Colloids and Surfaces A: Physicochem. Eng.

Aspects 430 (2013) 76–84

Contents lists available at SciVerse ScienceDirect

Colloids and Surfaces A: Physicochemical and

Engineering Aspects
journal homepage: www.elsevier.com/locate/colsurfa

Formulation and characterization of nanostructured lipid carriers

containing a mixed lipids core
Minying Zheng a , Mia Falkeborg b , Yan Zheng a,∗ , Tiankui Yang a , Xuebing Xu a,b
a
Wilmar (Shanghai) Biotechnology Research & Development Center Co., Ltd, 200137 Shanghai, China
b
Department of Engineering, Faculty of Science and Technology, Aarhus University, Gustav Wieds Vej 10, DK-8000 Aarhus C, Denmark

h i g h l i g h t s g r a p h i c a l a b s t r a c t

• We prepared drug free NLCs using

various solid fats and liquid oils.
• Effects of lipids on properties of NLCs
were investigated by PCS, DSC and
XRD.
• Rheological study indicated greater
elastic modulus over viscous modu-
lus of our NLC.
• Drug release model proved our NLC
formula to be a well-controlled deliv-
ery system.

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

Article history: The present study aimed to evaluate influences of mixed lipids and their proportions on formation and
Received 4 January 2013 properties of nanostructured lipid carriers (NLCs). The physicochemical parameters of drug-free NLCs
Received in revised form 27 March 2013 were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), differ-
Accepted 29 March 2013
ential scanning calorimetry (DSC), X-ray diffraction (XRD) and rheological measurements. In addition,
Available online xxx
a drug incorporation model was introduced into the NLCs for the application test. The hydrodynamic
diameter (Rd ) of the final NLCs (varying from 150 nm to 350 nm) was affected by diverse lipids core. DSC
Keywords:
results showed that both onset and melting points of NLCs declined due to the addition of the liquid
Nanostructured lipid carrier (NLC)
Particle size
oil, and the liquid oil enabled the melting peak wider, indicating a reduced crystallinity of NLC products
Crystallinity which almost coincided with the XRD analysis. Furthermore, rheological studies suggested stronger elas-
Rheological behavior ticity property than the viscosity property of the NLC systems. NLCs with high lipids content (20 wt%)
Drug incorporation had good thixotropy. Conjugated linoleic acid incorporation study showed that the NLC formulation had
the well-controlled release profile.
© 2013 Elsevier B.V. All rights reserved.

1. Introduction
As one of the main types of lipid nanoparticles, nanostructu-
red lipid carriers (NLCs) act as an alternative carrier system to
emulsions, liposomes and polymetric nanoparticles [1–3]. NLCs,
containing mixed lipids (solid fat and liquid oil), have many advan-
tages over solid lipid nanoparticles (SLNs). With more imperfection
∗ Corresponding author at: No. 118 Gaodong Road, Pudong New District, Shanghai
200137, China. Tel.: +86 21 31153159.
E-mail address: zhengyan1@wilmar-intl.com (Y. Zheng).
crystal structure compared to SLNs [4], NLCs have improved drug
loading capacity, and less drug expulsion during storage [2,5,6].
These benefits ensure wide application of NLC in delivery systems
for parenteral, peroral, dermal pharmaceuticals [7–9], as well as
other functional bioactives in food industry [10–12]. NLC-formulas
also appear in a number of cosmetic and skin care products [13,14].
However, most of the reports focus on a certain bioactive-loaded
NLC. Entrapment efficiency, loading capacity, release profile, and
stability of the bioactives in NLCs are mainly investigated. As for a
drug-free NLC system, effect of various surfactants has been stud-
ied by some experts [15,16]. Few researches have looked into the
influence of different lipids on the formulation of NLC and their

