IJPRD, 2011; Vol 4(06): August-2012 (137 152)

International Standard Serial Number 0974 9446

-------------------------------------------------------------------------------------------------------------------------------------------------NANOEMULSIONS AN EMERGING TREND: A REVIEW R.B.Desi reddy*1, Ch.T. Lalitha kumari1, G.Naga sowjanya1, S.L.Sindhuri1, P.Bandhavi1
1

Nalanda Institute of Pharmaceutical sciences, Kantepudi, Sattenapalli, India

ABSTRACT Nanoemulsions are one of the emerging trends in targeted & controlled drug delivery systems . Nanoemulsions are clear, thermodynamically stable, isotropic liquid mixtures of oil, water,surfactant and co-surfactant. These are oil-in-water (o/w) type of emulsions with the average droplet size ranging from 5nm to 100 nm. Reduction in droplet size to nanoscale leads to change in physical properties such as optical transparency & unusual elastic behavior. Nanoemulsions have widespread applications in different fields such as pharmaceutics, food technology . Nanoemulsion offers a promising vehicle for increasing the aqueous solubility of poorly water-soluble drugs considerably, which is usually necessary for parenteral application. Nanoemulsions have many advantages, for instance, enhance drug solubility, perfect thermodynamic stability, ease of manufacturing and permeation over conventional formulations that convert them to important drug delivery systems. Nanoemulsion can improve transdermal delivery of lipophilic and hydrophilic compound with different mechanisms. The design & development of nanoemulsions aimed at controlling or improving required bioavailability levels of therapeutic agents.. This review mainly discussed about the importance of nanoemulsions over other dosage forms, preparation methods ,current state of nanoemulsions in the delivery of drugs and other bioactives and characterization of nanoemulsions, applications. KEYWORDS : Nanoemulsion, parenteral, Characterization, Application in Drug Delivery. Preparation,

Correspondence to Author

R.B.Desi reddy Nalanda Institute of Pharmaceutical sciences, Kantepudi, Sattenapalli, India Email: suresindhuri@gmail.com

Available online on www.ijprd.com 137

International Journal of Pharmaceutical Research & Development INTRODUCTION Nanoemulsions are non-toxic lipid droplets with a few hundred nanometers in diameter and made from surfactants approved for human consumption and common food substances that are 'Generally Recognized as Safe' (GRAS) by the FDA. These emulsions are easily produced in large quantities by mixing a water-immiscible oil phase into an aqueous phase with a high-stress, mechanical extrusion process that is available worldwide. This process yields a uniform population of droplet particles that are stable for years even at elevated temperatures. The various nanoformulations are : Nanoemulsions (NE) (submicron sized emulsions), nanosuspensions (submicron sized suspensions), nanospheres (drug nanoparticles in polymer matrix), nanotubes (sequence of nanoscale C60 atoms arranged in a long thin cylindrical structure), nanoshells (concentric sphere nanoparticles consisting of a dielectric core and a metal shell), nanocapsules (encapsulated drug nanoparticles), lipid nanoparticles (lipid monolayer enclosing a solid lipid core) and dendrimers (nanoscale threedimensional macromolecules of polymer). Nanoemulsions are submicron sized emulsions that are under extensive investigation as drug carriers for improving the delivery of therapeutic agents. The small size of the particles in these kinds of delivery systems (r < 100 nm) means that they have a number of potential benefits for certain applications: enhanced long-term stability, high optical clarity and increased bioavailability. Nanoemulsions are increasingly being utilized in food and pharmaceutical industries to encapsulate, protect, and deliver lipophilic bioactive components. Nanoemulsions are formed when the interfacial tension at the oil/water interface is brought to a very low level and the interfacial layer is kept highly flexible and fluid. These two conditions are usually met by a careful and precise choice of the components and of their respective proportions and by the use of a co-surfactant which brings flexibility to the oil/water interface. These Available online on www.ijprd.com

ISSN: 0974 9446

conditions lead to a thermodynamically optimised structure, which is stable as opposed to conventional emulsions and does not require high input of energy (i.e. through agitation) to be formed. Components of Nano Emulsion The three main components of Nanoemulsions are as follows: 1. Oil (Table 1) 2. Surfactant/Co-surfactant (Table 2) 3. Aqueous phase (Table 3) Table 1. List of oils used in nanoemulsions Name Captex 355 Captex 200 Captex 8000 Witepsol Myritol 318 Isopropyl myristate Chemical Name Glyceryl Tricaorylate/Caprate Propylene Dicaprylate/Dicaprate Glycol Glyceryl Tricaprylate (Tricaprylin) 90:10 % w/w c12 Glyceride tri: diesters c8/c10 triglycerides Myristic acid isopropyl ester

Table 2. List of Surfactant used in nanoemulsions S.No 1 2 3 4 5 6 7 8 9 10 11 12 Solubilizing agents, surfactants, emulsifying agents adsorption enhancers Capryol 90 Gelucire 44/14, 50/13 Cremophor RH 40 Imwitor 191, 308(1), 380, 742, 780 K, 928, 988 Labrafil M 1944 CS, M 2125 CS Lauroglycol 90 PEG MW > 4000 Plurol Oleique CC 497 Poloxamer 124 and 188 Softigen 701, 767 Tagat TO Tween 80

138

International Journal of Pharmaceutical Research & Development Table 3. List nanoemulsions S.No 1 2 3 4 5 of Co-Surfactant used in

ISSN: 0974 9446

Co Surfactant TranscutolP Glycerin,Ethyle ne glycol Propylene glycol Ethanol Propanol

Trials for the treatment of Herpes Labialis with Good Manufacturing Procedures (GMP) nanoemulsion to be conducted. A B

