A rapid back flushed (BF) direct sample injection (DSI) high-performance liquid chromatography (HPLC) with UV detection (BF-DSI-HPLC-UV) has been developed to determine terbinafine (TERB) in human serum. For online solid phase extraction step, an isocratic mobile phase of phosphate buffer saline (pH 7.4) at 1 mL/min and a short protein-coated ODS column (PC-ODS-column) were used for the purification and enrichment of TERB. Two different chromatographic modes of PC-ODS-column were simultaneously operated. Macromolecular proteins were extracted by size-exclusion liquid chromatography, while TERB trapping and enrichment were achieved through reversed-phase liquid chromatography. The clear fraction containing TERB was transferred from the PC-ODS-column by BF mode onto the quantification step through a high pressure switching valve. An analytical mobile phase consisting of 80% methanol and 1% triethylamine in distilled deionized water (pH) 6 at 1 mL/min was used for the final separation on an ODS analytical column. TERB was quantified and detected by UV-detector at 224 nm. The proposed method showed high correlation coefficient (>0.999) over the concentrations range 4-1600 ng/mL with recoveries ranging from 98.48 to 93.86%. Measurement of TERB concentration in serum after administration of a single dose of 250 mg oral tablet was used to evaluate the applicability of the BF-DSI-HPLC-UV for pharmacokinetic study.
Keywords: Back flushed elution; Human serum; Pharmacokinetic study; Protein-coated ODS column; Terbinafine.
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