A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

Test solution: 10 mg per mL, in chloroform.

Mobile phase— Dissolve 27.2 g of monobasic potassium phosphate in 1000 mL of water, adjust with phosphoric acid to a pH of 2.4, and mix. Prepare a filtered and degassed mixture of this solution and acetonitrile (90:10). Make adjustments if necessary (see System Suitability under Chromatography 621).

System suitability standard solution— Dissolve an accurately weighed quantity of USP Ofloxacin RS in methanol to obtain a solution having a known concentration of about 1.0 µg per mL.

Test solution— Quantitatively dissolve an accurately weighed quantity of Ofloxacin in methanol to obtain a solution containing about 1.0 mg per mL.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 294-nm detector and a 4.6-mm × 10-cm column that contains packing L1. The flow rate is about 2 mL per minute. Chromatograph the System suitability standard solution, and record the peak responses as directed for Procedure: the column efficiency, determined from the ofloxacin peak, is not less than 1400 theoretical plates when calculated by the formula: and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Inject a volume (about 10 µL) of the Test solution into the chromatograph, record the chromatogram for a period of time that is 2.5 times the retention time of the main ofloxacin peak, and measure the areas for all the peaks, except to disregard the solvent peak. Calculate the percentage of desfluoroofloxacin in the portion of Ofloxacin taken by the formula: in which 1.13 is the response factor of desfluoroofloxacin relative to that of ofloxacin; rd is the area for the desfluoroofloxacin peak, if present, at a retention time of about 0.56 relative to that of ofloxacin; and rt is the total area of the peaks, except for the solvent peak: not more than 0.2% is found. Calculate the percentage of each other impurity with an area greater than that of the ofloxacin peak in the chromatogram of the System suitability standard solution obtained under Chromatographic system, by the formula: in which ri is the peak area for an individual impurity; and rt is the total area of the peaks in the chromatogram obtained from the Test solution, except for the solvent peak: not more than 0.3% of any individual impurity is found; and the sum of all impurities found is not more than 0.5%.

Internal standard solution— Prepare a solution in sodium hydroxide solution (1 in 100) containing 0.7 µL of n-propyl alcohol per mL. Transfer 2.0 mL of this solution to a 250-mL volumetric flask, dilute with the same sodium hydroxide solution (1 in 100) to volume, and mix.

Standard solution— Prepare a solution in Internal standard solution containing 10.0 µg each of methanol and dehydrated alcohol per mL. Transfer 2.0 mL of this solution to a vial fitted with a septum and crimp cap, and seal. Heat the sealed vial at 90 for 2 minutes, and shake for 6 minutes.

Test solution— Transfer 40 mg of Ofloxacin, accurately weighed, to a vial fitted with a septum and a crimp cap, add 2.0 mL of Internal standard solution, and seal the vial. Heat the sealed vial at 90 for 2 minutes, and shake for 6 minutes.

Blank— Transfer 2.0 mL of the Internal standard solution to a vial fitted with a septum and crimp cap, and seal. Heat the sealed vial at 90 for 2 minutes, and shake for 6 minutes.

Chromatographic system (see Chromatography 621)— The gas chromatograph is equipped with a flame-ionization detector, a 0.53-mm × 30-m fused silica column coated with a 3.0-µm film of stationary phase G43, and a fused silica precolumn. Helium is used as the carrier gas at a flow rate of about 7 mL per minute. The injection port and detector temperatures are maintained at about 170 and 250, respectively. Condition the column with the helium flowing at 200 for 2 hours or until a stable baseline is obtained. For analysis, the column temperature is programmed according to the following steps. It is maintained at 35 for 3 minutes, then increased to 90 at a rate of 20 per minute, then increased further to 200 at a rate of 40 per minute, and then maintained for 2 minutes. Chromatograph the headspace of the Standard solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.5 for methanol, 0.6 for ethanol, and 1.0 for n-propyl alcohol; the resolution, R, between the methanol peak and the ethanol peak is not less than 2.0; and the relative standard deviation for replicate injections is not more than 5%.

Procedure— Use a heated gas tight syringe to make injections of the headspace into the chromatograph. Separately inject equal volumes (about 1 mL) of the headspace of the Standard solution, the Blank, and the Test solution into the chromatograph, record the chromatograms, and measure the peak area responses. Calculate the percentage of methanol and ethanol in the Ofloxacin taken by the formula: in which W is the weight, in mg, of Ofloxacin taken to prepare the Test solution; and RU, RB, and RS are the peak response ratios of the relevant alcohol peak to the internal standard peak obtained from the Test solution, the Blank, and the Standard solution, respectively: not more than 0.005% of methanol and not more than 0.05% of ethanol are found.

(Official January 1, 2007)