Nanomaterials and nano- and microencapsulation represent exciting opportunities

for pharmaceutical and nutraceutical scientists. The nano- and microencapsule

consists of two components: the core or filling material and the shell or coat
material. Polymeric nano- and microparticles can be fabricated from biodegradable
or non-biodegradable polymers. There are different techniques for the
encapsulation of core materials, and all these techniques are generally categorized
into three types: (i) chemical methods; (ii) physico-chemical methods; and (iii)
physico-mechanical methods. These methods are widely used for nano- and
microencapsulation of various pharmaceutical and nutraceutical components. This
chapter highlights some of the main delivery routes where nano- and
microencapsulation systems have been studied as potential drug or nutrient
carriers, and indicates related advantages as well as their disadvantages.
Nutraceutical products are food ingredients, and may be nutritional supplements,
isolated nutrients, herbal products, specific diets, or genetically engineered foods,
and processed foods, such as cereals, beverages, and
soups------------------------------------------------------------------------

Currently, bioactive compounds are required in the design and production of functional foods,
with the aim of improving the health status of consumers all around the world. Various
epidemiological and clinical studies have demonstrated the salutary role of eicosapentaenoic acid
(EPA, 22:6 n3) and docosahexaenoic acid (DHA, 22:5 n3) in preventing diseases and reducing
mortality from cardiovascular diseases. The unsaturated nature of bioactive lipids leads to
susceptibility to oxidation under environmental conditions. Oxidative deterioration of omega-3
fatty acids can cause the reduction in their nutritional quality and sensory properties.
Encapsulation of these fatty acids could create a barrier against reaction with harmful
environmental factors. Currently, fortification of foods containing bioactive omega-3 fatty acids
has found great application in the food industries of different countries. Previous studies have
suggested that nano-encapsulation has significant effects on the stability of physical and
chemical properties of bioactive compounds. Considering the functional role of omega-3 fatty
acids, this study has provided a literature review on applications of nanoliposomal delivery
systems for encapsulation of these bioactive compounds.
Keywords: Omega-3 Fatty Acids, Stabilization, Nanoliposome, Bioactive
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1. Introduction
Polyunsaturated fatty acids (PUFA) with multiple double bonds exist in the form of -linolenic
acid, eicosapentaenoic acid, and docosahexaenoic acid. In recent decades, clinical studies have
verified that increasing the content of fatty acids in diet could be effective on the lipoproteinlipid index. Since 1970, various studies have been conducted to assess the effects of omega-3
fatty acids derivatives on preventing diseases, especially cardiovascular diseases, so that
manufacturers have been encouraged to formulate dietary supplements from these compounds
using different methods, including encapsulation. Various studies conducted in different
communities suggest that daily intake of 1 g fish oil (containing about 840 mg DHA+EPA) could

