Antibiotics_Industrial Microbiology.pptx

Antibiotics_Industrial Microbiology.pptx

• 1.
  Antibiotics Generation of IndustrialMicrobial Products
• 2.
  Introduction • Antibiotics arechemicals produced by microorganisms that in low concentrations can inhibit the growth of, or kill, other microorganisms. • Most antibiotics are secondary metabolites produced by filamentous fungi and bacteria, particularly the actinomycetes. • Higher plants and animals also produce anti-microbial substances. • Over 4000 antibiotics have been isolated from various organisms, but only about 50 are used regularly. • Different organisms may produce the same antibiotic or a single microorganism can produce different antibiotics. • The most well-known and medically important antibiotics are the β-lactams, including penicillins and cephalosporins, as well as aminoglycosides such as streptomycin, and the broad-spectrum tetracyclines.
• 3.
  Classification of Antibiotics •Based on chemical structure, antibiotics can be classified into 13 groups. • Here the most important groups are mentioned- Chemical group Name Producer organism Activity against Site or mode of action β - lactams Penicillin V Penicillium chrysogenum Gram (+) bacteria Wall synthesis Penicillin G Penicillium chrysogenum Gram (+) bacteria Cephalosporins Cephalosporium acremonium Broad spectrum Aminoglycosides Streptomycin Streptomyces griseus Gram (-) bacteria Protein synthesis Gentamycin Micromonospora purpurea Broad spectrum Neomycin Streptomyces fradiae Broad spectrum Tetracyclines Tetracycline Streptomyces species Broad spectrum Protein synthesis
• 4.
  Applications of Antibiotics •Mostly antibiotics are used in antimicrobial chemotherapy of humans and animals. • Antibiotics act as antitumor agents, for example: Actinomycin and mitomycin, produced by Streptomyces peucetius and S. caepitosus. • Useful for controlling microbial diseases of crop plants, or as tools in biochemistry and molecular biology research. • Several antibiotics are added to animal feed as growth promoters. For example: Penicillins and lincosamides, erythromycin and tetracyclines, etc.
• 5.
  Penicillin • Penicillin wasdiscovered by Sir Alexander Fleming in 1928 during World War Ⅱ, Penicillium notatum was responsible for penicillin production. • Penicillin is a beta-lactam antibiotic that has a four- membered lactam ring in its structure. The structure is termed as 6- aminopenicillanic acid ( 6-APA). • Penicillin inhibits the formation of the structure- conferring peptidoglycan of the bacterial cell wall. This component is absent in mammalian cells, so it has very low toxicity toward mammals. • Modification of the acyl group can provide new properties. Example of penicillin: Penicillin G (benzyl penicillin), Penicillin V (phenoxymethyl), methicillin, carbenicillin and ampicillin.
• 6.
  Production Steps ofPenicillin 1. Selection of microorganisms 2. Selection of raw materials 3. Preparation of inoculum 4. Fermentation process 5. Product recovery
• 7.
  1. Selection ofMicroorganisms • In the early days, Penicillium notatum was used for production but the yield was meager; 1 mg/L. • In 1943, Penicillium chrysogenum was discovered. In submerged culture, it gave a penicillin yield of up to 250 Oxford Units (1 Oxford Unit = 0.5988 of sodium benzyl penicillin) two to three times more than Penicillium notatum. • a ‘super strain’ Penicillium chrysogenum X 1612 was produced in 1953. After several ultraviolet irradiation treatments, new strains are created which can produce 50 g/L which is 50,000-fold higher than the initial strains. • Some other organisms- • Fungi: Aspergillus, Malbranchea, Cephalosporium, Emericellopsis, Paecilomyces, Trichophyton, Anixiopsis, Epidermophyton, Scopulariopsis, Spiroidium. • Bacteria: Actionomycete, Streptomyces.
• 8.
  Developmental Contributions toIncreasing Yields 1. Improvements in the composition of the medium. 2. Isolation of a better penicillin-producing mold species. 3. Development of submerged-culture technique, cultivation of the mold in large volumes of liquid medium through which sterile air is forced. 4. The production of mutant strains of P. chrysogenum, capable of producing large amounts of penicillin. A series of mutants created by exposure to X-ray and ultraviolet radiation resulted in strains with a remarkable capacity to produce penicillin. 5. The addition of chemicals to the medium served as precursors for the synthesis of penicillin. 6. Refinements in methods of recovering penicillin from the fermentation mixture.
• 9.
  2. Selection ofRaw Materials 1. Carbon source: Glucose, lactose, sucrose, beet molasses, ethanol, and vegetable oils. Glucose suppresses secondary metabolism and excess of it limits penicillin production. 2. Nitrogen source: Corn steep liquor, cotton seed, peanut, linseed, or soybean meals. Calcium carbonate and a phosphate buffer are added to neutralize the acidic medium. 3. Minerals like sulfur compounds are sometimes added for additional yields since penicillin contains sulfur. 4. Precursors of the appropriate side-chain are also added to the fermentation. For example, if benzyl penicillin is desired, phenylacetic acid is added.
• 10.
  3. Preparation ofInoculum • The aim is to develop a pure inoculum in sufficient volume. • Two types of inoculum 1. Seed stage inoculum: Lyophilized spores are added to a small fermenter at a concentration of 5⨉103 spores/ml. Fungal mycelium may then be grown through one or two further stages until it is sufficient to inoculate the production fermenter. 2. Production stage culture: Fungal mycelium is directly inoculated in a fermenter. Liquid medium is used.
• 11.
  4. Fermentation process •Penicillin production is usually via a fed-batch process carried out aseptically in stirred tank fermenters of 40000–200000 L capacity. • The fermentation of penicillin can be divided into three phases- • The first phase (trophophase) during which rapid growth occurs, lasts for about 2 days during which mycelia are produced. • The second phase (idiophase) lasts for 5 to 7 days; penicillin is produced. • In the third phase, carbon and nitrogen sources are depleted, antibiotic production ceases, and the mycelia lyses release ammonia and cause an increase in pH. • Oxygen is maintained at 25-60 mmol/L/h. • Temperature is maintained at 25 – 27°C • pH- 6.5 - 7.7
• 12.
  5. Product Recovery •Antibiotic recovery is often by solvent extraction of the cell-free medium, and this process gives a yield of up to 90%. 1. The fermentation broth is filtered with a rotary vacuum filter to remove mycelia and other solids, and pH is adjusted to 2 using sulfuric or phosphoric acid, followed by extraction at 0 – 3°C using amyl acetate or butyl acetate. Then the aqueous phase is removed using centrifugation. 2. The organic solvent containing the penicillin is then passed through charcoal to remove impurities, then it is extracted again with a 2% phosphate buffer at pH 7.5. 3. Repetition of the first step. 4. Then the product is transferred into smaller volumes of organic solvent during each extraction, concentrating the penicillin up to 80-100 times.
• 13.
  5. When itis sufficiently concentrated the penicillin may be converted to a stable salt form in one of several ways which employ the fact that penicillin is an acid: (a) It can be reacted with a calcium carbonate slurry to give the calcium salt which may be filtered, lyophilized, or spray-dried. (b) It may be reacted with sodium or potassium buffers to give the salts of these metals which can also be frozen or spray dried; (c) It can be precipitated with an organic base such as triethylamine. • The production of most other antibiotics follows a similar plan. The major differences relate to the organism, composition of the medium, and method of extraction.
• 14.
  Overview
• 15.
  Thank you!