» Bumetanide Tablets contain not less than 90.0 percent and not more than 110.0 percent of the labeled amount of bumetanide (C17H20N2O5S).
Packaging and storage Preserve in tight, light-resistant containers.
USP Reference standards 11
A: The relative retention time of the major peak in the chromatogram of the Assay preparation corresponds to that in the chromatogram of the Standard preparation, as obtained in the Assay.
B: The principal spot obtained from the chromatogram of the Test solution exhibits an RF value corresponding to that of the Identification solution, as obtained in the test for Related compounds.
Medium: water; 900 mL.
Apparatus 2: 50 rpm.
Time: 30 minutes.
pH 2.9 Glycine buffer Dissolve 7.505 g of glycine and 5.85 g of sodium chloride in water to make 1000 mL (stock solution). Dilute 80.0 mL of the stock solution and 20.0 mL of 0.1 N hydrochloric acid with water to 1000 mL. Adjust, if necessary, with 0.1 N hydrochloric acid or 0.1 N sodium hydroxide to a pH of 2.9.
Procedure Determine the amount of C17H20N2O5S dissolved, by employing a suitable fluorometer having an excitation wavelength of about 350 nm and a fluorescence emission of about 450 nm on filtered portions of the solution under test, suitably diluted with pH 2.9 Glycine buffer, in comparison with a Standard solution having a known concentration of USP Bumetanide RS in the same Medium.
Tolerances Not less than 85% (Q) of the labeled amount of C17H20N2O5S is dissolved in 30 minutes.
Uniformity of dosage units 905: meet the requirements.
Adsorbent: 0.25-mm layer of chromatographic silica gel mixture.
Test solution Transfer an accurately weighed portion of finely powdered Tablets, equivalent to 10 mg of bumetanide, to a 50-mL centrifuge tube, add 20 mL of acetone (spectrophotometric or HPLC quality), and shake by mechanical means for 10 minutes. Centrifuge for 10 minutes, decant the supernatant into a glass-stoppered, 25-mL conical flask, and evaporate with the aid of a stream of nitrogen to dryness. Dissolve the residue in 0.5 mL of methanol.
Identification solution Dissolve USP Bumetanide RS in methanol to obtain a solution having a concentration of about 20 mg per mL.
Standard solutions Dilute a volume of the Identification solution quantitatively, and stepwise if necessary, with methanol to obtain a solution having a known concentration of about 0.16 mg of USP Bumetanide RS per mL. Quantitatively dilute with methanol to obtain Standard solutions having the following compositions.
Standard solution 6 Dissolve an accurately weighed quantity of USP Bumetanide Related Compound A RS in methanol, and dilute quantitatively, and stepwise if necessary, with methanol to obtain a solution having a known concentration of about 0.04 mg per mL.
Application volume: 25 µL.
Developing solvent system: a mixture of chloroform, cyclohexane, glacial acetic acid, and methanol (80:10:10:2.5).
Procedure Proceed as directed for Thin-Layer Chromatography under Chromatography 621. Examine the plate under short-wavelength UV light. Any secondary spot obtained from the chromatogram of the Test solution having an RF value corresponding to the RF value of the principal spot obtained from the chromatogram of Standard solution 6 is not larger or more intense than the principal spot obtained from the chromatogram of Standard solution 6: not more than 0.2% of bumetanide related compound A is found. For all other secondary spots obtained from the chromatogram of the Test solution, compare the intensity of each spot with the principal spots obtained from the chromatograms of Standard solutions 1 through 5: not more than 0.2% of any individual other impurity is found; and not more than 0.8% of the sum of all other impurities is found (excluding bumetanide related compound A).
Mobile phase, Internal standard solution, Standard preparation, and Chromatographic system Prepare as directed in the Assay under Bumetanide Injection.
Assay preparation Weigh and finely powder not fewer than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 0.5 mg of bumetanide, to a 10-mL volumetric flask, add 2.0 mL of Internal standard solution, and sonicate for 5 minutes. Add 2.0 mL of water, and mix. Cool, and filter, discarding the first 1 mL of the filtrate.
Procedure Separately inject equal volumes (about 20 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. The relative retention times are about 0.7 for 4-ethylbenzaldehyde and 1.0 for bumetanide. Calculate the quantity, in mg, of bumetanide (C17H20N2O5S) in the portion of Tablets taken by the formula:
4C(RU / RS)in which C is the concentration, in mg per mL, of USP Bumetanide RS in the Standard preparation; and RU and RS are the peak response ratios obtained from the Assay preparation and the Standard preparation, respectively.
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USP35NF30 Page 2398Pharmacopeial Forum: Volume No. 27(6) Page 3253