significantly decrease rates of sudden death in patients with cardiovascular diseases. Therefore,
using this supplement is recommended for use alongside anticoagulant drugs in such cases (13).
The Food and Drug Administration (FDA) of America does not recommend acquiring these fatty
acids through only fish consumption, because of seafood containing compounds such as
polychlorinated biphenyls (PCBs), mercury, and other contaminants. In addition, the FDA has
approved the use of Lovasa (Omacor), which contains EPA and DHA, as a medicinal supplement
of omega-3 fatty acids because of its efficacy in preventing cardiovascular diseases in patients
with high triglyceride (46). The unsaturated nature of omega-3 fatty acids results in their
susceptibility to oxidation under environmental conditions. Hydroperoxides are products
generated from the primary oxidation of EPA and DHA, followed by degradation into secondary
oxidation compounds, including volatile aldehyde compounds. By encapsulation of these fatty
acids, a barrier can be created to prevent reaction of these compounds with oxidative factors.
When designing capsules, some factors should be considered when recognizing the target cell
and stability against pH changes of digestive system through adding chemical compounds or
other protective groups. As delivery systems, current liposomes include a wide range of bioactive
compounds with different applications in food, pharmaceutical, and agricultural industries (79).
Liposomes are made of natural lipids; therefore they are non-toxic and do not stimulate the
immune system, in addition to being biodegradable. Other benefits of liposomes include
solubility, capacity for controlled, directed and purposeful release of hydrophobic and
hydrophilic compounds, and ease of permeation and transmission through membranes.
Liposomes applications include acting as suitable carriers of different bioactive molecules in all
the nanomedicine platforms and liposomes have generated a great deal of recent interest. There
are several formulations approved by the Food and Drug Administration (FDA) for disease
treatment (1013). Nanoliposomes can be defined as lipid bilayers assemblies encompassing the
aqueous compartment within the nano-sized range. Nanoliposomes have been tried for various
applications as drug or gene delivery for treatment of diseases. Various methods typically used to
fabricate liposomes include the thin layer film hydration method, the ethanol injection method,
and the detergent removal method, as well as the heating method, the reverse phase evaporation
method, and homogenization (13, 14). Recently conducted studies suggest that the methods used
to provide liposomes have had an important effect on the stabilization of the physical and
chemical properties of bioactive compounds. In addition, by reducing the particle size of this
colloidal system, the specific surface area could lead to more favorable compliance and
bioavailability properties of bioactive compounds (3, 15, 16). Selecting the method of liposome
preparation depends on the following parameters: the physicochemical characteristics of
liposomal lipids and those of entrapped material; the kind of dispersion medium for lipid
vesicles; the concentration and potential toxicity of entrapped material; vesicle size,
polydispersity, and shelf-life intended for each application; batch-to-batch reproducibility; safe
and efficient liposome production on a large scale (13, 17, 18).
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2. Effects of Omega-3 Fatty Acids on Health

The main omega-3 fatty acids, in terms of nutrition, are -linolenic acid, eicosapentaenoic acid,
and docosahexaenoic acid. Technically, EPA is not essential fatty acid, since the body can
convert essential -linolenic acid into EPA. However, the physiologic conditions of the human

body cannot create DHA and EPA sufficiently to supply them as required for healthly living and
these compounds should be provided through dietary sources. Omega-3 fatty acids are obtained
from seafood, oils (algae oil, fish oil and flax seed oil), or dietary supplements. EPA is found in
fish and fish oil. DHA is of special importance for the body and is found mainly in fish and
commercial algae oil. ALA is mainly found in seeds, herbal oils, and green leafy vegetables. In
the body, ALA is first converted to EPA and then to DHA (19). Fish oil is often accessible in the
form of soft-gel capsule with different commercial names. Vitamins, antioxidants, and different
flavors are added to these products. Lovaza is a type of fish oil supplement with a high
concentration of omega-3 (465mg EPA and 375mg DHA), which was approved by the FDA in
2005 as a supplementary treatment for patients with hypertriglycemia (1, 20). DHA is the main
ingredient in the phospholipid membranes of the brain and retina cells. In addition, DHA
decreases serum triglyceride by inhibiting lipogenesis and simulates oxidation and lipid
metabolism. EPA and DHA, in omega-3, are significantly effective for reducing plasma TG. By
simulating peroxisome proliferators-activated receptor, DHA and EPA increase lipid metabolism
and decrease serum level of triglyceride. In addition, EPA is capable of reducing total cholesterol
(4, 8). The protective effects of unsaturated omega-3 fatty acids, such as EPA, in coronary heart
diseases are applied through different methods, some of which involve over activation of the
sympathetic nervous system, enhancement of vasodilation through reduction of endothelial
inflammatory response by inhibition of monocyte adhesion to the wall of arteries, and weakening
the production of inflammatory mediators and anti-thrombotic action. Overall, the beneficial
effects of omega-3 fatty acids on coronary arteries significantly reduces sudden death in patients
with cardiovascular diseases and specialists often recommend using supplements along with
antiplatelet drugs, beta blockers, and angiotensin converting enzyme inhibitors (20). In 2000, the
FDA approved the use of fish oil supplements for reducing the risk of cardiovascular diseases
and recommended patients with coronary artery disease to take 1g of supplements daily (21).
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3. Stabilization of Omega-3 Fatty Acids, Characterization

