When applied to intact skin, Transdermal Delivery Systems (TDS) are discrete, self-contained dosage forms designed to deliver a drug or drugs through the skin and into the systemic circulation. TDS operate through the process of diffusion, in which the medication enters the main circulation through the skin after diffusing from the drug reservoir either directly or through the rate-controlling membrane and/or contact adhesive¹ Dermal (topical) administration should be utilised solely to define a targeting to diseased areas inside the skin, with limited systemic absorption. This sort of drug localization is crucial in the cure of dermatological disorders such as eczema, microbial infection, skin cancer, psoriasis and eczema, because the disease’s seat is found in the skin.²

Through increasing cycle time, decreasing enzyme degradation, increasing drug stability and water solubility, and increasing target cell or tissue uptake rate, a class of nanomaterials called Nano-Drug Delivery Systems (NDDSs) can improve drug safety and efficacy. NDDSs have a high bioavailability and can be delivered through inhalation, oral administration, or intravenous injection.³

Touitou et al. developed ethosomes, a kind of UDV, in 1997.20 they are also known as elastic Nano vesicles due to their small size (150-200 nm) and remarkable deformability. Ethosomal systems are vesicles composed mainly of water, phospholipids, and a significant quantity of ethanol. Phospholipids are obtained from natural, semisynthetic, and synthetic sources, including soybeans and eggs, and can be used in concentrations ranging from 0.5% to 10%. Phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine, and hydrogenated phosphatidylcholine are a few examples of phospholipids. At amounts ranging from 20% to 45%, ethanol can be used as an effective skin promoter by interacting with the polar head group of SC lipid molecules, this substance minimizes their melting point and enhances the permeability of cell membranes and the fluidity of lipid bilayers.⁴ Ethosomes are safe drug delivery vehicles that enable medications to penetrate underneath the skin layers as well as systemic circulation. Since ethanol is known to disrupt the organization of the skin’s lipid bilayer, its high ethanol content sets the ethosomes apart. Consequently, it permits the vesicle to penetrate the stratum corneum when incorporated into a vesicle membrane. Additionally, because of the high ethanol content, the lipid membrane is enclosed a bit loosely than in typical vesicles, resulting in a more flexible structure and enhanced medication circulation capability in the stratum corneum lipids.^5–9 The structure of Ethosomes was depicted in Figure 1.

Figure 1:
The structure of ethosomes.

Quercetin is a plant flavonoid that is a member of the polyphenol flavonoid family. It can be found in many different fruits, vegetables, grains, seeds, and leaves. Common foods with significant amounts of it include red onions, kale, and capers. It has a bitter taste and is utilised in dietary supplements, drinks, and meals.¹⁰

It is most recognized for its anti-inflammatory and anti-allergy properties. It is frequently given for food and inhalant allergies, asthma, eczema, psoriasis, gout, and ulcerative colitis because it stabilizes mast cell membranes and limits the release of histamine and other inflammatory chemicals. Quercetin can also block inflammatory processes mediated by “leukotriene’s” (inflammatory effects a thousand times stronger than histamines), hyaluronidase (collagen-destroying enzymes), and lysosomal enzymes owing to its antioxidant impact.^11,12

Determination of absorption Maxima (λ_max) by UV-visible spectroscopy and Standard calibration curve of Quercetin

UV spectrophotometers (UV-1900) with matched 10 mm quartz cuvettes were used. Ethanol was used in the analysis of Quercetin. The following procedure was applied to determine the λ_max of quercetin, after accurately weighing 10 mg of the drugs; it was added to a 10 mL volumetric flask. Next, the volumetric flask was filled with 5 mL of ethanol and placed in an ultrasonicator for 5 min. Once a clear solution was obtained remaining volume was made up to mark with Phosphate Buffer pH 6.8. This is the stock solution that was used for the preparation of the standard calibration curve.

From the above stock solution, 1 mL was pippetted and diluted up to 10 mL by PBS to give a concentration of 100 μg/mL. At 200-400 nm, this solution was tested. To confirm the maximum absorbance of the drugs, the maximum absorbance was measured using a UV-vis spectrophotometer (UV-1900, Shimadzu, Japan).^13–19

The interaction and compatibility of pure drug, lipid, and surfactant used in TE preparation, as well as the physical mixture of drug and excipients, were studied using FTIR spectroscopic analysis. FTIR spectrometer was used for analyzing solid and liquid samples respectively. (SHIMADZU IR Affinity 1/8000, Japan and Bruker alpha II, USA). Inspecting was carried out using the transmission method, with a wave number range of 4000 to 400 cm¹.

