FERMENTATION Classical Biotechnology Humans have been using this technology for centuries Involves harnessing the wastes of bacteria and/or yeast for products that humans consume Big business

ENERGY transfer between enzymes, other molecules Cellular Respiration: the process of using glucose to make energy (ATP) for the cell. Aerobic Cellular Respiration (requires oxygen) enzymes 6O2 + C6H12O6 6CO2 + 6H2O oxygen glucose carbon dioxide water ENERGY transfer between enzymes, other molecules ADP + Pi ATP

Aerobic Cellular Respiration takes place in the mitochondria of cells. It can provide up to 38 molecules of ATP per molecule of glucose.

Anaerobic Cellular Respiration (without oxygen) also known as Fermentation Alcoholic Fermentation: C6H12O6 glucose 2CO2 + 2C2H5OH carbon dioxide ethanol provides 2 molecules ATP per glucose done by yeast

Products of Alcoholic Fermentation don’t drink alcohol

Making Rootbeer: 6 simple steps Heat Water to ~40 degrees C (yeast like it) Add sugar and dissolve Add root beer extract Add yeast Ferment @ ~27 degrees C Chill and enjoy!

Lactic Acid Fermentation: Glucose carbon dioxide + lactic acid provides 2 molecules ATP per glucose done by muscle cells done by bacteria cells

Products of Lactic Acid Fermentation

Other products of fermentation - some are fermented by both yeast and bacteria Idli, Dosas, Kimchee, Sausage, Kefir, sauerkraut, miso, tempeh, tamari, chutney

More products of fermentation

Yogurt Yogurt-like products have been made for millenia across Eastern Europe, North Africa, Central Asia and India. Contains bacteria that are “thermophilic” = heat loving

Two main types of Lactic Acid Bacteria (Identified around the year 1900): Lactobacillus meaning “milk” and “rod” over 50 different species found on plants and in the digestive system of animals such as cows and humans. Lactococcus meaning “milk” and “sphere” because of its shape found primarily on plants less common than lactobacillus Traditional spontaneously fermented milks contain species that can reside in the human digestive tract: Lactobacillus fermentum, L. casei L brevis L plantarum (from picked vegetables) L acidophilus

Commercial Yogurt Contains 2 species of bacteria specialized to grow well in milk (but can’t survive inside the human body): First, Streptococcus thermophilus is more active, then slows down when acidity reaches 0.5% Next, Lactobacillus bulgaricus is more acid tolerant and takes over until acidity >1% These bacteria work in symbiosis. Each bacterium stimulates the growth of the other => acidifies the milk more rapidly than either partner on its own.

Milk is Water Protein (casein and whey) Fat Sugar (lactose) Vitamins Minerals

How Does Milk Turn Into Yogurt? Lactic acid Bacteria -------------------------> Lactose Lactic Acid (Milk sugar) Acid causes casein (milk protein) to denature and hold water into a semi-solid gel = yogurt

Milk Yogurt Bacteria produce acid Fat globule Casein protein micelles (bundles) 10-7 meters in diameter Acid causes Casein bundles to fall apart into separate casein molecules. These rebind to each other in a network that traps water. => makes a gel Fat globule

Making Yogurt in 4 Simple Steps 1. Start with Cow, Sheep, or Goat milk. 2. Heat milk to 80 °C. Two purposes: destroy existing bacteria “condition” the proteins = begins the denaturing process (a whey protein molecule binds to a casein molecule which disrupts the casein bundles allowing them to make short branched micelle chains) 3. Cool milk to 40 °C and innoculate with bacteria 4. Incubate at 30 °C to 45 °C Casein after heat pre-treatment: Casein before heat pre-treatment: Casein after acid:

Incubation Temperature 40-45 °C takes 2-3 hours Produces a coarse protein network with thick strands give firmness but easily leak whey (a process called syneresis - the separation of liquid from the gel) 30 °C takes 18 hours Produces a finer more branched delicate network that holds the liquid whey