Method and Control of Autoxidation
The major factors in stabilization of lipids include the type of ingredients of fatty acids,
temperature, oxygen, humidity, heavy metals, light, and packaging. Various lipid oxidative
reactions also can occur, including non-enzymatic autoxidation, light oxidation and enzymatic
oxidation. In this regard, autoxidation is the main process and non-enzymatic autoxidation of
EPA and DHA occurs in ambient temperature, due to the high unsaturated nature of the
compounds, which causes further changes during processing. These fatty acids oxidize rapidly
with exposure to light, peroxidants, or high temperature, which causes primary oxidative
products to be formed. Sensory changes resulting from degradation of hydroperoxides into
secondary products, including aldehydes, ketones, acids, and alcohols, are proportional to the
carbon chain length and degree of unsaturation for the fatty acids. Some of these compounds
have olfactory thresholds. Therefore, they affect sensory quality at very low concentrations and
result in unpleasant taste and odor (22). Lipid oxidation is considered the main parameter
affecting the retention of different fish oils and products containing it. Autoxidation of lipids
results in formation of fatty acid based hydroperoxides and their degradation into secondary

oxidation products. Many of these compounds are the cause of unpleasant tastes and further
degradation. Some methods for preventing oxidation of unsaturated lipids include reduction of
oxygen through proper packaging, storage at low temperatures, and addition of antioxidants (23).
The use of nano-delivery systems, such as liposome, can stabilize and protect bioactive
compounds including omega-3 fatty acids against changes in environmental conditions and
chemical changes, along with improving their accessibility. Researchers believe that omega-3
fatty acids could be the best option of functional foods, if the oxidative deterioration difficulties
are overcome (16).
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4. Encapsulation Techniques of Omega-3 Fatty Acids

Encapsulation and controlled release techniques have developed very quickly in the last three
decades throughout the medical and pharmaceutical industries. In general, microencapsulation
technologies are divided into three groups, namely chemical methods of molecular inclusion and
interfacial polymerization, physical methods including spray drying, freeze drying, air
suspension coating and extrusion, as well as physicochemical methods such as coacervation,
inclusion complex, and liposome entrapment (16). In the food sector, the main objectives of
encapsulation technology are protecting active compounds, covering undesirable properties and
controlled release of materials. New microencapsulation technologies are continuously being
developed in functional food markets and are the main innovations in this field (24). Currently,
products from the nanotechnology market comprise over one billion dollars in the food industry
and this will develop to over 20 billion dollars in next decade. Two methods for accessing
nanoparticle systems include mechanical or top-down methods, which work by reducing particle
size and chemical processes, or bottom-up methods, where nanoparticles are created from
smaller molecules, such as lipids or proteins (25). Encapsulation methods are capable of
affecting the color, shape, weight, volume, solubility, sensitivity to pressure, and sensitivity to
heat and light of compounds. Nanotechnology is used for designing, characterization, production
and application of structures, devices and nanosized devices and systems (<100nm).
Nanoencapsulation is a new method to protect bioactive compounds (from harmful
environmental conditions, oxidation, pH change and enzymatic degradation) such as omega-3
fatty acids and keep them available as functional food compounds. Compared to
microencapsulation, nano-sized delivery systems provide more surface, increased solubility,
improved bio-availability, and improvement of controlled release, as well as active targeting (26
29). Using capsules or encapsulation of fatty acids with multiple double bonds, a barrier can be
created to prevent the oxidation of these compounds. Efficiency of encapsulation and stability
when maintaining microcapsules of omega-3 oil is a point of interest in developing encapsulation
systems. Accessing these features depends on selection of formulation and the processing
conditions used to produce microcapsules (28).

