(Ph. Eur. 11.6 update)
Action and use
Vasopressin analogue; treatment of diabetes insipidus; nocturnal enuresis; haemophillia; von Willebrand’s disease.
DEFINITION
Desmopressin Injection is a sterile solution of Desmopressin in Water for Injections.
The injection complies with the requirements stated under Parenteral Preparations and with the following requirements.
Content of desmopressin, C46H64N14O12S2
90.0 to 110.0% of the stated amount of the peptide.
CHARACTERISTICS
A colourless solution.
IDENTIFICATION
In the Assay, the principal peak in the chromatogram obtained with solution (1) corresponds to that in the chromatogram obtained with solution (2).
TESTS
Acidity
pH, 3.5 to 6.0, Appendix V L.
Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions and the normalisation procedure.
(1) Dilute the injection, if necessary, with water to produce a solution containing 0.0004% w/v of the peptide.
(2) Dissolve the contents of a vial of desmopressin impurity standard BPCRS in 2 mL of water.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (12 cm x 4.0 mm) packed with octadecylsilyl silica gel for chromatography (5 μm).
(b) Use gradient elution and the mobile phase described below.
(c) Use a flow rate of 1.5 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 220 nm.
(f) Inject 200 μL of each solution.
MOBILE PHASE
Mobile phase A 0.067M mixed phosphate buffer solution pH 7.0.
Mobile phase B A mixture of acetonitrile for chromatography and mobile phase A in equal volumes.
| Time (Minutes) | Mobile phase A% | Mobile phase B% | Comment |
| 0-4 | 76 | 24 | isocratic |
| 4-18 | 76→58 | 24→42 | linear gragident |
| 18-35 | 58→48 | 42→52 | linear gragident |
| 35-40 | 48→76 | 52→24 | linear gragident |
| 40-50 | 76 | 24 | re-equilibration |
The retention time of [4- L -glutamic acid]desmopressin is about 14 minutes and the retention time of desmopressin is about 15 minutes.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (2), the resolution between the two principal peaks is at least 3.4.
LIMITS
In the chromatogram obtained with solution (1):
the area of any secondary peak is not more than 4.0%;
the total area of any such peaks is not more than 5.0%.
Disregard any peak due to the solvent or with an area less than 2.0%.
Bacterial endotoxins
Carry out the test for bacterial endotoxins, Appendix XIV C. The endotoxin limit concentration is less than 10 IU per μg peptide.
ASSAY
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Dilute a volume of the injection in water to give a final concentration of 0.0004 % w/v of the peptide.
(2) 0.0004% w/v of desmopressin BPCRS in water.
(3) Dissolve the contents of a vial of desmopressin impurity standard BPCRS in 2 mL of water.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (12 cm x 4.0 mm) packed with octadecylsilyl silica gel for chromatography (5 μm) (Nucleosil C18 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 2 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 220 nm.
(f) Inject 200 μL of each solution.
MOBILE PHASE
20 volumes of acetonitrile for chromatography and 80 volumes of 0.067 M mixed phosphate buffer solution, pH 7.0.
The retention time of desmopressin is about 5 minutes; if necessary adjust the concentration of acetonitrile in the mobile phase to obtain a retention time of about 5 minutes.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution between the two principal peaks at least 2.8.
DETERMINATION OF CONTENT
Calculate the content of C46H64N14O12S2 in the injection from the chromatograms obtained and from the declared content of C46H64N14O12S2 in desmopressin BPCRS.
STORAGE
Desmopressin Injection should be protected from light and stored at a temperature of 2° to 8°.
