Simultaneous Estimation of Moxifloxacin and Tolfenamic Acid: Development and Validation of UV-visible, HPLC, and HPTLC Methods

Objectives

This research emphasizes validating the HPLC, HPTLC, and UV-visible techniques for the simultaneous quantitative measurement of Moxifloxacin (MOX) and Tolfenamic Acid (TA) in veterinary formulations.

Materials and Methods

The UV spectrophotometric analysis was performed at 291 nm for moxifloxacin and 272 nm for tolfenamic acid. For HPLC, chromatographic separation was achieved using a gradient technique on a reverse-phase C18 ODS column (250×4.6 mm, 5 µm particle size) with a mobile phase comprising 2% orthophosphoric acid and an acetonitrile with ratio of 80:20, with a 1 mL/min flow rate. Detection was accomplished at 254 nm. For HPTLC, separation was performed on pre-coated silica gel 60 F254 plates using a mobile phase of acetonitrile: water: formic acid: methanol:30% ammonia (8:1:1:1:2 v/v). Densitometric assessment for separate zones was performed at 230 nm.

Results

The HPLC analysis of the current method exhibits the linearity range from 2 to 12 µg/mL for moxifloxacin and 1 to 6 µg/mL for tolfenamic acid in HPLC, 8 to 20 µg/spot for moxifloxacin and 4 to 9 µg/spot for tolfenamic acid in HPTLC, and 6 to 42 µg/mL for MOX and 3 to 21 µg/mL for TA in UV spectrophotometry. The methods were found to be accurate; recovery was achieved from 98 to 102%. The Relative Standard Deviation (RSD) for precision was found to be less than 2%.

Conclusion

The proposed methods were successfully applied to the determination of MOX and TA in both bulk and veterinary tablet formulations. The methods were validated in accordance with ICH Q2-(R1) guidelines.