*Article* **Quality-by-Design-Assisted Optimization of Carvacrol Oil-Loaded Niosomal Gel for Anti-Inflammatory Efficacy by Topical Route**

**Mohammed Ghazwani 1,\* , Umme Hani <sup>1</sup> , Aftab Alam <sup>2</sup> and Mohammed H. Alqarni <sup>2</sup>**


**Abstract:** Niosomes are multilamellar vesicles that effectively transfer active ingredients into the skin's layers. To improve the active substance's penetration across the skin, these carriers are frequently utilized as topical drug delivery systems. Essential oils (EOs) have garnered significant interest in the field of research and development owing to their various pharmacological activities, cost-effectiveness, and simple manufacturing techniques. However, these ingredients undergo degradation and oxidation over time, leading to a loss of functionality. Niosome formulations have been developed to deal with these challenges. The main goal of this work was to create a niosomal gel of carvacrol oil (CVC) to improve its penetration into the skin for anti-inflammatory actions and stability. By changing the ratio of drug, cholesterol and surfactant, various formulations of CVC niosomes were formulated using Box Behnken Design (BBD). A thin-film hydration technique using a rotary evaporator was employed for the development of niosomes. Following optimization, the CVC-loaded niosomes had shown: 180.23 nm, 0.265, −31.70 mV, and 90.61% of vesicle size, PDI, zeta potential, and EE%. An in vitro study on drug release discovered the rates of drug release for CVC-Ns and CVC suspension, which were found to be 70.24 ± 1.21 and 32.87 ± 1.03, respectively. The release of CVC from niosomes best fit the Higuchi model, and the Korsmeyer–Peppas model suggests that the release of the drug followed the non-Fickian diffusion. In a dermatokinetic investigation, niosome gel significantly increased CVC transport in the skin layers when compared to CVC–conventional formulation gel (CVC-CFG). Confocal laser scanning microscopy (CLSM) of rat skin exposed to the rhodamine B-loaded niosome formulation showed a deeper penetration of 25.0 µm compared to the hydroalcoholic rhodamine B solution (5.0 µm). Additionally, the CVC-N gel antioxidant activity was higher than that of free CVC. The formulation coded F4 was selected as the optimized formulation and then gelled with carbopol to improve its topical application. Niosomal gel underwent tests for pH determination, spreadability, texture analysis, and CLSM. Our findings imply that the niosomal gel formulations could represent a potential strategy for the topical delivery of CVC in the treatment of inflammatory disease.

**Keywords:** carvacrol; essential oil; niosomes; confocal laser scanning microscopy; anti-inflammatory; dermatokinetics; in vitro drug release studies

### **1. Introduction**

Due to their low cost and simple manufacturing processes, the utilization of EO constituents for biomedical applications has attracted global interest [1]. Their numerous qualities, such as viricidal, bactericidal, fungicidal, local anaesthetic, anti-inflammatory, and analgesic properties, have made them widely utilized for centuries. Because they are relatively safer for the skin than synthetically made traditional drugs, EO are often chosen

**Citation:** Ghazwani, M.; Hani, U.; Alam, A.; Alqarni, M.H. Quality-by-Design-Assisted Optimization of Carvacrol Oil-Loaded Niosomal Gel for Anti-Inflammatory Efficacy by Topical Route. *Gels* **2023**, *9*, 401. https://doi.org/10.3390/ gels9050401

Academic Editor: Shige Wang

Received: 10 April 2023 Revised: 25 April 2023 Accepted: 6 May 2023 Published: 10 May 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

