Abstract
Essential oils (EOs) are aromatic compounds of plant origin with several bioactive properties. The current global essential oil production exceeds 300,000 tonnes, and the estimated EO market size in 2030 is approximately $ 39.5 billion USD. Such compounds undergo degradation upon exposure to environmental stresses during processing and storage. The nanocarrier systems primarily protect EO from environmental conditions, and the EO nanoformulations have demonstrated better performance compared to their free counterparts. For example, studies have reported a 2- to 9-fold increase in bioactivity, improved stability, and controlled release that extends from hours to days. The present review article is focused on the current state of knowledge of different advanced applications mediated through several nanoformulations of EO, such as nanoemulsions, nanoparticles, nanofibers, nanocapsules, nanoencapsulation, and nanoliposomes. The encapsulation efficiency of the formulations reaches up to 99% depending on the type of EO and the type of nanoformulation. It was seen that the usage of starch-based nanoformulation further enhances the nano-EO incorporation in aqueous matrices. Thereafter, the mechanism of action and release kinetics of EO nanoformulations are briefly discussed. Finally, the paper concludes with highlights on limitations that are frequently encountered, while emphasising the adaptability of nanoformulations for the target delivery of EO. Hence, the readers of this paper will gain a comprehensive knowledge through the information gathered from the literature of the latest years that will help advance future research on the safety evaluation, biodegradation, and long-term effects of the EO nanoformulation.
Keywords
Introduction
Essential oils (EOs) are aromatic compounds formed during secondary metabolism in plants. EOs possess a unique aroma and consist of monoterpenes, sesquiterpenes, and phenolics. The plants produce EOs as mixtures of organic components in the cytoplasm and plastids. The role of such volatile compounds is to defend the plants from invading microbes and to attract specific insects for plant pollination. Typically, EOs comprise about 20 to 60 constituents, or over 100 components, all at varying concentrations. Among these, two or three primary components are typically present at considerable concentrations (20% to 70%), in contrast to the other components present in trace quantities. 1 Driven by recent trends in natural product consumption, the global EO market was valued at USD 23.74 billion in 2023 and is projected to grow at a compound annual growth rate of 7.6 per cent from 2024 to 2030. 2 India is the major source of Indian basil oil and Japanese peppermint oil, while the USA is the largest buyer and producer of EOs, accounting for 14% of global imports and 17% of global exports. 3 EOs find applications in medicine, healthcare, agriculture, cosmetics, and the food industry due to their diverse biological functions. The spectrum of properties of EO includes antioxidant, antibacterial, antifungal, anti-biofilm, anticancer, antidepressant, anxiety reduction, skin anti-wrinkle and whitening abilities, insecticidal property, food preservative, chemopreventive, neuroprotective and anti-amnesic and pesticidal activity. 1
The EOs influence the environment through their functional abilities as chemical cues that help plants repel predators, draw in pollinating insects, restrict seed germination, and communicate with one another. Mostly, different parts of aromatic plants contain EOs, including the flowers (Nerium oleander, Lavandula hybribia L.), flower buds (Eugenia caryophyllata), bracts (Origanum vulgare L.), leaves (Cinnamomum osmophloeum), rhizomes (Zingiber zerumbet), roots (Cyathocline purpurea), seeds (Nigella sativa), fruits (Cuminum cyminum), and wood and bark (Cinnamomum verum). It was seen that secretory structures, such as granular hair, oil cells, oil canals, and oil chambers, accumulate EOs after production from plastids in cells. 4
Conventional extraction techniques, such as hydro distillation, steam distillation, solvent extraction, and cold pressing, have been extensively studied and widely used for many years. However, such methods are slow and associated with the possible loss of volatile compounds in the presence of residual solvents, causing toxicity. 5 Recently, innovative techniques, which are rapid, environmentally friendly, and preserve the bioactivity of EOs, have been developed for the extraction of essential oils from plants. The innovative techniques include supercritical Fluid extraction, ultrasound-assisted extraction, microwave extraction and the use of green solvents.
Nevertheless, the limited scope of applications of EOs is due to their physical and chemical properties. EOs have unique physicochemical characteristics, such as volatility, low solubility in water, and chemical instability, which have substantial implications for their therapeutic efficacy and, as such, demand advanced delivery approaches. The volatile components in EOs, such as carvacrol and thymol (due to low molecular weights), evaporate rapidly and have a low residence time on the application area. Being hydrophobic, they are poorly soluble in aqueous systems and possess low bioavailability. Furthermore, EOs are subject to oxidative and photo-degradation, which affect stability and therapeutic effectiveness. 6 This chemical instability, derived from the sensitivity to oxidative degradation, hydrolysis, and photolysis, may lead to the inactivation of the active ingredients, loss of activity, or unsafe metabolites. 7 The encapsulation of EOs can enhance pharmacokinetics in vivo, such as prolonged half-life, slower metabolism, and targeted accumulation, which increases their efficacy in both systemic and topical uses. These systems also play a role in reducing pronounced organoleptic characteristics and toxicity by facilitating sustained release and concealing undesirable tastes or odours. The remarkable introduction of nanoformulations in recent years continues to open up new avenues for problem-solving in EOs delivery. In different industries, similar technology can effectively reduce the volatility rate and losses, enhance the efficacy of its utilisation, and improve product quality. 8
The EO nanoformulations have shown better characteristics compared to the conventional EO delivery for application in different fields. In the biomedical applications, nanoformulations enhanced bioactivities and showed sustained EO release. For instance, Homalomen apineodora EO encapsulated in a chitosan nanoparticle showed enhanced antimicrobial activity and prolonged release up to 3 days, 9 Ginger EO loaded into nanoliposomes enhanced antioxidant activity, decreased cytotoxicity and sustained release characteristics, 10 thymol EO and β-cyclodextrin inclusion complex loaded in chitosan nanoparticles increased solubility and antioxidant activity. 11 Similarly, the use of EO nanoformulations in the storage and preservation of food products improved their shelf life and enhanced their properties. For example, the use of nano-encapsulated Heracleum persicum EO improved shelf life, sensory and rheological properties for dairy desserts. 12 Similarly, in agriculture, nanoformulation enhanced insecticidal activity and stability of EO. For instance, Piper nigrum EO encapsulated in chitosan nanoparticles enhanced insecticidal activity during grain storage. 13
At the outset, this review aims to provide a comprehensive understanding of EO-nanoformulations, focusing successively on their applications as an effective delivery system with improved physicochemical properties. The exploration extends to a detailed examination of different types of nanoformulations, evaluating the role of nanoformulations in enhancing bioactivity, stability, controlled release, and bioavailability across various sectors. Specifically, it highlights the use of starch for nanoformulation to address the challenges incurred by hydrophobic nanosystems, systematically covering the types of starch-based nanostructures and methodologies in nanoformulation. Most of the available reviews broadly focus on polymeric nanoformulations, and this material-specific review is underexplored. The safety/ toxicity insights provided may bridge the gap between lab-scale studies and consumer safety considerations. Hence, this review provides a consolidated overview of essential oil nanoformulations, offering detailed insights into their release mechanisms, benefits and emerging domain-specific applications.
