Niosomes: a review of their structure, properties, methods of preparation, and medical applications

Niosomes are spherical and consist of microscopic lamellar (unilamellar or multilamellar) structures (Figure 1). The bilayer is formed by nonionic surfactants, with or without cholesterol and a charge inducer [20, 21]. Different types of surfactants at variable combinations and molar ratios are used to form niosomes [22]. Examples of surfactants include alkyl ethers, alkyl glyceryl ethers, sorbitan fatty acid esters, and polyoxyethylene fatty acid esters [7]. Addition of cholesterol maintains the rigidity of the bilayer, resulting in less leaky niosomes. Meanwhile, charge inducers provide charge to the vesicles and increase vesicle size, increasing drug entrapment efficiency. Negative charge inducers, including dicetyl phosphate, dihexadecyl phosphate, and lipoamino acid, and positive charge inducers, including stearylamine and cetylpyridinium chloride, help to stabilize the vesicles [23,24,25,26]. Nonionic surfactants in niosomes tend to orient themselves in such a way that hydrophilic end faces outward (toward the aqueous phase), whereas the hydrophobic end faces inward to each other to form a closed bilayer structure, which encloses solutes in an aqueous solution [20]. As a result, the closed bilayer structure of niosomes has hydrophilic inner and outer surfaces, with a sandwiched lipophilic area in between [8, 26]. To form the closed bilayer structure, energy such as heat or physical agitation is required. Various forces inside the vesicles were found to play an important role in maintaining the vesicular structure, for example, van der Waals and repulsive forces that exist among the surfactant molecules. Varying the vesicle’s components (including type, composition, and concentration), size, surface charge, or volume will likely modify the properties of resultant niosomes [20, 21, 27].

Niosomes can be categorized into 3 groups based on their vesicle size, namely, small unilamellar vesicles (0.025–0.05 μm), multilamellar vesicles (>0.05 μm), and large unilamellar vesicles (>0.10 μm) [27].

Figure 1

Preparation of niosomes begins with the hydration of a surfactant and lipid mixture at elevated temperatures, followed by optional niosome size reduction in order to obtain a colloidal suspension [28]. There are several well-studied standard methods for the preparation of niosomes. Ether injection, hand shaking, sonication, and microfluidization methods are a few examples [7, 27, 29]. Subsequently, the unentrapped drug is separated from the entrapped drug by centrifugation, gel filtration, or dialysis [28].

The first step in niosome production by ether injection is through the dissolution of surfactant in diethyl ether. The solution is then injected through a 14-gauge needle into an aqueous solution of drug maintained at 60°C. Subsequently, single-layer vesicles with diameters ranging from 50 to 1000 nm are formed because of the vaporization of ether [20]. However, a small amount of residual ether frequently persists in the niosomal suspension [27].

In the hand-shaking method, also known as thin-film hydration technique, surfactant and cholesterol are dissolved in a volatile organic solvent and transferred to a rotary evaporator. After evaporation, a thin layer of solid mixture is deposited on the wall of the flask. The dried layer is then hydrated with an aqueous phase containing the drug of interest. This process may be carried out at room temperature with gentle agitation [15, 27].

Niosomes can also be produced through sonicating a mixture of surfactant, cholesterol, and aqueous phase containing the drug at 60°C for 3 min. The vesicles produced through this method are usually small and uniform in size [15, 27]. Microfluidization is another reproducible technique that achieves this size uniformity. Operationally, 2 fluidized streams move forward through a precisely defined micro-channel, and these 2 streams interact with each other at an ultrahigh velocity [20, 27, 30].

Alternative methods have since been defined for the preparation of niosomes. The multiple membrane extrusion method uses surfactant, cholesterol, and dicetyl phosphate in chloroform, and the mixture is evaporated to produce a thin film. The film is then hydrated with aqueous drug solution, and the suspension produced is extruded through polycarbonate membranes, which are placed in series for up to 8 passages [20, 27, 31].