4.1. Liposome as Delivery System of Functional Compounds

Biologically, liposomes can deliver bioactive compounds, since they are made from natural
lipids. Therefore, while being biodegradable, they are non-toxic and do not stimulate the immune
system. Their stability could be enhanced as well, since bioactive compounds are trapped inside

liposome (30). Currently, liposomes, as delivering systems, comprise a wide range of bioactive
compounds and have various applications in the food industry, the pharmaceutical industry, and
agriculture. The oldest studies using liposomes in the food industry date back to 1980 and
investigate liposomes as carriers of proteins, pigments, vitamins, antioxidants, and flavoring
agents. Other applications of liposomes in the food industry involve encapsulation of protease
enzyme that prevents premature breakdown of casein and reduction of enzyme loss.
Encapsulation of lactase is another application of liposome that releases lactase in the stomach
and, therefore, removes the sweet taste of hydrolyzed milk (9, 17, 27, 31). Because of its many
advantages, using liposome as carrier for bioactive compounds was approved by the FDA, in
addition to approval of loaded medical compounds inside liposomes, relative to using them as
free from. Many products in the form of liposome are available and have been approved by the
FDA for cancer treatment (32). Liposomes are created from one or more phospholipids, as one or
more bilayer membranes, are capable of encapsulating compounds with low molecular weight,
proteins, and genes. Various methods are used to produce liposomes. Some common methods
include thin layer film hydration, detergent depletion, ethanol/ether injection, reverse phase
evaporation, and emulsification methods (3, 18). In all methods, the lipid phase, which is
typically an organic phase, is mixed with the aqueous phase (3, 33). In most mentioned methods,
liposomes as multilamellar large vesicles (MLVs) are formed and to obtain suitable and uniform
particle size, various methods are used, including extrusion, sonication, and homogenization (3).
Selecting suitable method to produce liposomes and encapsulate bioactive compounds first
depends on the physicochemical properties of the material in question. Bioactive compounds are
divided into two classes depending on their interaction with liposome membranes: 1) Water
soluble compounds that are easily encapsulated inside the aqueous phase at the center of
liposomes or the aqueous phase between layers; 2) Amphiphilic and lipophilic compounds have
been shown to be trapped in the liposomal bilayers only if they formed a complex with the lipid
layers acyl chains. It is the coefficient of fat distribution or, in other words, the coefficient of
liposome distribution that causes a compound to be in the first or second group. In the case of
some materials, a method could be selected depending on the pKa, composition, and charge of
the phospholipids, so that the intended compound enters liposome aqueous phase or accumulates
in the lipid bilayer (34).
4.1.1. Stabilization of Liposomal Carrier Systems
Stabilization of colloid systems is typically described based on the presence or lack of particle
surface potential. Surface potential is created when dispersed solid particles have an electrical
charge relative to their ambient aqueous environment. Solid particles can obtain electrical charge
through a variety of methods. In brief, the main mechanisms involved in creating particle charge
are as follows: 1) Ionization of surface groups: Ionization is a common mechanism for surfaces
with ionizing groups. Some examples are carboxylic acid, sulfuric acid, sulfonic acid esters,
amino groups, and ammonium groups. In these cases, the amount of particles charge depends on
pH of the carrier environment; 2) Surface ion adsorption: In this state, adsorbed ions create
surface electric charge in particles. Surface adsorbed surfactants may create charge in particles
through ionization. In colloid systems such as liposomes, electrostatic repulsive forces, Van der
Waals attraction, and spatial repulsive forces play a more prominent role in stabilization of the
system (35). Stabilization of colloid systems is often performed based on electrostatic repulsion