Types of Nano-Delivery Systems for Essential Oils
A potential delivery system shields the EOs from unfavourable conditions during processing and administration. EO delivery system depends on its intended use and the site of application. Developing a suitable delivery system is challenging due to its properties, especially during its storage, transport, and application. 14
Nanoparticles
Nanoparticles (NPs) have sizes ranging from 1 to 100 nm in all dimensions. NP exhibits superior properties such as enhanced reactivity, surface-to-volume ratio, and stability compared to its precursor compound. NPs-EOs can achieve prolonged release, reduce evaporation losses, and, in particular, boost dispersion in an aqueous medium. 15 The methods for producing EO-loaded nanoparticles include co-precipitation, high-pressure homogenization, high-speed stirring, ultra-sonication, ionic gelation, mini-emulsion polymerisation, nanoprecipitation, spray drying, and the Stober process. 16 However, selecting an appropriate nanoparticle system is necessary for the effectiveness of the delivery system. Several nanoparticles, such as inorganic, biogenic and synthetic, have been used for incorporating the EO. For example, Chitosan nanoparticles loaded with EO suppressed the growth of human lung cancer cells A549 in a dosage-dependent manner. During the preparation, a negatively charged sodium tryptophane cross-linker and chitosan molecules interact ionically with positively charged amino groups. Following the procedure, the material improved diffusion properties, achieved a high dispersion around the target site, and successfully delivered a nanoscale medication within pathogenic cells. The morphological analyses revealed that the EOs containing chitosan caused damage to both the intracellular and extracellular components in treated cancer cells. 17 Similarly, protein-based NP (eg, zein) encapsulated orange EO by the nanoprecipitation method. Zein-NP is an antifungal agent that preserves agro-ecological maize seeds during storage. It was observed that the nanoparticle-encapsulated EO demonstrated effective fungal inhibition compared to the non-encapsulated form. 18 By using different nanoparticle materials, these formulations can be tailored for diverse applications. NPs provide an excellent platform for EO delivery, although challenges remain regarding scalability, instability over storage time and toxicity of the particles.
Liposomes
Liposomes are tiny vesicles formed by phospholipid dispersion in an aqueous solution and can encapsulate the aqueous moieties within their enclosed concentric spheres of phospholipid membranes. These vesicles can encapsulate lipophilic compounds such as EO within the lipid bilayer, making it suitable for EO delivery. Such liposomes with increased penetration and concentrations resolve issues like limited dissolution of medication in water and poor drug delivery. 19 The higher affinity of EO for lipids on a molecular scale and their interfacial interaction achieved stable dispersion in an aqueous condition. 20 Furthermore, liposomes improve the stability and solubility of the EOs and provide a shield to protect them from oxidation and evaporation. 21 Moreover, liposomes exhibit centrifugal, thermal, and storage stability, 22 higher solubility, bioavailability, pharmacological action, and lower toxicity and adverse effects. 23
Zingiber officinale EO-loaded nanoliposome was 100 nm in size with 66.24% loading capacity and demonstrated enhanced antioxidant effect, stability, and reduced cytotoxicity. It maintained the bioactivity of EO during storage and 90% of EO release under an oral, gastrointestinal environment. 10 Nanoliposomes loaded with Zhumeria majdae EO inhibited nosocomial infection-causing multidrug-resistant clinical pathogens. This formulation presented a 118 nm size, 92% encapsulation efficiency, and better antioxidant, anti-quorum-sensing, and antibiofilm effects than the pure EO. 24 Active packaging for meat products is another application of nanoliposomes in the food industry. The biodegradable film comprises a chitosan-based nanocomposite incorporated with nanoliposome-garlic EO to improve the film properties. Additionally, the film extended shelf life, improved the chemical properties, and preserved meat during storage at 4 °C. 25 In contrast to alternative encapsulation techniques for EO, liposomes exhibit superior biocompatibility and biodegradability, enable precisely regulated drug release, and facilitate distinct, simultaneous encapsulation of fat-soluble and water-soluble compounds.
Nanoemulsions
Emulsions are essentially ‘liquid in liquid colloids’ in which droplets of the dispersed phase scatter in the dispersion medium or continuous phase. Furthermore, they run the risk of destabilising over time due to thermodynamic instability, potentially leading to a loss of stability. Nanoemulsions (NE) contain dispersed-phase droplet sizes ranging from 10 to 100 nm. Due to their intrinsic ability to encase the drug in nanometric droplet dimensions, nanoemulsions are considered to be one of the most effective methods. Even though NEs resist settling because of their small size, Brownian motion overcomes gravity, and they can undergo flocculation, coalescence, and Ostwald ripening. 26 The exceedingly volatile nature of EOs limits the development of EO-based NEs. Additionally, EO-NEs are prone to losing some of their properties because of thermal and mechanical impact during their processing steps. Consequently, selecting the appropriate emulsifier, oil-to-surfactant proportion, treatment duration, and ambient conditions is critical in EO-NEs synthesis. 27
The primary application of EO-NE is in food preservation to increase the storage time by leveraging the antimicrobial and antioxidant properties of EO. For example, a fruit coating composed of Sicilian lemon EO potentially inhibited microbial growth on strawberries for at least 14 days. 28 Similarly, NE integration of edible pectin coatings with oregano EO and resveratrol elongated fresh pork shelf life under high oxygen-modified packaging storage at 4 °C. The NE size (50 nm) affected stability and preservative properties. 29 Another study prepared a wrapping edible composite film consisting of tamarind starch, whey protein concentrates, and nanoemulsions of thyme EO (20%). The addition of nanoemulsion with a particle size of 18.88 nm improved the film properties and tomato shelf life up to 14 days. 30 In the biomedical domain, specifically in cancer therapeutics, celery EO nanoemulsion reduced cell proliferation by inhibiting anchorage-independent cell division, which disrupted colony formation and caused apoptosis in cancer cells. Nanoemulsions were prepared with Tween 80 by the sonication method for 20 min, resulting in a droplet size of 23 nm. 31 The benefits of employing NEs over conventional emulsions for EOs derive from the small particle dimension, which offers kinetic stability and a large interfacial area, yet the production processes are complex, energy-intensive and limit large-scale production.
