Biotechnology Lecture 5: Products of Fermentation (University of Baghdad)

University of Baghdad Biotechnology4th stage College of Science ‫شيماء فاضل‬.‫د‬+‫ والء شوكت‬.‫ د‬+ ‫محمد لفته‬.‫د‬ Department of Biology Lecture 5: Productsof Fermentation Major Groups of Commercial Fermentation Products: 1- Microbial...

University of Baghdad Biotechnology4th stage College of Science ‫شيماء فاضل‬.‫د‬+‫ والء شوكت‬.‫ د‬+ ‫محمد لفته‬.‫د‬ Department of Biology Lecture 5: Productsof Fermentation Major Groups of Commercial Fermentation Products: 1- Microbial biomass or cells. 2- Microbial enzymes. 3- Microbial metabolites. 4- Bioconversion or Biotransformation. Microbial Biomass or cells Microbial biomass or cells may be subdivided into two major processes: a) Production of baker’s yeast by Saccharomyces cerevisiae. b) Production of microbial cells used as food for human or animal (Single- cell protein/SCP) which are in fact either: (i) whole cells of Spirullina (as algae), (ii) Candida or Saccharomyces (as yeast) and (iii) Lactobacillus (as bacteria).). Microbial Enzymes Enzyme have been produced commercially from plant, animal and microbial sources. However, microbial enzymes have enormous advantage of being able to be produced in large quantities by establishing fermentation techniques. Table below contains microbial enzymes used in production of commercial fermentation industries. Industry Enzyme Source (Genus) Baking, Flavours Amylase Aspergillus , Bacillus Beer, Protease Aspergillus, Bacillus Laundry detergents Lipase Aspergillus, Rhizopus, Bacillus Dairy Catalase Aspergillus,, Corynebacterium, Micrococcus Lactase (β- Aspergillus galactosidase) Pharmaceutical & Amylase Bacillus Clinical Streptokinase Heamolytic Streptococci Fruit Juice Pectinase Aspergillus , Penicillium Microbial Metabolites 1 Metabolites are the intermediates and products of metabolism. Metabolism is the sum of all the biochemical reactions carried out by an organism. Metabolism involves two pathways: a) Primary metabolic pathways (PMPs). Their products are called primary or central metabolites b) Secondary metabolic pathways (SMPs). Their products are called Secondary metabolites 1. A primary or central metabolites:  Theyare microbial products made during thetrophophase(exponential phase) of growth whose synthesis is an integral part of the normal growth process.  They include intermediates and end products of anabolic metabolism, which are used by the cell as building blocks for essential macromolecules (e.g., amino acids, nucleotides) or are converted to coenzymes (e.g., vitamins). Other primary metabolites (e.g., citric acid, acetic acid and ethanol) result from catabolic metabolism.  Industrially, the most important primary metabolites are amino acids, nucleotides, vitamins, solvents and organic acids. 2. Secondary metabolites:  They do not play a role in growth and are formed during the end or near theidiophase(stationary phase) of growth.  Usually has an important ecological function.  Many Secondary metabolites have antimicrobial activity, others are specific enzyme inhibitors, some are growth promoters and many have pharmacological properties.  Examples; antibiotics, pesticides, pigments, toxins. Examples on Primary Metabolites Organic Acid Production  Organic acids can be used both as: 1. Additives in the food industry. 2. Chemical feedstock.  Except for the production of citric acid which is manufactured entirely by fermentation, there is frequently great competition between microbiological & chemical processes for production of the various organic acids. Citric acid (C6H8O7) 2   Citric acid has been known as a natural plant substance since the end of the nineteenth century.  Since 1893 scientists have known that it is produced by filamentous fungi.  In 1923 the first practical microbial fermentation for the production of this organic acid was started.  Today over 99% of the world's output of citric acid is microbial produced.  The strains that are used for citric acid production are: Aspergillusniger, A.wentii ,A.clavatus ,Penicilliumluteum ,P. citrinum ,MucorpiriformiandSaccharomycopsislipolytica.  During the last 30 years the interest of researchers has been attracted by the use of yeasts (mostly Candida spp. and some Rhodotorula spp.) as citric acid producers.C. lipolytica has been developed as a microbial cell factory for citric acid production in recent years.  Compared to Penicillium strains, only mutants of Aspergillusniger are used for commercial production, Why? 1. Aspergilla produce more citric acid per unite time. 2. Production of the undesirable side products can be suppressed in these mutants. Uses of Citric acid: 1. As a food additive/preservative found in many different processed foods and soft drinks. 2. As an ingredient in cosmetic products to balance the pH levels, small amounts of citric acid can be found in shampoos, body wash, face cleansers, nail polish, hand soap and other cosmetics products. 3. As a powerful cleaning agent, it works well as both a cleaner and a deodorizer. 4. As a powerful water softener, it an ideal all-natural choice for treating hard water. 5. Citric acid is widely used as an acidulent in creams, gels, and liquids of all kinds. Amino Acid Production  Taste – enhancing properties of glutamic acid were discovered in 1908 in Japan.  Commercial production of sodium glutamate from acid hydrolysates of wheat & soy protein began soon after.  