INTRODUCTION:

Tinea pedis is the widespread form of superficial dermatophytic infection in the developing world, affecting 10 per cent of the population at any given time ¹. In the latter half of the 20th century, global tinea pedis rates rose alongside the clonal spread of the primary culprit, T. rubrum. Tinea pedis often spreads via autoinoculation, leading to related conditions such as tinea manuum, tinea inguinal, and tinea unguium. Improper treatment may further result in bacterial infections and allergic reactions ². Fungal skin infections are prevalent and often treated with topical or oral antifungal medications. Topical treatments are preferred due to the potential side effects associated with oral drugs. Developing an effective topical formulation requires consideration of several factors, including drug absorption through the skin, the formulation's adherence to the skin surface, its reservoir capacity, and patient acceptance³. Every formulation has drawbacks: aerosols and creams may cause mild, temporary stinging, gels require extended application times for effective penetration, thus potentially reducing patient compliance ⁴. So, to improve the penetration, solubility, and bio-availability and to minimize the side effect it could be formulated as liposome, ethosome, trasfersome and an inclusion complex can be made with cyclodextrin.

In the pharmaceutical industry, Cyclodextrins (CDs) primarily serve as complexing agents, forming inclusion complexes with organic and inorganic guest molecules of suitable size and polarity. This alters the physicochemical properties of the guest, offering advantages over conventional systems. These include mitigating issues like oxidative degradation, enhancing solubility of water-immiscible substances in aqueous solutions, protecting against UV damage, stabilizing volatile compounds, converting liquids into powders, and masking unpleasant tastes or odors ⁵. Inclusion complexes are formed when one molecule (the host molecule) encapsulates another. This interaction occurs between the guest molecule and cyclodextrin (CD), allowing partial or complete entry of the guest molecule into the CD's cavity ⁶.

Gels are semisolid formulations consisting of small inorganic or large organic particles suspended in a liquid. They are designed for application to the skin, have a translucent appearance, and can be easily rinsed off with water.

In this study, a gel containing a tolnaftate β-cyclodextrin inclusion complex was developed for the topical delivery of the antifungal drug tolnaftate. Tolnaftate is used to treat superficial dermatophyte infections, various forms of tinea, and pityriasis versicolor ^7,8. It works by inhibiting squalene epoxidase, an enzyme crucial for ergosterol biosynthesis in fungi, thereby preventing their growth. It is available in various forms, such as cream, paste, spray, aerosol, and gel⁹. Though it relieves discomfort from tinea pedis, tinea cruris, and tinea corporis, it also effectively avoids fungal symptoms, including itching and burns. Hence, in this study, an attempt has been made to prepare a topical gel of tolnaftate β-cyclodextrin inclusion complex.

MATERIALS AND METHODS:

Material:

Tolnaftate was obtained from Yarrow Chem Products, Mumbai. β-Cyclodextrin from HiMedia Laboratories Pvt. Ltd and Methanol, Carbopol 934, Disodium hydrogen phosphate, Triethanolamine, Potassium dihydrogen orthophosphate were obtained from Lobachemie, Mumbai.

Methodology:

Preparation of tolnaftate β-CD solid inclusion complex:

The tolnaftate-β-CD solid inclusion complex was prepared by using different techniques, Physical mixtures were also prepared for the comparative studies, which are explained below in detail.

Kneading method:

The tolnaftate-β-CD physical mixture at a 1:1 molar ratio was mixed with a small volume of water-ethanol (10:10 v/v) and kneaded thoroughly with pestle until a homogeneous slurry formed. Kneading continued until all solvent was evaporated. The resulting kneaded products were then placed in a desiccator to remove solvent traces and prepare them for use in gel formulation ¹⁰.

Co-precipitation method:

An inclusion complex of tolnaftate and β-CD was formed at a 1:1 molar ratio. Tolnaftate was dissolved in 5 ml acetone and added dropwise to an aqueous solution of β-CD (120 ml distilled water). The mixture was stirred continuously for 6 hours, and the solvent was evaporated using a water bath. The resulting product was dried in a desiccator to remove solvent traces. Stored in an airtight container for future use ¹¹.