0927-7757/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.colsurfa.2013.03.070
 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84
properties, especially in food grade level. Meanwhile, investiga-
tions into food grade NLC could provide useful information for
further application in food industry.
In this paper, the effects of different fats and oils on NLC
formation were investigated. Fully hydrogenated sunflower oil
(HSF), fully hydrogenated rapeseed oil (HRO) and medium chain
triglycerides (MCT) were chosen as main lipids not only for
their physical properties but also for their healthy, functional
and medical applications [17,18]. Contributions of the lipids to
the particle distribution, melting behavior, and crystalline state
of the final products were analyzed by dynamic light scatter-
ing (DLS), transmission electron microscope (TEM), differential
scanning calorimetry (DSC), and X-ray diffraction (XRD). Rheolo-
gical behaviors were studied via oscillation frequency sweep and
thixotropy experiments. In vitro release model was investigated for
the application experiment.
2. Materials and methods
2.1. Materials
Solid lipids: fully hydrogenated sunflower oil or saturated sun-
flower oil (HSF or sat. sunflower), fully hydrogenated rapeseed
oil or saturated rapeseed oil (HRO or sat. rapeseed), and mixture
of palm oil and palm stearin (PO + ST or palm mixture) were all
food-grade commercial products from Kerry Specialty Fats Co. Ltd.
(Shanghai, China). Liquid oils: soybean oil (SBO) was a food grade
commercial product from Kerry Food Industries Co. Ltd. (Shanghai,
China). Medium chain triglycerides (MCT) (mixed triacylglycerols
of medium chain length saturated fatty acids: 60% caprylic and 40%
capric acids) were from Cognis Oleochemicals Co., Ltd. (Shanghai,
China). Tween 80 (chemically pure) was from Sinopharm Chem-
ical Reagent Co., Ltd. (Shanghai, China). Conjugated linoleic acid
(CLA, major isomers: c9-t11 and t10-c12; food grade) was from
Guanghua Technology Co., Ltd. (Shanghai, China). Sucrose (A.R.)
was from Lingfeng Chemical Reagent Co., Ltd. (Shanghai, China).
All other reagents were mainly from Sinopharm Chemical Reagent
Co., Ltd. (Shanghai, China), and were of analytical grade. The water
used in all experiments was distilled water.
2.2. Methods
2.2.1. Preparation of NLC
The drug-free NLC was prepared by melt-emulsification and
ultrasonication process. A certain amount of solid fat and liquid
oil were blended and melted at 80 ◦ C to form a uniform and clear
oil phase. Meanwhile, the aqueous phase consisted of 2.5 wt% dis-
persing Tween 80 in distilled water was added dropwise to the oil
phase at the same temperature by the aid of agitation at 600 rpm
for 10 min. The coarse emulsion was further treated by probe-type
sonicator (SCIENTZ- II D, Xinzhi, China) for 15 min (active every 3 s
for a 3 s duration at 450 W) in hot water bath (80 ± 0.5 ◦ C). Sub-
sequently the dispersion was cooled in ice water bath to room
temperature and stored at 4 ◦ C.
2.2.2. Particle size analysis
Particle size analysis was performed by dynamic light scattering
(DLS), also known as photon correlation spectroscopy (PCS), using
a Malvern Zetasizer Nano ZS (Malvern Instruments, UK). Prior to
measurements, all samples were diluted using distilled water to
yield a suitable scattering intensity. Each measurement was per-
formed in triplicate at 25 ◦ C. DLS yields the hydrodynamic diameter
Rd (intensity weighted mean diameter).
77
2.2.3. Differential scanning calorimetry (DSC) of the NLC
suspensions
The melting behavior and crystallinity of the lipid nanoparti-