In 1957, L-glutamic acid was discovered as a product in the spent medium of Corynebacteriumglutamicum& this organism subsequently became the major source of sodium glutamate. 3 Commercial Uses of Amino Acids: 1. Food Industry: Amino acids are used either alone or in combination:  asflavor enhancers.  as an antioxidant for the preservation of fruit juices.  as a low-calorie artificial sweetener in soft-drink industry.  in the preparation of feed for animals. 2. Pharmaceutical Industry: The amino acids can be used as medicines. Essential amino acids are useful as ingredients of infusion fluids, for administration to patients in post-operative treatment. 3. Chemical Industry: Amino acids serve as starting materials for producing several compounds. For example:  Glycine is used as a precursor for the synthesis of glyphosate (a herbicide).  Poly-methyl glutamate is utilized for manufacturing synthetic leather.  Some amino acids are useful for the preparation of cosmetics.  With all theseapplications amino acids are on its way into the synthesis of biodegradable polymers. MicrobiologicalMethods of Production: There are three approaches to microbiological production: 1- Direct Fermentation of amino acids using different carbon sources, such as glucose, fructose, molasses, starch, hydrolysis …etc. 2- By converting inexpensive intermediate products via biosynthesis for example glycine which is inexpensive, can be converted to L-serine. 3- By the use of enzymes or immobilized cells, sometimes in continuous processes involving enzymes-membrane reactors. Glutamic Acid:  L-glutamic acid is manufactured predominantly by microbial means.  Japanese researchers began developing a direct fermentation process because the D,L-glutamic acid which is formed by chemical synthesis is the racemic mixture.  The most important industrial strains with high excretion of glutamic acid are Micrococcus glutamicus&Brevibacteriumflavum. There are similar, Gram-positive, nonsporulating, non-motile bacteria. Vitamins  Vitamins are defined as essential micronutrients that are not synthesized by mammals. 4   Most vitamins are essential for the metabolism of all living organisms, and they are synthesized by microorganisms and plants.  They are 2 types: Water-Soluble and fat -Soluble Vitamins.  Most vitamins and related compounds are now industrially produced and microorganisms can be successfully used for the commercial production of many of them.  Vitamins and related compounds are widely used as food or feed additives, medical or therapeutic agents, health aids, cosmetic and so on. Riboflavin (Vitamin B2)  It is a water soluble vitamin, essential for growth and reproduction in man and animals.  75% of the current world production of riboflavin is for feed additive and the remaining for food and pharmaceuticals.The crude concentrated form is also used for feed.  It is produced by both synthetic and fermentation processes.  Two closely related ascomycete fungi, Eremotheciumashbyii and Ashbyagossypii, are mainly used for the industrial production.  Yeasts (Candida flaeri,C. famata, etc.) and bacteria can also be used for the practical production. Example on Secondary Metabolites (Antibiotics)  Antibiotics are secondary metabolites produced by one type of microorganism that in low concentrations act against other organisms.  Antibiotics are elaborated by bacteria (predominantly by Actinomycetes in the genus Streptomyces) as well as fungi. For example, Penicilliumchrysogenum(a mold) produces penicillins; Streptomyces species (bacteria) produce streptomycins and tetracyclines.  Over 500 distinct antibiotic substances have been shown to be produced by streptomycetes. Penicillin  Penicillin, produced by Penicilliumchrysogenum, is an excellent example of a fermentation for which careful adjustment of the medium composition is used to achieve maximum yields.  Rapid production of cells, which can occur when high levels of glucose are used as a carbon source, does not lead to maximum antibiotic yields.  Provision of the slowly hydrolyzed disaccharide lactose, in combination with limited nitrogen availability, stimulates a greater accumulation of penicillin after growth has stopped. 5   The same result can be achieved by using a slow continuous feed of glucose. If particular penicillin is needed, the specific precursor is added to the medium. For example, phenylacetic acid is added to maximize production of penicillin G, which has a benzyl side chain.  Using genetic engineering techniques increased penicillin production up to 30-fold. Bioconversion or Biotransformationor Microbial Transformations  Microbial transformation is defined as the biological process of modifying an organic compound into a reversible product.  It involves the use of chemically defined enzyme catalyzed reactions in the living cells and it preferred over chemical transformation in industries. Bioconversion differs from chemical conversion in:  highly specificity  needing to low temperature  don't need to use the heavy metals  milder reaction condition Microbial transformation reactions are mainly categorized into oxidation/ reduction, hydrolysis, condensation & isomerization reactions (Fig. below).  One of the major applications of microbial transformation is in the production of secondary metabolites. Examples on biotransformation: 6  The industrial production of cortisone. One step is the bioconversion of progesterone to 11-alpha- Hydroxyprogesterone by Rhizopusnigricans.  The conversion of organic materials, such as plant or animal waste, into usable products or energy sources. 7