Thursday, June 18, 2020

Media Components- Nitrogen Sources

Nitrogen Sources

Most industrial microbes can utilize both inorganic and organic nitrogen sources.  Inorganic nitrogen may be supplied as ammonium salts, often ammonium sulphate and diammonium hydrogen phosphate, or ammonia.  Ammonia can also be used to adjust pH of the fermentation.  Organic nitrogen sources include amino acids, proteins and urea.  Nitrogen is often supplied in crude forms that are essentially by-products of other industries, such as corn steep liquor, yeast extracts, peptones and soya meal.  Purified amino acids are used only in special situations, usually as precursors for specific products.

 Corn Steep Liquor (CSL)

Corn steep liquor is a by-product of starch extraction from maize and its first use in fermentations was for penicillin production in the 1940s.  The extract composition of the liquor varies depending on the quality of the maize and the processing conditions. Concentrated extracts generally contain about 4% (w/v) nitrogen, including a wide range of amino acids, along with vitamins and minerals.  Any residual sugars are usually converted to lactic acid (9-20%, w/v) by contaminating bacteria.  Corn steep liquor can sometimes be replaced by similar liquors, such as those derived from potato starch production.

 Yeast Extracts

Yeast extracts may be produced from waste baker’s and brewer’s yeast, or other strains of S. cerevisiae.  Alternate sources are Kluveromyces marxianus (formerly classified as K. fragilis) grown on whey and Candida utilis cultivated using ethanol, or wastes from wood and paper processing.  Those extracts used in the formulation of fermentation media are normally salt-free concentrates of soluble components of hydrolyzed yeast cells.  Yeast extracts with sodium chloride concentrations greater than 0.05% (w/v) cannot be used in fermentation processes due to potential corrosion problems.

Yeast cell hydrolysis is often achieved by autolysis using the cell’s endogenous enzymes, usually without the need for additional hydrolytic enzymes.  Autolysis can be initiated by temperature or osmotic shock, causing cells to die but without inactivating their enzymes.  Temperature and pH are controlled throughout to ensure an optimal and standardized autolysis process.  Temperature control is particularly important to prevent loss of vitamins.   

Autolysis is performed at 50-55˚C for several hours before the temperature is raised to 75˚C to inactivate the enzymes.  Finally the cells are disrupted by plasmolysis or mechanical disruption.  Cell wall materials and other debris are removed by filtration or centrifugation and the resultant extract is rapidly concentrated.  Extracts are available as liquids containing 50-65% solids, viscous pastes or dry powders.  They contain amino acids, peptides, water-soluble vitamins and some glucose, derived from the yeast storage carbohydrates (trehalose and glycogen).

 Peptones

Peptones are usually too expensive for large scale industrial fermentations.  They are prepared by acid or enzyme hydrolysis of high protein materials: meat, casein, gelatine, keratin, peanuts, soy meal, cotton seeds etc.  Their amino acid compositions may vary depending upon the original protein source.  For example, gelatine-derived peptones are rich in proline and hydroxyproline, but are almost devoid of sulphur-containing amino acids; whereas keratin peptone is rich in both proline and cystine, but lacks lysine.  Peptones from plant sources invariably contain relatively large quantities of carbohydrates.

 Soya Bean Meal

Residues remaining after soya beans have been processed to extract the bulk of their oil are composed of 50% protein, 8% non-protein nitrogenous compound, 30% carbohydrates and 1% oil.  This residual soya meal is often used in antibiotic fermentations because the components are only slowly metabolized, thereby eliminating the possibility of repression of product formation.

References

  1. Industrial Microbiology: An Introduction. Michael J. Waites, Neil L. Morgan, John S
  2. Principles of Fermentation Technology- Peter Stanbury, Allan Whitaker, Stephen Hall

 

 


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