Nanofibers
Nanofibers and EOs combine for usage as environmentally friendly packaging, wound dressing and drug delivery systems. In the biomedical domain, Pickering emulsion electrospinning is a viable approach for producing nanofibers loaded with hydrophobic tea tree oil in a hydrophilic hydroxyethyl chitosan/polyvinyl alcohol nanofiber matrix, and the resulting nanofibers treat infected wounds. The nano-sized zinc oxide-argentum composites are the emulsion stabilisers. 32 In wound dressing application, electrospun chitosan nanofiber combined with Melissa officinalis L. and Anethum graveolens L. EO encapsulated in collagen hydrolysates was effective. The synergistic action of EOs enhanced antimicrobial activity, and the in vivo test indicated excellent biocompatibility for nanofibers. 33
Active food packaging with bioactive properties has been of interest in recent years, as it extends shelf life. The incorporation of angelica EO in gelatin nanofibers exhibited considerable antioxidant and antibacterial activity with no cytotoxic effects. The addition of EO resulted in enhanced hydrophobic properties and fibre diameter. 34 Another food packaging consists of the reinforced ZnO NPs, rosemary EO-loaded zein nanofiber with κ-carrageenan. The biocompatible nanofiber possessed a 672 nm diameter, exhibited homogenous morphology, antioxidant effect, antibacterial activity, and improved thermal and mechanical properties. 35 An electrospun nanofiber containing polylactic acid, guar gum, and thyme essential oil (30%) is suitable for food packaging. Properties such as smooth morphology, high surface hydrophobicity, antioxidants, antimicrobial effects, and biocompatibility are desirable for packaging applications. 36
In agricultural and food engineering, a blend of thyme and betel leaf EO has been incorporated in a nanofiber composed of polyvinyl alcohol for the control of anthracnose disease caused by Colletotrichum gloeosporioides in the post-harvest preservation of Manilkara achras mill fruits. The EOs demonstrated an additive effect for the control of pathogens, and the blend resulted in an encapsulation efficiency of more than 75%. The nanofiber-incorporated package inhibited the pathogen growth from 100% in the control to 40%. 37 Moreover, a nanofiber incorporated with EO is preferred as the free EO application leads to fruit peel burning. Incorporating essential oils into nanofibers results in enhanced stability, bioactivity and controlled release; however, challenges such as the volatility of EO during processing, leading to limited encapsulation efficiency, must be addressed.
Nanoencapsulation
Nanoencapsulation forms a protective barrier for EO, enabling its delivery at the target sites with a sustained release. Nanoencapsulation of EO within cross-linked biopolymer carbohydrate-protein improved its stability. For example, Arabic gum-gelatin cross-linked by citric acid nanoencapsulated savory EO (3 ml/L). The formulation showed improved stability and herbicidal activity against grain Amarnath germination. 38 Similarly, whey protein concentrate- pectin polymers encapsulated rose EO for food applications—the ratio of 4:0.5 of polymers obtained 96%encapsulation efficacy at pH 3. 39 Nanoencapsulation of eugenol in casein micelle enhanced the biopesticide property of eugenol. Under optimum conditions, the average size of the formulation was 179.83 nm, and an encapsulation efficiency of 79%. It also exhibited controlled release and increased pesticidal properties owing to its improved stability. 40 A gelatin nanocomposite hydrogel incorporated with thyme EO encapsulated in sodium caseinate nano micelle is an effective drug delivery platform for wound healing. The EO-loaded nanomicelle exhibited a size of 336 nm, encapsulation efficiency of 75%, and improved antibacterial activity. 41 The particle size and morphology are the primary parameters associated with the regulated release of bioactive chemicals at the specified site for improving the therapeutic rate of action and dosing. Nanoencapsulation safeguards EOs from heat and light stress, resulting in better stability, function, and flavour and preserves EOs from oxidation or evaporation. However, the rapid diffusion of EOs through the encapsulating material, if environmental conditions are not controlled, has to be addressed.
Nanocapsules
Nanocapsules (NCs) are vesicular nanostructures with an oily core encased by a polymeric wall. Polymers such as polycaprolactone (PCL) are biocompatible, biodegradable, and widely used in NC production. The NCs offer controlled and targeted drug delivery, withstand environmental conditions such as pH, light, and oxygen, 42 high hydrophobic ingredient encapsulation, and drug protection 43 and improved bioactivity. 44 In the food processing domain, NCs with rosemary EO are natural antioxidant agents in meat preservation and increase its stability. The increased heat stability of free oils enables such bioactives to be added to cooked food items. 45 In another study, interfacial deposition of the preformed polymer method synthesised a nanocapsule based on PCL loaded with Foeniculum vulgare Mill EO. The NC-EO is suitable for nutraceutical and pharmaceutical applications as it exhibited 210 nm, high encapsulation efficiency of 93%, and resistance to the gastric digestion environment. 46 NCs find application in the preservation of fresh-cut fruits. For example, nanocapsules containing lemon EO and curcumin coated on papaya are stored for 17 days. During storage, NCs regulated pH and acidity, reduced respiration, maintained colour, and decreased polyphenol oxidase, thereby preserving total phenolic content. 47 In insect management, a ready-to-use preparation comprising cedar wood EO (CWEO) combats mosquito breeding locations and is a vector control approach. The preparation is a cotton mini bag loaded with pectin-based nanocapsules incorporated with CW EO. It exhibited a 98% mortality rate of Anopheles culicifacies, the malaria vector, for four weeks. 48 The tailored surface properties of nanocapsules result in targeted delivery, enhanced physicochemical stability and increased bioavailability, but their effectiveness depends on the precise choice of material and processing methods.