The reverse-phase evaporation technique uses a mixture containing surfactant and cholesterol in a 1:1 ratio, in addition to ether and chloroform. An aqueous phase containing the target drug is added to the mixture followed by sonication at 4–5°C. Sonication is continued after adding a small amount of phosphate-buffered saline to the mixture. The organic solvent is removed at 40°C under a low pressure, and the remaining suspension is diluted with phosphate-buffered saline. After heating the mixture at 60°C for 10 min, the final product of niosomes is obtained [20, 27, 32]. The preparation of niosomes using the reverse-phase evaporation technique is illustrated in Figure 2.

Figure 2

Niosomes can be produced without the use of organic solvents using the “bubble” method. A “bubbling unit” consists of a round-bottomed flask with 3 necks positioned in a water bath; a water-cooled reflux condenser and thermometer are positioned in the first and second necks, respectively, while nitrogen is supplied through the third neck. Surfactant and cholesterol that are mixed at 70°C in a buffer are homogenized and “bubbled” at 70°C using the “bubbling unit” [20, 33]. The preparation of niosomes using this technique is illustrated in Figure 3.

Figure 3

In another variation of preparation of niosomes, a thin film resulting from evaporation of surfactant and cholesterol dissolved in chloroform is hydrated with 300 mM citric acid (pH 4.0). Then, the suspension is subjected to 3 successive freeze–thaw cycles. After sonication, the aqueous solution containing the drug is added to the suspension and vortexed. Disodium phosphate (1 M) is added to the mixture to increase the pH to 7.0–7.2. This mixture is later heated at 60°C for 10 min to produce niosomes. This method is known as the “transmembrane pH gradient (inside acidic) drug uptake process” [20, 27, 34]. Niosomes obtained by this method showed better entrapment efficiency and retention of drugs [35, 36]. Figure 4 shows the steps for the preparation of niosomes using this technique.

Figure 4

The emulsion method, which uses an oil-in-water emulsion prepared from an organic solution of surfactant, cholesterol, and an aqueous solution of drug, is another technique for preparation of niosomes. The organic solvent is evaporated to obtain the final product [8, 27, 37]. By contrast, a mixture of lipids and surfactant is melted and injected into a heated aqueous phase containing the drug in the lipid injection method [27].

Surfactant-based niosomes are biodegradable, biocompatible, and nonimmunogenic [23]. They act as a drug depot in the body, whereby they release drugs in a controlled manner through their closed bilayer structure, resulting in a sustained release of the enclosed drug to the target site [38]. The therapeutic effects of drugs enclosed in niosomes are improved by reduced clearance and specific targeting [15]. Because of their hydrophilic, amphiphilic, and lipophilic nature, niosomes are able to accommodate a wide variety of drugs with a wide range of solubility [39]. Bioavailability of otherwise poorly soluble drugs may be improved, and the efficacy of topical applications would be enhanced with the use of niosomes. Furthermore, labile and sensitive drugs may be delivered with greater ease as niosomes protect the encapsulated active pharmaceutical ingredients from deleterious conditions both inside and outside of the body [38].

The stability of niosomes is mainly affected by the type of surfactant, properties of the encapsulated drug, temperature of hydration, detergent, membrane-spanning lipids, polymerization of surfactant monomers, and charged molecules [20, 21, 27]. It is crucial for the surfactant used for the preparation of niosomes to consist of a hydrophilic head and a hydrophobic tail. Generally, surfactants possessing hydrophobic tails with an alkyl (chain length from C₁₂ to C₁₈), perfluoroalkyl, or steroidal groups are suitable for the preparation of niosomes [8]. Ester-type surfactants are less suitable because they are easily degraded by esterases in vivo, making them unstable in the body.

Comparatively, ether-type surfactants are a better choice. The size of niosomes increases proportionally with the increase in hydrophilic–lipophilic balance (HLB) of surfactants. If the HLB value falls between 4 and 8, niosome vesicle formation is considered relatively stable and optimal [8, 27, 40]. Addition of different types of additives together with the drug entrapped in niosomes is able to improve niosome stability. For instance, addition of cholesterol provides rigidity and reduces leaking in niosomes [41].

