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Pioglitazone (PGZ) is an oral anti-hyperglycemic agent, belongs to the class of thiazolidinediones, and is used for the treatment of diabetes mellitus type 2. In recent years, its anti-inflammatory activity has also been demonstrated in the literature for different diseases, including ocular inflammatory processes. Additionally, this drug belongs to Class II of the Biopharmaceutical Classification System, i.e., slightly soluble and highly permeable. The main objective of this study was to validate a new analytical HPLC-MS/MS method to quantify free-PGZ and PGZ from polymeric NPs to conduct nanoparticle application studies loaded with this active ingredient to transport it within ocular tissues. An accurate, sensitive, selective, reproducible and high throughput HPLC-MS/MS method was validated to quantify PGZ in cornea, sclera, lens, aqueous humor, and vitreous humor. The chromatographic separation was achieved in 10 min on a Kinetex C18 column. Linear response of PGZ was observed over the range of 5-100 ng/mL. The recovery of free-PGZ or PGZ from NPs was in the range of 85-110% in all tissues and levels tested. The intra-day and inter-day precision were &lt;5% and &lt;10%, respectively. The extracts were shown to be stable in various experimental conditions in all matrices studied. The range of concentrations covered by this validation is 80-1600 µg/kg of PGZ in ocular tissues. It is concluded that this method can be applied to quantify PGZ for in vivo and ex vivo biodistribution studies related to the ocular administration of free-PGZ and PGZ from nanoparticles.

Development and Validation of an HPLC–MS/MS Method for Pioglitazone from Nanocarriers Quantitation in Ex Vivo and In Vivo Ocular Tissues