You are on page 1of 7

Food Hydrocolloids 43 (2015) 540e546

Contents lists available at ScienceDirect

Food Hydrocolloids
journal homepage: www.elsevier.com/locate/foodhyd

Preparation and characterization of nanoemulsion encapsulating


curcumin
T.P. Sari a, Bimlesh Mann a, *, Rajesh Kumar a, R.R.B. Singh b, Rajan Sharma a,
Minaxi Bhardwaj a, S. Athira a
a
Dairy Chemistry Division, National Dairy Research Institute, Karnal, Haryana 132001, India
b
Dairy Technology Division, National Dairy Research Institute, Karnal 132001, India

a r t i c l e i n f o a b s t r a c t

Article history: Curcumin is the most active and least stable bioactive component of turmeric (Curcuma longa) plant. In
Received 7 March 2014 the present study, an attempt has been made to overcome the instability during processing and
Accepted 11 July 2014 bioavailability problems of curcumin by nanoencapsulation technology and the effect was evaluated by
Available online 22 July 2014
simulated digestion study. Curcumin was encapsulated in medium chain triglyceride oil droplets of
nanoemulsion prepared by ultrasonification using whey protein concentrate-70 and Tween-80 as
Keywords:
emulsifiers with an encapsulation efficiency of 90.56 ± 0.47%. The prepared nanoemulsion has particles of
Curcumin
average diameter 141.6 ± 15.4 nm and zeta potential of 6.9 ± 0.2 mV. In vitro release kinetics of cur-
Nanoencapsulation
Bioavailability
cumin from nanoemulsion by simulated gastrointestinal studies showed that the curcumin nanoemulsion
Nanoemulsion was relatively resistant to pepsin digestion but pancreatin causes release of curcumin from nanoemulsion.
Simulated gastrointestinal digestion The slow release of curcumin from the nanoemulsion was supposed to increase bioavailability. The total
antioxidant activity of curcumin nanoemulsion was reduced from 3.53 ± 0.11 mM Trolox/mg of curcumin
to 3.33 ± 0.02 mM Trolox/mg of curcumin after encapsulation. The prepared nanoemulsion was stable to
pasteurization, different ionic strengths (0.1e1 M) and pH ranging from 3.0 to 7.0. The study has
important implication in the formation and design of encapsulated bioactive systems.
© 2014 Elsevier Ltd. All rights reserved.

1. Introduction To improve the bioavailability of curcumin, numerous ap-


proaches have been undertaken by many researchers including,
Curcumin, the bioactive component obtained by the extraction preparation of nanocurcumin (Bisht et al., 2007), liposomal curcu-
and purification of ground rhizomes of Curcuma longa has been min (Li, Braiteh, & Kurzrock, 2005), curcuminephospholipid com-
found to exert wide range of beneficial biological and pharmaco- plex (Liu, Lou, Zhao, & Fan, 2006), and chelation with metals (John,
logical activities including antioxidant (Sharma, 1976), anti- Kuttan, & Krishnankutty, 2002). Micro/nanoencapsulation tech-
inflammatory (Chainani-Wu, 2003), antimicrobial (De et al., nology of poorly water soluble bioactives has attracted wide
2009) and anticancer (Aggarwal, Kumar, & Bharti, 2003) in clin- attention in the food and pharmaceutical industry for the past few
ical trials. But studies addressing the metabolism and uptake of years for various applications like protection of bioactivity and their
curcumin had shown that either no curcumin or little amount was controlled release for improving bioavailability. The encapsulation
detected in serum or tissue after administration (Dhillon et al., of polyphenols overcome the drawbacks of their instability, alle-
2008). The main reasons attributing to the low bioavailability are viate unpleasant tastes or flavors, as well as improve the half-life of
supposed to be the poor solubility of curcumin in aqueous media, the compound in vivo and in vitro (Augustin & Hemar, 2009;
rapid hydrolysis followed by molecular fragmentation at physio- Mozafari et al., 2008). Nanoencapsulation is generally employed
logical pH and inactivity of its metabolic products (Lin, Pan, & Lin- to improve solubility, stability and bioactivity of various oil-soluble
Shiau, 2000). phytochemicals due to their small droplet size and high kinetic
stability. Among the nanometric encapsulation systems, nano-
emulsions are particularly suitable and widely accepted for food
* Corresponding author. applications (McClements, Decker, & Weiss, 2007). Encapsulation
E-mail addresses: bimleshmann@gmail.com, bmann@rediffmail.com (B. Mann). into nanoemulsion-based delivery systems of bioactive compounds

http://dx.doi.org/10.1016/j.foodhyd.2014.07.011
0268-005X/© 2014 Elsevier Ltd. All rights reserved.
T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546 541