Polymeric Nanoparticles
The design of polymeric nanoparticles is based on the biomimetic concept of maintaining the in vitro stability of EO through the artificial carrier system, which substitutes for the natural conditions in plant cells. 49 The application of polymeric nanoparticles (PNPs) in medicine is attributed to their efficiency and bioavailability, improved delivery of medication, simple incorporation into physiological substrates, and drug transfer to the target site with the desired concentration, stability, and longer duration of action. 50 PNPs are based on polysaccharides and proteins. 51 Polysaccharides used in PNPs include plant-origin (eg, pectin, starch, and derivatives) or animal-origin (eg, xanthan gum, chitosan). While albumin, gelatin, and soy protein are protein sources used for PNP synthesis. Natural PNPs are biodegradable, biocompatible, and removed from the living system by metabolism. 52
Medium-molecular-weight chitosan (CS) is a superior encapsulating material that retains test bioactive compounds and demonstrates improved antibacterial properties. 53 Laurus nobilis EO encapsulation in the CS matrix by inter-molecular ionic cross-linkages improved the EO stability. 54 The composite nanoparticle comprising pectin and chitosan addressed the solubility and sensitivity challenges incurred by jasmine EO. The nanoformulation improved the properties, including thermal stability by 11.64 times, the antioxidant effect by 96%, and anticancer activity by 13 times. The formulation demonstrated improved cyto-compatibility against normal cells. 52 Similarly, bovine serum albumin nanoparticles loaded with carvacrol possess therapeutic and immunomodulatory activities in induced arthritis. The nanoparticles exhibited an encapsulation efficiency of 67% and a size of 148 nm. 55 Nettle EO-loaded chitosan nanoparticles exhibited higher antioxidant and antimicrobial effects than free EO. NEO-chitosan, with a retention rate of 59% to 68% and a size of 208.3–369.4 nm, is promising for food and pharmaceutical applications. 56 PNPs hold the benefit of being functionalized, releasing drugs at a regulated rate at target sites. Besides drug loading effectiveness, the drug release strategy determines PNP selection.
Mechanism of Action and Release Kinetics of EO Nanoformulations
Mechanism of Action of EO Nanoformulations
Essential oil nanoformulations are proving to be highly effective in boosting the antimicrobial, antioxidant, and preservative powers of volatile essential oils. The encapsulated EO compounds display antimicrobial properties through various mechanisms, as summarised in the Figure 1. Initially, numerous formulations depend on electrostatic attraction, where positively charged nanoparticles such as chitosan adhere to the negatively charged membranes of bacteria, resulting in membrane rupture, loss of cellular contents, and ultimately cell death. 57 Furthermore, metallic nanoparticles like ZnO and TiO2 boost antimicrobial effectiveness by producing reactive oxygen species (ROS) that harm microbial DNA, proteins, and lipids.58,59 Additionally, bioactive components of EO, including carvacrol, citral, and eugenol, directly compromise microbial membranes by modifying their permeability or disrupting enzymatic activities.60,61

Mechanism of Action of Antimicrobial EO Nanoformulations.
Controlled and Sustained Release
One of the primary benefits of nanoencapsulation is the ability to achieve controlled and sustained release of essential oils. The pattern of EO release through the matrix is represented in the Figure 2. This release usually exhibits a triphasic kinetic pattern: an initial burst phase, a steady diffusion phase, and a prolonged sustained release. During the burst phase, EO molecules that are adsorbed on the surfaces of nanoparticles are quickly released, providing an instant effect. The steady-state phase follows, allowing for a gradual release through the polymer matrix, mainly influenced by Fickian diffusion or swelling-controlled mechanisms. 62 For instance, chitosan nanoparticles infused with lemongrass EO exhibited approximately 45% release in the initial 50 min, followed by a slower release of an additional 40% over the course of several hours. 57 The final sustained phase guarantees that trace amounts of EO are accessible over prolonged durations, which decreases the frequency of dosing and maintains antimicrobial effectiveness over time. Importantly, the kinetics of release can be adjusted based on pH sensitivity; in acidic environments, the diffusion of EO is expedited due to increased swelling of the polymer. 58 Encapsulation within PEG, alginate, or zein matrices additionally aids in retention and controlled diffusion. 60 Mathematical models like the Higuchi, Weibull, and Korsmeyer Peppas equations are commonly used to forecast and enhance the release behaviour of essential oils, aiding in the creation of tailored delivery systems for applications in food or pharmaceuticals. 62

Release Profile of Essential Oil Nanoformulation Over Time.
Targeting Mechanisms
EO nanoformulations possess the ability for passive, active, and stimuli-responsive targeting as represented in Figure 3. These mechanisms improve the precision of antimicrobial and therapeutic effects at specific sites. Passive targeting has the advantage of the small size of these particles (less than 200 nm) to utilise the enhanced permeability and retention (EPR) phenomenon, particularly in inflamed or tumour-affected tissues. 1 Active targeting entails the modification of nanoparticle surfaces with ligands that specifically attach to receptors on microbial or host cells, thereby increasing selectivity. 57 Electrostatic targeting plays a crucial role in both food and biomedical fields. Cationic nanocarriers, such as chitosan nanoparticles (CSNPs) and zinc oxide nanoparticles, tend to adhere to negatively charged bacterial membranes, resulting in cell destruction. 58 Additionally, the release of stimuli-responsive agents is activated by shifts in pH or enzymatic conditions. For instance, the acidic environments often found in infections or spoiled food products boost the release of EO, making these systems well-suited for pathogen-responsive delivery. 16 In certain instances, hybrid nanocarriers like silver nanoparticles loaded with both cinnamaldehyde or ZnO infused with citronella oil demonstrate enhanced antimicrobial properties by integrating mechanical damage to membranes, oxidative stress, and the bioactivity of essential oils. 58

Targeting Mechanisms of EO Nanoformulations.
