Formulation and Evaluation of Stability of a Topical Niosomal Gel ContainingCiclopirox Olamine

Introduction

Fungal diseases can be life-threatening for malnourished and immunocompromised people. Infections of the nail and skin can be managed by topical antifungal drugs. However, in some cases such as primary and secondary bacterial infections, deeper penetration of the antifungal drug is important.^1,2 The drug particles in traditional topical formulations are not capable of penetrating into the skin’s deeper layers, such as dermis and epidermis.³ An extremely effective active pharmaceutical ingredient (API) that can permeate through the skin and nail is needed to maintain the quantity of the drug well above the required minimum inhibitory concentration (MIC) to ensure successful treatment.4 The transdermal route is the best form of drug delivery, but due to the powerful barrier posed by the stratum corneum (epidermis), very few drug candidates are capable of entering the systemic circulation. A variety of antifungal drug delivery strategies using nanocarriers such as niosomes, liposomes, ethosomes, etc., have been developed to promote transportation of the drug to the affected area, resulting in enhanced antifungal activity.⁵

Niosomes are nonionic surfactant-based vesicles with a bilayer structure containing lipophilic, amphiphilic, and hydrophilic moieties that self-assemble in aqueous solutions.³ Surfactants facilitate total penetration of compounds mainly through adsorption at interfaces, interaction with biological membranes, and changes in the barrier structure of the stratum corneum via reversible lipid modifications.

Ciclopirox olamine (CPO) is a broad-spectrum antifungal agent against yeasts, bacteria, dermatophytes, and fungal infections. It is a hydroxypyridone with a mechanism of action that is different from many other antifungal agents, such as azoles and allylamines.⁶

The assessment of physical and chemical stability of topical products, such as gel, cream, etc., is vital when planning the preparation, storage, and application of these formulations. In this present study, we focused on the development of a topical niosomal gel formulation containing CPO, and evaluation of its stability.

Materials and Methods

A gift sample of CPO was offered by Kumar Organic Products Ltd., Bangalore, India. Pure-grade reagents were purchased from Sigma Laboratories, India. Dialysis membrane-70 with an average flat width of 2229.31 mm was purchased from HI-Media Laboratories Pvt Ltd. All the chemicals and compounds were procured in reasonable quantities from authentic sources.

Preparation of CPO-packed niosomal gel

Preparation of vesicles

CPO niosomes were prepared using Span 60 as the surfactant with cholesterol with a drug: surfactant: cholesterol ratio of 1:2:0.2, using thin-film hydration technique. The lipid solution was transferred to a 100-mL round bottom flask at 55-65° C. The solvent was evaporated under reduced pressure with the help of a rotatory evaporator for 1 h until a thin lipid sheet was developed. The lipid sheet was hydrated with 20 mL phosphate-buffered solution having a pH of 7.4. The hydrated niosomes were sonicated with the help of a bath sonicator for 20 min to obtain a niosomal dispersion containing both entrapped and free drug particles of different sizes.^7-9

Preparation of niosomal gel

Carbopol 934P (2% w/w) was dissolved in an adequate amount of water and soaked overnight in the dark. It was then mixed with 1% CPO-encapsulated niosomal suspension along with 0.1% methylparaben as a preservative and 1% glycerol as a penetration enhancer. Later, it was neutralized with sodium hydroxide, changing the pH to 6.8 assisted by agitation.^10-12

Evaluation of niosomal gel formulation

Accelerated stability studies

The CPO niosomal gel was stored in sealed glass containers and preserved under refrigerated and room temperature conditions (25°C ± 2°C/60% ± 5% relative humidity [RH]) for 6 months. ^13-16 An accelerated stability evaluation of the gel was performed as per the ICH guidelines. Different parameters, which included physicochemical properties; in vitro drug release; and drug content on the first day and the last day of 4th and 8th weeks, as well as the 3rd and 6th months of the storage period, were used to assess stability.

Physical appearance

The gel was tested for clarity, color, uniformity, and the presence of foreign particles.^17,18 

pH of the gel

Exactly 2.5 g of the gel was weighed and dissolved in 25 mL of distilled water. The pH was recorded with a digital pH meter. Triplicates were used for measuring the performance.^17-19

Content uniformity

The drug content in the CPO niosomal gel was checked by dissolving correctly weighed gel quantity equal to 10 mg of the drug and 100 mL methanol in a 100 mL volumetric flask. The niosomal gel content was measured by recording its optical density (OD) at 301 nm against blank, using Shimadzu ultraviolet (UV)/ visible spectrophotometer. CPO calibration curve was used to calculate the contents of the drug.^17,19

Rheological study

Brookfield programmable DV III ultra-viscometer was used for determining the viscosity of the formulation.20 Experiments were performed in triplicates.