characterized by low solubility in aqueous phase represents an purchased from MERCK (Merck Specialities Pvt. Ltd., Worli, Mum-
effective approach to improve the dispersion of the bioactives into bai). Whey protein concentrate (WPC-70) was procured from
food products, to protect them against degradation or interaction Modern Dairy Pvt. Ltd. (Karnal, India). Pepsin and pancreatin were
with other ingredients, to reduce the impact on organoleptic purchased from Sisco Research Laboratories Pvt. Ltd. and Hi Media
properties of the food and to improve their bioavailability (Donsi, laboratories Pvt. Ltd., Mumbai, India, respectively.
Sessa, Mediounic, Mgaidic, & Ferrari, 2011). A kinetically stable
suspension can be obtained by adding stabilizers, emulsifiers or 2.2. Preparation and characterization of curcumin nanoemulsion
texture modifiers in the emulsion systems.
The slow release of curcumin from nanoparticulate curcumin Oil-in-water nanoemulsion was prepared in two stages using
formulation increased the bioavailability of delivered curcumin. the method described by Jafari, He, and Bhandari (2007) with
Unlike native curcumin, curcumin was detected in serum for a long modification. Curcumin (10e40 mg) was added to 100 ml of total
duration time period as observed after 24 h of its administration emulsion along with MCT-60 (0.5e2%), surfactant Tween-80
in vivo. Whereas, in native case, the curcumin level was subse- (2e10% w/w) and emulsifier WPC-70 (0e1% w/w) in Millipore
quently decreased in serum with time and not detectable after 1 h, water. The coarse emulsion was then prepared using magnetic
indicating rapid metabolism of native curcumin in physiological pH stirrer under ambient temperature for different time intervals. Fine
(Mohanty & Sahoo, 2010). Wang et al. (2008) prepared nano- emulsion was prepared by sonifying the coarse emulsion by opti-
emulsion of curcumin and found that the anti-inflammatory ac- mizing the sonification time using ultrasonicator (Fig. 1).
tivity improved than the conventional emulsion. Bioavailability of Emulsion stability was measured by centrifugation at 1300  g
curcumin was improved by encapsulation with polylactic-co- (Kubota, Tokyo, Japan) at 5  C for 30 min and heating at 80  C for
glycolic acid nanoparticles (Anand et al., 2010). 30 min. The stable sample was further analyzed for physico-
Emulsion based colloidal delivery systems are commercially chemical characterization. Particle size distribution was deter-
used in food, pharmaceutical and cosmetics industries. So, it is mined by dynamic laser light scattering method using Malvern
necessary to establish their stability to environmental stress and its Zetasizer Nano ZS90 (Malvern Instruments, UK). Zeta potential, the
effect on the physico-chemical characteristics. It has been reported electrical charge on the oil droplets in the emulsions was determined
that the physical and chemical stability of nanoemulsions greatly under holder temperature of 25  C and electrical voltage 3.9 V.
dependent on storage temperature, pH, and ionic strength (Qian,
Decker, Xiao, & McClements, 2012; Rao & McClements, 2011). 2.3. Determination of encapsulation efficiency
There is a need to optimize the right composition of nanoemulsions
such as decrease creaming, flocculation and coalescence for The encapsulation efficiency was determined and calculated by
different commercial applications. method of Surassmo, Min, Bejrapha, and Cho (2010) with modifica-
In order to design functional foods with potential to promote tion. Nanoemulsion was passed through Vivaspin concentrators of
human health, it is necessary to examine the digestibility and MWCO (Molecular Weight Cut Off) 100 kDa and centrifuged at
release of food components under simulated gastrointestinal (GI) 1300  g and 5  C for 30 min. The permeate was collected to calculate
conditions (Shani-Levi, Levi-Tal, & Lesmes, 2013). The delivery encapsulation efficiency by measuring its total phenolic content.
systems based on nanoencapsulation have been designed to control Total phenolic content of nanoemulsions and permeate was analyzed
the targeted release of encapsulated material at a specific location by FolineCiocalteu's method given by Zheng and Wang (2001).
in the human GI tract. The emulsion containing medium chain
triglyceride (MCT) as carrier lipid has been reported to substantially 2.4. Determination of stability of nanoemulsion under processing
increase the bioaccessibility of curcumin (Ahmed, Li, McClements, conditions
& Xiao, 2012). Knowledge of digestion kinetics of foods in human
GI tract is also required in establishing processing conditions at the The effect of different processing conditions like heating (63  C
manufacturing stage to achieve desirable release of nutrients. In the for 30 min and 95  C for 10 min), pH (3e7) and ionic strength
present study, we focused on the preparation, physico-chemical (0.1e1 M NaCl) was studied which will be applicable for their
characterization and in vitro digestion kinetics of curcumin commercial utilization. The particle size distribution, zeta potential
nanoemulsion. and stability were measured after exposing the nanoemulsions to
each treatment.
2. Materials and methods
2.5. Antioxidant activity of curcumin nanoemulsion
2.1. Materials
Radical scavenging activity of the curcumin emulsions before and
Curcumin (95% purity) was procured from Plant Lipids Pvt. Ltd. after encapsulation was determined by DPPH (2, 2 diphenyl-1-picryl
(Kerala, India) and used without further treatment. MCT-60 with hydrazyl) method given by Williams, Cuvelier, and Berset (1995).
predominantly MCT C8(58.6%) & C10(41.4%) was obtained from The results were expressed as trolox equivalent antioxidant capacity
Kamani Oil Industries Pvt. Ltd. (Mumbai, India). Tween-80 was (TEAC) values i.e. mM of trolox equivalent/mg of curcumin.

Fig. 1. Methodology of preparation of curcumin encapsulated nanoemulsion.