cles were investigated by DSC 823 (Mettler Toledo, Switzerland).
3–5 mg of each sample (nanoparticle dispersion or bulk lipid
material) was accurately weighted in 40 ␮L aluminum pan, and
hermetically sealed. The heating runs were performed from 20 ◦ C
to 85 ◦ C and cooled down to 20 ◦ C at the heating/cooling rate of
5 ◦ C/min. During thermal scans, the crucible was purged with nitro-
gen. An empty aluminum pan was used as the reference and each
thermograph was baseline-corrected. The resulting diagrams were
analyzed using Mettler StarE DB V9.10 software.
2.2.4. Transmission electron microscopy (TEM)
NLC dispersions were observed with a transmission electron
microscope (TEM) using the negative-staining method according
to Zheng et al. [19]. A drop of NLC dispersion was spread on a
200-mesh copper grid coated with carbon membranes, and excess
droplets were instantly removed. A droplet of phosphotungstic acid
solution (2%, w/w) was dripped on the copper grid and excess
droplets were removed. Finally the grid was dried for about 3 h
before examined by a JEM-2010F electron microscope (Jeol, Japan).
2.2.5. X-ray diffraction (XRD)
The XRD analysis (Bruker D8 Advance, Bruker, Germany) was
performed by wide angle X-ray scattering with a scan speed of
2◦ /min, in the range: 2 = 3.00–40.00◦ . The X-ray source was a
2.2 kW Cu anode (40 kV, 40 mA, Cu-K␣ radiation,  = 0.15406 nm).
Prior to the measurement, the NLC samples were lyophilized to
evaporate the water. The bulk lipid was analyzed without any pre-
treatment.
2.2.6. Rheological investigations
Rheological measurements were performed using a Rheometer
Physica MCR 101 (Anton Paar, Austria) equipped with a cone-and-
plate test geometry (plate diameter 50 mm, cone angle 1◦ ) or a plate
(diameter 25 mm). Samples were kept in storage at 4 ◦ C for 6 days
before analysis. All measurements were carried out at 20 ± 0.1 ◦ C.
Oscillation stress sweep tests were performed at a constant fre-
quency of 1 Hz with a strain between 0.01 and 100%. Oscillation
frequency tests were run over a frequency range of 0.01–10 Hz
at constant stress amplitude under the linear viscoelastic region
(0.5%) which was previously determined by the oscillation stress
sweep tests.
Thixotropy was studied by analyzing the hysteresis loop
obtained when plotting shear stress versus shear rate. Continuous
flow measurements were performed by increasing shear rate from
0.0 to 100.0 s−1 over 250 s followed by decreasing the shear rate
from 100 to 0.15 s−1 over 250 s.
2.2.7. Drug incorporation and release study
A release study was employed to investigate the application
of NLC as carrier system. Conjugated linoleic acid (CLA), typically
lipophilic and poor water soluble, was one or a mixture of positional
and geometric isomers of octadecadienoic fatty acids containing
two conjugated double bonds. Prior to preparation, CLA was first
dissolved in the lipid phase and then the same procedure was per-
formed as in Section 2.2.1. For the release experiment, 1 wt% and
2 wt% CLA-loaded samples were prepared.
During the release study, a dialysis system using Sango mem-
brance bags (cutting off: 10 kDa, Sango Biotech (Shanghai) Co., Ltd.,
China) was employed. Each bag was filled with 5 mL NLC suspension
and soaking in 200 mL water/ethanol (50/50, v/v) at room tem-
perature and the magnetic stirring speed was 300 rpm. At regular
intervals, 1 mL aliquots of the external medium were withdrawn
and restored with the same volume of water/ethanol mixture.
78 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84