In combination, EO nanoformulations improve the stability, release rates, and targeting efficiency of essential oils. By designing nanocarriers with tailored physicochemical and biological characteristics, scientists have gained precise control over the delivery of EOs, allowing for extended antimicrobial effects, minimized degradation, and effective targeting at specific sites. These innovations render EO nanoformulations significant in various fields, including food preservation, biomedicine, pharmaceuticals, and cosmetics.
Recent Applications of Nanoformulation-Mediated EO
The enhanced bioactivities of essential oils achieved through nanoformulation have led to the focus of recent studies on their practical applications. The intended application of EO-nano preparation determines the suitable nano-system and its preparation method. 63 Nano preparations of EOs have gained significant attention in various fields due to their unique benefits and the versatile applications, as shown in Figure 4. Some examples include extension of shelf life of food products, prevention of food spoilage, sustainable practices against plant pathogens, insecticide and pesticides in agriculture, sustained release of bioactive compounds in cosmetic formulations, enhanced bioactivity and reduced toxicity in drug formulations, drug delivery in therapeutics.

Medicinal, Health, Agricultural and Cosmetic Applications of Essential Oil Nanoformulations.
Cosmetic Application
EO is a common ingredient in cosmetics as it lowers the risk of oxidative damage, premature ageing and inflammation. EOs are permeation enhancers in cosmetic preparations; however, their allergic reactions limit the topical application of the cosmetics. EOs are effective for a short period because they oxidise on exposure to heat or light. Additionally, their limited solubility in water restricts their miscibility in biological fluids, which in turn limits their bioavailability. 64 EOs in cosmetics are a safer option than the synthetic compounds commonly used. Citrus, lavender, eucalyptus, tea tree, and other floral oils are some of the highly prized EOs used as perfumes; linalool, geraniol, limonene, citronellol, and citral are fragrance components used in various cosmetics. 65 Nanoformulations of EOs are vital in cosmeceuticals for the skin care, hair care, and perfumery industries. Nano preparations improve the skin penetration of EOs, increase stability, and offer regulated release. They are in demand in the market due to their excellent solubility in natural and synthetic cosmetic formulations. The formulations containing Helichrysum italicum EO loaded with chitosan NPs enhance skin hydration and preserve skin health conditions. 66 Nanoethosomal encapsulation of orange EO resulted in a stable fragrance formulation and exhibited the ability to emit aromatic molecules over an extended period, lasting around three hours and three times longer than the typical orange scent. 67 These EO nanoformulations have transformed the cosmetic industry by enhancing their stability, bioavailability and targeted delivery. Given their apparent advantages, factors including formulation compatibility and optimising dosage must be resolved to maximise EO potential.
Agricultural Field Application
EOs treat plant diseases in agriculture and are known to be safe, bioactive, and biodegradable. 68 Replacing the chemical plant protection agents with EOs has received increased attention in agriculture. The EO nanoformulation primarily focuses on pest control, disease management and enhanced plant growth. For example, Athyrium sinense EO-loaded chitosan nanoparticle is a potent antimicrobial agent in the management of Pectobacterium carotovorum, which causes soft rot disease in Chinese cabbage. 69 Similarly, for insecticidal uses, nanoformulation of EO is suitable for reducing the populations of 11-day-old larvae of Rhynchophorus ferrugineus, even though it showed lower larval mortality percentage compared to the bulk oils. 70 In a different approach, Pelargonium graveolens EO-loaded nanoemulsion is effective for pest prevention (Rice Weevil Sitophilus Oryzae) in Stored Wheat Grain. The nanoemulsion prepared using a high-energy ultra-sonication resulted in an average droplets size of 30.99 nm with lower LC50 value (2.298 ppm/cm2), indicating a higher toxicity level, compared to free oil (67.66 ppm/cm2). 71 These nanoformulations are eco-friendly, biodegradable, and a sustainable alternative to synthetic agrochemicals. Although EO nanoformulation demonstrates great promise in agriculture, it requires further studies, such as large-scale validation under varied agricultural conditions and evaluation of potential toxicity to humans and mammals.
Food Application
In response to customers’ demand for healthier diets, innovative methods are needed to incorporate minimally processed foods in daily diets without the use of preservatives. In food matrices (fish, meat, dairy, fruits, vegetables, and drinks), nano-preparations of EO were effectively applied as natural antioxidants and antimicrobials, as well as flavouring agents and shelf-life extenders. EO nanoformulations in food systems offer solutions to the limitations such as rapid evaporation, limited water solubility and instability within food systems. Different types of nanoformulations in food systems are nanoemulsions, nanoencapsulation, and nanoliposomes, and nanofibers. For example, edible coatings made from emulsions formed with EOs improved the storage quality of food. Nanoencapsulation of cinnamon-bark EO by utilising whey protein concentrate, followed by its incorporation into food-grade wax to coat apple surfaces, enhanced the visual charm of apples and demonstrated long-term antibacterial and antifungal effects. Nanoencapsulated EO of Nepeta crispa improved the sensory quality of yoghurt drinks and exhibited a preservative effect by inhibiting Escherichia coli and Staphylococcus aureus. 72 The nanoliposome formulation of Urtica dioica L. EO extended the shelf life and maintained the quality of minced camel meat in refrigerated storage. Eudragit® and collagen nanofibers enriched with bitter orange peel EO are effective for food packaging applications. These biocompatible nanofibers with EO incorporation inhibited the growth of E. coli and S. aureus bacteria and scavenged up to 50% of DPPH (2,2-Diphenyl-1-picrylhydrazyl) radicals. 73 These nanoformulations offer enhanced stability, bioavailability and controlled release of EOs, ensuing in different forms including food preservatives, food packaging and flavour enhancers. However, challenges related to the sensory acceptability of EO nanoformulation containing products must be addressed, and standardised protocols for testing these formulations in food applications need to be developed.