Freeze drying:

An inclusion complex of tolnaftate and βCD was prepared by dissolving tolnaftate in 30 ml methanol and β-CD in 60 ml distilled water at a 1:1 molar ratio. Then this solution was mixed and sonicated for 25 minutes. The solvent methanol was evaporated using a water bath, and the resultant solution was then frozen for 24 hours. Finally, the frozen solution underwent freeze-drying in a vacuum chamber ^12,13.

Characterization of prepared tolnaftate- β-CD solid inclusion complex:

UV visible spectroscopy:

Using a UV-visible spectrophotometer (Jasco-V-630), the absorption spectra of the Tolnaftate, β-CDs, and Tolnaftate/β-CDs inclusion complex sample solution were measured in the size range of 200-400nm ⁶.

Differential scanning calorimetry (DSC):

To validate the inclusion complexes, Differential scanning calorimetry (DSC) thermograms were obtained for Tolnaftate, β-CD, physical mixtures, and solid inclusion complexes (prepared via kneading, co-precipitation, and freeze-drying) using a Shimadzu® DSC 60 instrument. Samples (approximately 2 mg each) were tested in aluminum capsules under dynamic nitrogen (100 ml/min), with a temperature range of 30 to 300⁰C and a heating rate of 10⁰C/min. DSC-60 calibration was performed earlier using >99% pure indium ^14,15.

Fourier transform infrared spectroscopy (FTIR):

FT-IR spectra of CDs, free TNF, their physical mixtures, and their complexes (made from three different techniques) were captured over a scanning range of 4000-400 𝑐𝑚−1 using a Bruker FT-IR spectrometer (alpha brukerInc Germany). After cutting each sample into 1 mg fragments and immersed in 100mg of KBr ^16-18.

Formulation and characterization of a topical gel:

Preparation of topical gel containing an inclusion complex of tolnaftate β- CD:

A 1% Carbopol gel was formulated and to that freeze-dried inclusion complex of tolnaftate and β-CD were added. This was obtained by dissolving 0.5 g of Carbopol 934 in 50 ml of distilled water with magnetic stirring for 2 hours [49]. Subsequently, the freeze-dried inclusion complex of tolnaftate and β-CD, equivalent to 0.5 g of drug, was dissolved in the smallest amount of methanol or an aqueous solution containing a minimal quantity of methanol [46]. This suspension, containing 0.5 g of drug, was added to the 1% Carbopol gel at a 1:1 ratio (w/v). Triethanolamine was then added incrementally to achieve a gel-like consistency during continuous stirring ^19,20.

Characterization of topical gel incorporated with solid inclusion complex:

Physical appearance and measurement of pH:

Visual inspection was used to determine the physical appearance of the formulation. The pH of the gel was measured by dissolving the 1g of prepared gel in 20mL of purified water until a stable solution was obtained and then the pH value was measured using a digital pH meter ^21,22.

Measurement of viscosity:

The Brookfield viscometer (DV-11 +pro D220) measured viscosity by selecting the spindle number T-94 and varying the rpm fitted with an F96 “t bar spindle” with speeds of 5, 10, 20, 50, 100 rpm ^23,34.

Washability:

The washability of the prepared gel was checked by rubbing a small amount on the hand and then wiping it off with water, without using soap ²⁵.

Spreadability:

The gel’s spreadability was determined using a modified wooden block and glass slide apparatus.

𝑆 =M×L/T

Where S = is the spreadability, M = is the weight in the pan (attached to the upper slide), L = is the length transferred by the glass slide, and T = reflects the time required to remove the slide entirely from each other.

An excess amount of gel (about 1 gram) was weighed on this ground slide. The gel was then installed with the hook between this glass slide and a second glass slide that had a predefined ground slide length. A 100g weight was placed on each slide for five minutes in order to eliminate air and create a transparent gel coating in between slides. I scraped away the remaining gel in the corners. After that, the top plate was pulled down by thirty grammes. The duration (measured in seconds) required by the upper slide to move 5 cm with the string fastened to the handle.A shorter time is guaranteed by better spreadability ²⁶.

