Abstract: Recognizing the limited research on microneedles for the transdermal potent oral hypoglycemic drugs, we formulated, in this study, glimepiride nanocrystals and incorporated these nanocrystals in PVA/sucrose based dissolving microneedles. Nanocrystals were formulated using two different aqueous surfactant solutions as antisolvents; PVP (0.2% or 2.5%) and 0.2% SLS with grinding after formulating and drying the nanocrystals, or with grinding step. Glimepiride nanocrystals using 0.2% SLS with grinding showed the smallest particle size of 348 ± 27 nm with a PI of 0.29 ± 0.05, and those were confirmed with SEM imaging. Therefore, SLS with grinding formula was used to be incorporated into PVA (23% w/v)/sucrose (15% w/v) microneedles formula (GLIM_MN). GLIM_MN showed a microneedle height of 500 ± 14 and a sharp tip, as shown with SEM imaging. GLIM_MN showed ability to withstand an external force exerted by 500 g placed over the microneedle array. Moreover, GLIM_MN successfully penetrated 3 layers of parafilm, as analogue to the mechanical properties of the skin, and successfully penetrated in vivo healthy BALB/c mice skin to a depth of 480 ± 15 µm, as shown by the digital microscope image after the insertion of the microneedle array. Moreover, GLIM_MN showed full penetration of the array into excised human skin to a depth of 276 ± 65 µm, as appeared by the digital microscope image. Finally, we monitored the glucose level in healthy BALB/c mice for over 90 minutes. GLIM_MN showed significantly greater hypoglycemic effect compared to glimepiride gel of similar concentration, a comparable hypoglycemic effect to oral administration. This also corroborated with an increase in serum insulin in GLIM_MN treated groups compared to the untreated, after 5 hours from administration. Moreover GLIM_MN treated mice showed no significant change in the serum C-reactive protein than the untreated, indicating no inflammation with microneedle administration. These results collectively indicate that our glimepiride dissolving microneedles formulation could enhance the delivery of glimepiride with minimal invasiveness, and therefore can be a potential treatment in the management of type 2 diabetes mellitus.​