(DLVO theory), spatial repulsion (HVO: Hesselink-Vrij-Overbeek theory), and a combination of

electrostatic and spatial repulsion.
4.1.1.1. Physical Stability

By forming a liposome, part of aqueous phase is entrapped inside a close and continuous bilayer
construction and, therefore, water-soluble compounds could be encapsulated inside the liposome.
In addition to entrapment of polar compounds in the aqueous phase of the vesicles, entrapment of
hydrophobic and hydrophilic compounds can be solved via formation with the bilayer lipid or
reaction with this lipid layer. This trend can change vesicle properties, including the permeability
and stability of bilayer construction. Stability of liposome formulations during construction,
maintenance, and consumption is an important factor in selecting liposomes as proper carriers of
bioactive compounds. Stability of the molecular structure of liposome membrane and materials
entrapped by the liposome are both important to consider. In the case of compound formulation
of liposomes, 2-year stability in 4C is considered favorable. Stability of liposome membrane
depends on the arrangement of hydrocarbon chains of the lipid molecules (10, 37). Some of the
physical changes in liposomes include changes in particle size, due to aggregation or fusion
during maintenance because chemical or thermal decomposition reactions or leaking of
encapsulated material. Physical stability typically includes fixed particle size and ratio of loaded
compound into liposome lipids and it is observed that at optimized pH and suitable salt
concentration, the physical properties of liposome remain constant in refrigerator temperature
(38).
4.1.1.2. Chemical Stability

Chemical instability in liposomes is mainly the result of hydrolysis of ester groups and oxidation
of unsaturated lipid acyl chains. During hydrolysis, the ester bond is broken and hydrophobic
chains are separated from the lipid. In addition, pH, temperature, and buffer ingredients are
effective on hydrolysis kinetics. Studies reveal that changes in acidity or alkalinity accelerate
hydrolysis reactions. Oxidation occurs because of the presence of unsaturated phospholipids in
the liposome structure. Using natural or synthetic antioxidants, including -tocopherol, could
effectively solve this problem (38). During oxidation reactions, the structural and physical
behavior of bilayers can altered significantly. The peroxidation of phospholipids occurs in the
same way as peroxidation of triacylglycerol. This fact means that change in configuration of the
phospholipid acyl chain brings about important changes in the structure of the liposomal
membrane. Physical properties and arrangement of lipid molecules are factors that affect
oxidation of liposome bilayer membrane. Lipid peroxidation occurs through a series of
sequential reactions. It is obvious that the size and z-potential of liposomes are influenced by the
peroxidation reaction (39).
4.1.1.3. Biologic Stability

Changes occur in liposome structure that contact biologic systems in an undesirable manner. The
main biologic instability of liposomes is their high tendency to aggregate, as well as drug release
in plasma. This problem becomes more complicated in vivo because of the presence of surfaces
with negative charge, colloid systems, and main ingredients of serum (38). When liposomes
enter the body, they become diluted by biologic fluids, including blood, lymph, and extracellular

fluids. Plasma makes up about 55% of the bodys total blood volume and is composed of various
molecules, macromolecules, and different ions. In this environment, lipoproteins and
phospholipase have a greater effect on liposome stability. Liposome and protein interactions
result from electrostatic attraction between different surface charges or hydrophobic forces cause
weak binding of proteins to the surface of the liposome or penetration into membrane layers of
the liposome. Stability of liposomes in plasma depends on their relative concentration, size,
number of lipid layers, and coordination lipid composition with the biologic environment of
body and ambient temperature (40).