Health Care and Medicinal Application
Over the past decades, extensive studies have been carried out demonstrating the physiochemical and pharmacological properties of EO against various diseases. The pharmacological roles include antimicrobial, anti-inflammatory, antitumor, and antioxidant activities, and these natural products have the potential for new drug developments. The EO-based nano-drug delivery system mitigates the shortcomings of EOs by reducing their volatility and toxicity and improving the bioavailability and chemical stability. For example, nanoemulsions of Cymbopogon martinii EO are stable, antibacterial, and effective against Enterococcus faecalis biofilm, particularly within the affected tooth's root canal. 74 EOs of celandine roots and leaves loaded into chitosan nanoparticles using the emulsion-ionic gelation method demonstrated improved anticancer activity against the MCF-7 cell line. 75 In another work, a wound healing nanofiber consisting of type I collagen nanofibers (as the internal layer) and collagen/PLLA/Zataria Multiflora EO (as the external layer) demonstrated antibacterial and antifungal activity for wound dressing. 76 These EO nanoformulations are administered by different routes, including oral, topical, and inhalation. The nanoformulations of EOs have gained attention in healthcare and medicine due to their ability to improve solubility, stability and controlled and targeted drug delivery. However, further studies are needed to validate the clinical efficacy and safety of these approaches.
Safety and Toxicity of Essential Oil Nanoformulations
The potential of EO nanofomulation in different applications in the therapeutic, food and cosmetics domains is promising. However, these nanoformulations alter the biological interaction of EOs, varying the toxic reactions. A thorough evaluation of the toxicity and safety profile of such nanoformulations is necessary before commercialisation. In vitro studies evaluate the effects, including cytotoxicity and oxidative stress, on various cell lines. In vivo studies employ animal models to assess the impact, such as toxicity, organ histopathology, and biomarkers of inflammation. Table 1 represents the findings of different in vitro and in vivo studies of the EO nanoformulation toxicity studies. It depicts the EOs and their nanoformulations, which have been shown in vitro to exhibit concentration-dependent cytotoxic effects on various cell types. The sensitivity of different cell lines varies, and the toxicity of nanoformulations of EOs also varies. In several in vitro investigations, gene expression analysis has revealed the elevation of genes linked to oxidative stress, such as gst-4, sod-3, gpx-6, and cyp-14a3, indicating that essential oils activate cellular stress and xenobiotic response pathways. 77 Additionally, essential oil-based nanoemulsions have demonstrated concentration-dependent lethal effects on various cancer cell lines. For instance, it encourages oxidative stress and activates apoptotic regulators such as caspase-3 and BAX. In comparison to free essential oils, the nanoformulation also demonstrated improved bioavailability and decreased volatility. 78
Safety/ Toxicity Findings from in Vitro/ in Vivo Studies.
The toxicity of essential oils and nanoformulations is more complex in vivo, with varying effects depending on the oil, formulation, and exposure method. Systemic exposure to nanoemulsions resulted in oxidative stress, while optimised nanoemulsions indicated a safer treatment alternative. 79 For LNEO- NPs, an improved safety profile was observed for nanoformulation as histopathological analysis showed no observable damage to tissues in vivo. The PLGA-based encapsulation enhanced biocompatibility by limiting direct tissue exposure to the EO components. 80 Overall, the in vitro and in vivo studies have shown that EO nanoformulations are safer and effective than free EO. However, to address concerns regarding nanomaterial accumulation and immune responses, life cycle assessments are required to validate their commercial applications.
Despite the benefits offered by nanoformulations of EOs, they also raise regulatory, safety, and environmental concerns about possible effects on health and the ecosystem. As the safety data are limited, future studies could focus on addressing safety concerns and conducting risk assessments for the use of EO nanoformulations. In addition to the unclear safety profile, a regulatory gap exists in the use of nanoformulations of EOs in the food, pharma and cosmetics industry. The authorities have started considering the risks associated with the use of nanomaterials. For example, the European Food Safety Authority gave safety indications for nanotechnology in food in 2009 and updated them in 2020. 81 Similarly, in 2011, the US FDA issued guidance for the use of nanomaterials in animal feed. There still exists a global gap in the regulation, including rules for manufacturing of nanomaterials, labelling of the products containing them, SOPs for in vivo and in vitro toxicity testing and assessing environmental risks from nanoparticle residues.63,82 Encapsulated EOs may persist longer in the environment compared to free volatile EOs, possibly affecting microbial communities. Life cycle assessments need to be done to understand the long-term effects and environmental impact of encapsulated formulations. The nanomaterials, in general, cannot yet be regarded as long-term safe. Safety depends on various factors, including material properties and human genetics, and is therefore variable. The risks include barrier penetration and the possibility of permanent harm triggered by oxidative stress, leading to cumulative toxicity over time. 83 However, the EO nanoformulation often employs biodegradable carrier materials such as starch, lipid, chitosan, gelatin and other naturally derived biopolymers. These polymers aid in biodegradation through microbial, enzymatic or hydrolytic processes when the formulation is subjected to such conditions. For example, chitosan-alginate nanocapsules loaded with turmeric oil and lemongrass oil are hemocompatible, and the components are expected to degrade in biological milieus, indicating their suitability for safe biomedical applications. 84 In the context of food storage, a chitosan-ginger essential oil nanoemulsion loaded gelatin films that is used for food preservation was decomposed within 30 days under soil microbial action. 85 However, studies on biodegradation kinetics and their by-products are required to confirm that no toxic residues accumulate in tissues or environment.
Starch-Based Delivery Systems
Starch is an ideal biopolymer for use in encapsulation. The starch granules exhibit behaviours, such as swelling, gelatinisation, and retrogradation, depending on the temperature, moisture content, and heat. These behaviours have a significant impact on the material's practical use as a polymer.
89
Native starch granules are microscale granules with a dimension in the range of 1–100 μm. Starch is a desirable wall material due to its ease of enzymatic hydrolysis, and starch helps to preserve bioactive chemicals from being broken down during gastrointestinal digestion. Starch-based nanoformulations provide enhanced loading rate and accurate delivery of biological compounds that are sensitive and readily degraded because of their enormous specific surface area, superior stability and pH responsiveness.
90
Table 2 examines different starch-mediated nanocarriers for essential oils. The table illustrates various starch sources, including potato, corn, cassava, and avocado seed, used for nanoformulation preparation. These nanoformulations have applications in various fields, and a wide range of nanofabrication techniques are available, including nanoprecipitation, emulsification, inclusion complexation, and coacervation, among others. Figures 5 and 6 illustrate different approaches in nanoformulation preparation. Nanoformualtuond of different sizes, charges, compositions and physical properties can be prepared using these approaches. The standard methods include nanoprecipitation, coacervation, electrospinning, inclusion complexation, nanoemulsion, and ionic gelation. The nanoprecipitation (Figure 5a) process involves the addition of organic phase and EO to polymer solution (wall material), which leads to the precipitation of polymers on the nanoscale. In coacervation (Figure 5b), EOs are combined with a solution containing two different biopolymer molecules, typically a protein-polysaccharide. The initial pH prevents the interaction between the polymers. The polymers attract electrostatically once the pH is below the isoelectric point, resulting in the formation of a structure that encases EOs within it.