Hydration temperature plays an important role in the assembly of surfactants into vesicles and the shape and size formation of niosomes. Ideally, the temperature chosen should be above the temperature of the gel-to-liquid phase transition [8, 27, 42]. Encapsulated drugs usually interact with the head group of the surfactant and thus indirectly influence the charge and rigidity of the bilayer structure of the niosome. Hydrophobic drugs generally improve the stability of niosomes, while hydrophobic drugs decrease their stability. Interestingly, amphiphilic drugs have no obvious effect on the niosomes’ bilayer structure [27, 43].

Common problems related to preparation of niosomes include aggregation, fusion, and leaking, which are affected by the physiochemical properties of vesicles including their size, charge, lamellarity, elasticity, and thermodynamic phase. Preparation of proniosomes, which are a dried form of niosomes, might overcome the limitations mentioned because proniosomes hydrate immediately before use to yield an aqueous niosome dispersion [44]. Studies have found that proniosomes carried better therapeutic efficacy for anti-inflammatory drugs (flurbiprofen and piroxicam) administrated via a transdermal route [45, 46].

Niosomes are generally nontoxic to humans [27]. Researchers studied the toxic effect of surfactants in a topical niosome formulation on the proliferation of keratinocytes. The results show that ester was less toxic than ether because of enzymatic degradation of ester bonds. It had been proven that the physical form of niosomes (liquid crystal vs gel) did not influence their toxicity [47].

Niosomes possess a bilayer structure, which is similar to liposomes. However, the materials used to prepare niosomes confer better stability on them [20]. Niosomes are prepared from uncharged single chain surfactant and cholesterols. By contrast, liposomes are prepared from neutral or charged double chain phospholipids. The concentration of cholesterol is higher in liposomes than in niosomes. As a result, the drug entrapment efficiency of liposomes is less than that of niosomes. Niosomes are cost-effective for industrial manufacture and do not require special storage conditions, which are essential while manufacturing liposomes. The cost of liposome preparation is high because of the unstable chemical ingredients (phospholipids), which undergo oxidative degradation. Liposomes therefore require special handling methods [27]. Niosomes possess a longer shelf life than liposomes [47]. They prolong the circulation of encapsulated drugs and increase metabolic stability in an emulsified form, whereas liposomes have limited shelf life because of rancidification of their lipid components [7, 47,48,49,50].

Drug-delivery systems using niosomes through transdermal, parenteral, and ophthalmic routes are well studied [27, 51, 52]. Niosomal delivery via transdermal routes is able to overcome the slow penetration rate of conventional transdermal approaches. The bioavailability and therapeutic efficacy of drugs such as diclofenac, flurbiprofen, and nimesulide increase by their incorporation into niosomal formulations. For ophthalmic drug delivery, chitosan-coated niosomal formulation of timolol maleate exhibits a greater effect in reducing intraocular pressure compared with marketed formulations, with less cardiovascular side effects [52]. Because of their many desirable properties, niosomal formulations have been used in many other therapeutic applications, as discussed in the following sections.

Leishmaniasis is a parasitic disease invading the liver and spleen transmitted through the bite of a female sandfly. Therapeutic efficacy of amarogentin (an antileishmanial, secoiridoid glycoside isolated from Indian medicinal plant Swertia chirata) was examined in an experimental model of leishmaniasis in hamster. The efficacy of amarogentin was compared using liposomes and niosomes as tools of delivery to the infected site. Amarogentin encapsulated in niosomes was more efficacious than that in liposomes at the same level of membrane microviscosity [25]. Toxicity studies including blood pathology, histology, and specific enzyme levels related to normal liver function showed that the niosomal formulation had no apparent toxic effect [53].

Acquired immune deficiency syndrome (AIDS) is characterized by a severe damage to the immune system caused by the human immunodeficiency virus (HIV). Zidovudine (AZT), a pioneering anti-HIV compound, is approved for clinical use, either alone or in combination with other antiviral agents, for the treatment of AIDS and AIDS-related complex. However, there are some setbacks in AZT administration, including hematological toxicity, poor bioavailability, high first-pass metabolism, and short half-life [54]. A study revealed that niosomal AZT formulation prolonged the drug half-life in rabbit serum [55]. In another study, niosomes manufactured with Tween 80 were able to encapsulate a high amount of AZT, and addition of dicetyl phosphate enhanced drug release for a prolonged period (88.72% over 12 h). Changes in the micromolar ratios of nonionic surfactants with a constant ratio of cholesterol during the preparation of niosomes were associated with the changes in the encapsulation efficiency and controlled release of AZT [56].