4.2. Review of Literature

Encapsulation is a process where small solid particles, liquid droplets, or gas are trapped in
encapsulation materials. Encapsulation techniques have many applications in the food industry
and other industries. In the food sector, the main objectives of encapsulation techniques are
protecting active compound, covering undesirable properties, and controlled release of materials.
New microencapsulation methods are continually being developed in the market of functional
foods and are the main driving force for innovations in this field. Overall, encapsulation
techniques are divided into 3 groups: chemical, physical, and physicochemical techniques (24,
41). Transferring lipophilic bioactive compounds as liposome nanoparticles is an encapsulation
system developed in numerous different industries, but used especially often in the
pharmaceutical industry, and recently some applications of this method have been reported in
food industry as well. The oldest studies on applications of liposome in the food industry date
back to the 1980s (16). Some applications of nanotechnology in the food industry include
production of preservatives, oxidation-reduction factors, pigmentation compounds, sweeteners,
enzymes, antioxidants, acids, acids, alkalis, buffers, flavoring agents, and nutritional compounds.
The advantage of encapsulation with liposomes relative to other methods is that in the case of
excessive aqueous activity in the environment, encapsulated soluble materials are not released
and are only released in a target area or specific location (7). Nanoliposome technology provides
exciting opportunities for food technology in areas including encapsulation and controlled
release of food compounds, along with increasing bioavailability, stability, and increased shelflife of ingredients. Liposomes are used in the food industry to transfer flavoring and nutritional
compounds as well as antimicrobial materials. Currently, the market of nanotechnology products
in the food industry is about 1 billion dollars and in next decade it will develop to over 20 billion
dollars. Two ways of accessing nanoparticle systems include mechanical or top-down methods,
which work by decreasing particle size, and chemical or bottom-up processes, where
nanoparticles are generated from smaller molecules such as lipids or proteins (25). One of the
applications of liposome in the food industry is encapsulation of protease enzyme that prevents
premature breakdown of casein and reduces enzyme loss. Another application is lactase release
in the stomach that removes the sweet taste of hydrolyzed milk. Encapsulation of various
vitamins, such as vitamin D to fortify milk and its products, maintaining antioxidant activity of
vitamin C, and -topopherol, as well as encapsulation of antimicrobial materials such as nisin,
are among other applications of liposome in food industry (9, 27, 31). With regard to the
relationship between consuming omega-3 fatty acids and reduced risk of cardiovascular diseases,
researchers have conducted many studies on stabilization and bioavailability of omega-3 fatty
acids in liposome systems. Studies by Nara et al. (42) demonstrated that marine
phosphatidylcholine is very susceptible to degradation, in terms of oxidative stability. Oxidative

stability of phosphatidylcholine in salmon increases with addition of cholesterol, stearylamine,

and dicetyl phosphate. In addition, through using the proper amount of -tocopherol and
albumin, oxidation of phosphatidylcholine can be prevented. The most stabilized encapsulated
form of triglycerol with high DHA percentage was prepared with a liposome formulation
provided from salmon egg phosphatidylcholine. This study presents the potential of liposomal
formulation for encapsulation of the compounds prone to omega-3 oxidation (42). Nacka et al.
evaluated the physical and chemical stability of liposomes based on marine lipids under acidic
condition. Results demonstrated that acidic environments result in an increase in unilamellar
fusion of phosphatidylcholine vesicles, increase the number of large unilamellar liposomes, and
that the greatest effect was observed around the isoelectric point of phosphatidylcholine. In
addition, findings showed that increasing the surface charge of liposomes, as well as decreasing
electrolyte concentration, could interfere with decreasing fusion and aggregation (43). In a study,
Nacka et al. investigated fusion of -tocopherol in liposomes based on marine lipids in vivo and
in vitro. The results of their study demonstrated that the best oxidative stability was related to
liposomal formulation, where the -tocopherol content was 5 mol%. In addition, the production
of propanal was lower relative to other formulations. Liposome instability under acidic
conditions (pH 1.5) was evident along with hydrolysis (44).
Moussaoui et al. investigated the physical properties of liposomes provided by marine lipids
(Marionosomes). Liposome structural properties, including turbidity measurements via
spectrophotometry of optical density at 400nm (evaluation of morphology stability),
granulometric measurements to determine accurate size (equal to 300nm to 0.2 m after
filtration), and observing spherical shape using phase-contrast microscopy, were verified. In
addition, using freeze-fracture transmission electron microscopy, liposomes were evaluated in
terms of the number of lamella and whether they were all oligolamellar. Storing liposome
suspensions for 30 days at ambient temperatures indicated its stability. However, by reducing pH
(~4) and buffer, and using high concentration of calcium, liposome size decreased and liposomes
aggregated (45). After providing liposomes based on natural marine lipid by Cansell et al.,
availability of omega-3 fatty acids in animal models showed that marine phospholipid could be
an attractive compound to produce liposomes as food supplements with PUFA (46). Lyberg et al.
surveyed oxidation of different forms of docosahexaenoic acid within 28 days of storage at 28
30C. Based on the results of their study, phospholipid protects oxidation of carbon 4, 7, 8, and
11 of DHA. In addition, DPPC significantly prevents oxidation of carbon 20 of free DHA, which
is the most prone position to oxidation. This study shows the effective antioxidant ability of
DPPC phospholipid. This property is related to the arrangement of DHA atoms that create a
special bending and curvature in this molecule (47). Onuki et al. evaluated the effects of fatty
acids on thr physical properties of model bilamellar lipid membrane. Applying stearic acid (SA),
oleic acid (OA), docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA) in lipid
formulations decreased Van der Waals reactions between hydrocarbon chains of phospholipid
because of a twisted structure related to their double bonds (48). In another study, Namani et al.
demonstrated that when the molecular ratio of neutral DHA to anionic DHA is between 1:3 and
1:3 (pH between 8.5 and 9.2 and 10m DHA), it is shaped to form a vesicle. Investigating DHA
vesicles using cryo transmission electron microscopy (Cryo-TEM) demonstrated that the width
of a bilamellar vesicle was less than two times the length of a DHA molecule, which shows the
flexibility of DHA inside bilamellar vesicles. Comparison of vesicles prepared from saturated
fatty acids, including decanoic acid, with DHA vesicles demonstrated that the DHA vesicles