Drug content:

The drug content of the gel was determined by accurately dissolving 1g of prepared gels in 100 of a solution containing water and methanol in a 1:1 ratio. These solutions were transferred quantitatively to volumetric flasks, and enough dilutions were prepared using the water-methanol solution mentioned above. After that, the solution was filtered and examined at 257 nm using spectrophotometry. The basic curve of tolnaftate was used to calculate the drug content ²⁷.

Drug release studies:

In vitro drug release studies of formulations:

Franz Diffusion cells comprising two compartments were used in the in vitro experiments. There are two open ends to the donor compartment, one of which is sealed with a dialysis membrane dipped in a pH 5.5 phosphate buffer mixed with alcohol. In addition, the hydroalcoholic solutions of the inclusion complex and pure drug containing two mg of the medication each were added. Each dialysis membrane received 0.5 g (or 2 mg of drug) of the solid inclusion complex (GIC) gel and 0.5 g (or 2 mg of drug) of the traditional gel, in that order. Within the reservoir compartment, which contained 12 ml of the buffer mixture, a small magnetic bead spun steadily at 50 revolutions per minute. Operating at 37±0.5⁰C, the experiment lasted eight hours. Samples weighing 1 millilitre were taken from the reservoir compartment at predetermined intervals and appropriately diluted with a 1:1 methanol-to-water mixture. Next, spectrophotometric measurements of absorbance at 257 nm were made. The same volume of new pH 5.5 phosphate buffer containing alcohol was added to the reservoir compartment each time to maintain sink condition ^{3, 28}.

In vitro drug release kinetics:

Evaluation of the first-order kinetics (log average percentage of substance vs. time) and zero-order kinetics (cumulative amount of drug released vs. time) was done using the data from the drug release profile studies. The Korsmeyer-Peppas model, which plots the log cumulative percentage of drug release against log time, and the Higuchi matrix model, which plots the cumulative percentage of drug release against square root of time, were fitted for an in vitro kinetic analysis ²⁹.

Ex vivo drug permeation studies using porcine ear skin:

Ex vivo drug permeation studies used porcine ear skin instead of a dialysis membrane. The porcine ear skin was obtained from a slaughterhouse, and then the hair was separated from the skin and placed in phosphate buffer pH 5.5. The procedure remains the same as in the in vitro drug release studies ^{20, 30, 31}.

Calculation of skin permeation parameters:

The cumulative amount of drug permeated per unit area was plotted as a function of time. The flux was determined based on the linear portion’s slope. The permeability coefficient (Kp) of Tolnaftate through pig skin was determined using the equation below, which is taken from Fick’s first law of diffusion.

𝐾𝑝 = 𝐽/𝐶

Where J is the flux and C is the drug concentration in the donor compartment

Anti-fungal studies:

The agar cup diffusion procedure was used to test the antifungal activity of topical gel formulation conventional gel and GIC, solution of drug and marketed gel (Tinaderm) against Candida albicans. Marketed tolnaftate gel was used for the branded drug (standard). The standard, solution of drug, conventional geland GIC were taken into sterile sabroud dextrose agar cups which is previously seeded with candida Albicans obtained from the Nitte Science Education and Research Centre. After allowing the formulation to extend for 2 hours, the plate was incubated at 25 ⁰C for 48 hours. After 48 hours, the inhibition zones for the test and standard were measured in millimetres ³².

RESULTS AND DISCUSSION:

Formulation and characterization of tolnaftate- β-cyclodextrin solid inclusion- complex

Preparation of tolnaftate β-cyclodextrin solid inclusion complex

A 1:1 molar ratio solid inclusion complex of tolnaftate with β-CD was prepared using 3 different methods, i.e., kneading, co-precipitation and freeze-drying. The prepared complex was dried and kept in a desiccator for further use. Characterization of prepared tolnaftate- β-cyclodextrin solid inclusion complex

UV visible spectroscopy:

This study examined the absorption spectra of -CD, tolnaftate, and solid inclusion complex, which are typically confirmed using UV-visible spectroscopy. As shown in fig 1, the CD exhibited no absorption spectrum over the wavelength range. It was observed that in a methanol-water mixture tolnaftate has a significant maximum absorption peak at 257nm. The highest absorption peak of the solid inclusion complex has shifted from 257nm to 254-256nm, with a slight widening, confirming the formation of the tolnaftate-CD inclusion complex. Lower absorbance indicates that the tolnaftate is encased in the hydrophobic- CD cavity.

