Method development and validation of LC and densitometric-TLC for simultaneous determination of pitavastatin calcium and ezetimibe in a binary mixture.

Two simple and accurate methods to determine pitavastatin calcium (PIT) and ezetimibe (EZE), in a binary mixture, were developed and validated using LC and densitometric-TLC. The LC separation was achieved on a Phenomenex Luna C18 column (250 x 4.6 mm id, 5 microm), in the isocratic mode using 0.1% orthophosphoric acid-acetonitriletriethylamine (19.8 + 80 + 0.2, v/v/v), pH 3.00 +/- 0.05, at a flow rate of 1.4 mL/min. The retention times were about 6.98 and 2.36 min for PIT and EZE, respectively. Quantification was achieved with a photodiode array detector at 235 nm over the concentration range of 0.5-5 microg/mL for each drug, with mean recoveries of 99.35 +/- 0.19% and 99.51 +/- 0.23% for PIT and EZE, respectively. The TLC separation was achieved on silica gel 60 F254 HPTLC plates using toluene-ethyl acetate-glacial acetic acid (59.8 + 40.0 + 0.2, v/v/v), as the mobile phase. The R(f) values were about 0.38 and 0.58 for PIT and EZE, respectively. Quantification was achieved with UV detection at 235 nm over the concentration range of 50-600 ng/spot for each drug, with mean recoveries of 99.49 +/- 0.15% and 99.58 +/- 0.10% for PIT and EZE, respectively. Both methods were validated, and based on statistical results were found to be specific, accurate, precise, and robust. The methods were successfully applied for the determination of PIT and EZE in a binary mixture without any interferences from common excipients.