- British Pharmacopoeia Volume III
- Formulated Preparations: Specific Monographs
Gentamicin Injection |
Aminoglycoside antibacterial.
Gentamicin Injection is a sterile solution of Gentamicin Sulphate in Water for Injections.
The injection complies with the requirements stated under Parenteral Preparations and with the following requirements.
A. Carry out the method for thin-layer chromatography , Appendix III A, using a silica gel precoated plate (Merck silica gel 60 plates are suitable) and as the mobile phase the lower layer obtained by shaking together equal volumes of 13.5m ammonia , chloroform and methanol and allowing to separate. Apply separately to the plate (1) a volume of the injection containing the equivalent of 30 µg of gentamicin and (2) 0.1 mg of gentamicin sulphate BPCRS dissolved in a volume of water equal to the volume of the injection used. After removal of the plate, allow it to dry in air, spray with ninhydrin solution R1 and heat at 105° for 2 minutes. The three principal spots in the chromatogram obtained with solution (1) correspond to the three principal spots in the chromatogram obtained with solution (2).
B. In the test for Composition of gentamicin sulphate, the retention times of the four principal peaks in the chromatogram obtained with solution (2) correspond to those of the four principal peaks in the chromatogram obtained with solution (1).
pH, 3.0 to 5.5, Appendix V L.
Carry out the method for liquid chromatography , Appendix III D, using the following solutions. For solution (1) add 5 ml of methanol to 10 ml of a 0.065% w/v solution of gentamicin sulphate BPCRS, swirl and add 4 ml of phthalaldehyde reagent , mix, add sufficient methanol to produce 25 ml, heat in a water bath at 60° for 15 minutes and cool. If the solution is not used immediately, cool to 0° and use within 4 hours. Prepare solution (2) in the same manner as solution (1) but using 10 ml of a solution prepared by diluting a suitable volume of the injection with water to contain the equivalent of 0.045% of gentamicin.
The chromatographic procedure may be carried out using (a) a stainless steel column (10 to 12.5 cm × 4.6 to 5 mm) packed with octadecylsilyl silica gel for chromatography (5 µm) (Hypersil ODS is suitable), (b) 0.025m sodium heptanesulphonate monohydrate in a mixture of 5 volumes of glacial acetic acid , 25 volumes of water and 70 volumes of methanol as the mobile phase with a flow rate of 1.5 ml per minute and (c) a detection wavelength of 330 nm.
When the chromatograms are recorded under the prescribed conditions the retention time of component C2 is 10 to 20 minutes. The retention times relative to component C2 are: about 0.13 (reagent); about 0.27 (component C1); about 0.65 (component C1a); about 0.85 (component C2a).
The test is not valid unless, in the chromatogram obtained with solution (1), the resolution factor between the peaks due to components C2a and C2 is at least 1.3.
Using the chromatogram obtained with solution (2) calculate the percentage content of components C1, C1a, C2 and C2a in the injection by normalisation . The proportions are within the following limits: C1, 25.0 to 50.0%; C1a, 10.0 to 35.0%; C2 plus C2a, 25.0 to 55.0%.
Carry out the test for bacterial endotoxins , Appendix XIV C. Dilute the injection, if necessary, with water BET to give a solution containing the equivalent of 10 mg of gentamicin per ml (solution A). The endotoxin limit concentration of solution A is 16.7 IU per ml.
Carry out the microbiological assay of antibiotics , Appendix XIV A. The precision of the assay is such that the fiducial limits of error are not less than 95% and not more than 105% of the estimated potency.
Calculate the content of gentamicin in the injection, taking each 1000 IU found to be equivalent to 1 mg of gentamicin. The upper fiducial limit of error is not less than 97.0% and the lower fiducial limit of error is not more than 110.0% of the stated content.
The strength is stated in terms of the equivalent amount of gentamicin in a suitable dose-volume.