Fig. 1: Absorbance obtained by U.V Visible Spectroscopy

Differential scanning calorimetry (DSC):

The Differential Scanning Calorimetry (DSC) thermograms of pure Tolnaftate and its formulations prepared by co-precipitation and freeze-drying methods are shown in Figure 2. The pure drug exhibited a sharp endothermic peak at 115.40°C, corresponding to its melting point, confirming its crystalline nature. In the co-precipitation formulation, the drug showed a broadened melting peak at 112.39°C with decreased enthalpy and an additional minor endothermic transition at 222.92°C, indicating partial amorphization and possible drug–polymer interactions. The freeze-dried formulation displayed a further reduction in the melting peak intensity at 112.26°C and a broad endothermic transition around 213.78°C, suggesting a greater reduction in crystallinity and molecular dispersion of Tolnaftate within the polymer matrix. Overall, the DSC results confirm that both formulation methods reduced the crystallinity of Tolnaftate, with the freeze-drying method exhibiting a more pronounced amorphous transformation than the co-precipitation method, which could contribute to enhanced solubility and dissolution characteristics of the drug.

Fig 2: Differential Scanning Calorimetry (DSC) thermograms of (a) pure Tolnaftate, (b) co-precipitation method, and (c) freeze-drying method.

Fourier transform infrared spectroscopy (FTIR):

The FTIR spectra of pure Tolnaftate, β-cyclodextrin, physical mixture, and inclusion complexes prepared by kneading, freeze-drying, and co-precipitation methods are shown in Figure 3. The spectrum of pure Tolnaftate exhibited characteristic absorption bands at 1599 cm⁻¹ (C=C stretching), 1483 cm⁻¹ (aromatic ring vibration), 1212 cm⁻¹ (C–O stretching), and 752 cm⁻¹ (C–H bending), confirming its molecular structure. β-Cyclodextrin showed broad absorption around 3383 cm⁻¹ due to O–H stretching and peaks at 2921 cm⁻¹ corresponding to C–H stretching of glucopyranose units. In the physical mixture, most of the characteristic peaks of Tolnaftate and β-cyclodextrin were retained with slight intensity reduction, suggesting the absence of significant chemical interaction. However, in the spectra of the co-precipitated, freeze-dried, and kneaded inclusion complexes, the characteristic Tolnaftate peaks were found to be shifted, broadened, or reduced in intensity, particularly around 1599 cm⁻¹ and 1212 cm⁻¹, indicating possible hydrogen bonding and inclusion complex formation between Tolnaftate and β-cyclodextrin. Among the three, the freeze-dried method showed the most prominent changes in the drug’s characteristic peaks, suggesting stronger molecular interactions and more effective inclusion of Tolnaftate within the β-cyclodextrin cavity.

Fig 3: Comparative FTIR spectra of different formulations

Formulation and characterisation of a topical gel:

Preparation of a topical gel containing an inclusion complex of tolnaftate βcyclodextrin:

A topical gel containing a freeze-dried inclusion complex was formulated using 1% Carbopol 934. Carbopol 934 was used as a rate-retardant polymer for the gel preparation. To create a gel-like consistency, a small amount of triethanolamine is added.

Characterization of a topical gel incorporated with a solid inclusion complex:

Physical appearance:

The conventional gel was whitish in colour and whereas the gel consisting of the inclusion complex was transparent. Both the gels were odourless, easy to apply on the skin, smooth, homogeneous and free from grittiness or greasiness.

