21st Century Guidebook to Fungi, SECOND EDITION, by David Moore, Geoffrey D. Robson and Anthony P. J. Trinci

Table of Contents

1. Chapter 16: Fungi as pathogens of animals, including man
2. Chapter 17: Whole organism biotechnology
   1. Fungal fermentations in submerged liquid cultures
   2. Culturing fungi
   3. Oxygen demand and supply
   4. Fermenter engineering
   5. Fungal growth in liquid cultures
   6. Fermenter growth kinetics
   7. Growth yield
   8. Stationary phase
   9. Growth as pellets
   10. Beyond the batch culture
   11. Chemostats and turbidostats
   12. Uses of submerged fermentations
   13. Alcoholic fermentations
   14. Citric acid biotechnology
   15. Penicillin and other pharmaceuticals
   16. Enzymes for fabric conditioning and processing, and food processing
   17. Steroids and use of fungi to make chemical transformations
   18. The Quorn fermentation and evolution in fermenters
   19. Production of spores and other inocula
   20. Natural digestive fermentations in herbivores
   21. Solid state fermentations
   22. Digestion of lignocellulosic residues
   23. Bread; the other side of the alcoholic fermentation equation
   24. Cheese and salami manufacture
   25. Soy sauce, tempeh and other food products
   26. Chapter 17 References and further reading
   27. Buy a PDF of Chapter 17
3. Chapter 18: Molecular biotechnology

17.23 Bread; the other side of the alcoholic fermentation equation

Leavened bread is another product of the fermentation of sugars from cereal grains by Saccharomyces cerevisiae. Known, not surprisingly, as Baker’s yeast in this industry (though in the old days supplied to bakers by brewers), the yeast still uses the same chemistry of the ethanol fermentation we have summarised before in this equation:

C₆H₁₂O₆ → 2 C₂H₅OH + 2 CO₂

one hexose molecule is converted into two ethanol molecules and two molecules of carbon dioxide. For the brewer the key product is ethanol; for the baker the key product is the carbon dioxide. In summary, the fermentation process in bread dough depends on the breakdown of the starches in flour producing carbon dioxide which makes bubbles in the dough stabilised by gluten proteins from the flour; this causes the dough to expand. A small amount of alcohol is also produced, but this evaporates when the bread is baked. As with alcoholic fermentations, the process depends on the ability of S. cerevisiae to preferentially use fermentation even in the presence of oxygen, given appropriate nutrients.

Those ‘appropriate nutrients’ are contained in the culture medium for the Baker’s yeast, which of course is the bread dough; so perhaps we should look at a bread recipe. There are hundreds of bread recipes: white, brown, wholemeal and country French bread, baguettes, Bath buns, Chelsea buns, Scotch baps, barm cakes, granary bread, and so on, and so on (see, for example http://www.cookitsimply.com/category-0020-0e1.html; Hutkins, 2006; Montet & Ray, 2015). The ingredients for a basic white bread recipe are yeast, sugar, water, milk, white flour, salt and butter.

The yeast, sugar, water and milk are mixed and put in a warm place until fermentation makes it frothy, which only takes about 10 minutes; this foaming fermenting mixture is called barm and many years ago it was the practice to take the barm from beer fermentations to leaven bread. The flour and salt are mixed with the butter and the yeast liquid is added and the whole mixed (kneaded) to smooth dough. The dough is left to rise in a warm place until it has doubled in size; this takes 60 to 90 minutes. The dough is then kneaded again for 5 minutes, then shaped into a baking tin and left for the second rising (called proving) for about 1 hour. Finally, the loaf is baked in an oven at 230°C for 30-40 minutes.

So let’s look at the biology of this process. After all the ingredients are mixed together, forming the dough, this doubles in volume in 60 to 90 minutes in a warm place (remember, this is a bakery we’re talking about, so warm places are not hard to find).

The dough does this because the yeast fermentation produces carbon dioxide and because the wheat flour contains the protein mixture known as gluten. Gluten is a combination of the proteins gliadin and glutenin, which are stored in the endosperm of wheat, rye, and barley seeds to nourish the germinating plants along with starch. Together, gliadin and glutenin comprise about 80% of the protein contained in wheat seeds. The gluten they form is an important source of nutrition in foods, including bread, prepared from these cereal grains. But it is crucial to the structure of leavened bread which depends on the sticky and viscous properties of the gluten protein in the dough. Gliadin is a glycoprotein and forms a three-dimensional network with glutenin by inter- and intra-molecular sulfur cross linkages which develop in the dough during the kneading process.

