HPLC ASSAY METHOD FOR CEFUROXIME

Chromatographic System :

Column               : 25cm x 4.6mm, packed with particles of silica(5µm)

Flow rate             : 1.2ml / minute , so that the resolution factors between the peaks  corresponding to the Cefuroxime axetil diastereoisomers A & B  in solution(4)and between the peaks corresponding to Cefuroxime axetil diastereoisomer A & the Cefuroxime axetil A³ -isomer in  solution (2) are each not less than 1.5.

Wavelength                   : 278nm

Temperature       :Ambient

Injection volume : 20µL

Mobile Phase       : A mixture of 38 volumes of Methanol & 62 volumes of

                               0.2M Ammonium dihydrogen orthophosphate.

 Solution (1) :- Disperse 10 tablets in 0.2M Ammonium Dihydrogen Orthophosphate

previously adjusted to pH 2.4 with Orthophosphoric acid using 10ml per gm of the stated content of cefuroxime . Immediately add sufficient methanol to produce a solution containing the equivalent to 0.5%w/v of cefuroxime and shake vigorously.

Filter and dilute a quantity of the filtrate with sufficient mobile phase to produce a

solution containing the equivalent to 0.025% w/v of cefuroxime.

Solution (2) :- Heat a quantity of solution(1) at 60⁰ C for 1hour or until sufficient impurities (   ⁰   -isomers) have been generated.

 Solution (3) :- Expose  a quantity of solution(1) to ultraviolet light (254nm) for            24 hours or until sufficient impurities (E-isomers) have been generated.             Solution (4) contains 0.03% w/v of Cefuroxime Axetil CRS in mobile phase.

  All solutions containing Cefuroxime Axetil, if not to be used for immediate

  analysis, should be stored in the dark at a temperature 2º & 8º before analysis.

The retention times relative to Cefuroxime axetil diastereoisomer A are approximately 0.9 for Cefuroxime axetil diastereoisomer B , 1.2 for the Cefuroxime

axetil  A³-isomers and 1.7 & 2.1 for E-isomers .The relative standard deviation for the

response factor of Cefuroxime axetil for replicate injections of solution (1) is not more than 2.0%.

Calculate the content of C₂₀H₂₂N₄O₁₀S as the sum of the areas of the two peaks  corresponding to diastereoisomers A & B of Cefuroxime axetil using the declared

Content of C₂₀H₂₂N₄O₁₀S in Cefuroxime axetil CRS. Each mg of Cefuroxime axetil

Is equivalent to 0.8313mg Cefuroxime.