Mobile phase, Diluting solution, System suitability solution and Chromatographic system —Prepare as directed in the test for Related compounds under Furosemide.

Standard solution— Prepare a solution in Diluting solution containing 10.0 µg of USP Furosemide Related Compound B RS per mL.

Test solution— Transfer an accurately measured volume of Injection, equivalent to about 10 mg of furosemide, to a 10-mL volumetric flask, add Diluting solution to volume, and mix.

Procedure— Separately inject equal volumes (about 20 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the peak responses. The response at 254 nm obtained for any peak observed in the chromatogram of the Test solution at a retention time corresponding to that of the Reference Standard in the Standard solution is not greater than the response at 254 nm obtained for the peak in the chromatogram of the Standard solution, corresponding to not more than 1.0% of furosemide related compound B. Where the Injection is labeled as intended solely for veterinary use, the response at 254 nm obtained in the chromatogram of the Test solution at a retention time corresponding to that of the Reference Standard in the Standard solution is not greater than 2.5 times the response at 254 nm obtained for the peak in the chromatogram of the Standard solution, corresponding to not more than 2.5% of furosemide related compound B.

Mobile phase, Diluting solution, System suitability solution, and Chromatographic system— Prepare as directed in the test for Related compounds under Furosemide.

Standard preparation— Dissolve an accurately weighed quantity of USP Furosemide RS in Diluting solution to obtain a solution having a known concentration of about 1.0 mg per mL.

Assay preparation— Transfer an accurately measured volume of Injection, equivalent to about 10 mg of furosemide, to a 10-mL volumetric flask, add Diluting solution to volume, and mix.

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 peak responses. Using the response at 254 nm, calculate the quantity, in mg, of furosemide (C12H11ClN2O5S) in each mL of the Injection taken by the formula: in which C is the concentration, in mg per mL, of USP Furosemide RS in the Standard preparation; V is the volume, in mL, of Injection taken; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.