Limit of cephalexin—
Using the chromatogram of the
Assay preparation obtained in the
Assay, calculate the percentage of cephalexin (C
16H
17N
3O
4S) in the portion of Cephradine taken by the formula:
100(rUx / rU),
in which
rUx is the cephalexin peak response in the chromatogram obtained from the
Assay preparation, and
rU is the sum of the cephalexin and cephradine peak responses in the chromatogram obtained from the
Assay preparation: not more than 5.0%, calculated on the anhydrous basis, is found.
Assay—
Mobile phase—
Prepare a mixture of water, methanol, 0.5 M sodium acetate, and 0.7 N acetic acid (782:200:15:3). Make adjustments if necessary (see
System Suitability under
Chromatography 
621
). Filter the solution through a filter of 1 µm or finer porosity, and degas before use.
Standard preparation—
Dissolve an accurately weighed quantity of
USP Cephradine RS quantitatively in
Mobile phase to obtain a solution having a known concentration of about 0.5 mg per mL.
Assay preparation—
Transfer about 50 mg of Cephradine, accurately weighed, to a 100-mL volumetric flask, add about 30 mL of Mobile phase, and sonicate. Dilute with Mobile phase to volume, and mix.
Chromatographic system
(see
Chromatography 621)—The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the
Resolution solution, and record the peak responses as directed under
Procedure: the relative retention times are about 0.8 for cephalexin and 1.0 for cephradine; and the resolution,
R, between the cephalexin peak and the cephradine peak is not less than 2.0. Chromatograph the
Standard preparation, and record the peak responses as directed under
Procedure: the relative standard deviation for replicate injections is not more than 2.0%.
Procedure—
Separately inject equal volumes (about 10 µL) of the
Standard preparation, and the
Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in µg, of total cephalosporins (sum of cephradine and cephalexin) in each mg of the Cephradine taken by the formula:
100(CP / M)(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Cephradine RS in the
Standard preparation;
P is the designated potency, in µg per mg, of
USP Cephradine RS;
M is the quantity, in mg, of Cephradine taken to prepare the
Assay preparation; and
rU and
rS are the sums of the cephradine and cephalexin peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
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