Microemulsions are used for controlled release and targeted delivery of different pharmaceutics agents. For instance, microemulsions were used to deliver oligonucleotides (small fragments of DNA) specifically to ovarian cancer cells. In contrast to microemulsions, Nanoemulsions consist in very fine oil-in-water dispersions, having droplets diameter smaller than 100 nm. Compared to microemulsions, they are in a metastable state and their structure depends on the history of the system. Nanoemulsions are very fragile systems. The nanoemulsions can find applications in skin care due to their good sensorial properties (rapid penetration, merging textures) and their biophysical properties (especially their hydrating power). This technology has a great advantage over the other dosage forms that the formulation can be delivered by various routes including oral , ocular and transdermal . Nanoemulsions have broad-spectrum antimicrobial activity against bacteria, enveloped viruses, fungi, protozoa and spores, due to their ability to lyse these organisms. In contrast, studies of nanoemulsions in animals have shown these compounds to be very well tolerated on the skin and mucous membranes. These attributes provide a broad therapeutic index when the nanoemulsions are used in humans as topical treatments for disorders including Herpes Labialis, cutaneous fungal infections, vaginitis, and respiratory infections. This material holds such unique promise and low risk that the FDA allowed Phase II Clinical Available online on www.ijprd.com

A:Microemulsions,B:Nanoemulsions Preparation methods of Nanoemulsions Several methods have been suggested for the preparation of nanoemulsion. The basic objectives of the nanoemulsion preparation is to achieve the droplet size range of 100-600 nm and another is to provide the stability condition. Formation of nanoemulsion system required a high amount of energy. This energy can be provided either by mechanical equipment or the chemical potential inherent within the component . Here some methods are discussed which are freely used for the nanoemulsion preparation. 1. Phase inversion method : In this method fine dispersion is obtained by chemical energy resulting of phase transitions taking place through emulsification path. The adequate phase transitions are produced by varying the composition at constant temperature or by varying the temperature at constant composition. Phase inversion temperature (PIT) method was introduced by Shinoda et al. based on the changes of solubility of polyoxyethylene type surfactant with temperature. This surfactant becomes lipophilic with increase in temperature due to dehydration of polymer chain. But at low temperature, the surfactant monolayer has a large positive spontaneous curvature forming oil-swollen 139

International Journal of Pharmaceutical Research & Development micellar solution phase .

ISSN: 0974 9446

2.Sonication method: Sonication method is another best way to prepare nanoemulsion. In this method the droplet size of conventional emulsion are reduced with the help of sonication mechanism. This method is not suitable for large batches, only small batches of nanoemulsion can be prepared by this method .

chamber,consisting of small channels called "microchannels." The product flows through the microchannels on to an impingement area resulting in very fine particles of submicron range. The two solutions (aqueous phase and oily phase) are combined together and processed in an inline homogenizer to yield a coarse emulsion. The coarse emulsion is into a microfluidizer where it is further processed to obtain a stable nanoemulsion. The coarse emulsion is passed through the interaction chamber of the microfluidizer repeatedly until desired particle size is obtained. The bulk emulsion is then filtered through a filter under nitrogen to remove large droplets resulting in a uniform nanoemulsion. Production With High-Amplitude Ultrasound High-amplitude ultrasound is a viable alternative to high-pressure homogenization. Intense shear forces necessary for the nanoemulsification are generated by ultrasonic cavitation, which produces violently and asymmetrically imploding vacuum bubbles and causes micro-jets that disperse and break up particles down to the nanometer scale. Known for many decades, this effect has been extensively studied and successfully used in smallscale production of pharmaceutical nanoemulsions and liposomes. However, prior to the introduction of Barbell Horn Ultrasonic Technology (BHUT), ultrasonic liquid processors could not effectively compete with high-pressure homogenizers in this market because they were not able to generate sufficiently high-amplitude (70 - 120 microns) ultrasonic vibrations on the industrial scale. Conventional high-power ultrasonic technology inherently forces all processes to run either at a small scale and high amplitude or a large scale and low amplitude, not allowing for the possibility of implementing high amplitudes on industrial scale. Thus, despite its potential, the ultrasonic method has mainly been restricted to laboratory investigations. Ultrasonic Technology:

Sonication method 3.Highpressurehomogenizer: This method is performed by applying a high pressure over the system having oil phase, aqueous phase and surfactant or co-surfactant. The pressure is applied with the help of a special equipment know as homogenizer. There are some problems which are associated with homogenizer such as poor productivity, component deterioration due to difficult mass production and generation of much heat. With this method only oil in water (o/w) liquid nanoemulsion of less than 20% oil phase can be prepared and cream nanoemulsion of high viscosity or hardness with a mean droplet diameter lower than 200 nm cannot be prepared . 4.Microfluidization: Microfluidization is a patented mixing technology, which makes use of a device called microfluidizer. This device uses a high-pressure positive displacement pump (500-20000 psi), which forces the product through the interaction Available online on www.ijprd.com

140

International Journal of Pharmaceutical Research & Development

ISSN: 0974 9446

ultrasonic devices utilize high-gain Barbell horns, which make it possible to reproduce any highamplitude laboratory-optimized process in a commercial production setting. These flow-through processors provide extremely high ultrasonic amplitudes and very uniform exposure patterns, ensuring that all treated liquid is exposed to tremendous ultrasonic cavitation-induced shear forces and that no part of the liquid is able to bypass the active cavitation zone in the reactor. Each ISMs industrial ultrasonic processor incorporates a calibrated amplitude sensor and is able to display and record ultrasonic amplitudes in microns peak to peak during operation. Characterization of nanoparticles Nanoemulsions are not thermodynamically stable, because of that their characteristics will depend on preparation method. Here some parameters are discussed which should be analyzed at the time of preparation of nanoemulsion . (i) Phase behaviour study: This study is a characterization and optimization of ingredients (surfactant, oil phase and aqueous phase). Generally the study is necessary in case of nanoemulsion formulation prepared by phase inversion temperature method and self emulsification method in order to determine the phase of nanoemulsion and dispersibility. Study is done by placing the different ingredients of nanoemulsion by varying the concentration in glass ampules and thoroughly homogenized at a certain temperature for a time until equilibrium. Anisotropic phase can be identified by polarized light. (ii) Particle Size Analysis: Formulated nanoemulsion should be analyzed for their hydrodynamic particle size and particle size distribution. Generally in case of nanoemulsion dynamic light scattering (DLS) method are used for the measurement of particles and further particle size distribution.