542 T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546

2.6. Release kinetics of curcumin from nanoemulsion were tried alone and in combination to get stable formulations.
Preliminary studies in our laboratory (Sari et al., 2013) have indi-
The release mechanism of encapsulated curcumin under simu- cated that individual use of both Tween-80 (1e10%) and WPC-70
lated GI conditions was evaluated by mimicking the physiological (1e5%) did not show a positive result with respect to their stabil-
situation in the upper tract (stomach and small intestine) of human ity as studied during heating followed by centrifugation.
body. Simulated gastric fluid (SGF) was prepared by the method Researchers have successfully prepared emulsions of curcumin
given by Sanchez, Fernandez-Garcia, Margolles, Reyes-Gavilan, and with relatively higher concentration of Tween-80 (Nakagawa,
Ruas-Madiedo (2010). Curcumin nanoemulsion (adjusted to pH 1.5) Sowasod, Charinpanitkul, Soottitantawat, & Tanthapanichakoon,
was mixed with SGF in the ratio of 1:3 in a series of test tubes. The 2011; Wang et al., 2008). There are two types of emulsifiers being
mixture was placed in a shaking water bath at 37  C for 2 h which used in foods i.e. low molecular weight (e.g. Tween-80) and high
represents the average duration of GI transit. The entire samples molecular weight (e.g. proteins) surfactants. At similar bulk con-
were subdivided in different test tubes with equal volume. During centrations (W/V), low molecular weight surfactants decrease the
the course of experiment the samples were removed after every surface or interfacial tension to a greater extent than the macro-
hour for testing the percentage of curcumin released with respect molecular surfactants. This difference mainly related to differences
to time. The release of curcumin from nanoemulsion was evaluated in orientation and configuration of these surfactants at the inter-
after deactivation of enzyme by keeping the emulsions at 90  C/ face. In the case of low molecular mass surfactants, the entire
10 min. Simulated intestinal fluid (SIF) was prepared by the method molecule adsorbs and instantaneously orients itself and this par-
given by Liang et al. (2012) with slight modifications. The SIF con- titioning of the entire molecule between the two phases facilitates
tained 4 mg/ml pancreatin and 25 mg/ml bile salts. The pH was a maximum reduction of interfacial tension. Although proteins are
adjusted to 7.5 using 1 N NaOH. The sample after 2 h of gastric less surface active than low molecular mass surfactants, but
digestion was mixed with SIF and allowed to incubate for further emulsions formed by low molecular mass surfactants are mostly
3 h at 37  C. Samples were removed from the water bath as unstable. This is because proteins, in addition to lowering interfa-
mentioned above after each hour and release of curcumin from cial tension, can form a continuous viscoelastic membrane like film
nanoemulsion was measured as the total phenolic contents after around oil droplets via noncovalent intermolecular interactions
deactivating of enzymes. and via covalent disulphide cross-linking whereas low molecular
mass surfactants cannot form such a viscoelastic films. Thus,
2.7. Statistical analysis emulsions which contain both low molecular and macromolecular
surfactants, the stability of colloidally dispersed phases is primarily
All statistical analysis was done using MS-EXCEL-2010 package. dependent on protein films adsorbed at the interfaces (Damodaran,
All measurements were done for at least three replicates. Results 1997). Moreover, in case of water soluble non-ionic surfactant,
are presented as means ± standard error of means (SEM). whether the addition occurs before or after emulsification has
relatively little influence on the competitive adsorption behavior
3. Results and discussion (Courthaudon, Dickinison, Matsumura, & Williams, 1991). In light
of the above discussion we may say that in the present formulation
3.1. Optimization and characterization of curcumin nanoemulsion both are present at the interface.
The zeta potential roughly characterizes the surface charge of
Different concentration of both core (curcumin and oil) and the emulsion particles. High absolute values lead to repulsive forces
coating materials (emulsifiers, and/or co-surfactant) at different between particles, which may improve the physical stability of
mass ratios were used to optimize the stable formulations (Fig. 1). multiphase systems (Muller, 1996). The zeta potential of the
The conditions for the preparation of nanoemulsions were opti- emulsion has been presented in Table 1. The relatively low value of
mized and stable nanoemulsions were obtained by sonification at zeta potential in the prepared nanoemulsion may be due to the
4  C for 15 min. The most stable formulation was obtained with presence of relatively higher amount of Tween-80 used in formu-
40 mg curcumin dissolved in a suitable solvent (2% MCT) þ 2% lations so that the surface charge is contributed only by whey
Tween-80 þ 0.5% WPC-70 having particle size of 141.6 ± 15.4 nm, proteins. Electrostatic and steric repulsion are the two main sta-
zeta potential of 6.9 ± 0.2 mV and poly dispersity index (PDI) of bilizing forces for whey protein stabilized emulsions. Electrostatic
0.273. The pH of the nanoemulsion was 5.57 ± 0.1 and it was stable interactions between similarly charged droplets are repulsive, and
for 27 ± 2 days at 25  C (Table 1). The relatively lower value of PDI may be either short range or long range depending on ionic
for nanoemulsion can be correlated with more stability on storage. strength (Demetriades, Coupland, & McClements, 1997). When
The reason for the aggregation of nanoemulsion during storage in droplets come so close together that the adsorbed emulsifier layer
globular protein stabilized emulsions is flocculation characterized begins to overlap, some short-range interactions become impor-
by the exposure of non-polar groups in the protein (Kim, Decker, & tant, such as steric, thermal fluctuation and hydration forces
McClements, 2002). In the present study, both WPC and Tween-80 (Israelachvili, 1992). So it indicates the emulsion droplets stabilized
by protein due to electrostatic and steric repulsion but electrostatic
repulsion predominates due to the surface charge on the proteins.
Table 1
Whereas the oil droplets of emulsions are stabilized by Tween-80
Physico-chemical characteristics of curcumin nanoemulsion.
against aggregation by steric (rather than electrostatic) repulsion
Nanoemulsion composition 2% MCT* þ 2% Tween-80 þ 0.5% due to relatively large hydrophilic (polyoxyethylene) head groups
WPC-70** þ 40 mg curcumin
of adsorbed Tween molecules (Klang & Valenta, 2011).
Size (nm) 141.6 ± 15.4
Zeta potential (mV) 6.9 ± 0.2
3.2. Encapsulation efficiency of curcumin in its nanoemulsion
Poly dispersity index 0.273
Stability at room temperature (days) 27 ± 2
pH 5.57 ± 0.1 The encapsulation efficiency of curcumin into nanoemulsion
Encapsulation efficiency (%) 90.56 ± 0.47 was 90.56 ± 0.47% (Table 1) which was evaluated by considering
MCT*: medium chain triglycerides and WPC-70**: whey protein concentrates-70. total phenolic content as a marker. To determine the encapsulation
Values mentioned above is mean ± SEM (n ¼ 3). efficiency the samples (native and nanoencapsulated) were passed
T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546 543