Measurement of pH , spreadability, washability and drug content:

The pH measurement is an important requirement for topical formulations because it is a significant parameter for enhancing skin penetration. If it differs from the normal skin pH, it might cause skin injury. The pH of the conventional gel was found to be 4.9 and gel containing the inclusion complex was 5.0, which is closer to the skin’s pH. The spreadability of the gel containing the inclusion complex was 14.7 g/cm². Meanwhile, the spreadability of the conventional gel was found to be 17.97g/cm². As a result, the prepared gel is easily spread with a modest amount of shear, indicating good spreadability. The washability test for gel formulations was done three times, and the average was recorded. The gel formulations were easily washable and left no residue on the skin. The drug content of the gel formulations was estimated, and the result showed 88.07% for the gel containing the inclusion complex and 85.03% for the conventional gel. The drug content analysis further indicated that the medication was evenly distributed throughout the formulation. For semi-solid preparation, uniformity of the drug content is required to ensure homogeneity of the distributed drug throughout the formulation.

Measurement of viscosity:

The Brookfield viscometer was utilised to determine the viscosity of the formulated gel at 5, 10, 20, 50, and 100rpm. The viscosity of conventional gel and gel containing inclusion complex was 830.77cps-5230cps and 900.45cps-5190.88cps, respectively. A rheogram was created based on the data collected, demonstrating that each formulation had shear-thinning effects, as shown in Fig 4. With an increase in shear rate, the viscosity of the formulation decreases, indicating pseudoplastic behaviour.

Fig 4: Viscosity of plain gel and gel using freeze-dried inclusion complex

Drug release studies:

In vitro drug release of different formulations:

By the result of the in vitro drug release profile of different formulations and the pure drug shown in Fig 5, we can conclude that the reduced percentage of drug release was triggered by biodegradable lipids owing to a more compact wall around the drug and it displayed a persistent pattern of release over a prolonged period. The percentage cumulative drug release after 480 minutes was greater for the suspension relative to the gel formulation. The percentage cumulative drug release from the solution of the inclusion complex (SIC) was 88.78%, whereas it was 80.49% from the drug solution and 75.27% from the gel formulation consisting of the inclusion complex (GIC). The percentage cumulative drug release from the conventional gel was 80.49% within 480 minutes.

Fig. 5: Comparative in vitro cumulative drug release study of different formulations

In vitro drug release kinetics:

Various kinetic models analysed the release kinetics of the different formulations and pure drugs, and the result is shown in Table 1. The data analysis was focused on the corresponding significance of the regression coefficients. The plain gel formulation showed zero-order kinetics. The pure drug solution, a solution containing an inclusion complex also gel formulation consisting of an inclusion complex showed first-order release kinetics since the regression coefficient (R²) of all the formulations have higher values in comparison with a zero-order kinetic model, which indicates they are capable of providing controlled release profile. The drug release from all formulations was analyzed by fitting data in the exponential model Korsmeyer peppas and the model Higuchi. Good linearity was observed for the final gel formulation with a regression coefficient (R²) of 0.9972 as per the release plotted for Higuchi. The mechanism of drug release followed the Higuchi model having regression coefficient (R²) value higher than korsemeyer peppas model. The Higuchi model showed drug release by swelling of polymers and diffusion through the matrix.

Ex vivo drug release study:

An Ex vivo study has been performed to evaluate the amount of drug permeated through the porcine ear skin. The obtained release profile is seen in Fig 6 and table 2. The drug release exhibited a different trend at the end of 480 minutes compared to the in vitro release model. It thus revealed that the release of the medication from the gel consisting of an inclusion complex (GIC) had a higher permeation of the drug through the skin at the end of 480 minutes. Calculation of Skin permeability parameter The cumulative amount of drug permeated by porcine ear skin after 480 minutes for both hydro alcoholic solution of drug and conventional gel (498.75, 450.3049μg/cm2) was significantly lower compared to a solution consisting of inclusion complex (851.023μg/cm2), suggesting that inclusion complexation could improve the delivery of hydrophobic drugs such as Tolnaftate to the skin. It was found that the improved permeation of Tolnaftate from the solution of the inclusion complex was greater compared to the drug solution, which may be due to the presence of CD, known as a penetration enhancer, along with increased solubilization of tolnaftate resulting from inclusion complexation. As a result, they can easily pass through the stratum corneum's intracellular lipid and bypass the skin's barrier function. When an inclusion complex was present in the solution, the steady-state flux was greater than when the drug was present. Permeability coefficients and steady-state flux were found to be directly correlated. The permeability coefficient of a solution containing an inclusion was also higher than that of the drug solution. The results could be attributed to the high deformability and flexibility of the inclusion complex, which enabled it to overcome the skin barrier properties. The cumulative amount of drug permeated, steady-state flux and permeability coefficient was found to be less in the case of all the gel formulations when compared to the solution of the inclusion complex. It may be due to the viscous nature of the gel, retarded the release of the drug from the formulation. The cumulative amount of drug permeated from the gel containing the inclusion complex was significantly higher than that from the conventional gel.

