Action and use
Benzodiazepine.
DEFINITION
Nitrazepam Oral Suspension is a suspension of Nitrazepam in a suitable flavoured vehicle.
The oral suspension complies with the requirements stated under Oral Liquids and with the following requirements.
Content of nitrazepam, C15H11N3O3
90.0 to 105.0% of the stated amount.
IDENTIFICATION
A. Carry out the method for thin-layer chromatography, Appendix III A, using the following solutions.
(1) Shake a quantity of the oral suspension containing 2.5 mg of Nitrazepam with 10 mL of acetonitrile, centrifuge and use the supernatant liquid.
(2) 0.025% w/v of nitrazepam BPCRS in methanol.
(3) 0.025% w/v each of nitrazepam BPCRS and diazepam BPCRS in methanol.
CHROMATOGRAPHIC CONDITIONS
(a) Use as the coating silica gel.
(b) Use the mobile phase as described below.
(c) Apply 10 μL of each solution.
(d) Develop the plate to 15 cm.
(e) After removal of the plate, dry in air, spray with dilute potassium iodobismuthate solution and examine in daylight.
MOBILE PHASE
5 volumes of concentrated ammonia, 10 volumes of methanol and 85 volumes of ethyl acetate.
SYSTEM SUITABILITY
The test is not valid unless the chromatogram obtained with solution (3) shows two clearly separated spots.
CONFIRMATION
The principal spot in the chromatogram obtained with solution (1) corresponds in position and colour to that in the chromatogram obtained with solution (2).
B. In the Assay, the chromatogram obtained with solution (1) shows a peak with the same retention time as the principal peak in the chromatogram obtained with solution (2).
TESTS
Carry out the following tests protected from light.
Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Shake a quantity of the oral suspension containing 5 mg of Nitrazepam with 60 mL of acetonitrile, add sufficient acetonitrile to produce 100 mL and filter.
(2) Dilute 1 volume of solution (1) to 100 volumes with acetonitrile.
(3) 0.01% w/v of nitrazepam BPCRS and 0.0002% w/v of clonazepam BPCRS in acetonitrile.
(4) Dilute 1 volume of solution (2) to 10 volumes with acetonitrile.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (25 cm × 4.0 mm) packed with octylsilyl silica gel for chromatography (5 μm) (Licrospher RP8 is suitable) and a stainless steel guard column (4 mm × 2 mm) packed with octylsilyl silica gel for chromatography (5 μm) (Phenomenex C8 is suitable).
(b) Use gradient elution and the mobile phase described below.
(c) Use a flow rate of 1 mL per minute.
(d) Use a column temperature of 40°.
(e) Use a detection wavelength of 270 nm.
(f) Inject 10 μL of each solution.
MOBILE PHASE
Mobile phase A: 0.05M sodium dihydrogen orthophosphate, adjusted to pH 3.0 with orthophosphoric acid.
Mobile phase B: 20 volumes of mobile phase A and 80 volumes of acetonitrile.
When the chromatograms are recorded under the prescribed conditions the retention times relative to nitrazepam (retention time about 9 minutes) are: impurity A, about 1.3 and impurity B, about 1.6.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution between the peaks due to nitrazepam and clonazepam is at least 1.3.
LIMITS
In the chromatogram obtained with solution (1):
the area of any peak corresponding to impurity A is not greater than 1.8 times the area of the principal peak in the chromatogram obtained with solution (2) (1.8%);
the area of any peak corresponding to impurity B is not greater than 3.2 times the area of the principal peak in the chromatogram obtained with solution (2) (3.2%);
the area of any other secondary peak is not greater than half of the area of the principal peak in the chromatogram obtained with solution (2) (0.5%);
the sum of the areas of all secondary peaks is not greater than 6 times the area of the principal peak in the chromatogram obtained with solution (2) (6%).
Disregard 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.
(1) Add a weighed quantity of the oral suspension containing 12.5 mg of Nitrazepam to 25 mL of water, extract with three 50-mL quantities of chloroform, filtering each extract through phase-separating paper and add sufficient chloroform to produce 200 mL.
(2) 0.00625% w/v of nitrazepam BPCRS in chloroform.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (20 cm × 4.6 mm) packed with silica gel for chromatography (5 μm) (Lichrosorb or Partisil is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 2 mL per minute.
(d) Use a column temperature of 40°.
(e) Use a detection wavelength of 254 nm.
(f) Inject 20 μL of each solution.
MOBILE PHASE
10 volumes of absolute ethanol and 90 volumes of n-hexane.
Under the prescribed conditions the retention time of the peak due to nitrazepam is about 8 minutes.
DETERMINATION OF CONTENT
Determine the weight per mL of the oral suspension, Appendix V G, and calculate the content of C15H11N3O3, weight in volume, using the declared content of C15H11N3O3 in nitrazepam BPCRS.
STORAGE
Nitrazepam Oral Suspension should be protected from light.
IMPURITIES
The impurities limited by the requirements of this monograph include those listed under Nitrazepam.
