An antibiotic is a medical preparation, containing a sufficient amount of chemical entity that is caused to produce naturally (by a microorganism) or artificially (semi-synthetic way) and that possesses the inherent ability to either destroy (bactericidal) or inhibit (bacteriostatic) microorganisms in relatively dilute solutions.
There are mainly three important points for standardization
of antibiotics:
• FDA (Food Drug Administration) regulations governing all
aspects of antibiotics testing are completely detailed and are subject to
periodic amendment.
• FDA regulations need to be referred to with regard to
prescribed methods for the assay of individual antibiotics and their
preparations.
• During the evaluation of the potency of antibiotic
substances, the actual and apparent measured effect is the degree of inhibition
of the growth of a suitable strain of microorganism i.e. the prevention of the
multiplication of the test organisms.
Two methods are usually applied, the cylinder-plate (or
cup-plate) method and the turbidimetric method. The cylinder-plate method
depends upon the diffusion of the antibiotic from a vertical cylinder through a
solidified agar layer in a Petri dish. The growth of the added micro-organism
is prevented entirely in a zone around the cylinder containing a solution of
the antibiotic. The turbidimetric method depends upon the inhibition of the growth
of a microbial culture in a uniform solution of the antibiotic in a fluid
medium that is favorable to its rapid growth in the absence of the antibiotic.
Media Preparation: All the required ingredients are
dissolved in sufficient water to produce 1000 ml and added sufficient amount of
1 M Sodium hydroxide or 1 M Hydrochloride acid is, as required so that after
sterilization the pH is between 6.5 to 7.5.
Buffer Solution Preparation: Buffer solutions are
prepared by dissolving the required quantities of dipotassium hydrogen
phosphate and potassium dihydrogen phosphate in sufficient water to produce
1000 ml and pH adjusted with 8 M phosphoric acid or 10 M potassium hydroxide.
Standard Preparation: Standard preparation is an
authentic sample of the appropriate antibiotic for which the potency has been
determined by reference to the appropriate international standard. The potency
of the standard preparation is expressed in µg per mg of an international unit
of the pure antibiotic.
A stock solution is prepared by dissolving a required amount
of weighed quantity of the standard preparation of a given antibiotic.
Preparation of the Sample Solution: Dilutions of
sample or unknown samples are prepared at the range of dilutions of standard
prepared dilutions.
Test Organisms: Some of the test organisms for each
antibiotic are listed in Table.
Table: Test organisms for microbial assay of antibiotics
|
Test organism |
Antibiotics |
*ATCC no. |
|
Amikacin |
Staphylococcus
aureus |
29737 |
|
Amphotericin
B |
Saccharomyces
cerevisiae |
9763 |
|
Gentamicin |
Styaphylococcus
epidermidis |
12228 |
|
Kanamycin |
Bacillus
pumilus |
14884 |
|
Neomycin |
Styaphylococcus
epidermidis |
12228 |
|
Streptomycin |
Bacillus
subtilis |
6633 |
|
Tetracycline |
Bacillus
cereus |
11778 |
*American Type Culture Collection, 21301 Park Lawn Drive,
Rockville, MD20852, USA
Assay Method of Streptomycin: Streptomycin is a bactericidal
antibiotic that belongs to the class aminoglycosides. It is derived from
actinobacterium Streptomyces griseus. It is used against gram-negative
bacteria, mainly against tuberculosis by inhibiting protein synthesis of
microorganisms. It is water soluble. Bacillus subtilis is used for the
microbiological assay of Streptomycin.