Table 3.

Experimental evidence derived from acetazolamide-loaded nanoparticulate systems

Hypotensive drug/nanosystem	Pharmaceutical form	Study design/model	Results	References
Eudragit nanoparticles (NPs) of ACZ incorporated into an ocular insert	Eudragit nanoparticles	In vitro drug diffusion study Ex vivo transcorneal permeability study (excised fresh goat corneas) In vivo ocular tolerability and IOP reduction study (albino rabbits): animals received drinking water (40 ml/kg of body weight of rabbit) to increase IOP 3 groups: (1) ACZ reference solution, (2) Eudragit NPs dispersion, and (3) ocular insert of Eudragit NPs	Ex vivo transcorneal permeation study showed the following results. Flow across corneal tissue (µg/min): drug suspension. 0.671 ± 0.020; NPs suspension. 2.460 ± 0.028; ocular insert, 2.402 ± 0.032, which means that ACZ-loaded Eudragit NPs displayed better permeability and flow across corneal tissue than the drug suspension In vivo studies with optimized ACZ loaded Eudragit NPs and ocular insert demonstrated substantial IOP lowering and improved ocular tolerability when compared to ACZ suspension	[125]
ACZ-loaded nanoparticulate in situ gels (NP-ISG)	Polymeric nanoparticles with Eudragit RL100, Eudragit RS100, or poly(lactide-co-glycolide) 75:25 (PLGA)	Ex vivo transcorneal permeability study (fresh goat cornea)	Ex vivo transcorneal permeation study displayed higher ACZ permeation at 8 h with NP10 (93.5 ± 2.25 mg/cm2) and with NP-ISG5 (74.50 ± 2.20 mg/cm2) than with ACZ eye drops (20.08 ± 3.12 mg/cm2) and ACZ suspension (16.03 ± 2.14 mg/cm2)	[126]
		Ex vivo corneal toxicity study (fresh goat cornea)	NP-ISG did not display harmful effects on any corneal layer
		In vivo pharmacodynamic activity study (normotensive rabbit)	1% ACZ nanoparticulate in situ gel exhibited greater IOP-lowering effect 1 h after administration, which was sustained for up to 8 h, while 1% ACZ eye drops only sustained its action for approximately 2 h
ACZ-loaded water-soluble mucoadhesive carbosilane dendrimers	C–Si backbone (carbosilane) cationic dendrimers	In vitro (cytotoxicity and cell viability) investigation (using telomerase-immortalized, human corneal-limbal epithelial cell line, HCLE)	Generations 1 and 2 of the cationic dendrimers and all 3 generations of the anionic dendrimers were well tolerated at 10 μM, with higher than 80% cell survival for all of them, except for the G3-C (from the 3rd generation of carbosilane cationic dendrimers)	[127]
		In vivo (ocular tolerability) study (normotensive New Zealand rabbits): specular microscopy, slit lamp examination, and IOP measurements	Formulation containing ACZ 0.07% (289.4 mOsm; 5.6 pH; 41.7 mN/m) and G3 cationic carbosilane dendrimers (5 μM) demonstrated the best IOP-lowering effect. It obtained a rapid (1 h post-instillation) and sustained (up to 7 h) hypotensive effect, reaching a peak 22.6% IOP reduction
ACZ-loaded hyper branched poly(propylene imine) (PPI) dendrimers	PPI dendrimers	In vitro drug release studies Ex vivo studies (effect on the morphology of human erythrocytes) In vivo studies (normotensive New Zealand rabbits): determination of ocular irritation index, ocular residence time, IOP reduction (25 µL of dendrimer formulation was administered into the lower cul-de-sac of the eye)	Hemolytic toxicity study showed a slightly higher hemolysis rate of dendrimer formulations (D1 = 4.8%, D2 = 5.6%, D3 = 7.2%) when compared to plain drug (3.3%) and plain 5.0G PPI dendrimer (7.9%) Plain ACZ solution produced IOP reductions up to 2 h after instillation, whereas the dendrimer-based formulation lowered IOP for longer (4 h)	[128]
ACZ-loaded ion-activated nanoemulsion-based in situ gelling systems using gellan gum polymer alone and in combination with other polymers (xanthan gum, hydroxymethylcellulose, or Carbopol)	Nanoemulsion Gellan gum (in various concentrations: 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, and 0.6%)	In vitro release studies	Optimized formula of ion-induced nanoemulsion-based in situ gels demonstrated a significantly sustained drug release profile	[129]
		In vivo studies: ocular irritation and pharmacodynamic studies in adult male New Zealand albino rabbits	Ocular irritation study revealed no damage to the ocular surface and other parts of the eye Area under the curve of percentage decrease in IOP versus time from 0 to 10 h: nanoemulsion formula = 189.15 ± 10.18; Azopt® = 82.51 ± 7.53, and Cidamex® = 79.77 ± 7.58
ACZ-loaded Eudragit RL100 nanoparticle suspension (ACZ-E-NPs)	Polymeric nanoparticles with Eudragit RL100	In vitro drug release study In vivo studies (albino rabbits):   Group 1 received 0.5% ACZ solution   Group 2 received formulation E3 (drug polymer ratio 1:10; organic aqueous phase ratio 1:4; organic phase acetone)   Group 3 received formulation E8 (drug polymer ratio 1:10; organic aqueous phase ratio 1:3; organic phase acetone and ethanol)	Plain solution of ACZ was able to lower the IOP for about 3 h after instillation, whereas ACZ-E-NP solutions showed a greater efficacy (IOP lowering for up to 8 h after instillation). The peak IOP reduction by plain ACZ solution was 2.98 ± 0.11 mmHg at 2 h after topical administration, whereas best ACZ-E-NP formulations (E3 and E8) progressively reduced IOP, peaking at 8 h after instillation (F3 = 5.32 ± 0.07 mmHg; E8 = 5.19 ± 0.06 mmHg; mean ± SD)	[130]