Specific determination of unbound oxacillin in protein solution with cefoperazone by high-performance frontal analysis with chemiluminescence detection

Unbound oxacillin concentrations in human serum albumin (HSA) solutions in the presence or absence of cefoperazone were determined using high-performance frontal analysis coupled with chemiluminescence detection (HPFA-CL). The HPFA was performed on an ISRP column with 67 mM potassium phosphate buffer of pH 7.4 and ionic strength of 0.17 as the mobile phase. The luminol-H(2)O(2)-Co(2+) system was employed in the chemiluminescence detection. The detection was highly specific for oxacillin in the presence of cefoperazone. Although both drugs in HSA solutions co-eluted in the same region in HPFA, cefoperazone did not interfere with the determination of unbound concentration of oxacillin. In the solution of 100 microM HSA and 11.33 micro M oxacillin the bound percentage of oxacillin to HSA was estimated as 80.5%. Addition of 30.98 micro M cefoperazone into the HSA-equilibrated solution produced little effect on the protein binding of oxacillin. In the presence of 154.9 micro M cefoperazone, however, the bound percentage of oxacillin was significantly reduced. This specific method could be applied to the investigation of drug-drug interaction in protein binding.