ISMs Ultrasonic Processor Industrial Sonomechanics, LLC, (ISM) has successfully overcome the aforementioned limitation by developing BHUT, which permits constructing industrial ultrasonic systems able to operate at extremely high ultrasonic amplitudes (up to about 200 microns). The output tip areas of the incorporated Barbell horns and the resulting productivity rates of the systems are more than 10 times higher than those of any conventional ultrasonic device operating at high amplitudes. ISM's Barbell horn-based high-amplitude industrial ultrasonic processors can be used for the commercial-scale production of the highest-quality nanoemulsions and liposomes, while offering many advantages over high-pressure homogenizers. These include significantly lower equipment costs, smaller number of wetted parts (easier cleaning, less wear and simpler servicing), no need to use a separate rotor-stator high-shear mixer to prepare a preliminary emulsion, as well as a much more practicable aseptic processing. In addition, it is much easier to create an ultrasonic system design that eliminates the need for multiple passes of the liquid through the system, which has not been possible with any high-pressure homogenizer. ISM offers directly scalable bench-top and industrial ultrasonic processors for the manufacture of high-quality pharmaceutical nanoemulsions and liposomes. Our patented Available online on www.ijprd.com

141

International Journal of Pharmaceutical Research & Development (iii) Surface charge measurement: Surface zeta potential of nanoemulsion droplets will be measured with the help of mini electrode to predict the surface properties of nanoemulsion. (iv) Transmission Electron Microscopy (TEM): This method is used to observe the morphology in the nanoemulsion. (v) Drug content: This method is used to determine the amount of drug contained in the formulation. Various methods (especially Western Blot method) are used in this order. (vi) Viscosity: Viscosity will be measured to ensure the better delivery of the formulation. Stability of Nanoemulsions : Stability of a dosage form refers to the chemical and physical integrity of the dosage unit and when appropriate, the ability of the dosage unit to maintain protection against microbiological contamination . Stability of drug product is one of the problems associated with the development of emulsions, microemulsions and nanoemulsions. Nanoemulsions have been known to enhance the physical as well as chemical stability of drugs . Stability studies are performed on nanoemulsions by storing them at refrigerator and room temperatures over a number of months. The viscosity, refractive index and droplet size are determined during this period of storage. Insignificant changes in these parameters indicate formulation stability. Accelerated stability studies can also performed. In this instance, nanoemulsion formulation are kept at accelerated temperatures and samples withdrawn at regular intervals and analyzed for drug content by stability indicating HPLC methods . The amount of drug degraded and remaining in nanoemulsion formulation is determined at each time interval. Morphology of Nanoemulsions Oral Delivery: Available online on www.ijprd.com

ISSN: 0974 9446

The morphology of nanoemulsions can be determined by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). SEM gives a three-dimensional image of the globules . The samples are examined at suitable accelerating voltage, usually 20 kV, at different magnifications. A good analysis of surface morphology of disperse phase in the formulation is obtained through SEM. Image analysis software, (e.g., Leica Im- aging systems, Cambridge, UK), may be employed to obtain an automatic analysis result of the shape and sur- face morphology . ApplicationsOfNanoemulsions: Parenteral Delivery: Parenteral administration (especially via the intravenous route) of drugs with limited solubility is a major problem in industry because of the extremely low amount of drug actually delivered to a targeted site. Nanoemulsion formulations have distinct advantages over macroemulsion systems when delivered parenterally because of the fine particle size. Nanoemulsion is cleared more slowly than the coarse particle emulsion and , therefore, have a longer residence time in the body. Both O/W and W/O nanoemulsions can be used for parenteral delivery. The literature contains the details of the many nanoemulsion systems, few of these can be used for the parenteral delivery because the toxicity of the surfactant and parenteral use. An alternative approach was taken by Von Corsewant and Thoren in which C3-C4 alcohols were replaced with parenterally acceptable co-surfactants, polyethylene glycol (400) / polyethylene glycol (660) 12hydroxystearate / ethanol, while maintaining a flexible surfactant film and spontaneous curvature near zero to obtain and almost balanced middle phase nanoemulsion. The middle phase structure was preferred in this application, because it has been able to incorporate large volumes of oil and water with a minimal concentration of surfactant.

142

International Journal of Pharmaceutical Research & Development Nanoemulsion formulations offer the several benefits over conventional oral formulation for oral administration including increased absorption, improved clinical potency and decreased drug toxicity. Therefore, Nanoemulsion have been reported to be ideal delivery of drugs such as steroids, hormones, diuretic and antibiotics. Pharmaceutical drugs of peptides and proteins are highly potent and specific in their physiological functions. However, most are difficult to administer orally. With on oral bioavailability in conventional (i.e. non-Nanoemulsion based) formulation of less than 10%, they are usually not therapeutically active by oral administration. Because of their low oral bioavailability, most protein drugs are only available as parenteral formulations. However, peptide drugs have an extremely short biological half life when administered parenterally, so require multiple dosing. A Nanoemulsion formulation of cyclosporine, named Neoral has been introduced to replace Sandimmune, a crude oil-in-water emulsion of cyclosporine formulation. Neoral is formulated with a finer dispersion, giving it a more rapid and predictable absorption and less inter and intra patient variability. Topical Delivery: Nanoemulsion formulation provides a rapid penetration of active ingredients through skin due to the large surface area of droplets. Even sometimes it is found that nanoemulsion penetrate easily through rough skin. This property of nanoemulsion minimizes the additional utilization of special penetration enhancer which is responsible for incompatibility of formulation. Topical administration of drugs can have advantages over other methods for several reasons, one of which is the avoidance of hepatic first pass metabolism of the drug and related toxicity effects. Another is the direct delivery and targetability of the drug to affected area of the skin or eyes. Both O/W and W/O Nanoemulsions have been evaluated in a hairless mouse model for the delivery of prostaglandin E1. The Nanoemulsions Available online on www.ijprd.com