Inhalation therapy utilizing glucocorticosteroids such as beclomethasone dipropionate (BDP) in chronic obstructive pulmonary disease (COPD) is promising, but the common drawback of the medication is poor permeation through hydrophilic mucus to reach the glucocorticoid receptors of bronchial epithelial cells [74]. Niosomes developed from polysorbate 20 containing BDP are well suited as a drug-delivery system for COPD patients through pulmonary delivery. This is because of the advantages offered by the niosomes, such as high drug encapsulation efficiency, strong mucus permeation, and sustained delivery to the target site [75]. Thus, the use of niosomes overcame weaknesses of the drug and other conventional inhalation therapies. Niosomes may offer a better targeted-delivery system for patients with COPD as these nonionic vesicles remarkably increased the permeation rate of BDP through the mucosal membrane barrier. Clinical efficacy of an inhalation therapy is usually dependent on the aerodynamic size distribution of the aerosol and drug output from a nebulizer. Niosomal dispersion provides a better aerodynamic diameter than the commercial products’ dispersion. Under all conditions of nebulization, niosomes provide a higher efficacy, thus offering better targeting of corticosteroids in the treatment of COPD [74].

Niosomal encapsulation of isoniazid was found to effectively treat tuberculosis, with 62% of cellular uptake by macrophages. In addition, the niosomal formulation decreased the dose required and reduced the level of toxicity; these contributed to improved patient compliance. The additional advantages of the niosomal formulation are that it was site specific to where tuberculosis bacteria were harbored and it was able to maintain steady drug concentrations for up to 30 h [76]. Another study showed that intravenous and intraperitoneal administration of rifampicin encapsulated in niosomes prepared using surfactants of sorbitan ester class:cholesterol (50:50 percent mole fraction ratio) could be used for the treatment of tuberculosis [77]. Positive results were noted with rifampicin encapsulated in Span 85 (sorbitan trioleate)-based niosomes. The rifampicin was found accumulated in the lungs of mice, therefore offering possibility of improved antituberculosis therapy [8, 78].

Gentamycin sulfate used in the treatment of nosocomial pneumonia displays short half-life and has various side effects. Niosomal formulations may ensure that an efficient concentration of the drug is achieved in the lungs without inducing systemic effects. Gentamycin sulfate in niosomes prepared with polyoxyethylene sorbitan esters showed a significantly higher accumulation in lungs than the plain drug, indicating that niosomes are an enhanced drug-delivery system for this therapy [79].

Niosomal formulations of diclofenac sodium, nimesulide, and flurbiprofen exhibit greater anti-inflammatory activity than the free drugs [52]. Niosomal formulations of diclofenac diethylammonium, aceclofenac, meloxicam, and lornoxicam used for topical application also show good anti-inflammatory activity because of the penetration of niosomes into the deeper layers of the skin [7, 80,81,82]. Additionally, mefenamic acid-loaded niosomes prepared by Kamboj et al. [83] demonstrated enhanced inhibition of inflammation in vivo.

Serratiopeptidase is controversially used in the treatment of arthritis, fibrocystic breast disease, chronic bronchitis, and carpal tunnel syndrome to alleviate pain and inflammation. Orally administered serratiopeptidase has been reported to show systemic side effects. Therefore, a topical formulation of serratiopeptidase that potentially reduces adverse effects and increase local effects has been developed by Shinde and Kanojiya using niosomes. The results showed that the newly formulated serratiopeptidase niosomal gel had anti-inflammatory activity comparable to that of diclofenac gel [84].

