Edition: BP 2025 (Ph. Eur. 11.6 update)
Action and use
Tetracycline antibacterial.
DEFINITION
Oxytetracycline Cutaneous Spray contains Oxytetracycline Hydrochloride in a suitable vehicle.
The cutaneous spray complies with the requirements stated under Veterinary Liquid Preparations for Cutaneous Application and with the following requirements.
Content of oxytetracycline hydrochloride, C22H24N2O9,HCl
90.0 to 110.0% of the stated amount.
IDENTIFICATION
In the Assay, record the UV spectrum of the principal peak in the chromatograms obtained with solutions (1) and (2) with a diode array detector in the range of 210 to 400 nm:
the UV spectrum of the principal peak in the chromatogram obtained with solution (1) is concordant to that of the peak in the chromatogram obtained with solution (2);
the retention time of the principal peak in the chromatogram obtained with solution (1) is similar to that of the peak in the chromatogram obtained with solution (2).
TESTS
Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions in a mixture of 20 volumes of acetonitrile and 80 volumes of 0.01M oxalic acid (solvent A). Prepare the solutions immediately before use.
(1) Shake a quantity of the cutaneous spray containing 0.16 g of Oxytetracycline Hydrochloride in 150 mL of solvent A and dilute to 200 mL. Filter the resulting solution (Whatman GF/C filter is suitable).
(2) Dilute 1 volume of solution (1) to 100 volumes.
(3) 0.08% w/v of oxytetracycline for system suitability A EPCRS.
(4) 0.0008% w/v of sulfan blue.
(5) Dilute 1 volume of solution (2) to 10 volumes.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (15 cm × 4.6 mm) packed with end-capped octylsilyl silica gel for chromatography (5 µm) (Inertsil C8 is suitable).
(b) Use gradient elution and the mobile phase described below.
(c) Use a flow rate of 1.3 mL per minute.
(d) Use a column temperature of 50°.
(e) Use a detection wavelength of 254 nm.
(f) Inject 10 µL of each solution.
MOBILE PHASE
Mobile phase A 0.05% v/v trifluoroacetic acid.
Mobile phase B 5 volumes of tetrahydrofuran, 15 volumes of methanol and 80 volumes of acetonitrile.
| Time (Minutes) | Mobile phase A (% v/v) | Mobile phase B (% v/v) | Comment |
| 0-5 | 90 | 10 | isocratic |
| 5-20 | 90→65 | 10→35 | linear gradient |
| 20-30 | 65 | 35 | isocratic |
| 30-31 | 65→90 | 35→10 | linear gradient |
| 31-37 | 90 | 10 | re-equilibration |
When the chromatograms are recorded under the prescribed conditions, the relative retentions with reference to oxytetracycline (retention time about 7 minutes) are: impurity A, about 0.9; impurity B, about 1.2; impurity C, about 1.3; impurity D, about 1.4; impurity E, about 2.2; impurity F, about 2.3.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the peak-to-valley ratio is at least 3.0, where Hp is the height above the baseline of the peak due to impurity A and Hv is the height above the baseline of the lowest point of the curve separating this peak from the peak due to oxytetracycline.
The test is not valid unless, in the chromatogram obtained with solution (3), the peak-to-valley ratio is at least 3.0, where Hp is the height above the baseline of the peak due to impurity B and Hv is the height above the baseline of the lowest point of the curve separating this peak from the peak due to oxytetracycline.
LIMITS
Identify any peak corresponding to impurities A, B, C, D, E and F in the chromatogram obtained with solution (1), using the chromatogram obtained with solution (3). Identify the peak due to patent blue V if present in the chromatogram obtained with solution (1), using the chromatogram obtained with solution (4). Multiply the areas of the peaks due to Impurity D and E by a correction factor of 0.4.
In the chromatogram obtained with solution (1):
the area of any peak corresponding to impurity A is not greater than the area of the principal peak in the chromatogram obtained with solution (2) (1.0%);
the area of any peak corresponding to impurity B is not greater than 2.5 times the area of the principal peak in the chromatogram obtained with solution (2) (2.5%);
the area of any peak corresponding to impurity C is not greater than twice the area of the principal peak in the chromatogram obtained with solution (2) (2.0%);
the area of any other secondary peak is not greater than the area of the principal peak in the chromatogram obtained with solution (2) (1.0%);
the sum of the areas of all the secondary peaks is not greater than 7 times the area of the principal peak in the chromatogram obtained with solution (2) (7.0%).
Disregard any peak due to patent blue V and any peak with an area less than the area of the principal peak in the chromatogram obtained with solution (4) (0.1%).
ASSAY
Carry out the method for liquid chromatography, Appendix III D, using the following solutions in a mixture of 20 volumes of acetonitrile and 80 volumes of 0.01M oxalic acid (solvent A). Prepare the solutions immediately before use.
(1) Dilute a quantity of the cutaneous spray containing 0.16 g of Oxytetracycline Hydrochloride with 150 mL of solvent A and shake. Dilute to 200 mL and filter (Whatman GF/C filter is suitable). Dilute 1 volume of the filtrate to 10 volumes.
(2) 0.0074% w/v of oxytetracycline BPCRS.
CHROMATOGRAPHIC CONDITIONS
The chromatographic conditions stated under Related substances may be used.
DETERMINATION OF CONTENT
Calculate the content of C22H24N2O9,HCl in the spray using the declared content of C22H24N2O9 in oxytetracycline BPCRS. Each mg of C22H24N2O9 is equivalent to 1.079 mg of C22H24N2O9,HCl.
IMPURITIES
The impurities limited by the requirements of this monograph include those listed under Oxytetracycline hydrochloride.
