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ASSAY OF FERMENTATION PRODUCT Name Saurabh mandal Roll R2002A01 Course Code- BTY459 Course Name- Industrial biotechnology

gy Submitted To- Dr. Joginder Panwer

Fermentation technology is the oldest of all the biotechnological processes. The term derived from the Latin verb fevere means to boil.
It

the processes of chemical change caused by microorganism or production of product by the means of microorganism.
In

fermentation the assay of its product is very important aspect along with its other aspects

Assay

may be defined as qualitative and/or quantitative determination of any chemical compound from a simple or even complex material with the use of microbes. Thus, microbes are exploited as analytical tools in an analytical laboratory. The principal of assay were developed in around 1920s In assay there are so many categories- physicalchemical, biological, chromatographic, enzymatic etc.

Physical - chemical assay include titration and gravitimetric analysis


By

Titration- a common laboratory method of quantitative chemical analysis that is used to determine the unknown concentration of an identified analyte .
For

example- The amount of an organic acid such as lactic acid produced is roughly determined by adding a pH indicating dye such as bromo-thymol blue to the sample of fermentation broth followed with alkali of known strength.

In gravimetric methods, growth is determined by harvesting the mycelium, drying in an oven at 90C. for at least 2 hours or overnight to constant weight and weighing on a suitable balance. Dry weight measurements are laborious and of limited precision. Either a micro-balance or a large sample of cells is needed.

In turbidimetric methods, absorption of light by the cellular mass is usually determined. This may be accomplished by a suitable instrument such as photoelectric colorimeter or spectrophotometer. A system for using a particular wavelength of light is highly desirable, particularly of the assay sample. Ordinary light filters are usually satisfactory for this purpose.

Various wavelengths have been used ranging from 400 to 760 m, depending some cases on instruments and filters available.
The use of lower wavelengths increases the sensitivity of the assay, since turbidity readings increase as the wavelength of light used is decreased. linear relationships that are preferably used in assays

The principle of the biological assay is microbiological assay that of Antibiotics The growth of a sensitive test microorganism does not occur in the presence of an antibiotic, provided a sufficient quantity of the anti-biotic is present. There are several methods for quantitative determination of antibiotics in test samples. The important methods may be listed as under: (A) Agar diffusion (B) Serial dilution technique (C) Photometric assay

Agar Diffusion: In this method, the diameter of the inhibition zone is directly proportional to the quantity of an antibiotic. The antibiotic present in the solution diffuses radially through the agar medium layers and exerts its toxic effect on the sensitive test micro-organism present in the seed agar medium. Test micro-organisms like Staphylococcus aureus or Sarcina lutea, stock cultures sample preparation measuring of Response in such assays in terms of the diameter of the inhibition zone with the help of Calipers, Millimeter ruler, Scale projectors, Automatic and semi-automatic measuring devices

Serial Dilutions The inoculums is diluted out in a series of dilution tubes which are plated out. The number of colonies on the plate are counted and corrected for the dilution to calculate the number of organisms in the original inoculums.

Like the serial dilution technique, this assay method is very simple in principle. The test substance is added to a suspension of the test micro-organism in a nutrient medium the mixture incubated and the response (in terms of turbidity) of the test micro-organism measured. The factors governing this assay method include concentration of cells, size of cells, opacity of cells medium, incubation and response of the test micro-organism to the test substance.

Partition chromatography has allowed marked stride in fermentation research and technology.
Paper and thin layer chromatography are form of partition chromatography. The solute or sample is partitioned continuously between a stationary phase such as paper or silica gel of the thin layer plates, and a mobile phase consisting of a mixture of solvent as these solvent migrate across the paper or silica gel layer.

Paper chromatography has particular application water-soluble compound and thin layer chromatography for the more hydrophobic compounds.

ENZYMATIC ASSAY
Enzymatic assay are highly specific and they quantatively detect minutes amount of a fermentation products. For example lglutamic acid is a small sample of fermentation broth can be assayed by adding washed cells certain strains of E.Coli which contain the enzyme glutamic acid decarboxylase

Assay is the vibrant and fast growing fermentation technology. Development and improvement in the assay technique contribute to analyze the fermented product and help us to improve their quality.
References:
-industrial microbiology ; L.E Clasida, JR new age interntional publishers,2010 -industrial microbiology; A. H Patel Macmillan publication (2nd edition) Weblinks-www.biomed.com/1472-6904/9/1 -www.pharmacopedia.co.in/v29240/usp29nf24300_c81