the food has been sterilized.
Smoking preserves food by heating, drying, and sealing the outside of the food with substances that do not promote the growth of spoilage organisms. Smoking foods also controls oxidation to some extent, which is another way that food becomes spoiled.
Since the smoke does not penetrate very far into the food, the main preserving effect comes from the salting, drying, and heating of the food during the smoking process.
Alcohol sterilization
We have seen how alcohol helps to preserve grains and fruit juices when they are fermented, but alcohol is also sometimes used to preserve other foods. In brandied fruit, alcohol is added to chopped fruits that are then allowed to ferment. The alcohol prevents bacteria from competing with the fermentation yeasts in the early stages of the process. Once the process has been started, new fruit can be added as the brandied fruit is eaten, without needing to add extra alcohol. In this way, the original brandied fruit acts like the starter used in making sourdough bread.
Antimicrobials in spices
It is no accident that the use of spices in food correlates with the average temperature of the region from which the recipe is derived. Temperatures that promote the growth of bacterial and fungal spoilage organisms are prevalent in the countries where foods are heavily spiced.
The essential oils in sage, mint, hyssop, and chamomile have bacteriostatic effects (growth prevention) on Gram negative bacteria such as E. coli, and Gram positive bacteria such as Listeria innocua. The essential oils from oregano have bactericidal (killing) effects that are most pronounced on Gram negative bacteria.
Strong antimicrobial activity is found in cinnamon, cloves, and mustard. Less pronounced effects in allspice, bay leaf, caraway, coriander, cumin, thyme and rosemary are similar to those of sage and oregano. Black pepper, red pepper, and ginger have only weak activity.
Garlic and onion are effective against salmonella, E. coli, staphylococcus, and Candida albicans.
Cinnamon, cloves, and sage all contain eugenol, an effective antimicrobial and mold inhibitor that is used in mouthwashes and baked goods. Sage and oregano contain thymol, another effective antimicrobial, often used in toothpastes.
The oleoresins in rosemary also act as antioxidants, and prevent rancidity of fats and oils in foods.
Acids
We have seen how making foods more acid helps to preserve them in the case of yogurt and sourdough bread. But raising the acidity much higher, by pickling foods in vinegar, is an ancient and effective way of preserving foods from olives and cucumbers to boiled eggs and herring.
Besides yogurt and sourdough bread, other foods are pickled by fermentation that produces lactic acid. The most recognizable is probably pickled cucumbers or peppers. But sauerkraut and kimchi are also fermented in this way, after adding salt to make a brine.
Microbial competition
We also use friendly microbes to help fight harmful ones. Some produce toxic chemicals like alcohol or lactic acid, but others produce more targeted antibacterial agents. The penicillin molds in some cheeses kill bacteria. Bacteria in other cheeses produce propionates that kill molds and fungi.