Fig 6: Ex vivo penetration of drug from various formulations through pig skin

Table 2. Permeated amount of Tolnaftate at 480 minutes, flux and permeability coefficient

Form. code	Permeated amount at 480 minutes (µg/cm²)	Flux (µg/ cm². min)	Permeability constant (Kp) × 10^-3 (cm/min)
Pure drug	919.2045	1.4152	2.8304
SIC	1008.864	1.6978	3.3956
Plain gel	585.2273	1.2378	2.4756
GIC	855.3409	1.7151	3.4302

Anti-fungal studies:

Figure 7 illustrates the antifungal activity of various tolnaftate formulations against Candida albicans using the cup plate method. The zones of inhibition for the marketed tolnaftate gel, Tinaderm (A), drug solution (B), conventional gel (C), and inclusion complex-based gel (D) were recorded as 24.9 mm, 20.9 mm, 22.8 mm, and 24.4 mm, respectively. The inclusion complex-loaded gel exhibited an inhibition zone comparable to that of the marketed formulation, demonstrating enhanced antifungal efficacy. This improvement can be attributed to the formation of the inclusion complex, which likely increased the aqueous solubility and dispersion of tolnaftate, promoting better drug diffusion through the agar medium. Moreover, the in-situ gel matrix may have facilitated sustained drug release and prolonged contact with the fungal cells, thereby maintaining the inhibitory effect. These findings confirm that the inclusion complex-based gel offers a promising approach for enhancing the topical delivery and therapeutic performance of tolnaftate.

Fig. 7: Zone of inhibition A) Tinaderm B) solution of drug C) Conventional gel D) Gel containing inclusion complex

CONCLUSION:

An inclusion complex containing β-cyclodextrin (β-CD) and the antifungal medication tolnaftate was prepared. Significant changes to tolnaftate's chemical, physical, and release properties may result from this inclusion complex. CDs have the potential to improve the drug's absorption and release while also indicating that the drug's effectiveness will be improved.

ACKNOWLEDGEMENT:

The authors thank Nitte (Deemed to be University) and NGSM Institute of Pharmaceutical Sciences for providing the necessary facilities to carry out the study.

CONFLICT OF INTEREST:

The authors declared no conflict of interest.

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Received on 16.07.2025 Revised on 27.11.2025

Accepted on 11.02.2026 Published on 01.07.2026

Available online from July 04, 2026

Research J. Pharmacy and Technology. 2026;19(7):3333-3340.

DOI: 10.52711/0974-360X.2026.00474

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.

Formulation code	Kinetic models
	Zero-order		First order		Higuchi		Korsmeyer-peppas
	R ²	K	R ²	K	R 2	K	R 2	K	n
Pure drug	0.7529	0.0025	0.8726	0.0007	0.904	3.3583	0.9958	0.7362	0.5587
SIC	0.8918	0.003	0.9691	0.002	0.9787	3.8518	0.9978	0.8966	0.4212
Conventional gel	0.9255	0.0022	0.8726	0.0014	0.978	2.755	0.9158	-0.4465	0.8554
GIC	0.9675	0.003	0.9944	0.0012	0.9972	3.7707	0.9258	-0.6383	1.0151