For the physical mixture, the samples were taken in a 1:1 ratio and kept at room temperature. FTIR spectra were obtained for pure drug (QUERCITIN), lipid, and an actual combination of drug and lipid after a particular quantity of solid sample was combined with Potassium bromide (KBr).

For liquid samples, (Surfactant, mixture of drug and surfactant, and physical mixture of drug, lipid, and surfactant) specified amount is directly placed on the ATR crystal and analyzed.²⁰

DSC, a thermal analytical process, was employed to get the thermogram of quercetin by using aluminum pans. In this technique, an accurate weight of the sample was taken in the aluminum pans and sealed tightly. To determine the thermal behavior of the quercetin, the aluminum pans containing the sample were heated at a temperature range of 40-300°C, with a scanning rate of 20°C/min. To provide an inert atmosphere nitrogen gas was purged continuously at a 40 mL/min flow rate.

Most likely the technique most frequently employed to prepare ethosomal formulation. This method uses a mixer and vigorous stirring to disperse drugs, phospholipids, and other types of lipid materials in ethanol in a wrapped vessel at room temperature. Propylene glycol or another polyol is added while stirring. The mixture is heated to 300°C in a water bath. After adding the water that has been heated to 300°C in a separate vessel, the mixture is stirred for 5 min in a covered vessel. The ethosomal formulation’s vesicle size can be reduced to the appropriate degree by extrusion or sonication. Finally, the mixture is stored in a refrigerator.It was given in Tables 1 and 2.

The drug-loaded ethosomes (which are equal to 1%) were added to gels by slow mechanical mixing for 30 min at 600 revolutions per minute (REMI-type BS stirrer). The perfect mixture was added to a 1% w/w concentration of carbopol, and 0.001% of propyl paraben was used as a preservative.

The preparation of the quercetin hydrogel was achieved by gradual addition of gelling agent 1% Carbopol 940 by mechanical mixing at 600 rpm (REMI type BS stirrer) for 30 min.²¹

Using a particle size analyzer (Nanopartica SZ-100, HORIBA Scientific, USA), the prepared quercetin-loaded TE formulations’ particle size and PDI were measured using the malvan zeta sizer technique. The nanovesicles were diluted with distilled water and then analyzed at room temperature. The particle size and PDI of the diluted samples were measured in triplicate, and the results were reported as an average±standard

It’s the efficiency of the drug to entrap the drug, can be measured using the centrifugation method. The unentrapped drug is separated by centrifugation method, at the end of the method supernatant liquid is obtained. The amount of drug entrapped is calculated.

Zeta potential is the measurement of the electric charge on a nanoparticle’s surface that represents the system’s physical stability. The Zeta potential of an optimized batch of transethosome was determined by the electrophoretic light scattering technique using a particle size analyzer (Nanopartica SZ-100, HORIBA Scientific, USA Before analysis, the samples were diluted with double distilled water. The findings were presented as mean ± standard deviation, and the analysis was done in triplicate.

A scanning electron microscope was used to study the morphology of the optimum ethosomes.

Samples of nanoparticles were put into dialysis bags, sealed, and submerged in a dissolution medium. The USP Dissolve Test Apparatus, Type II, was used to conduct a 2 hr drug release study at 37±0.5° and 100 rpm. 5 mL of sample were taken at each interval and replaced with brand-new buffers. Following suitable dilutions, spectrophotometric evaluation was performed on the samples.²²

The digital pH meter was used to determine the gel’s pH. After agitating 1 g of ethosomal gel in distilled water until a homogenous suspension was achieved, the pH of the mixture was determined.

To compress the sample to a consistent thickness, an excess of gel (2 g) was sandwiched between petri dishes, and 50 g of weight was placed on slides for 5 sec. The tin (sec) required to separate the two slides, was taken as a measure of spreadability.

It was calculated using the formula,

Where, S=spreadability

M=weight tied to upper slide

L=length of glass slide

T=time taken in min

The gel’s viscosity was measured with a Brookfield viscometer (LV) of the Di type. As spindle number four is non-Newtonian in nature, it is utilized. Viscosity was measured at 0.3 rpm for a predetermined duration of 2 min.²³

Formulas were applied to the skin before being manually assessed for the ease and extent of water cleansing.²⁴

Color matters when it comes to patient compliance. We visually examined the prepared gels to ensure they were stable, uniform, and had a pleasing texture.²⁵

Absorption maxima of Quercetin was carried out using wavelength from 800-200nm. At 373nm, the highest absorption was recorded and employed. λ_max was depicted in Figure 2. The Quercetin standard calibration curve was estimated by plotting concentration v/s Absorption. The standard calibration curve was found to be linear at a maximum distance of 373nm in the beer’s range. Since y=0.073-0.0048, the obtained correlation coefficient R₂=0.9993.