Fig. 1. The hydrodynamic diameter (Rd ) data of NLC formulations based on particle size measurements. (a) Influence of oil content on Rd of NLCs made from palm (mix) and
SBO or MCT; (b) influence of oil content on Rd of NLCs made from sat. sunflower and SBO or MCT; (c) influence of oil content on Rd of NLCs made from sat. rapeseed and SBO
or MCT; (d) effects of different lipid constituents on the size when the fat to oil ratio was fixed at 80 to 20. In this study, the total amount of lipid phase (fat and oil) was kept
constant being 5% (w/w) with regard to the total formulation.

The amount of the released drug was measured by UV analysis at
230 nm (Biophotometer, Eppendorf, Germany). The diffusion pro-
file of pure drug solution through a dialysis bag was examined as
control. The pure drug solution was prepared by dissolving 1 mg
CLA in 1 mL water ethanol solution (50/50, v/v). The pure CLA solu-
tion undertook the same dialysis process as NLC suspension.
2.3. Statistical analysis
For the particle size analysis, each sample was measured in trip-
licate. Both mean value and SD were present in the results.
3. Results and discussion
3.1. Effect of different bulk lipids on the particle size
In this study, the total amount of lipid phase (fat and oil) was
kept constant being 5% (w/w) with regard to the total formulation,
while the oil content varied. Oil content refers to the percentage of
liquid oil in the lipid matrix (fat and oil). The hydrodynamic diam-
eter (Rd ) data of NLC samples were presented in Fig. 1. It could be
clearly observed that the average size of the NLC decreased with
the increase of liquid oil amount (Fig. 1a–c). As shown in Fig. 1d,
palm and MCT made the smallest NLC suspension (around 158 nm);
while HRO and soybean oil-derived NLC formulation had the largest
particle size (around 283 nm).
In order to explain this phenomenon, the fatty acid composi-
tion (FAC) of all the lipids was determined by gas chromatography
(results in Table 1). According to Table 1, HRO contained 40.1%
(weight percentage of the total fatty acid) stearic acid (C18:0)
and 37.7% behenic acid (C22:0). HSF mainly contained stearic acid
(C18:0), while palm (mix) contained 45.3% palmitic acid (C16:0)
and 37.9% oleic acid (C18:1). As far as different solid lipids were con-
cerned (for example, the MCT-based NLCs in Fig. 1d), HRO made the
biggest NLC particles due to high content of behenic acid (C22:0).
On the other hand, NLC comprised of palm (mix) had the smallest
particle size owing to fatty acids composition of palmitic and oleic
acid. The size relationship of NLCs from these three solid fats was
as follows: HRO > HSF > palm (mix).
MCT contained 60% caprylic acid (C8:0) and 40% capric acid
(C10:0). Compared to MCT, soybean oil was composed of much
longer chain fatty acids (C18:2, C18:1, C18:0). Thus based on the
same solid lipid and the same oil content, MCT tended to form
smaller size of NLC than soybean oil.
Due to their different fatty acid compositions, bulk lipids
affected the particle size of final NLCs significantly. For both solid
 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84 79

Table 1
FAC analysis data of lipid materials.

Fatty acids Sat. sunflower [% of total FA] Sat. rapeseed [% of total FA] Palm fat [% of total FA] SBO [% of total FA]

C12:0 0.528 1.481 0.160 0.000

C14:0 0.191 0.667 0.778 0.000
C16:0 5.740 5.729 45.289 9.903
C18:0 91.376 40.130 4.717 3.583
C18:1 0.000 1.335 37.889 27.064
C18:1 T 0.543 2.327 0.666 0.000
C18:2 0.000 0.000 9.095 51.451
C18:2 T 0.000 0.049 0.356 0.000
C20:0 0.468 7.830 0.000 0.000
C20:1 0.000 0.000 0.000 5.621
C22:0 0.680 37.655 0.000 0.000
Others 0.475 2.796 1.049 2.376