Subsequently, the carbon dioxide gas produced during fermentation is trapped into bubbles by the gluten and as more and more bubbles are formed in the dough they make it ‘rise’. The second kneading process redistributes the bubbles and forms more disulphide cross links, helping more carbon dioxide gas bubbles to form during the proving stage. After this final fermentation period the dough is cooked, the alcohol evaporates and the bubbly structure of the dough is fixed into the open structure of bread.

The first records of any sort of bread are in ancient Egyptian hieroglyphs from over 5000 years ago that show bakeries with dough rising next to bread ovens. Wine-making, brewing and baking occurred alongside one another in ancient Egypt. An Egyptian wooden model dating from 2000 BC of a combined brewery and bakery can be seen at the British Museum and we can presume (on the basis of optical and scanning electron microscopy of desiccated bread loaves and beer remains preserved in tombs from about 2000 to 1200 BC; Samuel, 1996) that bread was made with flour from raw grain and also with malt and yeast suggesting that fermenting brewing liquor, what we now call barm, was mixed with bread dough for the first leavened bread.

The exact nature of yeast was unknown until Louis Pasteur, in 1859, demonstrated that wine yeast is a living organism and that only active living cells can cause fermentation. There seems always to have been a direct relation between brewing and baking. The Faculty of Medicine in 17th Century Paris debated for months whether bakers should be allowed to use beer barm for their bread. They eventually decided to ban the practice but the bakers didn’t take much notice and continued to use barm for the fine light bread that the upper classes liked so much. British cookery books included recipes and instructions for brewing as well as baking as a matter of course until the 19th century; beer-making being the only dependable source of baking yeast. Wine-making barm is too bitter to be used in baking. Even beer barm needs to be washed to reduce the bitterness of hops used in brewing ale (Montet & Ray, 2015).

However, there are different requirements for the yeasts in the two industries. The baker needs yeast that tolerates higher temperatures with no particular preference for alcohol production; the brewer needs yeast that tolerates and produces high alcohol concentrations with no particular preference for performance at higher temperatures (and strains used for dried yeast production (see below) must be tolerant to the drying process) (Hutkins, 2006; Gibson et al., 2007; Ali et al., 2012; Montet & Ray, 2015).

Today, reliable and highly specialised yeasts are produced commercially and marketed around the world as dried or compressed preparations. Commercial yeast production starts with a pure culture tube or frozen vial of the appropriate yeast strain. This sample is the inoculum for the first of a series of progressively larger cultures that amplify the volume of the yeast suspension. The early stages are grown as batch fermentations, using a medium comprised of molasses, phosphate, ammonia and minerals; later stages are conducted as fed-batch fermentations during which molasses and other nutrients are fed to the yeast at a rate that maximises yeast multiplication but prevents the production of alcohol. From the last of these seed fermentations the vessel contents are pumped to separators that separate the yeast from the spent molasses. After being washed with cold water the yeast cream is held at 1 to 2°C until being used to inoculate the final commercial fermentation tanks.

These commercial fermenters have working volumes up to 250 m³. Water is first pumped into the fermenter, then this is pitched with yeast cream from the seed store. Aeration (sparging with about one fermenter volume of air per minute), cooling (through an external heat exchanger to maintain culture temperature at 30°C) and nutrient additions initiate a 15-20 hour fermentation. At the start of the fermentation, the culture occupies 30% to 50% of the fermenter volume. The unit is operated as a fed-batch and additions of nutrients at increasing rate during the course of fermentation (to support growth of the increasing cell population) and maintenance of pH in the range of 4.5-5.5 bring the fermenter to its final volume. The number of yeast cells increases about 5 to 8 times during this fermentation.

At the end of fermentation, yeast is separated from the fermenter broth by centrifuges, washed with water and re-centrifuged to make a yeast cream with a solids concentration of approximately 18%. The yeast cream is cooled to about 7°C and stored in refrigerated stainless steel tanks. Cream yeast can be delivered to customers directly with tanker trucks. Alternatively, the yeast cream can be pumped to a filter press and sufficient water removed to produce a press cake having 30-32% yeast solids content. The press cake yeast can be crumbled into pieces and bagged for distribution to customers in refrigerated trucks. Harvesting and processing wet yeast can take several hours and chilling is needed to store the resultant yeast products; cream and compressed yeast products have to be used within 10 to 28 days, respectively.

Dried yeast manufacture involves drying the yeast with evaporative cooling on fluidised beds (in which air passing through crumbled press cake yeast forms a bed-of-air, which suspends and tumbles the product to dry it). Spray drying is also effective in preserving viable yeast, but spray drying is a high energy-demand operation (in practice requiring 7,500 to 10,000 J g^-1 of evaporated water) so it needs to be optimised carefully (Luna-Solano et al., 2005). The final dried product is vacuum packed and can be stored at normal room temperatures for months to years.

Updated July, 2019