Table 2
Effect of pH, ionic strength and heating on particle size and zeta potential of cur-
cumin nanoemulsions.

Particle Zeta
size (nm) potential (mV)

Effect of pH
Control nanoemulsion 141.6 ± 15.4 6.9 ± 0.2
(pH: 5.57 ± 0.1)
pH 3.0 163.4 ± 7.03 þ2.03 ± 0.1
pH 5.0 143 ± 8.08 6.9 ± 0.2
pH 7.0 140.5 ± 5.74 6.26 ± 0.4

Effect of ionic strength


0.1 M NaCl 137.9 ± 11.7 1.83 ± 0.1
0.5 M NaCl 140.3 ± 9.2 1.25 ± 0.3
1.0 M NaCl 140.7 ± 7.4 0.523 ± 0.1

Effect of heating
63  C/30 min (pasteurization) 139.0 ± 11.7 4.19 ± 0.7
95  C/10 min (boiling) 531.9 ± 6.76 1.65 ± 0.2

Values mentioned above is mean ± SEM (n ¼ 3).

the conformation of the b-lactoglobulin monomer. These changes


Fig. 2. Images of curcumin solution (15 ml) passed through 100 kDa MWCO mem- lead to the exposure of hydrophobic groups and the free sulphydryl
brane; a) native curcumin dissolved in ethyl alcohol, b) nanoencapsulated curcumin group (Cys121) (Relkin, Meylheuc, Launay, & Raynal, 1998), resulting
(2% oil þ 2% Tween-80 þ 0.5% whey protein concentrate-70). in a reactive monomer which can propagate an aggregation reac-
tion leading to the formation of non-native dimers, trimmers, tet-
through 100 kDa MWCO membrane. From Fig. 2, it can be depicted ramers and larger aggregates by polymerization. b-Lactoglobulin
that native curcumin dissolved in ethyl alcohol passed through the being the most abundant whey protein, dominates the overall
membrane while, nanoencapsulated curcumin retains in the behavior of whey protein products. Qian et al. (2012) investigated
retentate. the influence of temperature on the physical and chemical stability
of b-carotene enriched nanoemulsions and concluded that the
3.3. Changes in size and zeta potential of nanoemulsion under nanoemulsions were prone to droplet aggregation at elevated
different processing conditions temperatures (>37  C).
The zeta potential of the nanoemulsion was also measured as a
3.3.1. Effect of heating function of heat treatments. The denaturation of whey protein also
The influence of different heat treatment on the particle size caused the change in the surface charge of particles. The zeta po-
distribution of the curcumin nanoemulsion was evaluated at 63  C tential was 6.9 ± 0.2 mV (Table 1) initially and the magnitude of
for 30 min (pasteurization) and 95  C for 10 min (boiling) and re- the charge decreased appreciably with increasing temperature.
sults are presented in Fig. 3 and Table 2. The average size of the After pasteurization and boiling, the particles had relatively low net
particles changed slightly after pasteurization while boiling causes charges, with the magnitude of the zeta potential being 4.19 ± 0.7
gradual increase in the average size due to aggregation (Fig. 3). The and 1.65 ± 0.2 mV (Table 2), respectively.
nanoemulsion was unstable to aggregation of the particles at
elevated temperature most likely due to the denaturation of whey 3.3.2. Effect of pH
protein which causes aggregation. On heating to 70  C, the dimers The particle size and zeta potential of the nanoemulsion were
of b-lactoglobulin dissociate to form monomers (Hambling, evaluated after adjusting to different pH (2.0e7.0) (Table 2 and
McAlpine, & Sawyer, 1992) followed by some critical change in Fig. 4). Average particle size increased with decrease in acidity of
the emulsion i.e. from 163.4 to 140.5 nm at pH 7.0. The higher mean

