Quantitative analysis

A reverse-phase high-performance liquid chromatography (HPLC) method was developed to simultaneously detect sumatriptan succinate and metoclopramide HCL in the samples. The method was developed using an ODS-AQ SS 120 Å column (4.6 × 250 mm) with an injection volume of 10 µL and a flow rate of 1 mL/min. The mobile phase was composed of methanol with 0.3% THF as an organic phase and 10 mM potassium phosphate buffer at pH 4 as buffer phase, injected in a 40:60 ratio. The detection wavelength for both, was 279 nm. The method was validated through inter-day and intra-day accuracy and precision test, as well as linearity, limit of detection (LOD), and limit of quantification (LOQ).

Saturation solubility

To determine the optimal donor concentration and the most suitable receptor solution for sumatriptan succinate and metoclopramide HCL, their saturation solubility was evaluated in various vehicles. First an excess amount of the drug added to the vehicle and placed on a platform shaker to be shaken overnight. After that, it was centrifuged, filtered, and analyzed in HPLC to determine the amount of drug dissolved in the vehicle. However, if the solubility of the drug in the vehicle is found to exceed the intended donor concentration, this process would not be performed. Various vehicles, including 10% dimethyl isosorbide (DMI), 15% propylene glycol (PG), 15% isopropyl myristate (IPM), 15% dimethyl sulfoxide (DMSO), 20% IPM, 20% PG, 25% PG, 15% DMI, and their combinations in 10 mM phosphate-buffered saline (PBS), were evaluated.

Preparation of hydrophilic patch

To prepare the base matrix, all components of the patch formulation, excluding the active pharmaceutical ingredients (metoclopramide HCL and sumatriptan succinate), were combined in a scintillation vial with 15 mL of deionized water. This mixture was then placed on a rotary shaker overnight to ensure complete dissolution of all components. The resulting homogeneous solution was carefully poured into 19.63 cm2 Petri dishes. The optimal drying time and temperature for the patches were subsequently determined through experimentation, testing a range of temperatures and durations in a drying oven. Once the optimal drying conditions were identified, drug-loaded patches were prepared. This was achieved by adding the pre-determined amounts of metoclopramide HCL and sumatriptan succinate to the previously prepared base solution.

However, incorporating the positively charged metoclopramide HCL and sumatriptan succinate into the initial aqueous solution resulted in an undesirable interaction with the negatively charged polyacrylic acid (PAA) and sodium polyacrylate (SPA) present in the matrix, leading to agglomeration. To mitigate this issue, orthophosphoric acid was added to acidify the solution. This approach was based on the principle that PAA and SPA become protonated and thus lose their negative charge in acidic environments [21]. While this strategy successfully prevented agglomeration, it was observed that a pH of 2.5 was required for complete dissolution, which is too acidic for a topical patch formulation. Consequently, to ensure proper drug incorporation and maintain a suitable pH for skin application, PAA and SPA were removed from the primary patch matrix formulation. To create the final drug-loaded patch solution, all liquid components, including PG, DMI, PEG, glycerin, and water, were combined with the metoclopramide HCL and sumatriptan succinate in a scintillation vial. This mixture was placed on a rotary shaker overnight to ensure thorough mixing. In a separate step, the powdered components, including mannitol, gelatin, and the chosen polymers (such as HPMC, CMC-Na, or PVP) were weighed. The following day, these pre-weighed powdered components were added to the liquid mixture and the entire solution was returned to the rotary shaker for overnight mixing. The resulting homogenous solution was then poured into Petri dishes and dried in an oven under the pre-determined optimal conditions. Once the patches were dried, a 15% (w/w) PAA solution in methanol was uniformly applied to the exposed surface of each patch. This PAA solution was prepared in advance by combining equal weights of 50 wt% and 25 wt% PAA solutions in methanol. For formulations containing PVP, a modified procedure was employed. PVP was initially dissolved in water by heating at 60 °C for 2 h. Following complete dissolution of the PVP, the remaining liquid components (PG, DMI, PEG, glycerin, and water) were added to the PVP solution and the same procedure as described above was applied. Table 1 details the composition of each patch matrix, with percentages showed as w/w, excluding the weight of water.

Table 1 Composition of the different hydrophilic patch formulations with polyacrylic acid(PAA) solution in menthol cast in both side

To identify a suitable backing membrane, a 15% (w/w) PAA solution in methanol was cast onto various membranes, including CoTran™ 9720, CoTran™ 9728, CoTran™ 9702, CoTran™ 9707, CoTran™ 9706, CoTran™ 9722, and ScotchPak™ 9733, using a Gardner casting knife (BYK-AG-4300 series, Columbia, MD, USA). Both 20 µm and 40 µm thicknesses were evaluated for casting the PAA solution. The optimal membrane was selected based on its ability to allow the PAA solution to form a uniform, stable film without any aggregation, and then it was allowed to dry at room temperature for 30 min. Subsequently, the dried patch matrix was removed from the Petri dish using forceps and applied to the chosen backing membrane. To ensure proper adhesion between the patch matrix and the backing membrane, the patch was applied with its bottom side (the non-PAA-coated side) facing the backing membrane which already had a layer of dried PAA on its surface. The assembled patch was then left at room temperature for 2 h to allow for complete adhesion. Next, to identify a suitable release liner, various liners, including ScotchPak™ 9744, ScotchPak™ 1022, 9709 YDS sample roll SKU, and 27,323 PET, were evaluated. Each liner was placed on top of the PAA-coated surface of the assembled patch, and gentle pressure was applied to ensure complete contact without any air gaps. After 30 min, the release liners were removed using forceps. The optimal release liner was chosen based on its ability to detach cleanly from the patch without disrupting the PAA layer or the patch matrix itself.

