(Ph. Eur. monograph 0717)
C16H12FN3O3 313.3 1622-62-4
Action and use
Benzodiazepine.
DEFINITION
5-(2-Fluorophenyl)-1-methyl-7-nitro-1,3-dihydro-2H-1,4-benzodiazepin-2-one.
Content
99.0 per cent to 101.0 per cent (dried substance).
CHARACTERS
Appearance
White or yellowish, crystalline powder.
Solubility
Practically insoluble in water, soluble in acetone, slightly soluble in ethanol (96 per cent).
IDENTIFICATION
Infrared absorption spectrophotometry (2.2.24).
Comparison Ph. Eur. reference spectrum of flunitrazepam.
TESTS
Related substances
Liquid chromatography (2.2.29). Prepare the solutions immediately before use.
Test solution: Dissolve 100.0 mg of the substance to be examined in 10 mL of acetonitrile R and dilute to 50.0 mL with the mobile phase.
Reference solution (a): Dilute 1.0 mL of the test solution to 100.0 mL with the mobile phase. Dilute 1.0 mL of this solution to 10.0 mL with the mobile phase.
Reference solution (b): Dissolve the contents of a vial of flunitrazepam for system suitability CRS (containing impurities B and C) in 200 μL of acetonitrile R and dilute to 1 mL with the mobile phase.
Column:
— size: l = 0.15 m, Ø = 4.6 mm;
— stationary phase: end-capped octadecylsilyl silica gel for chromatography R (5 μm).
Mobile phase methanol R, acetonitrile R, water for chromatography R (50:305:645 V/V/V).
Flow rate 1.0 mL/min.
Detection Spectrophotometer at 254 nm.
Injection 20 μL.
Run time 6 times the retention time of flunitrazepam.
Identification of impurities: Use the chromatogram supplied with flunitrazepam for system suitability CRS and the chromatogram obtained with reference solution (b) to identify the peaks due to impurities B and C.
Relative retention: With reference to flunitrazepam (retention time = about 13 min): impurity B = about 0.6;
impurity C = about 2.5.
System suitability Reference solution (b):
— resolution: minimum 3.0 between the peaks due to impurity B and flunitrazepam.
Calculation of percentage contents:
— correction factor: multiply the peak area of impurity C by 2.44;
— for each impurity, use the concentration of flunitrazepam in reference solution (a).
Limits:
— impurity C: maximum 0.10 per cent;
— unspecified impurities: for each impurity, maximum 0.10 per cent;
— total: maximum 0.3 per cent;
— reporting threshold: 0.05 per cent.
Loss on drying (2.2.32)
Maximum 0.5 per cent, determined on 1.000 g by drying in an oven at 105 °C.
Sulfated ash (2.4.14)
Maximum 0.1 per cent, determined on 1.0 g in a platinum crucible.
ASSAY
Dissolve 0.250 g in 20 mL of anhydrous acetic acid R and add 50 mL of acetic anhydride R. Titrate with 0.1 M perchloric acid, determining the end-point potentiometrically (2.2.20).
1 mL of 0.1 M perchloric acid is equivalent to 31.33 mg of C16H12FN3O3.
STORAGE
Protected from light.
IMPURITIES
Specified impurities C.
Other detectable impurities (the following substances would, if present at a sufficient level, be detected by one or other of the tests in the monograph. They are limited by the general acceptance criterion for other/unspecified impurities and/or by the general monograph Substances for pharmaceutical use (2034). It is therefore not necessary to identify these impurities for demonstration of compliance. See also 5.10. Control of impurities in substances for pharmaceutical use) A, B, D.
A. 7-amino-5-(2-fluorophenyl)-1,3-dihydro-2H-1,4-benzodiazepin-2-one (7-aminodemethylflunitrazepam),
B. 5-(2-fluorophenyl)-7-nitro-1,3-dihydro-2H-1,4-benzodiazepin-2-one (demethylflunitrazepam),
C. 3-amino-4-(2-fluorophenyl)-1-methyl-6-nitroquinolin-2(1H)-one,
D. (2-fluorophenyl)[2-(methylamino)-5-nitrophenyl]methanone.