Fig. 3. Average particle size and particle size distributions of curcumin nanoemulsion Fig. 4. Average particle size and particle size distributions of curcumin nanoemulsion
after pasteurization (63  C/30 min) and boiling (95  C/10 min). maintained at different pH (3.0, 5.0 and 7.0).
544 T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546

value of particle size at low pH was may be due to the aggregation activity of curcumin nanoemulsion determined by DPPH radical
of particles at pH values close to the isoelectric point of whey scavenging assay was 3.33 ± 0.02 mM Trolox/mg of curcumin,
protein. This can also be correlated with the reduction in the net whereas radical scavenging activity of native curcumin was
negative charge at low pH values which leads to particle aggrega- 3.53 ± 0.11 02 mM Trolox/mg of curcumin. To exert the bioactivity,
tion and corresponding increase in the particle size. antioxidants have to be bio-accessible, i.e., released from the food
There was a significant difference in the zeta potential of the matrix and solubilized (Bouayed, Hoffmann, & Bohn, 2011).
nanoemulsion with change in pH from 3.0 to 7.0. The particles at pH Although the antioxidant activity of the encapsulated compound is
3.0 have net positive charge (þ2.03 ± 0.1 mV) compared to particles lower than unencapsulated, the fact is that, the encapsulation not
at pH 7.0 (6.26 ± 0.4 mV) (Table 2). This is due to the fact that the only protects curcumin from degradation but may also preserve the
surface charge of the milk proteins shifts to positive at pH below antioxidant activity.
the isoelectric point. This shift in the zeta potential indicates that Donsi et al. (2011) evaluated the effect of the delivery systems of
the surface charge of the prepared nanoemulsion is contributed by curcumin by comparing the antioxidant activity of the encapsu-
milk proteins. Rao and McClements (2011) studied the impact of lated compound with that of unencapsulated curcumin. The anti-
nano/microemulsions on various environmental stress and found oxidant activity of curcumin encapsulated in solid lipid
that relatively stable nanoemulsions were formed at pH 6.0 and 7.0, nanoemulsion using FRAP assay was 0.996 ± 0.07 mM L-ascorbic
but extensive particle growth/aggregation occurred at lower and acid equivalent while that of unencapsulated curcumin shows a
higher pH values, which was attributed to either chemical (hy- value of 2.504 ± 0.06 mM L-ascorbic acid equivalent.
drolysis) or physical (electrical charge) effects. The reduction in the
electrical charge on the particles at lower pH value would reduce 3.5. In vitro digestion of curcumin nanoemulsion
the electrostatic repulsion between them, thereby leading to ag-
gregation (Rao & McClements, 2012). The digestion of encapsulated bioactives in the gastrointestinal
tract is a complex process and its impact on the release of bioactive
3.3.3. Effect of ionic strength component plays a major role in the uptake, distribution as well as
Salts are common food additives and are also present in the bioavailability of the component. The gastric and intestinal digest
human gastrointestinal tract. Salts may therefore affect the func- was analyzed for % release of total phenolic content (curcumin)
tional performance of colloidal delivery systems in a product or from nanoemulsion (Fig. 6). The results indicated that the encap-
after ingestion. For these reasons, the influence of salt (0.1e1 M sulated curcumin was very stable and not released from the de-
NaCl) on properties of nanoemulsion was examined. The addition livery system by the action of pepsin. During simulated gastric
of salt did not cause any significant change in the particle size digestion (2 h), over 90% of the encapsulated curcumin was
(Fig. 5). The zeta potential of the nanoemulsion shifted toward zero retained in emulsion. The resistance of nanoemulsion
due to the decrease in the electrostatic repulsion between the toward pepsin digestion may be attributed to the resistance of b-
molecules upon addition of salt and consequently the charge on the lactoglobulin against pepsin digestion. Since WPC-70 is composed
molecules was reduced (Table 2). Thus addition of salt may lead to of 50e60% b-lactoglobulin and native b-lactoglobulin has been
destabilization of the nanoemulsions. The destabilization of the reported to be almost resistant to breakdown in the gastric
nanoemulsions at high salt concentrations can be attributed to compartment following simulated digestion (Fu, Abbott, & Hatzos,
screening of the electrostatic repulsion between the protein coated 2002; Schmidt, Meijer, Slangen, & Van Beresteijn, 1995) due to its
droplets by the salt ions. At relatively low salt levels, the electro- compact globular structure (Guo, Fox, & Flynn, 1995), 8.48% release
static repulsion is still sufficiently strong to overcome the weak van of curcumin after gastric digestion in the present study may be due
der Waals and hydrophobic attraction, but above a critical salt level, to highly acidic condition.
it is no longer strong enough so that the attractive forces dominate, The synergistic interactions between enzymes and acidic-
leading to droplet aggregation (McClements, 2005). alkaline pH conditions result in a significant breakdown of food

3.4. DPPH scavenging activity measurements of curcumin


nanoemulsion

The antioxidant activity of the encapsulated curcumin is slightly


lower than that of unencapsulated curcumin. The total antioxidant

Fig. 6. Release of curcumin from the nanoemulsion under simulated gastro-intestinal


