Separation and Determination of Diflunisal and its Impurity by Two Chromatographic Methods: TLC-Densitometry and HPLC.

Abstract

Diflunisal (DIF) has analgesic and anti-inflammatory activity. It is a pharmacopeial drug found in the British Pharmacopoeia (BP), and its major pharmacopeial impurity is biphenyl-4-ol (BPL). DIF has not previously been determined together with BPL. The presence of BPL could significantly affect the dose of DIF in its dosage forms; hence it is crucial to determine DIF and BPL in each other's presence. TLC is the first proposed method, where DIF and BPL were separated on silica gel TLC F254 plates. The eluent was toluene-acetone-acetic acid solution (3.5:6.5:1, v/v). Reversed-phase (RP) HPLC is the second suggested method, where a mixture of DIF and BPL was separated on a C18 (5 µm ps, 250 mm and 4.6 id) column using phosphate buffer pH 4 (0.05 M)-acetonitrile (40:60, v/v). Detection was carried out at 254 nm in both methods. For the TLC method, concentration ranges of 0.5-3 and 0.3-1.7 µg/band were used, with mean percentage recoveries of 100.22% (SD 0.893) and 100.52% (SD 0.952) for DIF and BPL, respectively. The RP-HPLC method was carried out over a concentration range of 5-30 and 2-9 μg/mL, with mean percentage recoveries of 100.10% (SD 1.259) and 98.88% (SD 0.822) for DIF and BPL, respectively. The TLC and RP-HPLC methods were successfully applied for the determination of DIF and BPL, quantitatively, whether in bulk powder or in pharmaceutical formulations. Two chromatographic methods were developed and validated according to International Council on Harmonization guidelines for the assay of DIF and its pharmacopeial impurity.