as priority [2]. The components derived from plants still have the potential to cause allergies, but they are used in much lower concentrations than directly applying the oils to the skin, making them a safer option [3]. EOs are mostly made up of volatile substances such as aliphatic, aromatic, terpenoids and phenolic compounds. However, these chemicals could slowly deteriorate and oxidize, which would reduce their functionality [4]. Additionally, the bioavailability, flavour, and colour of the other ingredients may change in response to one another [5]. EOs are substances that, from a physiological standpoint, rapidly evaporate and/or decay even while the temperature is milder than the room temperature. This is because their boiling temperatures are extremely low, and they break down when directly exposed to light or oxygen, in general. Additionally, EO formulations frequently result in skin sensitivity and irritation [5]. The plant species *Thymus vulgaris*, *Origanum vulgare*, *Ocimum gratissimum*, *Carum copticum*, *Oliveria decumbens*, *Trachyspermum ammi*, *Monarda didyma*, *Nigella sativa*, *Origanum onites*, *Origanum syriacum*, *Plectranthus amboinicus*, *Lavandula multifida*, and *Satureja thymbra* are the primary sources of thymol (2-isopropyl-5-methylphenol; C10H14O), an isomeric form of carvacrol (5-isopropyl-2-methylphenol; C10H14O) [6,7]. Carvacrol and thymol have similar chemical structures and biological functions, which may indicate that they have related action mechanisms and therapeutic benefits. The CVC possesses a wide range of therapeutic activity, including those of an immunomodulator, an antioxidant, an antiseptic, an anti-diabetic, a cancer prophylactic agent, an anti-inflammatory compound, an anti-spasmodic molecule, and a growth promoter [8–15]. CVC is a promising drug for the treatment of local skin damage [16,17]. However, its pharmaceutical use is severely constrained by its lower bioavailability as a result of its poor water solubility, high volatility, and instability [10]. CVC exhibited antiinflammatory activity by reducing cyclooxygenase-2 (COX-2) and inhibiting interleukin-1β (IL-1β), and potentially induced interleukin-10 (IL-10) release [18]. The release of IL-10 would result in an anti-inflammatory impact because IL-10 is an anti-inflammatory cytokine that inhibits several cell surface molecules, such as the MHC class II proteins and co-stimulatory molecules [19,20]. Many strategies have been put up to overcome this solubility and bioavailability. One of the flexible approaches involves encasing the EOs in other nanoparticles, such as liposomes [21], niosomes [22], graphene oxide [23], and others. Niosomes can extend drug retention duration, increase solubility, and improve targeted delivery [24].

Niosome is a biocompatible bilayer system based on non-ionic surfactants and cholesterol (as a lipid) [25]. The amphiphilic non-ionic surfactant self-assembles to produce niosomes in aqueous fluids. Niosomes also have several benefits, such as the ability to encapsulate both hydrophilic and hydrophobic components, improved therapeutic performance, long storage times, easy surface modification, biodegradability, and nonimmunogenicity [26]. Entrapping EOs in the hydrophobic areas of niosomes can increase their permeability and stability qualities, lowering their potential for skin irritation [27]. These nonionic surfactant vesicles enhance the potential of the entrapped drug when applied topically by modifying the properties of the stratum corneum, facilitating the transdermal transport of trapped substances and decreasing trans-epidermal water loss [28]. Niosomal formulations have been created for a variety of targeted drug delivery purposes throughout the past decade [29]. To produce small-sized niosomes with maximal encapsulation efficiency for any given drug, however, the size and drug encapsulation efficiency of the niosomes must be optimized by altering the formulation (surfactants and lipids) [30,31]. Niosomal formulations of compounds containing CVC oil have been developed for antiinflammatory and other studies, but a CVC-loaded niosomal formulation has not yet been developed [32–36]. The primary objective of this study was to investigate the potential of CVC for the development of niosomes. Niosome efficacy can be affected by many factors related to formulation and processing. Investigation into these aspects of niosome preparation would thus significantly improve the formulation. The quality by design (QbD) strategy involves the design and development of a product with manufacturing procedures that meet predetermined product criteria [37]. To ensure that selection criteria for desired

responses are established for a range of vesicle sizes (<200 nm), polydispersity index (PDI) (<0.3), and entrapment efficiency (>70%) for formulation optimization, response surface methodology (RSM) was employed [38]. For the preparation of niosomes in drug delivery, a variety of nonionic surfactants, including polysorbates, alkyl esters, and alkyl ethers, are usually used. An important factor influencing the physicochemical properties of niosomes, such as bilayer rigidity, vesicle size, stability, and drug release rate, is the type of surfactant used [39]. Tween 80 and Span 80 have undergone testing as drug delivery systems, and specifically, they can be used to create highly stable niosomes [40]. In this formulation of niosomes, tween 80 was used as a surfactant. The stabilization of niosomes that have both hydrophilic and lipophilic domains was accomplished using the surfactant Tween 80. Its longer aliphatic tail causes adsorption on the drug's surface and allows its hydrophilic portion to potentially reach the aqueous phase. This would completely cover the surface and lessen the interfacial friction between hydrophobic drug vesicles [41]. The larger unsaturated side chain of the tween 80's oleate component may increase the poorly water-soluble drug CVC's ability to be enclosed more densely in niosome bilayers [42]. The developed niosomal formulation was incorporated into the gel for better retention time. The vesicle morphology, in vitro drug release, CLSM, dermatokinetic and optimized gel formulation texture analysis was also assessed. It can be anticipated that, the use of quality-by-design approach to optimise the CVC-loaded niosomal gel formulation is a novel approach that can help to ensure the formulation's quality, safety, and efficacy. Furthermore, using a topical niosomal formulation can improve carvacrol skin penetration and absorption, making it more beneficial for inflammatory skin disorders that require targeted therapy without systemic absorption. This approach may lead to the development of a more efficient and effective treatment for inflammatory skin disorders.