Fig. 5. Average particle size and particle size distributions of curcumin nanoemulsion digestion condition by using simulated gastric fluid (SGF) and simulated intestinal fluid
maintained at different ionic strengths (0.1e1.0 M NaCl). (SIF).
T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546 545

materials including emulsions (Sarkar, Goh, Singh, & Singh, 2009). Kerala (India) and Modern Dairy Pvt. Ltd., Haryana (India) for
Stability of emulsion droplets in the gastric environment is a providing curcumin and WPC-70, respectively, for the emulsion
desirable attribute for an encapsulation system in order to protect preparation.
the encapsulant from the harsh gastric environment (Anal & Singh,
2007; Vandenberg, Drolet, Scott, & De la Noue, 2001). But in case of References
intestinal digestion, the incubation with SIF resulted in destabili-
zation of the emulsion and approximately 77% of the encapsulated Aggarwal, B. B., Kumar, A., & Bharti, A. C. (2003). Anticancer potential of curcumin:
curcumin was released within 2 h of incubation (Fig. 6). Fu et al. preclinical and clinical studies. Anticancer Research, 23(1A), 363e398.
Ahmed, K., Li, Y., McClements, J. D., & Xiao, H. (2012). Nanoemulsion- and emulsion-
(2002) and Schmidt et al. (1995) reported that b-lactoglobulin is based delivery systems for curcumin: encapsulation and release properties.
almost completely digested by pancreatic enzymes. This indicates Food Chemistry, 132, 799e807.
that the prepared nanoemulsions of curcumin are stable to gastric Anal, A. K., & Singh, H. (2007). Recent advances in microencapsulation of probiotics
for industrial applications and targeted delivery. Trends in Food Science and
enzymes but can be destabilized with subsequent release of cur-
Technology, 18(5), 240e251.
cumin in presence of pancreatin. Bossios et al. (2011) reported that Anand, P., Nair, H. B., Sung, B., Kunnumakkara, A. B., Yadav, V. R., & Tekmal, R. R.
only b-lactoglobulin was broken down to smaller peptides after (2010). Design of curcumin-loaded PLGA nanoparticles formulation with
gastro-duodenal digestion although a sizable amount of intact enhanced cellular uptake and increased bioactivity in vitro and superior
bioavailability in vivo. Biochemical Pharmacology, 79(3), 330e338.
protein still remained. Simulated intestinal fluid (SIF) is a mixture Augustin, M. A., & Hemar, Y. (2009). Nano- and micro-structured assemblies for
of pancreatin and bile salts. Bile salts may change the interface encapsulation of food ingredients. Chemical Society Reviews, 38, 902e912.
which facilitates activity of lipase present in pancreatin and helps in Benzariaa, A., Maresca, M., Taiebc, N., & Dumay, E. (2013). Interaction of curcumin
with phosphocasein micelles processed or not by dynamic high-pressure. Food
the release of curcumin. Similar observation was made by Sarkar, Chemistry, 138, 2327e2337.
Horne, and Singh (2010) that bile salt preferentially adsorbs at Bisht, S., Feldmann, G., Soni, S., Ravi, R., Karikar, C., Maitra, A., et al. (2007). Poly-
the oilewater interface displacing the protein or binding on to the meric nanoparticle-encapsulated curcumin (nanocurcumin): a novel strategy
for human cancer therapy. Journal of Nanobiotechnology, 5(3), 1e18.
protein coated droplets, thereby allowing lipase to act. In the Bossios, A., Theodoropoulou, M., Mondoulet, L., Rigby, N. M., Papadopoulos, N. G.,
presence of bile salts, coalescence between emulsion droplets Bernard, H., et al. (2011). Effect of simulated gastro-duodenal digestion
seemed to occur, resulting in oiling-off and breakdown of the on the allergenic reactivity of beta-lactoglobulin. Clinical and Translational Al-
lergy, 1(6).
b-lactoglobulin stabilized emulsion system. This was probably due Bouayed, J., Hoffmann, L., & Bohn, T. (2011). Total phenolics, flavonoids, anthocya-
to the lipolytic activities of pancreatin, being favored by the bile nins and antioxidant activity following simulated gastrointestinal digestion and
salts displacing the interfacial b-lactoglobulin. Apart from dialysis of apple varieties: bioaccessibility and potential uptake. Food Chemistry,
128(1), 14e21.
the possible lipolytic action of the pancreatin causing coalescence,
Chainani-Wu, N. (2003). Safety and anti-inflammatory activity of curcumin: a
the proteolysis of the adsorbed protein layer (due to the tryp- component of tumeric (Curcuma longa). The Journal of Alternative & Comple-
sinechymotrypsin fractions present in pancreatin) at the droplets mentary Medicine, 9(1), 161e168.
surface of the emulsion would possibly have resulted in reduced Courthaudon, J. L., Dickinison, E., Matsumura, Y., & Williams, A. (1991). Influence of