ISSN: 0974 9446

were based on oleic acid or Gelucire 44/14 as the oil phase and were stabilized by a mixture of Labrasol (C8 and C10 polyglycolysed glycerides) and Plurol Oleique CC 497 as surfactant. Although enhanced delivery rates were observed in the case of the O/W Nanoemulsion, the authors concluded that the penetration rates were inadequate for practical use from either system. The use of lecithin/IPP/water Nanoemulsion for the transdermal transport of indomethacin and diclofenac has also been reported. Fourier transform infra red (FTIR) spectroscopy and differential scanning calorimetry (DSC) showed the IPP organogel had disrupted the lipid organisation in human stratum corneum after a 1 day incubation. The transdermal delivery of the hydrophilic drug diphenhydramine hydrochloride from a W/O Nanoemulsion into excised human skin have also been investigated. The formulation was based on combinations of Tween 80 and Span 20 with IPM. However two additional formulations were tested containing cholesterol and oleic acid, respectively. Cholesterol increased drug penetration whereas oleic acid had no measurable effect, but the authors clearly demonstrated that penetration characteristics can be modulated by compositional selection. Ocular and Pulmonary Delivery: For the treatment of eye diseases, drugs are essentially delivered topically. O/W Nanoemulsions have been investigated for ocular administration, to dissolve poorly soluble drugs, to increase absorption and to attain prolong release profile. The Nanoemulsions containing pilocarpine were formulated using lecithin, propylene glycol and PEG 200 as co-surfactant and IPM as the oil phase. The formulations were of low viscosity with a refractive index lending to ophthalmologic applications. The formation of a water-in-HFA propellent Nanoemulsion stabilized by fluorocarbon non-ionic surfactant and intended for pulmonary delivery has been described. Nanoemulsions in Biotechnology:

143

International Journal of Pharmaceutical Research & Development Many enzymatic and biocatalytic reactions are conducted in pure organic or aqua-organic media. Biphasic media are also used for these types of reactions. The use of pure apolar media causes the denaturation of biocatalysts. The use of waterproof media is relatively advantageous. Enzymes in low water content display and have Increased solubility in non-polar reactants. Possibility of shifting thermodynamic equilibria in favour of condensations. Improvement of thermal stability of the enzymes, enabling reactions to be carried out at higher temperatures. Many enzymes, including lipases, esterases, dehydrogenases and oxidases often function in the cells in microenvironments that are hydrophobic in nature. In biological systems many enzymes operate at the interface between hydrophobic and hydrophilic domains and these usually interfaces are stabilized by polar lipids and other natural amphiphiles. Enzymatic catalysis in Nanoemulsions has been used for a variety of reactions, such as synthesis of esters, peptides and sugar acetals transesterification; various hydrolysis reactions and steroid transformation. The most widely used class of enzymes in microemulsion-based reactions is of lipase. Nanoemulsions in vaccine development: Nanoemulsions can be used as a mucosal vaccine adjuvant. Nasal spray nanoemulsion vaccine fuses with antigen and is then sprayed into a nostril. Nanoemulsion droplets with antigen penetrate the nasal mucosa. Diagram of nanoemulsion droplet with antigen in its interface. Blue dots are antigen present in emulsion. Antigen delivery by nanoemulsion into nasal submucosa where fusion with dendritic cells delivers the antigen to the immune system. Dendritic cells can then transport the antigen to other parts of the body to trigger the desired immune response.

ISSN: 0974 9446

Antimicrobialnanoemulsions: Antimicrobial NEs are oil-in-water droplets that range from 200 to 600 nm. They are composed of oil and water and are stabilized by surfactants and alcohol. The NE has a broad-spectrum activity against bacteria (e.g.E.coil, salmonella, S. aureus), enveloped viruses (e.g. HIV, Herpes simplex), fungi (e.g. Candida, Dermatophytes), and spores (e.g. anthrax). The NE particles are thermodynamically driven to fuse with lipid-containing organisms. This fusion is enhanced by the electrostatic attraction between the cationic charge of the emulsion and the anionic charge on the pathogen. When enough nanoparticles fuse with the pathogens, they release part of the energy trapped within the emulsion. Both the active ingredient and the energy released destabilize the pathogen lipid membrane, resulting in cell lysis and death. In the case of spores, additional germination enhancers are incorporated into the emulsion. Once initiation of germination takes place, the germinating spores become susceptible to the antimicrobial action of the NE. A unique aspect of the NEs is their selective toxicity to microbes at concentrations that are nonirritating to skin or mucous membrane. The safety margin of the NE is due to the low level of detergent in each droplet, yet when acting in concert, these droplets have sufficient energy and surfactant to destabilize the targeted microbes without damaging healthy cells. As a result, the NE can achieve a level of topical antimicrobial activity that has only been previously achieved by systemic antibiotics. 144

Available online on www.ijprd.com

International Journal of Pharmaceutical Research & Development

ISSN: 0974 9446

Nanoemulsions as a mucosal vaccines .: Nanoemulsions are being used to deliver either recombinant proteins or inactivated organisms to a mucosal surface to produce an immune response. The first applications, an influenza vaccine and an HIV vaccine, can proceed to clinical trials. The Nanoemulsions causes proteins applied to the mucosal surface to be adjuvanted and it facilitates uptake by antigen-presenting cells. This results in a significant systemic and mucosal immune response that involves the production of specific IgG and IgA antibody as well as cellular immunity. Initial work in influenza has demonstrated that animals can be protected against influenza after just a single mucosal exposure to the virus mixed with the emulsion. Research has also demonstrated that animals exposed to recombinant gp120 in NE on their nasal mucosa develop significant responses to HIV, thus providing a basis to examine the use of this material as an HIV vaccine. Additional research is ongoing to complete the proof of concept in animal trials for other vaccines including Hepatitis B and anthrax A novel technique for vaccinating against a variety of infectious diseases-using an oilbased emulsion placed in the nose, rather than needles-has proved able to produce a strong immune response against smallpox and HIV in two new studies. Nanoemulsion as non-toxic disinfectant cleaner : A breakthrough nontoxic disinfectant cleaner for use in commercial markets that include healthcare, hospitality, travel, food processing, and military applications has been developed by EnviroSystems, Inc. that kills tuberculosis and a wide spectrum of viruses, bacteria and fungi in 5-10 min without any of the hazards posed by other categories of disinfectants. The product needs no warning labels. It does not irritate eyes and can be absorbed through the skin, inhaled, or swallowed without harmful effects. The disinfectant formulation is made up of nanospheres of oil droplets #106 mm that are suspended in water to create a NE requiring only miniscule amounts of the active ingredient, PCMX (parachlorometaxylenol). The Available online on www.ijprd.com