A number of compounds isolated from natural products have recently emerged as potential anti-inflammatory agents. Compounds isolated from Glycyrrhiza glabra L. and Zingiber cassumunar Roxb. are a few examples. Extracts of G. glabra are purported to be useful in the treatment of dermatitis, eczema, and psoriasis [85, 86]. The plant’s active compound, ammonium glycyrrhizinate, showed strong anti-inflammatory effects in various models [87, 88]. When ammonium glycyrrhizinate was encapsulated in niosomes prepared by using Tween 85, Span 20, and cholesterol in various molar ratios, improved anti-inflammatory activities were observed in chemically induced skin erythema in mice and humans [89]. (E)-4-(3′,4′-dimethoxyphenyl)but-3-en-1-ol (also known as compound D) is the active compound isolated from Z. cassumunar. Encapsulation of compound D in niosomes enhanced its chemical stability and skin permeation with anti-inflammatory effects comparable to commercial anti-inflammatory drugs when applied on the skin of mice [90]. Z. cassumunar-encapsulated niosomes were also effective for local subcutaneous inflammation when used in combination with therapeutic ultrasound [91].

Resveratrol is a polyphenolic compound found in grape skin, peanuts, and the roots of Polygonum cuspidatum [92]. Topical application of resveratrol has shown to improve some types of skin conditions such as inflammation. However, the therapeutic potential of resveratrol is limited because of its poor bioavailability, low water solubility, chemical instability, and limited skin permeability [93, 94]. Recently, a resveratrol-loaded niosomal hydrogel system, which could enhance the permeation and deposition of resveratrol in skin, had been developed by Negi et al. Administration of this niosomal gel reduced edema with prolonged therapeutic action, as evaluated with a model of edema induced in rat paw by carrageenan injection [95].

Flurbiprofen is commonly used in the treatment of ocular inflammatory diseases [96]. However, challenges arise with the use of the ocular drugs, including limited ocular bioavailability because of nasolacrimal drainage, the corneal barrier, and unintentional absorption of the drugs to the systemic circulation [97]. As a result, not more than 5% of applied drugs can reach intraocular tissues [98]. Therefore, to enhance the ocular bioavailability of flurbiprofen, the drug was encapsulated in niosomes consisting of nonionic surfactant Span 60 using the thin-film hydration technique. The encapsulated flurbiprofen was found to rapidly reduce eye inflammation of New Zealand albino rabbits, with a higher ocular bioavailability than that of a flurbiprofen solution [99].

Itraconazole is the drug of choice for treating fungal infections caused by Blastomyces dermatitidis, Histoplasma capsulatum, Paracoccidioides brasiliensis, and Coccidioides immitis. It is used in the treatment of pseudallescheriasis, sporotrichosis, tinea corporis, tinea versicolor, and toenail onychomycosis. However, it has poor solubility and permeation that hinder its absorption through skin. Incorporation of the drug in niosomes enhanced its therapeutic efficacy, as evidenced by a study in which the itraconazole niosomes demonstrated greater antimycotic activity against Candida albicans than marketed formulations [100]. Another potentially antifungal drug called diallyl disulfide was found to be entrapped in niosomes prepared efficiently using Span 80. When the niosomal diallyl disulfide was administered to C. albicans-infected mice, it cleared the fungal burden and increased the survival rate of the mice; the effect was better than the drug in free form [101]. Fluconazole, which is commonly available in parental and oral dosage forms, is used in the treatment of cutaneous candidiasis. However, the drug is well known for its adverse effects such as taste disturbances and gastrointestinal irritation. When the drug is given in an oral form, a high dose is usually required to reach therapeutic concentrations. Therefore, a topical formulation of the drug was developed. Fluconazole-loaded niosomal gels were formulated as a topical ocular drug-delivery system for corneal fungal infections. Fluconazole was incorporated into poloxamer or chitosan niosomal gel, and the antifungal effects of the 2 gels were compared. Fluconazole in the poloxamer gel showed a better effect than it did in the chitosan gel [102]. In another study, miconazole loaded in proniosomal vesicles was proven to be effective against Trichophyton rubrum in the treatment of tinea infections [103].