91
Electrospinning (Figure5c) provides easy regulation in the generation of continuous and consistent nanofibers, and the consequent nanofibers are characterised by a high specific surface area and high porousness, which renders them especially appropriate for the loading and regulated release of compounds.
92
Through electrospinning, starch can be fabricated into nanocapsules or nanofibers with regulated dimensions. Starch with a high amylose content is strongly related to electrospinnability because of the linearity of this macromolecule, which makes it susceptible to entanglement through the electrospinning process.
93
The synthesis parameters, like the usage of surfactants during the process, impact the mean diameters of starch nanoparticles. Inclusion complexation (

EO Nanoformulaton Fabrication Approaches and Their Merits (a) Nanoprecipitation, (b) Coacervation, (c) Electrospinning.

EO Nanoformulation Fabrication Approaches and Their Merits (a) Inclusion Complexation, (b) Nanoemulsion, (c) Ionic Gelation.
Starch-Mediated Nanotechnology-Based Delivery Systems for Essential Oils.
Starch has found widespread applications in pharmaceuticals, cosmetics, food, and agriculture due to its characteristics and nature, including biodegradability, renewability, affordability, and availability. Starch is a GRAS biopolymer and a promising drug carrier due to its abundance, inexpensiveness, and non-allergenic nature.
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The benefits of starch-based encapsulation systems over alternative wall materials (such as proteins, lipids, and other polysaccharides) can be related to the following:
Resistant starch evades small intestine digestion and delivers the bioactive chemicals to specific locations. Amylose frequently forms V-type inclusion complexes with hydrophobic molecules, which are resistant to digestion and have high thermal endurance. Gelatinised starch is readily converted into a paste and can form films.
Because starch comes from various affordable sources, it is a popular ingredient in food and can encapsulate other food additives, with customer acceptance. 95
Chemical Modification of Starch
Starch is a popular polymer choice for the encapsulation of essential oil. Nonetheless, the starch particles are ineffective in emulsion stabilisation due to their enormous hydroxyl groups. 103 The degree of polymerisation in starch can be altered to modify hydrogen bonds, which, in turn, enhances polymer structure and ability to retain volatile substances. 105 To improve the ability of starch to disperse in water, modifications through processing, disintegration, or compounding can yield a variety of derivatives, including modified starch, dextrin, oligosaccharides, and sugar alcohols. Such changes improve its stability and provide additional benefits. Chemical alteration with octenyl succinic anhydride (OSA) results in the hydrophobic groups in the starch molecule, increasing its suitability with the oil-in-water interface. 106
OSA-Modified Starch
Octenyl succinylated starch (OS-starch), a hydrocolloid with amphiphilic characteristics, is produced when starch is esterified using OSA. Because of its extraordinary emulsification capacity, amphiphilicity, and non-toxicity, OS starch has been extensively employed as an encapsulating material in starch-based encapsulation systems. However, encapsulating particles made solely of OS-starch typically showed less oxidative stability and more wrinkles. 107 The use of a blend of OSA-modified starch with other polymers, such as proteins (gelatin and whey protein isolate) or polysaccharides (including maltodextrin and alginate) for encapsulation solves the existing challenge. Moreover, nanosized materials tend to agglomerate spontaneously to reach a thermodynamic equilibrium state, and consequently, modified nano-starch with enhanced stability, hydrophobicity, and dispersion properties is of interest. 89 OSA-S, a hydrophobically altered starch, has increased surface activity and adequate thickening potential beyond pH 4.5. The generated hydrophobic groups improve the molecule's interaction with the oil-in-water interaction. However, high molecular masses in OSA-modified starch may result in over-aggregation and poor encapsulation effectiveness because of its low gravitational stability and high viscosity. 108
Majeed et al prepared a food preservative formulation using OSA starch encapsulation. Ultrasonic preparation of nanoemulsions composed of eugenol, carvacrol, 2% succinylated starch, 10% oil phase, and sonication for 10 min has broad-spectrum antimicrobial activity. The formulation is a green preservative in the food industry, as it exhibited sustained release and bioaccessibility up to 80% in the mice serum. 109 Similarly, the natural antibacterial cinnamaldehyde-EO-loaded electrospun nanofibers of OSS and pullulan (PUL) have applications in food packaging and wound dressings. A greater degree of substitution and lower molecular size of OSS are advantageous for electrospinning OSS/PUL into superior nanofiber mats. 110 Biodegradable food packaging films of pullulan incorporated with cinnamon EO (CEO) emulsified by OSA starch improved their properties. CEO nanoemulsion addition improved the thickness (29%), elongation at break (37%), and antibacterial activity of the film. 106
Cyclodextrins
Cyclodextrins (CDs) are suitable to encapsulate a wide range of compounds because of their properties, such as their ability to enhance the dissolution of lipophilic substances in both the solid and liquid states and to generate weak intermolecular interactions. 111 The enzymatic breakdown of starch synthesises cyclic oligosaccharides and includes glucose units connected by α-(1,4)-glycosidic linkages. They can form inclusion complexes with compounds of limited water solubility. Such complexes of EO in cyclodextrin enhance its solubility and stability and offer a prolonged release of these compounds. 112 In CD, hydrogen atoms and glycosidic oxygen linkages rotate inwardly while the hydroxyl group turns outward to offer a lipophilic environment. The amount of glucose determines the cavity size and ability to accommodate foreign molecules. CDs form inclusion complexes wherein the hydrophobic cavity encapsulates the foreign compound in solid and liquid phases. 113
Recent research on the use of cyclodextrin in EO encapsulation demonstrates superior properties. For example, β-cyclodextrin loaded with oregano EO and incorporated into polylactic acid and polycaprolactone electrospun nanofibers is an active packaging material. The material exhibits improved thermal stability, anti-deformation properties, and antimicrobial capabilities. It is a safe material and delays blackberries’ post-harvest decay and quality deterioration. 114 The use of the methyl-β-cyclodextrin EO complex for wood preservation is owing to the antifungal potential of EO. The EOs used are eugenol, trans-cinnamaldehyde, thymol, and carvacrol, and they showed high inclusion yield. The complex-treated pine sapwood logs demonstrated enhanced decay resistance. 69 Eugenia brejoensis EO inclusion complex formation with β-cyclodextrin in cervical cancer cells enhanced the cytotoxic effect. In addition, the complex formation improved the thermal stability and selectivity of the tumour lineage. 115 Edible antimicrobial films of chitosan incorporating clove oil/β-cyclodextrin complex are suitable food packaging materials to extend shelf life. 116 Orange EO combined with zein and the β-cyclodextrin inclusion complex has antifungal properties. The formulation is a food preservative that slows the microbiological deterioration of cakes from 30 to 150 days. Overall, cyclodextrins serve as highly effective carriers for EO; however, limitations, including size selectivity and high cost of production, must be addressed to realise their applications.