nanospheres carry surface charges that efficiently penetrate the surface charges on microorganisms' membranes much like breaking through an electric fence. Rather than "drowning" cells, the formulation allows PCMX to target and penetrate cell walls. As a result, PCMX is effective at concentration levels 1-2 orders of magnitude lower than those of other disinfectants; hence, there are no toxic effects on people, animals, or the environment. Other microbial disinfectants require large doses of their respective active ingredients to surround pathogen cell walls, which cause them to disintegrate, fundamentally "drowning" them in the disinfectant solution. The formulation is a broad-spectrum disinfectant cleaner that can be applied to any hard surface, including equipment, counters, walls, fixtures, and floors. One product can now take the place of many reducing product inventories and saving valuable storage space. Chemical disposal costs can be eliminated, and disinfection and cleaning costs can be reduced. It is marketed as a EcoTru&#8482; (EnviroSystems, Inc.) Nanoemulsions in cell culture technology Cell cultures are used for in vitro assays or to produce biological compounds, such as antibodies or recombinant proteins. To optimize cell growth, the culture medium can be supplemented with a number of defined molecules or with blood serum. Up to now, it has been very difficult to supplement the media with oil-soluble substances that are available to the cells, and only small amounts of these lipophilic compounds could be absorbed by the cells. NEs are a new method for the delivery of oil-soluble substances to mammalian cell cultures. The delivery system is based on a NE, which is stabilized by phospholipids. These NEs are transparent and can be passed through 0.1 mm filters for sterilization. NE droplets are easily taken up by the cells. The encapsulated oil-soluble substances therefore have a high bioavailability to cells in culture. The advantages of using NEs in cell culture technology are better uptake of oil-soluble supplements in cell cultures; improve growth and vitality of cultured cells, and allowance of toxicity studies of oil-soluble drugs in cell cultures. 145

International Journal of Pharmaceutical Research & Development

ISSN: 0974 9446

Nanoemulsion in cancer therapy and in targeted drug delivery : It is also reported that nanoemulsion may be used for the target delivery of active ingredient especially in cancer therapy.The effects of the formulation and particle composition of gadolinium (Gd)-containing lipid NE (Gd-nanoLE) on the biodistribution of Gd after its intravenous (IV) injection in D1-179 melanoma-bearing hamsters were evaluated for its application in cancer neutron-capture therapy. Biodistribution data revealed that Brij 700 and HCO-60 prolonged the retention of Gd in the blood and enhanced its accumulation in tumors. Among the core components employed, soybean oil yielded the highest Gd concentration in the blood and tumor, and the lowest in the liver and spleen. When each Gd-nanoLE was IV injected once or twice at a 24-h interval, the Gd concentration in the tumor correlated well with the total dose of Gd, and it reached a maximum of a 189 mg/g wet tumor. This maximum Gd level was greater than the limit required for significantly suppressing tumor growth in neutron-capture therapy. In order to achieve penetration of Paclitaxel (PCL) into deeper skin layers while minimizing the systemic escape, a NE (NE) was formulated and its in vivo pharmacokinetic performance was evaluated. Further, the same formulation was explored for peroral bioavailability enhancement of PCL. Upon dermal application, the drug was predominantly localized in deeper skin layers, with minimal systemic escape. This has amounted to an absolute bioavailability of 70.62%. Inhibition of Pglycoprotein efflux by D-tocopheryl polyethyleneglycol 1000 succinate and labrasol would have contributed to the enhanced peroral bioavailability of PCL. This investigation provides direct evidence on the localization of highmolecular-weight, lipophilic drug, PCL, in dermis. Further, the NE formulation has enhanced the peroral bioavailability significantly to more than 70%. The developed NE formulation was safe and effective for both. Available online on www.ijprd.com

Camptothecin is a topoisomerase I inhibitor that acts against a broad spectrum of cancers. However, its clinical application is limited by its insolubility, instability, and toxicity. The aim of the present study was to develop acoustically active NEs for camptothecin encapsulation to circumvent these delivery problems. The NEs were prepared using liquid perfluorocarbons and coconut oil as the cores of the inner phase. These NEs were stabilized by phospholipids and/or Pluronic F68 (PF68). The NEs were prepared at high drug loading of approximately 100% with a mean droplet diameter of 220-420 nm. Camptothecin in these systems showed retarded drug release. Camptothecin in NEs with a lower oil concentration exhibited cytotoxicity against melanomas and ovarian cancer cells. Confocal laser scanning microscopy confirmed NE uptake into cells. Using a 1 MHz ultrasound, an increased release of camptothecin from the system with lower oil concentration could be established, illustrating a drug-targeting effect. The advantages of formulating various lipophilic anti-cancer drugs in submicron O/W emulsion are obvious. The oil phase of the emulsion systems can act as a solubilizer for the lipophilic compound. Therefore, solubility of lipophilic drugs can be significantly enhanced in an emulsion system, leading to smaller administration volumes compared to an aqueous solution. In addition, because lipophilic drugs are incorporated within the innermost oil phase, they are sequestered from direct contact with body fluids and tissues. Lipid emulsions can minimize the pain associated with intravenously administered drugs by exposing the tissues to lower concentrations of the drug or by avoiding a tissue-irritating vehicle. This has been demonstrated with propofol, diazepam, methohexital, clarithromycin, and etomidate. Another study reported the formulation of filter sterilizable emulsion formulation of paclitaxel using -tocopherol as the oil phase and tocopherylpolyethyleneglycol-1000 succinate (TGPS) and poloxamer 407 as emulsifiers. The formulation exhibited better efficacy and was more 146

