2.2.4. Rheological Evaluation of the HA-FS-NTF Hydrogel

The rheological property, viscosity, of the HA-FS-NTF and the plain hyaluronic acid gel was measured using a Brookfield Viscometer fitted with a spindle 52. A quantity equivalent to 1 g of the swollen tested hydrogel sample was used to determine the desired parameters, and the run of the spindle was performed at 25 ± 5 ◦C [20]. The findings were recorded over a range of shear rates (2, 10, 20, 30, 40, 50, and 60 s−<sup>1</sup> ) to identify the flow pattern of the formulated gels. Thus, the shear rate (s−<sup>1</sup> ), shear stress (dyne/cm<sup>2</sup> ), and viscosity (cP) were recorded and plotted. The viscosity and degree of pseudoplasticity (Farrow's constant) were determined at a minimum rate of shear (ηmin) and a maximum rate of shear (ηmax) by Farrow's equation (Equation (3)) [21]. *Pharmaceutics* **2021**, *13*, x 7 of 23 cosity (cP) were recorded and plotted. The viscosity and degree of pseudoplasticity (Farrow's constant) were determined at a minimum rate of shear (ηmin) and a maximum rate of shear (ηmax) by Farrow's equation (Equation (3)) [21]. *Pharmaceutics* **2021**, *13*, x 7 of 23 cosity (cP) were recorded and plotted. The viscosity and degree of pseudoplasticity (Farrow's constant) were determined at a minimum rate of shear (ηmin) and a maximum rate

$$\text{LogG} = \text{N}\,\text{LogF} - \text{Log}\,\text{ }\mathbf{I}\tag{3}$$

In vitro release of the entrapped drug from the developed HA-FS-NTF containing

One gram of each sample was loaded in the cylindrical glass tube assembly as described above. The release study was performed in 250 mL phosphate buffered saline (pH 6.8), which was maintained at a physiological temperature (37 ± 0.5 °C) with the rotation of the stirring shaft fixed at 50 rpm. The release study was performed for 3 h, during which time 5-mL aliquots were withdrawn at intervals of 0.0, 0.25, 0.5, 1.0, 1.5, 2.0, 2.5, and 3.0 h. Fresh media were replaced during each withdrawal and maintained at room temperature. The collected samples were filtered using a 0.45-m membrane filter, and the rate of drug release was determined using a UV-visible spectrophotometer at 261.6 nm, with phos-

the 0.5% marketed formulation of fluconazole and the suspension containing 0.5% fluconazole was studied using a Type I USP dissolution apparatus (basket type) (DT 700 LH device, ERWEKA GmbH DT 700, Heusenstamm, Germany). The samples were loaded into the respective cylindrical tubes (10 cm in length and 2.7 cm in diameter) and were attached to the apparatus instead of the basket, with the lower end of the tube closed tightly by a semipermeable membrane with a pore size of 100 µm. Before use, the dialysis membrane was activated by placing the dialysis membrane in boiling water containing 1M NaHCO3 for 1 h. Thereafter, it was washed thoroughly using tap water and stored in phosphate buffer (pH 7.4) for 1 h. The dialysis membranes were kept in the refrigerator in the phosphate buffer (pH 7.4) media overnight prior to performing the release study.

where the shear rate, viscosity, and shear stress are expressed as G, ղ, and F, respectively, and Farrow's constant is expressed as N. where the shear rate, viscosity, and shear stress are expressed as G, LogG = N LogF − Log ղ (3) chamber was calculated using Equation (4): , and F, respectively, and Farrow's constant is expressed as N.

In vitro release of the entrapped drug from the developed HA-FS-NTF containing

the 0.5% marketed formulation of fluconazole and the suspension containing 0.5% fluconazole was studied using a Type I USP dissolution apparatus (basket type) (DT 700 LH device, ERWEKA GmbH DT 700, Heusenstamm, Germany). The samples were loaded into the respective cylindrical tubes (10 cm in length and 2.7 cm in diameter) and were attached to the apparatus instead of the basket, with the lower end of the tube closed tightly by a semipermeable membrane with a pore size of 100 µm. Before use, the dialysis membrane was activated by placing the dialysis membrane in boiling water containing 1M NaHCO3 for 1 h. Thereafter, it was washed thoroughly using tap water and stored in phosphate buffer (pH 7.4) for 1 h. The dialysis membranes were kept in the refrigerator in

the phosphate buffer (pH 7.4) media overnight prior to performing the release study.