Starch-Based Nanostructures
Starch Nanoparticles
Starch is hydrophilic, and loading hydrophobic medications, or EOs, onto nanoparticles is particularly challenging. Starch nanoparticles (SNPs) are not used as encapsulating protective wall material, but rather as adsorbent carriers for essential oil. 117 Qiu and coworkers produced SNPs by nanoprecipitation of debranched waxy corn starch and loaded with menthone. The aqueous environments enhanced the solubility of menthone loaded onto SNPs since the SNPs are of greater surface area and hydrophilic. 97 Liu et al (2017) established a rapid and environmentally friendly ultrasonic method for fabricating starch nanoparticles for encapsulating peppermint essential oil. This NP-EO enhanced thermal stability and enabled sustained release. 118 SNPs offer promising applications in the medical field, functional food, and cosmetics; however, challenges such as scalability and structural uniformity must be addressed.
Starch Nanofibers
Nanofibers are suitable for the regulated release and encapsulation of bioactive compounds due to their high porosity and enormous specific surface area. 92 In starch, high amylose content enables nanofiber synthesis by electrospinning, while amylopectin is ineffective in electrospinning. Native starch needs organic solvents for complete dissolution and is unsuitable for electrospinning fibers. OSS can produce molecular entanglements that promote nanofiber production in an aqueous solution. 119 OSS with greater levels of substitution and lower dimensions are advantageous for electrospinning into superior nanofiber mats. The key factors influencing the electrospinning of OSS/Pullunan aqueous solution are lower surface tension, enhanced conductivity, and moderate apparent viscosity. For electrospun OSS/Pullulan nanofiber mats, cinnamon EO loading via physical adsorption altered the fiber microstructure and demonstrated antibacterial properties against Staphylococcus aureus, Escherichia coli, and Aspergillus flavus.
When compared to nanofibers formed of neat starch, the addition of thyme EO appeared to improve the characteristics (ie, homogenous and continuous fibres) of the electrospun nanofibers, decreasing bead formation and altering the diameter. 120 The carvacrol-loaded starch nanofibers are suitable for food packaging applications and exhibit superior qualities such as homogeneous morphology, 73–95 nm in size, and enhanced antibacterial and antioxidant activity of 83%. 100
Starch in Nanoemulsions
Developing oil-in-water nanoemulsions is a common approach for encasing lipophilic functional compounds to improve their activity and stability. For instance, the black cumin essential oil nanoemulsions with canola and flaxseed oils as ripening inhibitors were prepared and stabilised with OSA modified waxy maize starch. Nanoemulsions showed monomodal size distributions, which suggested a significant electrostatic repulsion between the scattered oil droplets. It exhibited improved stability, regulated release, and capacity to self-assemble with the gram-positive bacterial cell membrane before destroying the constituent cells, and NE demonstrated longer-lasting bactericidal activity. Nanoemulsion prepared with cinnamon EO was emulsified with OSA-modified starch. 121 Further, prepared nanoemulsion was added to pullulan film to improve its properties, such as water vapor permeability, tensile strength, antimicrobial effect, and reduction in water content. The OSA-modified starch increased the loading capacity of cinnamon EO in pullulan film. 106 These nanofiber incorporated with essential oil is a promising material in food packaging applications; however, further studies are required for their large-scale fabrication methods.
Conclusions and Future Perspectives
In the present paper, a comprehensive state-of-the-art review of the recent progress in different phenomenal aspects of nano-delivery systems of essential oils is considered, particularly emphasising the information gathered from the reported literature in recent years. It is observed that essential oils nanoformulations are of great significance because they overcome characteristic challenges of EOs, such as volatility, poor water solubility, and susceptibility to degradation. Such nanoformulations, with enhanced stability, sustained release, targeted delivery, and improved bioactivity, are well-suited for applications in the pharmaceutical, agricultural, food, and cosmetic industries. The various nanocarriers include nanoparticles, nanoemulsions, nanofibers, nanoencapsulations, polymeric nanoparticles, and nanocapsules. However, the type of nanocarrier system depends on its application and the nature of its target. The hydrophilic starch-based nanocarrier is suitable for a variety of formulations in various applications. Compared to synthetic polymers, starch nanocarriers offer a more sustainable approach due to their biocompatibility and biodegradability.
Although EO nanoformulations exhibit beneficial outcomes, further research is necessary to determine their fabrication and application in multiple industries on a large scale. Standardisation of methods of preparation, optimisation, and surface modification would enhance the reproducibility and commercialisation. Future studies should also address safety issues by performing risk assessment for the use of EO nanoformulations. The regulatory validation, specifically through in vivo studies, needs to determine the pharmacokinetics, nanomaterial accumulation, biodegradation kinetics, possible residues that accumulate in tissues or the environment and long-term safety profiles. The transition of EO nanoformulations from lab scale to the real world requires collaborative efforts between biotechnologists, biomaterial scientists, pharmacologists, and regulatory bodies.
Footnotes
Acknowledgment
The authors are thankful to Karunya Institute of Technology and Sciences, Coimbatore, India for providing support for this study.
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The authors declare that no funds, grants, or other support were received during the preparation of this manuscript.
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