were chemically less stable as a result of the 6 allylic double bonds. Using calcein illustrated that
water-soluble molecules could be encapsulated inside DHA vesicles. Therefore, their application
in medicine and foods was suggested (49). Shaw et al. (2007) evaluated emulsions as carriers of
omega-3 fatty acids in food systems. Characterization of emulsion formulations composed of
lecithin or lecithin-chitosan, along with menhaden fish oil and 120% corn syrup, at 6%
humidity was conducted during different storage times. After 5 days of storage, hydroperoxide
value increased in samples containing fish oil and corn syrup (1%, 2%, and 10%) and after 10
days it ranges between 120170 m for each kg of oil. This increase was observed in powders
regenerated in water and was observed again after 12 days, when levels were between 140200
m for each kg of oil. The lowest peroxide index was related to powdered emulsion samples with
20 and 5% corn syrup, which were about 7 and 40 mmol for each kg of oil, respectively, after 30
days. The propanal value in regenerated samples with 20% corn syrup was determined after 3
and 6 days and after 12 days it was determined to be less than 25g for each kg of oil. Peroxides
in these samples ranged between 100200 mmol for each kg oil (50). Onuki et al. investigated
the special effect of unsaturated fatty acids on bilayer phospholipid membranes with small
amounts of cholesterol. The results of their study indicate that omega-3 fatty acids increase
separation of regular and irregular phases in membranes and that liposomes made of DPPC are
very susceptible to fluidity effect of omega-3 fatty acids. Effects of omega-3 fatty acids on DPPC
liposomes are significantly higher than oleic acid and stearic acid, which had no effect on DPPC
liposomes. By adding cholesterol to the mix, fluidity effects of omega-3 fatty acids weakened.
No evidence was presented to show an increase in lipid raft formation caused by omega-3 fatty
acids (48).
In a recent study, Barrow et al. stated that the main problem of fortifying food products with fish
oil containing EPA and DHA is their instability and that coacervation could be used
commercially to fortify food products and drinks with oils containing omega-3 fatty acids.
Reports provided by these authors on the comparison of equivalence for bioavailability of
microencapsulated omega-3 fatty acids with complex coacervation. Fish oil soft-gel capsules in
human demonstrated that bioavailability of microencapsulated omega-3 fatty acids is equal to the
method of complex coacervation by soft-gel fish oil capsules (51). The effect of absorption and
interaction of omega-3 fatty acids, including docosahexaenoic acid, docosapentaenoic acid, and
linoleic acid, on membrane models of dimyristoylphosphatidylcholine (DMPC) through
differential scanning calorimetry was investigated by Sarpietro et al. MLVs resulted from pure
DMPC, with different molar values of 0.0150.18 of omega-3 fatty acids with two endothermic
peaks; a small peak in 15C and main peak in 24.8C. The first peak was transferring from the
primary gel stage to the ripple state known as the primary stage, and the second or main peak
was transferring from the ripple state to the liquid crystal state. All fatty acids under study
resulted in shortening of the primary transfer peak and widening of the calorimetric peaks
according to the concentration used. The highest reduction was related to DHA acid, followed by
DPA acid and LNA respectively (52). The report from Rasti et al. on the physical and chemical
stability of liposomes and nanoliposomes containing fish oil, with 40% DHA and EPA, prepared
by conventional and thermal methods in a 10-month period after storage at 4C revealed that the
surface charge and stability of liposomes decreases with increasing size. They observed
differences in oxidative stability for PUFA because of the use of inorganic solvents for preparing
the liposomes (14). In the following study, the effects of spray and freeze drying was compared
with spray-freeze-drying methods in microencapsulation of DHA by Karthilk et al. Efficiency of