International Journal of Pharmaceutical Research & Development tolerable when studied in B16 melanoma tumor model in mice. Emulsion formulations also show promise in cancer chemotherapy as vehicles for prolonging the drug release after intramuscular and intratumoral injection (W/O systems) and as a means of enhancing the transport of anti-cancer drugs via the lymphatic system. The perfluorochemical Nanoemulsions (PFCE) have opened interesting opportunities in cancer therapy. It is suggested that fluorocarbon emulsions might find a role in photodynamic therapy, both as carriers for sensitizing dyes and to maintain tissue oxygenation in hypoxic regions of solid tumors. The high solubility of oxygen in fluorocarbon emulsions maintains solution oxygen tension, optimizing photo-oxidative damage. The hydrophobic anti-cancer drugs can be delivered to the tumor mass by dissolving them in a hydrophobic core of the emulsion. Furthermore, PFCE can be used as an adjuvant to radiation therapy and/or chemotherapy in the treatment of solid tumors. The preclinical studies have shown very positive effects with single dose and fractionated radiation in several rodent solid tumor models. Many widely used anticancer drugs, including anti-tumor alkylating agents and doxorubicin, have shown improved response by PFCE coadministration. Also, local application of toxic doses of PFCEs resulted in the necrosis of cancer cells. This is especially promising in the treatment of cancers of the head and neck regions that are currently difficult to treat. Nanoemulsion in the treatment of various other disease conditions: Pharmos' (US-based company) has developed the nanoemulsion topical diclofenac cream as a potential treatment for osteoarthritis (OA) pain. Topical diclofenac is also being considered as treatment for soft tissue injuries, sprains, and strains. It is estimated that 20% of OA patients are Available online on www.ijprd.com

ISSN: 0974 9446

not receiving treatment, mainly due to gastrointestinal side effects of oral NSAIDs and cardiovascular risk of COX-2 inhibitors. A topical NSAID offering adequate pain relief targeted to the site of injury with an improved safety profile could become a treatment alternative for these patients. One of the unique characteristics of the Nanoemulsion technology is the relatively high percentage of total particle volume occupied by the internal hydrophobic oil core of the droplets. This provides high solubilization capacity for lipophilic compounds compared to other lipoidal vehicles such as liposomes. Viscosity-imparting agents are used for nanoemulsion thickening to produce creams with the desired semisolid consistency for application to the skin. The skin penetrative properties of the solvent-free NE delivery technology and its low irritancy make this novel topical nanovehicle a promising candidate for effective transcutaneous delivery of lipophilic drugs. A topical application of the nanotechnology has already demonstrated excellent targeted delivery of lipophilic drugs to muscle and joints in animal models. Preclinical data using a paw edema animal model showed enhanced anti-inflammatory activity with NSAIDs encapsulated in nanoemulsion creams compared to commercial formulations. Pharmacokinetic studies using nanoemulsion topical creams containing radiolabeled diclofenac and ketoprofen were performed to assess drug penetration through skin and to determine local tissue (muscle and joint) and plasma levels of drugs following topical administration. Compared to oral administration, diclofenac and ketoprofen administered via nanoemulsion topical creams demonstrated 4- to 6-fold lower drug concentration in plasma, 60-to 80-fold more drug in muscle tissue, and about 9-fold more drug in jointsThe NE technology consists of spheric oily droplets (in the range of 100-200 nm) uniformly dispersed in an aqueous medium. The emulsion droplet size reduction is essential to generate drug formulations with high stability. The NE technology has been successfully applied in the formulation of ophthalmic preparations showing improved drug delivery and reduced ocular irritation in humans in 147

International Journal of Pharmaceutical Research & Development Phase I/II clinical studies.

ISSN: 0974 9446

Primaquine (PQ) is one of the most widely used antimalarial and is the only available drug till date to combat relapsing form of malaria especially in case of Plasmodium vivax and Plasmodium ovale. Primaquine acts specifically on the pre-erythrocytic schizonts that are concentrated predominantly in the liver and causes relapse after multiplication. However, application of PQ in higher doses is limited by severe tissue toxicity including hematological and GI-related side effects that are needed to be minimized. Lipid NE has been widely explored for parenteral delivery of drugs. Primaquine when incorporated into oral lipid NE having a particle size in the range of 10-200 nm showed effective antimalarial activity against Plasmodium bergheii infection in Swiss albino mice at a 25% lower dose level as compared to conventional oral dose. Lipid NE of primaquine exhibited improved oral bioavailability and was taken up preferentially by the liver with drug concentration higher at least by 45% as compared to the plain drug. Nanoemulsion formulations for improved oral delivery of poorly soluble drugs : NE formulations were developed to enhance oral bioavailability of hydrophobic drugs. Paclitaxel was selected as a model hydrophobic drug. The oil-inwater (o/w) Nanoemulsions were made with pine nut oil as the internal oil phase, egg lecithin as the primary emulsifier, and water as the external phase. Stearylamine and deoxycholic acid were used to impart positive and negative charge to the emulsions,respectively. Coenzyme Q10 (CoQ10), also known as ubiquinone, is used for energy production within cells and acts as an anti-oxidant. Since CoQ10 is highly lipophilic, the topical and oral bioavailability is very low. Several attempts have been made to improve absorption. Latest technical developments reveal that encapsulation of CoQ10 in NEs results Available online on www.ijprd.com