One gram of each sample was loaded in the cylindrical glass tube assembly as de-

An ex vivo permeability study was performed using sheep buccal mucous membrane

). The permeation characteristics were evaluated for the devel-

An ex vivo permeability study was performed using sheep buccal mucous membrane

USA). The freshly excised buccal mucous membrane was obtained from the slaughterhouse and kept in phosphate buffered saline. A prepared buccal mucous membrane of 2 × 2 cm was carefully arranged between the acceptor and donor compartments of the dif-

oped HA-FS-NTF containing the 0.5% marketed formulation of fluconazole and the suspension containing the 0.5% fluconazole. Eight milliliters of phosphate buffered saline (pH 6.8) prepared from disodium hydrogen phosphate (Na2HPO4 anhydrous) = 14.4 g sodium dihydrogen phosphate (KH2PO4 anhydrous) = 2.4 g sodium chloride (NaCl) = 80.0 g, and potassium chloride (KCl) = 2.0 g, was kept in the receptor chamber, which was maintained at 32 ± 2 °C with the help of a warm water jacket and stirred at 410 to 430 rpm using the magnetic bead. The samples were autosampled at predetermined time intervals and quantified in parallel using high-performance liquid chromatography (HPLC) [22]. Diffusion of the fluconazole from the donor compartment to the receptor compartment through the mounted mucous membrane was determined by plotting the cumulative release pattern of the drug per unit of time and area. We calculated the diffusion coefficient (D), enhancement factor (EF), permeability coefficient (Pc), and steady-state flux (Jss) to determine whether the developed formulation was superior. Finally, the percentage of fluconazole permeated and the total amount of fluconazole diffused across the receptor

USA). The freshly excised buccal mucous membrane was obtained from the slaughterhouse and kept in phosphate buffered saline. A prepared buccal mucous membrane of 2 × 2 cm was carefully arranged between the acceptor and donor compartments of the dif-

oped HA-FS-NTF containing the 0.5% marketed formulation of fluconazole and the suspension containing the 0.5% fluconazole. Eight milliliters of phosphate buffered saline (pH 6.8) prepared from disodium hydrogen phosphate (Na2HPO4 anhydrous) = 14.4 g sodium dihydrogen phosphate (KH2PO4 anhydrous) = 2.4 g sodium chloride (NaCl) = 80.0 g, and potassium chloride (KCl) = 2.0 g, was kept in the receptor chamber, which was maintained at 32 ± 2 °C with the help of a warm water jacket and stirred at 410 to 430 rpm using the magnetic bead. The samples were autosampled at predetermined time intervals and quantified in parallel using high-performance liquid chromatography (HPLC) [22]. Diffusion of the fluconazole from the donor compartment to the receptor compartment through the mounted mucous membrane was determined by plotting the cumulative release pattern of the drug per unit of time and area. We calculated the diffusion coefficient (D), enhancement factor (EF), permeability coefficient (Pc), and steady-state flux (Jss) to determine whether the developed formulation was superior. Finally, the percentage of fluconazole permeated and the total amount of fluconazole diffused across the receptor

, Teledyne Hanson, Chatsworth, CA,

). The permeation characteristics were evaluated for the devel-

, Teledyne Hanson, Chatsworth, CA,

scribed above. The release study was performed in 250 mL phosphate buffered saline (pH 6.8), which was maintained at a physiological temperature (37 ± 0.5 °C) with the rotation of the stirring shaft fixed at 50 rpm. The release study was performed for 3 h, during which time 5-mL aliquots were withdrawn at intervals of 0.0, 0.25, 0.5, 1.0, 1.5, 2.0, 2.5, and 3.0 h. Fresh media were replaced during each withdrawal and maintained at room temperature. The collected samples were filtered using a 0.45-m membrane filter, and the rate of drug release was determined using a UV-visible spectrophotometer at 261.6 nm, with phos-

2.2.5. In Vitro Release of the FS-NTF-Loaded Hydrogel

2.2.5. In Vitro Release of the FS-NTF-Loaded Hydrogel

phate buffered saline as the blank.

fusion cell area (1.75 cm<sup>2</sup>

2.2.6. Ex Vivo Skin Permeation Studies

chamber was calculated using Equation (4):

by the Franz diffusion cell method (MicroettePlus®

fusion cell area (1.75 cm<sup>2</sup>

phate buffered saline as the blank.

2.2.6. Ex Vivo Skin Permeation Studies

by the Franz diffusion cell method (MicroettePlus®