Skin source and preparation

Porcine ear skin was obtained and dermatomed using Dermatome 75 µm (Nouvag AG, Goldach, Switzerland). The skin samples were stored at −80 °C until use. Before each study, the skin samples were thawed in 10 mM PBS. The hair on the skin was trimmed with scissors, and the thickness of each sample was measured using a thickness gauge (0–1 in/0–25 mm, Electromatic Equipment Co., Inc., Cedarhurst, NY, USA). Only skin samples with an average thickness of 550 µm were selected for the study.

Measurement of skin barrier integrity

To ensure the integrity of the skin samples, transepidermal electrical resistance (TEER) measurements were performed using a multi-instrument setup. This setup consisted of silver/silver chloride electrodes, an Agilent 33220 A function generator, and an Agilent 34410 A multimeter (Agilent Technologies, CA, USA). Skin samples were mounted onto vertical Franz diffusion cells, and 300 µL of PBS was added to the donor chamber. The TEER values were measured using protocol previously used in our lab [22]. Skin samples with TEER values below 10 kΩ were excluded from the study. To calculate the TEER, a load resistor (RL = 100 kΩ) was connected in series with the skin sample. A constant voltage drop (Vo = 100 mV) was applied across the circuit. The resulting voltage (Vs) was measured, and the skin resistance was calculated. TEER analysis provides a quantitative assessment of barrier function, with higher values indicating tighter cellular junctions and lower permeability, while lower values reflect compromised barriers and increased permeability [23].

In vitro permeation testing (IVPT)

In vitro permeation studies were conducted using vertical static Franz diffusion cells (PermeGear, Inc., PA, USA) with a permeation area of 0.64 cm2. Dermatomed porcine ear skin was mounted onto the cells, and 10 mM PBS was used as the receptor medium at a volume of 5 mL to maintain sink conditions. The skin surface temperature was maintained at 32 ± 1 °C by regulating the receptor chamber temperature to 37 ± 1 °C, using a water jacket connected to a circulating water bath. At predetermined time intervals (0, 1, 2, 4, 8, 22, and 24 h), 300 µL samples were withdrawn from the receptor chamber and immediately replaced with fresh PBS to maintain consistent receptor volume. The drug concentrations in the collected samples were analyzed using HPLC to evaluate drugs permeation profile. For studies involving chemical enhancers, 100 µL of the drug-loaded solution containing the selected chemical enhancers was applied to the donor chamber. In studies with patches, circular patches with a diameter of 0.62 cm2 were punched, and the release liner was removed with forceps. The patches were then applied to the skin surface placed on parafilm, and gentle pressure was applied using a glass rod rolled over the patch to ensure uniform adhesion and contact with the skin. The skin samples with the applied patches were clamped between the donor and receptor compartments of the Franz diffusion cells.

Characterizations of optimized patchesIn vitro release testing (IVRT)

In vitro release testing (IVRT) was conducted using vertical static Franz diffusion cells with a permeation area of 0.64 cm2. Dialysis membranes were used as the synthetic barrier in this study. After mounting the membrane onto the Franz diffusion cell, a 0.54 cm2 section of the patch was placed in the donor compartment. At predetermined time points (0.1, 1, 2, 4, 6, 8, 22, and 24 h), 300 µL of the receptor medium, PBS, was withdrawn and immediately replenished with an equal volume of fresh PBS to maintain sink conditions. The study was conducted in triplicate (n = 3). The collected samples were analyzed using HPLC to quantify the amount of drug released over time.

Coat weight

To calculate the coating weight of the patch, three samples were punched from different areas of the patch using a 0.62 cm2 punch. The punched patches, which included the backing membrane, were weighed. Next, the backing membrane alone was punched three times using the same punch size and weighed separately. The weight of the coating was calculated by subtracting the weight of the backing membrane from the total weight of the patch (including the coating). The results were shown as the mean ± standard error (SE).

Drug content and uniformity

Patches were punched three times from different areas using a punch size of 0.62 cm2. Each punched sample was placed in a scintillation vial, and 15 mL of 10 mM PBS was added. The vials were then placed on a platform shaker overnight to dissolve. After shaking, the solution was filtered through a 0.22 µm filter, diluted 100 times, and analyzed by HPLC.

Tack testing

Tack, a measure of the adhesive's ability to form an initial bond with a different substrate after brief contact and minimal pressure [24] was evaluated using a Texture Analyzer (TA. XT Express, Texture Technologies Corp. and Stable Micro Systems, Hamilton, MA, USA). A stainless-steel cylindrical probe was used to measure the force required for debonding, the positive area, and the separation distance at a speed of 0.5 mm/s, with a return speed of 5 mm/s, and a hold time of 10 s [25].

Folding endurance

Folding endurance was evaluated to determine the mechanical strength and flexibility of the transdermal patches. A strip measuring 2 × 2 cm2 was repeatedly folded at the same point until it broke. This test was performed in triplicate for each formulation, and the average folding endurance along with the standard deviation was calculated [26].

Slide crystallization studies

A slide crystallization study was conducted to determine whether the drug in the patch would crystallize upon vehicle evaporation [27]. The drug-loaded patch solution was cast onto a microscope slide and allowed to dry under a fume hood. The dried sample was then examined under polarized light microscope to assess the presence of any drug crystals.

Statistical analysis

The results from the studies were analyzed using GraphPad Prism software (GraphPad Software, San Diego, CA; version 9.4.1). Data were presented as mean ± standard deviation (SD) with a sample size of 3 or 4 (n = 3 or 4). One-way ANOVA was used to compare the groups. A significant difference between the test groups was determined when the p-value was less than 0.05 [28].