in a significantly enhanced bioavailability. The application of CoQ10 has been further improved by the development of novel CoQ10 double NEs containing tocopherol and CoQ10 in individual nanodroplets. In addition, the CoQ10 concentration in these NEs could be increased by the development of a supersaturated CoQ10 NE. Nanoemulsions as a vehicle for transdermal delivery: From in vitro and in vivo data, it was concluded that the developed NEs have great potential for transdermal drug delivery of aceclofenac. The NEs of the system containing ketoprofen evidenced a high degree of stability. Ketoprofen-loaded NEs enhanced the in vitro permeation rate through mouse skins as compared to the control. The study was developed to evaluate the potential of NEs for increasing the solubility and the in vitro transdermal delivery of carvedilol. The prepared NEs were subjected to physical stability tests. Transdermal permeation of carvedilol through rat abdominal skin was determined with the Keshary-Chien diffusion cell. Significant increase (P < 0.05) in the steady state flux (Jss) and permeability coefficient (Kp) was observed in NE formulations as compared to control or drugloaded neat components. The irritation studies suggested that the optimized NE was a non-irritant transdermal delivery system. Celecoxib, a selective cyclo-oxygenase-2 inhibitor, has been recommended orally for the treatment of arthritis and osteoarthritis. Long-term oral administration of celecoxib produces serious gastrointestinal side effects. Skin permeation mechanism of celecoxib from NE was evaluated by FTIR spectral analysis, DSC thermogram, activation energy measurement, and histopathological examination. The optimized NE was subjected to pharmacokinetic (bioavailability) studies on Wistar male rats. Photomicrograph of a skin sample showed the disruption of lipid bilayers as distinct voids and empty spaces were visible in the 148

International Journal of Pharmaceutical Research & Development epidermal region. Results of skin permeation mechanism and pharmacokinetic studies indicated that the NEs can be successfully used as potential vehicles for enhancement of skin permeation and bioavailability of poorly soluble drugs. Solid self-nanoemulsifying delivery systems as a platform technology for formulation of poorly soluble drugs : New drug discovery programs produce molecules with poor physico-chemical properties, making delivery of these molecules at the right proportion into the body, a big challenge to the formulationscientist. The various options available to overcome the hurdle include solvent precipitation, micronisation or nanonization using high-pressure homogenization or jet milling, salt formation, use of microspheres, solid dispersions, cogrinding, complexation, and many others. Selfnanoemulsifying systems (SNES) form one of the most popular and commercially viable approaches for delivery of poorly soluble drugs exhibiting dissolution rate limited absorption, especially those belonging to the Biopharmaceutics Classification System II/IV. SNES are essentially an isotropic blend of oils, surfactants, and/or cosolvents that emulsify spontaneously to produce oil in water NE when introduced into aqueous phase under gentle agitation. Conventional SNES consist of liquid forms filled in hard or soft gelatin capsules, which are least preferred due to leaching and leakage phenomenon, interaction with capsule shell components, handling difficulties, machinability, and stability problems. Solidification of these liquid systems to yield solid self-nanoemulsifying systems (SSNES) offer a possible solution to the mentioned complications, and that is why these systems have attracted wide attention. Use of nanoemulsions in cosmetics: Evaluation of Nanoemulsions: NEs have recently become increasingly important as potential vehicles for the controlled delivery of cosmetics and for the optimized dispersion of active ingredients in particular skin layers. Due to Available online on www.ijprd.com

ISSN: 0974 9446

their lipophilic interior, NEs are more suitable for the transport of lipophilic compounds than liposomes. Similar to liposomes, they support the skin penetration of active ingredients and thus increase their concentration in the skin. Another advantage is the small-sized droplet with its high surface area allowing effective transport of the active to the skin. Furthermore, NEs gain increasing interest due to their own bioactive effects. This may reduce the trans-epidermal water loss (TEWL), indicating that the barrier function of the skin is strengthened. NEs are acceptable in cosmetics because there are no inherent creaming, sedimentation, flocculation, or coalescence that are observed with macroemulsions. The incorporation of potentially irritating surfactants can often be avoided by using high-energy equipment during manufacturing. NanoGel technology provides a simple process and system to create submicron emulsions from an easy-to-use, oil-in-water concentrate. The formula is particularly suited to minimizing transepidermal water loss, enhanced skin production, and penetration of active ingredient. These characteristics suggest that it would be particularly useful for sun care products as well as moisturizing and anti-aging creams-particular areas where nanotechnology is already being incorporated into a host of products currently on the market. Likewise, it is also highlighted that it helps to give skin care formulations a good skin feel, an increasingly important characteristic for formulators. NEs have attracted considerable attention in recent years for application in personal care products as potential vehicles for the controlled delivery of cosmetics and the optimized dispersion of active ingredients in particular skin layers.

In Vitro Skin Permeation Studies (for transdermal drug delivery system):

149

International Journal of Pharmaceutical Research & Development Franz diffusion cell is used to obtain the drug release profile of the nanoemulsion formulation in the case of formulations for transdermal application. The extent or depth of skin penetration by the released content can be visualized by confocal scanning laser microscopy. In vitro drug release can be determined by dispersing an amount of the preparation in the donor compartment of a Franz cell having a membrane as barrier and monitoring the appearance of the encapsulated drug in the reception medium, usually PBS (pH 7.4) and stirring on a magnetic stirrer at 100 rpm at 37C 1C. Samples (1 ml) of the dispersion are withdrawn from the medium and replaced with an equivalent amount of the medium at definite intervals. The withdrawn sample is then filtered using a 0.22 - 50 m filter (e.g., Millipore, USA) and the drug released then analyzed using UV-visible spectroscopy at wavelength of peak absorption of the drug . An alternative and popular method of ex-vivo release study is performed using diffusion cell. The skin is cut from the ear or abdomen and underlying cartilage and fats care- fully removed. Appropriate size of skin is cut and placed on the diffusion cell which had earlier been filled with receptor solution. Samples of the vesicular preparation are then applied on the dorsal surface of the skin and the instrument started. At intervals, up to 24 h, samples are withdrawn from the receptor medium and replaced with equal amounts of the medium and the withdrawn samples analyzed for the drug permeated using HPLC or UV spectroscopy. Semipermeable membrane such as regenerated cellulose could be used in place of skin for in vitro release studies. The flux J, of the drug across the skin or membrane is calculated from the formula: J = D dc/dx (2) where D is the diffusion coefficient and is a function of the size, shape and flexibility of the diffusing molecule as well as the membrane resistance, c is the concentration of the diffusing species, x is the spatial coordinate . Available online on www.ijprd.com