emulsifiers on the competitive adsorption of whey proteins in emulsions. Food
surface viscosity compared to intact interface and facilitates Structure, 10, 109e115.
coalescence. Damodaran, S. (1997). Protein-stabilized foams and emulsions. In S. Damodaran, &
Benzariaa, Maresca, Taiebc, and Dumay (2013) reported that A. Paraf (Eds.), Food proteins and their applications (pp. 57e59). New York:
Marcel Dekker, Inc.
enzymatic digestion of PC (phosphor casein) curcumin complexes De, R., Kundu, P., Swarnakar, S., Ramamurthy, T., Chowdhury, A., Nair, G. B., et al.
was quite resistant to pepsin treatment in simulated stomach (2009). Antimicrobial activity of curcumin against Helicobacter pylori isolates
digestion. Pancreatin treatment caused greater and faster release of from India and during infections in mice. Antimicrobial Agents and Chemo-
therapy, 53(4), 1592e1597.
curcumin, showing 38e77% of total curcumin released within Demetriades, K., Coupland, J. N., & McClements, D. J. (1997). Physical properties of
5e60 min incubation, and 81% after 3 h incubation. Incubation in whey protein stabilized emulsions as related to pH and NaCl. Journal of Food
simulated intestinal environment resulted in destabilization of the Science, 62(2), 342e347.
Devadasu, V. R., Wadsworth, R. M., & Kumar, M. N. V. R. (2012). Tissue localization of
emulsion and approximately 60% of the encapsulated curcumin
nanoparticles is altered due to hypoxia resulting in poor efficacy of curcumin
was released within 2 h of incubation (Tikekar, Pan, & Nitin, 2013). nanoparticles in pulmonary hypertension. European Journal of Pharmaceutics
Devadasu, Wadsworth, and Kumar (2012) reported that curcumin and Biopharmaceutics, 80(3), 578e584.
Dhillon, N., Aggarwal, B. B., Newman, R. A., Wolff, R. A., Kunnumakkara, A. B.,
is mainly found in the intestinal tract after oral administration of
Abbruzzese, J. L., et al. (2008). Phase II trial of curcumin in patients with
nanoparticles. Our study also supported this statement as majority advanced pancreatic cancer. Clinical Cancer Research, 14(14), 4491e4499.
of curcumin got released with intestinal enzymatic action. Donsi, F., Sessa, M., Mediounic, H., Mgaidic, A., & Ferrari, G. (2011). Encapsulation of
bioactive compounds in nanoemulsion based delivery systems. Procedia Food
Science, 1, 1666e1671.
4. Conclusion Fu, T. J., Abbott, U. R., & Hatzos, C. (2002). Digestibility of food allergens
and nonallergenic proteins in simulated gastric fluid and simulated intestinal
Curcumin being highly unstable and hydrophobic is difficult to fluid e a comparative study. Journal of Agricultural and Food Chemistry, 50(24),
7154e7160.
incorporate in aqueous food systems. For this reason the curcumin Guo, M. R., Fox, P.,F., & Flynn, A. (1995). Susceptibility of b-lactoglobulin and sodium
was encapsulated inside carrier oil in an emulsion form. The results caseinate to proteolysis by pepsin and trypsin. Journal of Dairy Science, 78(11),
in the present study showed that curcumin releases slowly from 2336e2344.
Hambling, S.,G., McAlpine, A. S., & Sawyer, L. (1992). b-Lactoglobulin. In P. F. Fox
nanoemulsion under simulated digestion. These results have (Ed.), Advanced dairy chemistry (Vol. 1, pp. 141e190). London: Elsevier Applied
important implication in the design of nanoemulsions of curcumin Science Publishers.
for commercial applications. It is concluded that the nano- Israelachvili, J. N. (1992). Intermolecular and surface forces. London: Academic Press.
Jafari, S., He, Y., & Bhandari, B. (2007). Production of sub-micron emulsions by
encapsulation of highly lipophilic and unstable compounds is an ultrasound and microfluidization techniques. Journal of Food Engineering, 82(4),
effective platform to increase the hydrophilicity, bioaccessibility 478e488.
and to protect them from degradation. John, V. D., Kuttan, G., & Krishnankutty, K. (2002). Anti-tumour studies of metal
chelates of synthetic curcuminoids. Journal of Experimental & Clinical Cancer
Research, 21(2), 219e224.
Acknowledgments Kim, H. J., Decker, E. A., & McClements, D. J. (2002). Role of postadsorption
conformation changes of beta-lactoglobulin on its ability to stabilize oil drop-
We are grateful for the financial support given by National Dairy lets against flocculation during heating at neutral pH. Langmuir, 18(20),
7577e7583.
Research Institute, Karnal, Haryana and NAIP, ICAR for this work. Klang, V., & Valenta, C. (2011). Lecithin based nanoemulsions. Journal of Drug De-
(Grant No. C30029), We would like to thank Plant Lipid Pvt. Ltd., livery Science and Technology, 21(1), 55e76.
546 T.P. Sari et al. / Food Hydrocolloids 43 (2015) 540e546