fats and liquid oils, high proportion of long chain triacylglycerols
contributed to large NLC particles. On the other hand, the size of
NLC particles decreased with the increase of liquid oil for the same
kind of bulk lipids. This was due to that liquid oil could be eas-
ily dispersed into the aqueous phase and contributed to smaller
particles.
3.2. DSC investigation
DSC was frequently used to measure the heat loss or gain result-
ing from physical or chemical changes within a sample as a function
of the temperature. It had been used to characterize the state and
the crystallinity of lipid dispersions, semi-solid systems, polymers
and liposomes. It also allowed the study of the melting and crys-
tallization behavior of crystalline material like lipid nanoparticles
[15,20]. In the case of NLC, DSC experiments were useful to under-
stand the mixing behavior of solid fat with liquid oil.
DSC analysis was employed for all the 24 samples mentioned in
Section 3.1, and Fig. 2 depicted DSC curves of several NLC samples.
In the range of 20–85 ◦ C, no melting and crystallization peaks were
observed for palm (mix) derived NLCs, indicating that no solid lipid
particle existed in the final NLC product. Palm (mix) actually formed
emulsion system instead of NLC at room temperature. On the other
hand, HSF and HRO related NLCs rendered melting and crystalliza-
tion behavior during DSC treatment. In Fig. 2a, HRO-derived NLCs
tended to have lower melting temperature compared with HSF-
derived samples. And the HRO-derived NLC crystallized at higher
temperature than HSF-derived samples.
For the HSF- and HRO-derived samples, the influence of liq-
uid oil on the melting behavior was further investigated. Fig. 2b
compared the melting process of NLCs made from HRO and MCT
with different fat to oil ratios. The number above each curve repre-
sented the weight ratio of HRO and MCT in the bulk material. For
example, 9:1 means 90% HRO and 10% MCT mixed together as the
bulk lipid to make the NLC formulation. The melting peaks reflected
the crystal structure in our NLC systems. With solid lipid only (the
black curve, actually SLN), the peak seemed like steep slope. With
the increase of oil content (from 9:1 to 8:2), the peak gradually
broadened (increase in width of melting area) and the peak height
decreased, hence the crystallinity was reduced. This suggested that
the liquid oil was molecularly dispersed in the solid fat matrix.
According to Fig. 3, a decreased melting point and onset temper-
ature were observed with increasing oil content in the lipid blends
(both HRO & MCT and HSF & MCT), which also occurred in similar
NLC systems [16]. Width of melting refers to the temperature span
of each melting peak, mostly from the onset point to the ending
point of the melting process. It broadened with increasing oil per-
centage in the lipid blends, indicating a more imperfection crystal
structure.
As we known, the melting point depression occurred if one
compound was dissolved in another compound. With increasing
concentration of the first compound, the melting point decreased
in a linear fashion. In Fig. 3a, the decrease was linear up to 20% oil in
the lipid blend, indicating good miscibility when the oil content was
below 20%. When MCT increased up to 30%, the curves for melting
point and onset temperature declined sharply. It was deduced that
excessive oil not only made the melting start earlier, but also made
the width of melting process become much larger. In Fig. 3b, linear
decline was up to 30% oil in the lipid blend, indicating that HSF and
MCT had better miscibility compared to HRO and MCT.

3.3. TEM observation

TEM images of the dried suspension of NLC were presented in

Fig. 2. DSC diagrams of several NLC samples. (a) 10% (w/w) MCT in total lipids core
with different solid fats, the DSC procedure was as follow: 20–85 ◦ C, heating rate
5 ◦ C/min; 85 ◦ C, 3 min; 85–20 ◦ C, cooling rate 5 ◦ C/min. (b) Samples made from HRO
and MCT (numbers above each curve mean the ratio of solid and liquid oil) DCS
procedure: 20–80 ◦ C, heating rate 5 ◦ C/min.
Fig. 4. The four pictures represented the views from different pos-
itions with different amplification factors. All of the four pictures
indicated that the NLC particles were spherical. In picture A and
B, the diameter was around 200–300 nm, while in C and D, the
diameter was smaller (50–100 nm).
80 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84

Fig. 3. The influence of oil content on the melting temperature, onset temperature, and the width of melting. Here, width of melting refers to the temperature span from the
onset point to the ending point. (a) Sat. rapeseed and MCT; (b) sat. sunflower and MCT.

The sample for TEM imaging was 5% lipid (HSF:MCT = 7:3) dis-
persed in 2.5% Tween 80 aqueous solution. According to the size
data (based on DLS method) in Section 3.1, the diameter was
about 200 nm. The size difference obtained by TEM and DLS mainly
came from the different sample preparation processes and differ-
ent principles [6,21]. The DLS did not ‘measure’ the particle sizes
but detected the light scattering effects which were used to cal-
culate the particle size. The diameter from DLS would be a little
bigger than that observed from TEM. In addition, the resultant NLC
by our ultrasonication method was not a homogeneous system. A
few no-spherical particles may cause uncertainties.
3.4. X-ray diffraction
The molecules composing of long chain fatty acid had been
known to possess polymorphism [22]. The crystallinity study was
performed by X-ray diffraction for bulk lipid HSF, NLC 1 (5 wt%
lipid), NLC 2 (3.5 wt% lipid and 1.5 wt% MCT). The aqueous phase for
NLC 1 and NLC 2 was 2.5 wt% Tween80 solution. Before analysis, two
NLC samples were lyophilized to facilitate XRD sample preparation.
Fig. 5 showed the scattering pattern of the three samples.
According to Larsson, short spacings of triglycerides could be
described as following [23], ˛: hexagonal (H) subcell with a lattice
spacing of 0.42 nm; ˇ : orthorhombic perpendicular (O⊥ ) subcell
with strong lattice spacings of 0.42–0.43 and 0.37–0.40 nm; ˇ: tri-
clinic parallel (T) subcell with a strong lattice spacing of 0.46 nm.
Further polymorphic forms were found with complex glycerides
like mixed acid triacylglycerides or partial glycerides. Multiple ˇ
and ˇ, sub ˛ or intermediate forms were described [24]. Table 2 pre-
sented the d-spacing values of bulk lipid and NCL formulations from
XRD pattern. When sat. sunflower (HSF) was made into NLC formu-
lation, increasing relative intensities with the same d-spacing (0.38
 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84 81