Figure 2:
Determination of λ_max.

The drug shows a linear relationship between absorbance and drug concentration level. The synthesized impurity shows maximum absorbance at 373 nm. The equation of the straight line for quercetin is y=0.073-0.0048 and the regression coefficient is R₂= 0.9997 which is given in Figure 3.

Figure 3:
Standard calibration curve of Quercetin.

The FTIR spectra of pure quercetin, physical nature of quercetin, carbopol, HPMC, is shown in Figure 4. The quercetin FTIR spectra clearly showed all of the characteristic peaks. Thus, all the drug’s properties remained largely unchanged, and quercetin was effectively incorporated into the gel formulation.

Figure 4:
FTIR Spectra of quercetin.

DSC thermograms of pure quercetin, physical mixture of quercetin, carbopol. Sharp endothermic peaks at 270.67°C were visible on the thermal graph of pure quercetin, signifying the compound’s melting point (Figure 5).

Figure 5:
DSC thermograms of quercetin.

Particle size was found to be 57.4 nm and PDI was found to be 1 which was depicted in Figure 6.

Figure 6:
Determination of particle size and PDI.

Entrapment efficiency was found to be 92%.

The optimized formulation’s zeta potential was found to be 0.38 after the analysis was conducted in triplicate.

A scanning electron microscope was used to study the morphology of the optimum ethosomes. The particles was found to be sphere shape which was given in Figure 7.

Figure 7:
SEM of the quercetin loaded Ethosomes.

The graph of the cumulative percentage of the drug released over time was plotted to obtain the in vitro drug release profiles of quercetin from optimized batch at pH 7.4 is shown in figure. The drug release study indicates that the optimized batch of quercetin ethosomes releases with an initial burst release of roughly 21.48%, respectively, in one hour, followed by maximum amount of drug released was observed up to 24 hr (Figure 8).

Figure 8:
In vitro drug release study.

The pH of Quercetin loaded ethosomal gel was found to be 6.4 (Figure 9) and pH of hydrogel was found to be 6.8. The pure gel was found to be 7 which may avoid any skin irritation.

Figure 9:
Determination of pH.

The therapeutic effect of the Quercetin loaded ethosomal gel depends on the spreading value. The spreadibility of Quercetin loaded ethosomal gel was 2.31 g cm/sec and of hydrogel was 1.85 g cm/sec (Figure 10).

Figure 10:
Spreadability studies.

The viscosity of Quercetin loaded ethosomal gel was found to be 665 CPE and viscosity of hydrogel was found to be 748 CPE (Figure 11).

Figure 11:
Determination of viscosity of the quercetin.

The washability of Quercetin loaded ethosomal gel was found to be 5.70 sec and that of hydrogel was 8.23 sec (Figure 12).

Figure 12:
Determination of washability of quercetin.

The colour of Quercetin loaded ethosomal gel and hydrogel was pale yellow.

Hydrogel showed better physical stability and ease of preparation compared to the ethosomal gel. Hydrogels are also biocompatible and is readily taken away from the skin without leaving any residue. However, the hydrogel showed lower skin permeation and retention of quercetin that can be confirmed through better spreadability, viscosity and washability of ethosomal gel when compared to quercetin hydrogel.

Therefore, the choice of formulation depends on the specific needs and requirements of the intended application. Ethosomal gel may be preferred when high skin permeation and retention of quercetin are desired, whereas hydrogel may be preferred for ease of preparation and removal, and better physical stability.

Based on the comparative study of quercetin ethosomal gel and hydrogel, it can be concluded that both formulations have their own advantages and limitations. Quercetin ethosomal gel showed higher skin permeation and retention of quercetin compared to hydrogel. This can be attributed to the enhanced penetration ability of ethosomes due to its low particle size and high entrapment efficiency, which allows easy passage of drug through the stratum corneum and reach deeper skin layers. The study’s optimal formulation was chosen to be the ethosomal formulation F5, which displayed lower particle size and higher entrapment efficiency. The ethosomal gel of optimized F5 formulation was prepared that exhibited better antioxidant and anti-inflammatory activity, which can be beneficial in treating various skin disorders.

The authors express heartfelt gratitude towards the KLE College of Pharmacy, KLE Academy of Higher Education and Research, Belagavi, for providing all the obligatory facilities for completion of this research.