In-vitro permeation studies: The optimized CPO niosomal gel (10 mg) was kept between the lower (receptor) and donor compartments of the Franz Cell apparatus. The diffusion cells were filled with phosphatebuffered saline (pH 7.4, containing 10% v/v methanol) maintained at 37°C ± 0.5°C throughout the experiment, with continuous stirring at 50 rpm on a magnetic stirrer. After 24 h, the samples were taken out, and the quantity of the drug in the gel was calculated based on the OD at 306.20 nm on a UV spectrophotometer. Again, the experiments were performed in triplicates.^21-23

Fourier Transform Infrared Spectroscopy (FTIS)

The potassium bromide dispersion procedure was used to analyze CPO’s infrared (IR) spectrum on the FTIS (Model Name: FT/IR-4100 type-A). Correction of the baseline was performed with dry potassium bromide.

The IR absorption spectrum, within the wavelength range of 4000-400 cm^-1, of the potassium bromide and drug combination was tested, followed by that of the drug with excipients.²⁴

The key goal was to ensure the safe storability of a solid-state drug and also to determine the appropriate excipients in the formulation. The integration of the excipient and drug in the formulation is required for assessing the similarity between the two. The actual mixture of the medication and other excipients was taken as a test sample in the IR studies.^25,26 

Results and Discussion

Stability tests were conducted by storing niosomal gel in glass containers, which were sealed and preserved for 6 months under refrigerated and room temperature conditions. The samples were taken out and analyzed for different parameters on the 28th day (4 weeks), 56th day (8 weeks), and end of the 3rd and 6th months. No significant changes were observed in the physicochemical properties such as viscosity, release profile, and drug content of the niosomal gel when stored in different temperature and humidity conditions, indicating its stability (Tables 1 and 2).

Physicochemical analysis of niosomal gel

The niosomal gel was kept under normal temperature and refrigerated conditions for the evaluation of its appearance, homogeneity, texture, grittiness, and pH. It showed good stability after 4th and 8th weeks, and 3rd and 6th months in the accelerated conditions. There was no statistically significant difference in any of these parameters after the tests (Tables 1 and 2).

Spreadability study

The consistency of a semi-solid formulation determines its ability to spread evenly on a surface after a certain amount of time. Topical products should be simple to apply on the skin surface. Assessment of the spreading potential of the niosomal gel formulation at accelerated conditions revealed no significant variations before and after the storage duration. 

Rheological study

The viscosity of a formulation will depend on its physicochemical characteristics and temperature conditions. This parameter was analyzed to determine the required quality and fluidity and also the product output overtime. The results showed that there was no notable change in the viscosity of the niosomal gel at the end of the 4th week, 8th week, 3rd month, and 6th month of storage.

Content uniformity

The residual drug content was determined on the 28th day (4 weeks), 56th day (8 weeks), and the end of the 3rd and 6th months. Minimal drug leakage from the vesicles was observed at 4°C±2°C and 25°C±2°C. This could be due to lipid phase transition and surfactanttriggered vesicle loss during preservation at higher temperatures. Based on these findings, it was concluded that the optimal storage condition for niosomal gel was 4o C (Tables 1 and 2). 

In-vitro permeation studies

There was no noticeable change in the in-vitro drug diffusion study of the niosomal gel at 4°C±2°C and at normal temperature (25°C±2°C/60%±5% RH) after 6 months (Tables 1 and 2). However, after stability at 25°C±2°C/60%±5% RH, there was a decrease in the in vitro diffusion when compared to 4°C±2°C. This can be attributed to the influence of temperature, as well as the significant chemical degeneration of the drug during the lipid bilayer gel-to-liquid transition.

IR analysis

Characteristic peaks in the IR spectrum of the pure form of CPO were observed for:  C-H stretch at 2852.97 cm^-1, O-H stretch at 3415.00 cm^-1, C=C stretch at 1592.38 cm^-1, C-C stretch at 1450.22 cm^-1, N-H wag at 893.24 cm^-1, and C=O stretch at 1634.31cm^-1. Similar characteristics were also observed in the IR spectrum of the CPO-containing niosomal gel (peaks at: 3424.96 cm^-1 for O-H stretch, 2919.70 cm^-1/ 2851.24 cm^-1 for C-H stretch, 1586.27 cm^-1 for C=C stretch, 1649.80 cm-1 for C=O stretch, 1465.63 cm^-1 for C-C stretch, and 888.05 cm^-1 for N-H wag) (Table 3). The FTIR spectrum of niosomal gel formulation clearly shows the retention of these peaks of the pure CPO drug (Figs 1 and 2). If the structure of the drug is not disturbed during development of the niosomal gel formulation, there will be no interaction between selected polymers and the drug.

Conclusion

The development of CPO niosomal gel formulation as a topical drug delivery system with the use of 2% Carbopol 934P as the gel base with 1% glycerol and 0.1% methylparaben was successful. It has also been also concluded that storage and preservation at different temperatures and conditions does not changes its stability and there is no interaction between selected drug and polymer.