» Chlorambucil Tablets contain not less than 85.0 percent and not more than 110.0 percent of the labeled amount of C14H19Cl2NO2.
Packaging and storage Preserve coated Tablets in well-closed containers; preserve uncoated Tablets in well-closed, light-resistant containers.
USP Reference standards 11
Identification Shake a quantity of finely powdered Tablets, equivalent to about 16 mg of chlorambucil, with 20 mL of carbon disulfide. Filter, evaporate to dryness, and dissolve the residue in 2 mL of carbon disulfide: the resulting solution responds to Identification test A under Chlorambucil.
Disintegration 701 Place 1 Tablet in each of the six tubes of the basket, and if the Tablet has a soluble external coating, immerse the basket in water at room temperature for 5 minutes. Operate the apparatus, using simulated gastric fluid TS maintained at 37 ± 2 as the immersion fluid. After 30 minutes of operation in simulated gastric fluid TS, lift the basket from the fluid, and observe the Tablets. If the Tablets have not disintegrated completely, substitute simulated intestinal fluid TS maintained at 37 ± 2 as the immersion fluid, and continue the test for a total period of time, including previous exposure to water and simulated gastric fluid TS, equal to 45 minutes. Lift the basket from the fluid, and observe the Tablets: all of the Tablets have disintegrated completely. If 1 or 2 Tablets fail to disintegrate completely, repeat the test on 12 additional Tablets: not less than 16 of the total of 18 Tablets tested disintegrate completely.
Uniformity of dosage units 905: meet the requirements.
Internal standard solution Transfer about 20 mg of USP Propylparaben RS to a 50-mL volumetric flask, dissolve in and dilute with alcohol to volume, and mix.
Standard preparation Dissolve in alcohol a suitable quantity of USP Chlorambucil RS, accurately weighed, and prepare, by quantitative dilution, a solution in alcohol having a known concentration of about 1 mg per mL. Transfer 2.0 mL of the solution to a 100-mL volumetric flask containing about 50 mL of alcohol and, while gently swirling, add 5.0 mL of 0.1 N hydrochloric acid and 2.0 mL of Internal standard solution. Dilute with alcohol to volume, and mix.
Assay preparation Weigh and finely powder not fewer than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 2 mg of chlorambucil, to a 100-mL volumetric flask containing about 50 mL of alcohol and, while gently swirling, add 5.0 mL of 0.1 N hydrochloric acid and 2.0 mL of Internal standard solution. Sonicate for 5 minutes, dilute with alcohol to volume, and mix. Filter through a medium-porosity, sintered-glass filtering funnel, maintaining reduced pressure for the minimum necessary time in order to avoid solvent loss of evaporation.
Mobile phase Mix 500 mL of alcohol with 1.0 mL of glacial acetic acid in a 1-liter volumetric flask, dilute with water to volume, and mix. The alcohol concentration may be varied to meet system suitability requirements and to provide a suitable elution time for chlorambucil. Degas the solution at a pressure of approximately 250 mm of mercury for 2 minutes.
Chromatographic system Typically, a high-pressure liquid chromatograph, operated at room temperature, is fitted with a 25- or 30-cm × 2-mm stainless steel column packed with spherical silica microbeads, 5 µm to 10 µm in diameter, to which is bonded a nominal 10% or 20% (w/w) octadecyl silane. The mobile phase is maintained at a flow rate capable of giving the required resolution (see System suitability test) and a suitable elution time. An UV detector that monitors absorption at the 254-nm wavelength is used.
System suitability test Chromatograph 6 to 10 injections of the Standard preparation, and measure the peak responses as directed under Procedure. Calculate the response factor, PS, and the resolution factor, R (see Chromatography 621). The response factor for 6 to 8 injections does not vary by more than 2.0% relative standard deviation and the resolution factor is not less than 2.0.
Procedure Introduce equal volumes (10 to 12 µL) of the Standard preparation and the Assay preparation into the high-pressure liquid chromatograph by means of a suitable sampling valve or high-pressure microsyringe, and measure the peak responses at identical retention times obtained with each preparation. Calculate the quantity, in mg, of C14H19Cl2NO2 in the portion of Tablets taken by the formula:
0.1C(PU / PS)in which C is the concentration, in µg per mL, of USP Chlorambucil RS in the Standard preparation, and PU and PS are the response factors of the Assay preparation and the Standard preparation, respectively.
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USP35NF30 Page 2606