ISSN: 0974 9446

In vivo release study otherwise referred to as dermatopharmacokinetics, is carried out by applying or admin- istering the preparation to whole live animal. Blood samples are then withdrawn at intervals, centrifuged and the plasma analyzed for the drug content using HPLC. Results obtained from in vitro and in vivo studies are extrapolated to reflect bioavailability of the drug formulation. ADVANTAGES OF NANOEMULSIONS OVER OTHER DOSAGE FORMS Increase the rate of absorption. Eliminates variability in absorption. Helps solublize lipophilic drug. Provides aqueous dosage form for water insoluble drugs. Increases bioavailability. Various routes like tropical, oral and intravenous can be used to deliver the product. Rapid and efficient penetration of the drug moiety. Helpful in taste masking. Provides protection from hydrolysis and oxidation as drug in oil phase in O/W Nanoemulsion is not exposed to attack by water and air. Liquid dosage form increases patient compliance. Less amount of energy requirement. Nanoemulsion has a transparent and fluidy property which improves the formulation patient compliance and safe for administration due to the absence of any thickening agent and colloidal particles. Nanoemulsions are thermodynamically stable system and the stability allows selfemulsification of the system . Nanoemulsion formulation required low amount of surfactant compared to microemulsion. For example about 20- 25 % surfactant is required for the preparation of microemulsion but 5-10 % surfactant is sufficient in case of nanoemulsion.

150

International Journal of Pharmaceutical Research & Development Limitations of nanoemulsions: Even though nanoemulsions provide great advantages as a delivery system, but sometimes the re duced size of droplets are responsible for the limited use of nanoemulsion formulation. Some limitations of nanoemulsions are as follows : The manufacturing process of nanoemulsion formulation is expensive, because size reduction of droplets is very difficult as it required a special kind of instruments and process methods. For example, homogenizer arrangement, microfluidization &ultrasonification require high financial support. Nanoemulsion stability creates a big problem during the storage of formulation for the longer time period. Ostwald ripening is the main factor associated with unacceptability of nanoemulsion formulations. This is due to the high rate of curvature of small droplet show greater solubility as compared to large drop with a low radius of curvature. Less availability of surfactant and co-surfactant required for the manufacturing of nanoemulsion is another factor which marks as a limitation to nanoemulsion manufacturing. Limited solubilizing capacity for high-melting substances. Commercial nanoemulsion formulations. Drug/Bioactive Palmitate alprostadil Ibuprofen Category

ISSN: 0974 9446

CONCLUSION: Nanoemulsions have now-a-days become an answer for the questions regarding targeted delivery . Because of their submicron size , they can be easily targeted . Moreover, the possibility of surface functionalization with a targeting moiety has opened new avenues for targeted delivery of drugs, genes, photosensitizers& other molecules of tumer area. So,nanoemulsions can set a better mark for targeted drug delivery system . REFERENCES 1. Lachman L, Lieberman HA, Kanig JL. The Theory and Practice of Industrial Pharmacy; 3 rd ed. p. 510-1. 2. P.shah e tal , Nanoemulsion: A pharmaceutical review , article info, Volume : 1 , Issue : 1 , Page : 24-32, Feb:2010 3. Nitin Sharma, Mayank Bansal, Sharad Visht, PK Sharma, GT Kulkarni, Chron Young Sci;1:2-6, oct 2010 4. Charles Lovelyn, Anthony A. Attama, Current State of Nanoemulsions in Drug Delivery, Journal of Biomaterials and Nanobiotechnology, Nov2011, 2, 626-639. 5. Koewn JH, Chi SC and Park ES. Transdermal delivery of diclofenac using nanoemulsions. Arch. Pharm. Res. 2004) 27: 351-356. 6. Lawrence MJ and Rees GD. Nanoemulsionbased media as novel drug delivery systems.Adv. Drug Del. Rev. (2000) 45: 89-121. 7. T. P. U. Ravi and T. Padma, Nanoemulsions for Drug Delivery through Different Routes, Research in Bio-technology, Vol. 2, No. 3, 2011, pp. 1-13.

Dexamethasone Indomethacin, (IND) Propofol Flurbiprofenaxtil Vitamins A, D, E and K

Vasodilator, platelet inhibitor non-steroidal antiinflammatory drug (NSAID) Steroid NSAID Anaesthetic NSAID Parenteral nutrition

Available online on www.ijprd.com 151

International Journal of Pharmaceutical Research & Development 8. N. Anton and T. Vandamme, The Universality of Low- Energy Nano-Emulsification, International Journal Phar- maceutics, Vol. 377, No. 1-2, 2009, pp. 142-147. 9. T. G. Mason, S. M. Graves, J. N. Wilking and M. Y. Lin, Extreme Emulsification: Formation and Structure of Nanoemulsions, Journal of Physics and Condensed Matter, Vol. 9, No. 1, 2006, pp. 193-199.

ISSN: 0974 9446

10. C. Quin and D. J. Mc Clement, Formation of Nano- emulsions Stabilized by Model Food Grade Emulsifiers Using High Pressure Homogenization: Factors Effecting Particle Size, Food Hydrocolloids, Vol. 25, No. 5, 2011, pp. 1000-1008. 11. I. Sole, C. M. Pey, A. Maestro, C. Gonzalez, M. Porras, C. Solans and J. M. Gutierrez, Nanoemulsions Prepared by Phase Inversion Composition Method: Preparation Vari-ables and Scale up, Journal of Colloid and Interface Science, Vol. 344, No. 2, 2010, pp. 417-423.

*****

Available online on www.ijprd.com 152