[Determination of cefotaxime in human serum by high performance liquid chromatography (HPLC)].
A reversed-phase HPLC assay has been developed for measuring cefotaxime (CTX) in human serum. A solid-phase extraction column (Bond Elute C18) was used for the pre-purification of sample. Serum (200 microL) was mixed with 100 microL of internal standard (acetylamino phenol) solution and 200 microL of sodium acetate buffer (0.01 mol/L, pH 5.8), loaded onto the conditioned column and washed with 2 mL of acetate buffer, the compounds were eluted with 0.5 mL x 2 of methanol-acetate buffer (40:60, V/V ). The eluate (20 microL) was injected directly into an Apex ODS column (250 mm x 4.6 mm i.d., 5 microm) with a mobile phase of acetate buffer (0.01 mol/L, pH 5.8)-methanol (80:20, V/V ), detection was achieved at 254 nm. The average extraction recoveries were 96.7% and 97.7% for CTX and internal standard respectively, which were higher than that of CTX (91.0%) obtained by protein precipitation with methanol. The solid-phase extraction method also provided much clearer samples for HPLC analysis. The concern of acid degradation of CTX when an acid is used as a protein precipitation agent was avoided. The calibration curve of CTX was linear between 10 and 150 mg/L of serum concentration with a correlation coefficient of 0.9992, and the minimum detection limit was 2 mg/L. The precision was tested with three sample concentrations, and within-day RSD below 3.0% and day-to-day RSD below 4.1% were achieved. Maximum serum concentrations of 25.95 and 22.89 mg/L were measured at 45 min after intramuscular injection of 1 g CTX to two healthy volunteers respectively. The chromatographic behaviour of CTX was also studied.
Li
F
F
Shenyang Pharmaceutical University, Shenyang, 110015.
Guo
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L
D000328:Adult; D000900:Anti-Bacterial Agents / Q000008:administration & dosage / Q000097:blood*; D002439:Cefotaxime / Q000008:administration & dosage / Q000097:blood*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D052616:Solid Phase Extraction
D000900:Anti-Bacterial Agents; D002439:Cefotaxime