encapsulation (oil retention) in freeze, spray and spray-freeze-drying methods was reported as
73%, 83%, and 71%, respectively. The amount of peroxide found in samples powdered by the
spray-freeze method after 36 days of storage in 30C was about 8 meq O2/kg and the results for
each of samples prepared using freeze and spray drying were similar for approximately 12 meq
O2/kg. Therefore, the amount of peroxide in samples prepared using this method after this period
of storage in 4C differs slightly and can reach approximately 5 meq O2/kg (53). The study
conducted by Hadian et al. (54) revealed that liposomes can play a key role as solubilizing agents
in addressing the need for improved delivery of polyunsaturated FAs. DHA/EPA encapsulated
nanoliposomes were found to be promising and capable of effective reversal, which merits
further investigation. Using the thin layer hydration method (a schematic flow diagram is shown
in Figure 1), they produced DHA and EPA-loaded nanoliposomes. Probe ultrasound treatment of
pre-formed liposomes facilitates significant loading of DHA and EPA into the nanoliposomal
membrane. Transmission electron microscopy (TEM) image of liposomes showed that the
liposomes were spherical in shape and maintain high structural integrity (Figure 2) (54).

Background
Nanoencapsulation is the coating of various substances within another material at sizes on the
nano scale. This technique is already commonplace within a range of industries but it is accepted
that only around 10% of potential applications are being exploited.

Microencapsulation
Microencapsulation is similar to nanoencapsulation aside from it involving larger particles and
having been done for a greater period of time than nanoencapsulation. Nanoencapsulation has
evolved from and can be considered to be the miniaturisation of microencapsulation.

Encapsulation Terms
The encapsulated material is commonly referred to as the internal phase, the core material, the
filler or the fill. The encapsulation material is known as the external phase, the shell, coating or
membrane.

Nanocapsule Appearance
Common macro sized capsules used for off the shelf pharmaceuticals and vitamins are smooth
uniformly sized object but they are vastly different to micro and nanoencapsulated materials. As
the core material for different nanocapsules may vary greatly in size, shape and composition, the
encapsulated particle can be have an appearance that ranges from having regular, uniform shape
through to being jagged and irregular.

Nanoencapsulation Techniques
A multitude of techniques are used in nanoencapsulation and as the field is an emerging one, new
techniques are constantly being developed. The more popular techniques include:

Fluid bed coating

Wax and lipid coating

Spray drying

Spray congealing

Hydrogel encapsulation

Melt extrusion

Application
The basic reason for nanoencapsulation is to protect the core material and to then release it when
it is required. Applications for this include:

Targeted drug delivery systems that release the drug only when the drug has arrived at the
site in the body where it is required.

Timed release drug delivery where the nanoencapsulation material slowly allows the drug
to be released into the body such as nasal delivery of insulin. The coating material can be
customised to determine the rate of delivery

Embedded fragrances for branded perfumed clothing

Food additions and food enhancements such as Omega-3 fatty acid additions to bread that
do not alter taste

Increasing shelf life and stability of products like vitamins