Media for Industrial Fermentation Chapter 7 Media for Industrial Fermentation

Steps to Medium Development Design – nutritional requirements for growth and product formation Formulation – mass composition of cells, stoichiometry of growth and product formation Optimization

Media must meet the following criteria It will produce a maximum yield of product at biomass per gram of substrate used. It will produce a maximum concentration of product or biomass. It will permit the maximum rate of product formation. It will be the minimum yield of undesired product. It will be cheap and of a consistent quality and is readily available throughout the year. It will cause minimal problem in other aspects of production and agitation, extraction, purification and waste treatment.

Macroelements Constitutes 90-95% of the dry weight of microbial biomass C, N, O, S, P, K, Mg Microorganisms are heterotroph (obtain the cell carbon from organic compound) and autotrophs (obtain cell carbon from CO2) Assimilated to provide cell carbon and energy – to synthesize cell material, osmotic work, cell motility Eg Penicillin production: of the total amount of glucose consumed, 10% for carbon in biomass, 10% for energy for biomass synthesis, 10% penicillin carbon and energy, 70% for maintenance energy

Macroelements [cont] Range of C compounds: carbohydrates, alcohols, organic acids, hydrocarbons, organic nitrogen compounds N requirements: generally supplied as (NH4)2SO4 Protein and peptides can be utilized if the microbes are able to hydrolyze Yeast extract function??

Microelements Ca, Mn, Fe, Cu, Co and Zn are essential B, Cr and Mo are rarely required Commonly added as mineral salts Trace elements deficiencies in batch cultures are reflected by limitation of the growth rate In continuous culture, by the decrease observed in the steady state biomass conc. Must consider the interactions because optimum conc of any ion is dependent on the conc of the others.

Growth Factors Some microbes require specific nutritional factors which they cannot synthesize - growth factors 3 groups – vitamins, amino acids and miscellaneous compounds Water soluble vitamins – mostly coenzymes In complex media, eg cane molasses already contain several vitamins

Other Additives Precursors, eg phenylacetic acid in penicillin biosynthesis Stimulants, eg methanol in citric acid production Protectants Antifoams Chelators – eg EDTA Stabilizers – to avoid the loss of plasmids Neutralizing agent – eg. Phosphate buffers

Influence of Physical Factors Temperature and water activity Eg. To grow S.cereviciae at 37C instead of 30C, the pantothenic acid content must be increased. Pasteurella pestis requires more amino acids and vitamins for growth at 37C than at 28C.

The medium selected will affect the design of the fermenter to be used. A laboratory medium may not ideal in a large fermenter with a low gas-transfer pattern. Media with a high viscosity will also need a higher power input for effective stirring. Besides the requirement for growth and product formation, medium may also influence pH variation, foam formation, oxidation-reduction potential and the morphological form of the organisms. It may also be necessary to provide precursors or metabolic inhibitors.

Guide to Design and Formulation for Plant Cell Suspension Media The use of two-step process, first using a growth medium and second, a production medium C source – sucrose is preferred. Add phytohormones to improve yield or productivity Adding precursors to enhance secondary metabolite formation

Growth Medium Select a basal medium such as MS, B5 or other Test the medium in shake flasks and in a bioreactor and optimize it for maximal biomass production

Production Medium Maintain the same carbohydrate conc of the growth media and then test the following: Reduce the phosphate and nitrogen content Modify the vitamins and iron (chelated) content Find adequate phytohormones level and their ratios Add a precursor and find the correct time for its addition Test a biotic or an abiotic elicitor Optimize the medium by fractional factorial statistical methodology

Large Scale Media Take into account: - fermenter size - fermentation time - purification procedure - medium composition before implementation!!

Can you detect the differences between plant and animal culture media?