Li, L., Braiteh, F. S., & Kurzrock, R. (2005). Liposome-encapsulated curcumin: in vitro nchez, B., Fern
Sa andez-García, M., Margolles, A., Reyes-Gavila n, C. G. D., &
and in vivo effects on proliferation, apoptosis, signaling, and angiogenesis. Ruas-Madiedo, P. (2010). Technological and probiotic selection criteria of a bile-
Cancer, 104(6), 1322e1331. adapted Bifidobacterium animalis subsp. lactis strain. International Dairy Journal,
Liang, L., Wu, X., Zhao, T., Zhao, J., Li, F., Zou, Y., et al. (2012). In vitro bioaccessibility 20, 800e805.
and antioxidant activity of anthocyanins from mulberry (Morus atropurpurea Sari, T. P., Mann, B., Sharma, R., Kumar, R., Vikrant, & Minaxi. (2013). Process opti-
Roxb.) following simulated gastro-intestinal digestion. Food Research Interna- mization for the production of nanoencapsulated curcumin and analysis for
tional, 46, 76e82. physicochemical characteristics and antioxidant mechanism. International
Lin, J. K., Pan, H. M., & Lin-Shiau, S. (2000). Recent studies on the biofunctions and Journal of Biotechnology and Bioengineering Research, 4, 581e586.
biotransformations of curcumin. Biofactors, 13, 153e158. Sarkar, A., Goh, K. K. T., Singh, R. P., & Singh, H. (2009). Behaviour of an oil-in-water
Liu, A., Lou, H., Zhao, L., & Fan, P. (2006). Validated LC/MS/MS assay for curcumin emulsion stabilized by beta-lactoglobulin in an in vitro gastric model. Food
and tetrahydrocurcumin in rat plasma and application to pharmacokinetic Hydrocolloids, 23(6), 1563e1569.
study of phospholipid complex of curcumin. Journal of Pharmaceutical & Sarkar, A., Horne, D. S., & Singh, H. (2010). Pancreatin-induced coalescence of oil-in-
Biomedical Analysis, 40(3), 720e725. water emulsions in an in vitro duodenal model. International Dairy Journal, 20,
McClements, D. J. (2005). Food emulsions: Principles, practice, and techniques. Boca 589e597.
Raton, FL: CRC Press. Schmidt, D. J., Meijer, R. J., Slangen, C. J., & Van Beresteijn, E. C. (1995). Raising the
McClements, D. J., Decker, E. A., & Weiss, J. (2007). Emulsion-based delivery systems pH of the pepsin catalyzed hydrolysis of bovine whey proteins increases the
for lipophilic bioactive components. Journal of Food Science, 72(8), 109e124. allergenicity of the hydrolysates. Clinical and Experimental Allergy, 25(10),
Mohanty, C., & Sahoo, K. S. (2010). The in vitro stability and in vivo pharmacoki- 1007e1017.
netics of curcumin prepared as an aqueous nanoparticulate formulation. Bio- Shani-Levi, C., Levi-Tal, S., & Lesmes, U. (2013). Comparative performance of milk
materials, 31, 6597e6611. proteins and their emulsions under dynamic in vitro adult and infant gastric
Mozafari, M. R., Khosravi-Darani, K., Borazan, G. G., Cui, J., Pardakhty, A., & digestion. Food Hydrocolloids, 32, 349e357.
Yurdugul, S. (2008). Encapsulation of food ingredients using nanoliposome Sharma, O. P. (1976). Antioxidant activity of curcumin and related compounds.
technology. International Journal of Food Properties, 11, 833e844. Biochemical Pharmacology, 25(15), 1811e1812.
Muller, R. H. (1996). Zeta-potential and particle charge in the laboratory practice e Intro- Surassmo, S., Min, S., Bejrapha, P., & Cho, M. (2010). Effects of surfactants on the
duction to theory, practical and data interpretation (pp. 104e108). Stuttgart: Wis- physical properties of capsicum oleoresin-loaded nanocapsules formulated
senschaftliche Verlagsgesellschaft. through the emulsionediffusion method. Food Research International, 43, 8e17.
Nakagawa, K., Sowasod, N., Charinpanitkul, T., Soottitantawat, A., & Tikekar, V. R., Pan, Y., & Nitin, N. (2013). Fate of curcumin encapsulated in silica
Tanthapanichakoon, W. (2011). Encapsulation of curcumin loaded oil droplets nanoparticle stabilized Pickering emulsion during storage and simulated
by cryotropic gel formation from O/W emulsion. Procedia Food Science, digestion. Food Research International, 51, 370e377.
1973e1979. Vandenberg, G. W., Drolet, C., Scott, S. L., & De la Noue, J. (2001). Factors affecting
Qian, C., Decker, A. E., Xiao, H., & McClements, D. J. (2012). Physical and chemical protein release from alginateechitosan coacervate microcapsules during pro-
stability of b-carotene-enriched nanoemulsions: influence of pH, ionic strength, duction and gastric/intestinal simulation. Journal of Controlled Release, 77(3),
temperature, and emulsifier type. Food Chemistry, 132(3), 1221e1229. 297e307.
Rao, J., & McClements. (2012). Food-grade microemulsions and nanoemulsions: role Wang, X., Jiang, Y., Wang, Y., Huang, M., Ho, C., & Huang, Q. (2008). Enhancing anti-
of oil phase composition on formation and stability. Food Hydrocolloids, 29, inflammation activity of curcumin through O/W nanoemulsions. Food Chemis-
326e339. try, 108(2), 419e424.
Rao, J., & McClements, D. J. (2011). Food-grade microemulsions, nanoemulsions Williams, W. B., Cuvelier, M. E., & Berset, C. (1995). Use of a free radical method
and emulsions: fabrication from sucrose monopalmitate and lemon oil. Food to evaluate antioxidant activity. LWT e Food Science and Technology, 28(1),
Hydrocolloids, 25, 1413e1423. 25e30.
Relkin, P., Meylheuc, T., Launay, B., & Raynal, K. (1998). Heat-induced gelation of Zheng, W., & Wang, S. Y. (2001). Antioxidant activity and phenolic compounds in
globularprotein mixtures. A DSC and a SEM study. Journal of Thermal Analysis, selected herbs. Journal of Agricultural and Food Chemistry, 49, 5165e5170.
51, 747e755.

You might also like