Fermentation notes for M.Pharm pharmaceutical chemistry (Fermentation)).pptx

M.Pharm pharmaceutical chemistry notes

1. FERMENTATION Vitamins BY A. SUREGA M.PHARM FIRST YEAR department of pharmaceutical chemistry

2. INTRODUCTION Fermentation may be defined as the process of growing a culture of microorganism in a nutrient media and thereby converting feed into a desired end product. It is the process for the production of products by the mass culture of microorganisms.

3. Types of fermentation Fermentation can be broadly classified into 2 types 1. aerobic fermentation 2. anaerobic fermentation Aerobic fermentation Aerobic means “presence of oxygen” Here the fermentation process takes place in the presence of oxygen oxygen is required for the reproduction and growth of microorganisms

4. Anaerobic fermentation Anaerobic means in the absence of oxygen It is the process of chemical breakdown of organic substrates into ethanol or lactic acid by microorganisms. Occurs in yeast, parasites and bacteria Occurs in cytoplasm. GLYCOLYSIS Before any of the anaerobic fermentation takes place a process is called as glycolysis It is the conversion of complex organic compounds like glucose into simpler 3 carbon containing compound pyruvic acid. 2 molecules of ATP are produced. 4

5. Major fermentation products GROUP PRODUCTS PRODUCING MICRO-ORGANISM Antibiotics penicillin streptomycin Penicillium chrysogenum Streptomyces griseus Enzymes amylases proteases Aspergillus oryzae bacillus species vitamins riboflavin vitamin B 12 Ashbya gossypii Pseudomonas dentrificans

6. VITAMINS Vitamins are organic substances, that are needed in trace quantities, for the growth and normal metabolic functioning of most forms of life. vitamins can be obtained from natural sources. These include, 1.animal products eg fish ox or calf liver oil, eggs, etc 2.vegetable products eg cabbage, carrot, orange, lemon, corn (or) maize oil, and corn(or)maize bran,etc . 3.micro-organisms eg brewers yeast, bakers yeast, etc

7. Among many vitamins, i.e , thiamine, riboflavin, biotin, folic acid, pantothenic acid, pyridoxine and vitamin B12 produced microbiologically, processes for only riboflavin and vitamin B12 were commercialized. In fact, vitamin b12 is the only vitamin produced by fermentation. For the commercial production of riboflavin, chemical or semisynthetic procedures are feasible.

8. Structure of vitamin b2(riboflavin)

9. Vitamin b2 Riboflavin is an important vitamin required by man and animals . Kuhn, Gyorgy and Wagner Jauregg in 1933 isolated riboflavin from whey of milk. It was first isolated from milk and then prepared by fermentation in 1935. It is a water soluble, yellow-orange crystalline powder . It is also present in other foods as flavoproteins which contain the prosthetic group flavin mononucleotide(FMN) or flavin adenine dinucleotide(FAD). Chemically, riboflavin is an alloxazine derivative which consists of a pteridine ring condensed to a benzene ring. Mostly it is absorbed in small intestine .

10. The side chain consists of a c5-polyhydroxyl group. Riboflavin is prepared by a fermentation process as well as by a synthetic method. It is produced by a number of micro-organisms including BACTERIA YEAST YEAST LIKE Clostridium butyricum Clostridium acetobutylicum Clostridium felsineum Mycobacterium smegmatis Aerobacter aerogenes Anasco sporogenous Candida arborea Candida flareri Candida utilis Asco sporogenous Hansenula suaveolens Saccharomyces species Ashbya gossypii Eremothecium ashbyii

11. Production by ashbya gossypii Ashbya gossypii is the causative organism of a disease of cotton bolls (especially in south Africa ) which produces rooting and staining. it may also cause infection to beans, citrus, fruits, coffee, okra and tomatoes. Since the organism is highly pathogenic, it is essential to sterilize all fermentation residues and cultures before they are discarded.

12. Nutrient requirement carbon source sucrose maltose nitrogen source peptone animal stick liquor protein source corn-steep liquor For optimum growth of micro-organism biotin thiamin meso-inositol

13. Stock culture Asbya gossypii may be transferred at weekly intervals on a medium containing, peptone : 0.5 % yeast extract : 0.3 % malt extract : 0.3 % commercial glucose : 1.0 % agar : 2.0 % incubation temperature : 27-30 ° c Inoculum development pfeifer et al (1950) described a method for preparing an inoculum for pilot-plant fermentation. A 24hr old ashbya gossypii NRRL Y-1056 placed in 100ml of the following medium in a 500 ml flask and incubated for 24hrs on a reciprocating shaker at 26 to 30 °c. glucose : 2.0 % peptone : 0.5 % corn-steep liquor : 1.0 % water : to 100 ml

14. Fermentation process The hyphae of Ashbya gossypii can produce large amount of riboflavin . Carbon sources, nitrogen sources and other nutrients are used to produce maximum yield of riboflavin. Medium is prepared according to the type of microorganisms . Inoculum is prepared from slants or spores of particular cultures. Glucose, corn steep liquor, soyabean and glycine are most suitable for the production of vitamin b2 from Ashbya gossypii. A laboratory culture of the Ashbya gossypii is inoculated into a sterilized medium containing the required ingredients.

15. The fermentation process is carried out at 28 °c temperature and at pH 6.8 for 4 to 5 days under aerobic conditions . The excess aeration is not suitable for growth of cells because it inhibits mycelia production and reduces the yield. An aeration rate at 0.25 to 0.30 volumes of air per volume of medium per minute is satisfactory for maximum yield. The fermentation process proceeds through 4 phases.

16. Recovery of riboflavin The riboflavin is present in solution as well as bound form to the mycelium. Acetone, petroleum ether, and butanol are used for extraction of riboflavin from fermentation broth. It is precipitated by using reducing agents such as hydro sulphite and finely divided diatomaceous earth .