Nisin is an antimicrobial agent used in food and pharmaceutical applications [104]. However, its effectiveness is limited because of its inaccessibility to an inner membrane of bacteria. Hence, ethylenediaminetetraacetic acid (EDTA) is used to improve the efficacy of nisin [105]. Niosomes encapsulating both nisin and EDTA were evaluated for their antibacterial activity. The encapsulated form of nisin and EDTA showed a better and longer-lasting effect in inhibiting Staphylococcus aureus than their free forms [106]. Gallidermin, an antibiotic that has effects similar to erythromycin or fusidic acid, is effective against endocarditis, abscesses, skin infections, and acne [107]. When gallidermin is loaded in anionic niosomes and incorporated in gel, it accumulated in the skin with no risk of a systemic effect and displayed better antibacterial effects, particularly against Propionibacterium acnes and S. aureus [108]. Propolis is considered to have broad-spectrum antimicrobial properties [109]. Its antimicrobial effect was enhanced when it was incorporated into niosomes that were prepared with varying concentrations of Span 60 and cholesterol. This is because niosomes are able to interact with the bacterial cell envelope, thereby facilitating the diffusion of constituents of propolis across the cell wall [110].

Gatifloxacin is widely used for ocular infections. Patients are required to apply the medicine several times a day, and this greatly inconveniences the patients. Therefore, Zubairu et al. designed and formulated a novel delivery system for gatifloxacin that could help in reducing the dosing frequency. A chitosan-coated niosomal formulation of gatifloxacin developed by researchers showed a longer retention time on the eyes, suggesting the potential use of the drug in the form of transcorneal delivery [111]. Another group of scientists developed chitosan gel-embedded moxifloxacin niosomes, and the drug formulation was used for topical microbial infections [112].

Pseudomonas aeruginosa causes highly resistant infections through biofilm formation, which inhibits the entry of antibiotics [113]. Synergistic combination therapy is believed to be more effective in the treatment of infections related to biofilms. Hence, a niosomal drug formulation consisting of bismuth-ethanedithiol and tobramycin was developed and its antibacterial effect was tested through the inhibition of biofilm formation and reduction of N-acyl homoserine lactone in P. aeruginosa. Results showed that the formulation may be useful in the treatment of infections caused by this pathogen [114]. Antibacterial and antifungal effects of the niosomal drugs/compounds mentioned are summarized in Table 2.

Table 2

Drugs/compounds encapsulated in niosomes and their antibacterial or antifungal effects

Niosomal formulations have been used extensively in many other applications such as transdermal delivery. For example, 8-methoxypsoralen, a compound used in psoralen ultraviolet A therapy [115], had been formulated into niosomal vesicles and delivered topically to increase its local efficacy and safety [116]. Another group of investigators has attempted to develop a niosomal gel formulation with acyclovir that has a superior topical bioavailability [117]. Acyclovir is used in the treatment of herpes simplex, varicella zoster, and herpes zoster viral infections. Its usual route of oral administration is plagued by poor bioavailability and short elimination half-life [118]. Usual topical delivery is limited by the poor intrinsic permeation of acyclovir into the basal epidermis. A goal of topical delivery is that the drug is transported through the stratum corneum, thereby effectively reaching the target tissue. The niosomal gel developed by Jacob et al. [117] demonstrated successful delivery of acyclovir through topical administration.

Niosomes could also potentially serve as gene delivery system. Niosomes prepared by using cationic lipid N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride and the nonionic surfactant polysorbate 60, with or without lycopene, were used to encapsulate promoter of cytomegalovirus-enhanced green florescent protein (pCMS-EGFP)-based plasmids. The niosomes generated were then used in the transfection of ARPE-19 human retinal pigment epithelial cells. The higher transfection efficiency obtained indicates the potential use of this gene delivery system for various inherited retinal diseases [119]. Opanasopit et al. also investigated the use of niosomes using this approach. They reported that cationic niosomes composed of Span 20, cholesterol, and spermine-based cationic lipids encapsulating pDNA encoding EGFP contributed to a high transfection efficiency of HeLa human cervical cells [120].

Other uses of niosomes include the treatment of glaucoma using dorzolamide niosomes [121], specific niosomal drug-delivery systems that target the liver [122] and induction of angiogenesis using growth factor-loaded nano-niosomal gel formulations [123]. Moghassemi et al. [124] formulated bovine serum albumin-loaded niosomes that have potential use in pharmaceutical and cosmetic applications.