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Ceftriaxone stability and sensitive analysis by fully validated ion-pair HPLC assay in human plasma

Rajaa Farhan Hussein, Muhammad M.Hammami

We validated a sensitive, precise, and accurate HPLC assay for ceftriaxone measurement in human plasma and studied the stability of ceftriaxone.After protein precipitation with methanol at 4ºC, ceftriaxone and cefazolin (internal standard)were eluted onXTerra® RP18, 5 µm steel column at room temperature (RT). The mobile phase consisted of 0.02 M cetyltrimethylammonium bromide in 0.01 M dipotassium hydrogen orthophosphate buffer (pH = 6.5, adjusted with phosphoric acid), acetonitrile, and triethylamine (70:30: 0.001, v:v:v)with a run time of 14min. The analytes were detected using 2998 photodiode array detector set at 272 nm. The response was linear over the range of 0.2–200 µg/ml. Extraction recovery and inter-run bias and precession were = 94% (mean 96%), = 8%, and = 5.7%, respectively. Ceftriaxone was stable in plasma for 24 hours at RT (= 95%), 8 weeks at –200C (= 97%), and after 3 cycles of freeze at –200C and thaw at RT (= 91%). In processed samples, ceftriaxone was stable for 24 hours at RT (= 96%) and 48 hours at –200C (= 91%). Stock solution of ceftriaxone (1 mg/ml in water) was stable for 48 hours at RT (100%) and 8 weeks at 200C(96%).