Development of Practical HPLC Methods for Analysis and Quality Assessment of the Novel Carbonic Anhydrase Inhibitor MK-0507 and the Acetamidosulfonamide Intermediate

Abstract

MK-0507 (Dorzolamide HCl, (4S,6S)-4-Ethylamino-5,6-dihydro-6-methyl-4H-thieno-[2,3-b] thiopyran-2-sulfonamide 7,7-dioxide hydrochloride) is the first topically active, water-soluble, carbonic anhydrase inhibitor to be developed for the treatment of glaucoma and ocular hypertension. Dorzolamide HCl is an effective and well tolerated agent as monotherapy for patients who cannot tolerate ophthalmic beta-blockers, and for those who need add-on therapy to beta-blockers.¹ The steps taken in the development and validation of a fast and rugged reverse-phase HPLC method for the analysis and quality assessment of MK-0507 drug substance and acetamidosulfonamide intermediate are described. Four columns were used during the development: Du Pont Zorbax C-18, Rainin Microsorb C-8, Perkin Elmer CR-C8 and YMC4. The last two columns showed excellent resolution, peak shape, and precision. The selected HPLC method for acetamidosulfonamide, using the Perkin Elmer CR-C8 column, was validated with respect to linearity, limit of detection (LOD) and limit of quantitation (LOQ). The selected HPLC method for the MK-0507 drug substance using the Perkin Elmer CR-C8 column was validated with respect to linearity, LOD, LOQ, precision on an injection-to-injection basis, and on a day-to-day basis, selectivity and accuracy.

The method is rugged with respect to solution stability, variations in buffer concentration, flow rate, column-to-column variability and column temperature. The relative response factor for the cis-isomer of MK-0507 and the desethyl impurity were also determined in order to quantitatively measure the levels of these impurities in MK-0507. There are three possible stereoisomers of MK-0507 (4S,6S): the enantiomer 4R,6R and the cis-diastereoisomers 4S,6R and 4R,6S. The chiral HPLC method separates the two trans-diastereoisomers, and the two cis-diastereo-isomers; however, in the same chiral method the 4R,6R and 4R,6S stereoisomers coelute.