Validated HPTLC and HPLC methods for determination of fluorometholone and sodium cromoglycate in presence of their impurities and degradation products; application to kinetic study and on rabbit aqueous humor

ABSTRACT

Fluorometholone (FLM) and Sodium Cromoglycate (CMG) are co-formulated in ophthalmic preparation and showed marked instability under different conditions. Two specific, sensitive and precise stability-indicating chromatographic methods have been developed and validated for their determination in the presence of their degradation products and FLM impurity. Ten components were efficiently separated by them. The first method was HPTLC-spectrodensitometry, where the separation was achieved using silica gel 60 F₂₅₄ HPTLC plates and developing system of ethyl acetate: methanol (9:1, v/v). The second method was a reversed phase HPLC associated with kinetic study of the degradation process and was successfully applied for determination of the studied compounds in spiked rabbit aqueous humor. The mobile phase was acetonitrile: methanol: 0.05 M potassium dihydrogenphosphate (0.1% trimethylamine); pH 2.5, adjusted with orthophosphoric acid (20: 30: 50, by volume). In both methods, the separated components were detected at 240 nm and system suitability was checked. Good correlation was obtained in the range of 0.10–24.00 and 0.20–48.00 µg band⁻¹, for FLM and CMG by HPTLC. While for HPLC, the linearity ranges from 0.01–50.00 and 0.05–50.00 µg mL⁻¹ for both drugs. The methods were applied in pharmaceutical formulation, where they were compared to the reported method with no significant difference.

GRAPHICAL ABSTRACT

KEYWORDS:

• Fluorometholone
• HPLC
• HPTLC
• sodium cromoglycate
• stability indicating