Fig. 4. TEM microphotographs of NLCs. A, B, C and D represented the views from different positions with different amplification factors.

Fig. 5. X-ray diffraction patterns. (a) NLC 2 made from 3.5 wt% HSF and 1.5 wt% MCT; (b) NLC 1 made from 5 wt% HSF; (c) the bulk lipid HSF.
82 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84
Fig. 6. Storage modulus (G ), loss modulus (G ) and complex viscosity (*) of NLC aqueous dispersion with different oil content as a function of the frequency (0.1–100 s−1 )
at constant stress amplitude of 0.5%. (a) Oil content 0% (actually SLN); (b) oil content 10%; (c) oil content 20%; (d) oil content 30%.
and 0.37 nm, ˇ -form) indicated that the ˇ crystal became greater.
While liquid oil was employed into the NLC (from NLC 1 to NLC
2), ˇ form peak (0.46 nm) was no longer with the biggest intensity.
In summary, the polymorph of our NLC with a mixed lipids core
turned to less ˇ (stable) and greater ˇ (meta-stable). This feature
could facilitate the drug incorporation [25].
3.5. Rheological measurements
3.5.1. Oscillation frequency sweep test
Rheological measurements are useful for the characterization
of the viscoelastic properties of aqueous NLC dispersions. Fig. 6
showed the results of oscillation frequency sweep test of aque-
ous NLC dispersions. For all the four samples, the storage modulus
G (elastic component) was far greater than the loss modulus G
(viscous component), indicating a more elastic rather than viscous
behavior. Besides, the * depended on the frequency, i.e. decreased
with the frequency which is a typical for plastic materials [26]. At
extremely low frequency (bellow 0.5 s−1 ), storage modulus G , loss
modulus G , and complex viscosity * had a similar ascending stage
in Fig. 6a–c. While in the same frequency range, such sharp slope
did not occur in Fig. 6d. At high frequency range (mainly from 1
Table 2
XRD d-spacing values of bulk lipid and NCL formulations.
Samples d-Spacing (nm) (relative intensity, %)
HSF 0.456 (100), 0.383 (64.8), 0.367 (57.6)
NLC 1 0.454 (100), 0.382 (82.6), 0.367 (78.1)
NLC 2 1.478 (100), 0.456 (49.7), 0.383 (47.1)
to 100 s−1 ), the moduli values showed weak dependence on the
applied frequency.
As Fig. 6a and b demonstrate, 10% oil in the lipid core resulted
in a sharp decrease in both G and G , but this decrease tended to
be more smooth when the oil content change from 10% (Fig. 6b)
to 30% (Fig. 6d). It could also be deduced that when the oil con-
tent reached a certain degree (for example 30% in Fig. 6d), G and
G tended to more easily become equilibrium (at low frequency).
Moreover, the equilibrium value of both G and G declined with
the increasing oil content. Oil in the mixed lipid core ensured
better miscibility (DSC results) between lipids, and made smaller
particle size of NLC dispersion (PCS results). The better dispersed
formulation had smaller viscosity and decreased loss modulus
G . The elastic property weakened due to addition of liquid oil
[27].
3.5.2. Thixotropic study
Thixotropy was studied by analysing the hysteresis loop
obtained when plotting shear stress versus shear rate using a trian-
gular protocol. This method was acceptable for quality control and
could provide useful information [28].
Fig. 7 indicated the rheograms of two NLC samples with dif-
ferent lipids proportion. The lipids meaned the mixed lipids core
(sat. sunflower: MCT = 7:3), and the percentage meaned the weight
proportion in the final NLC products. The hysteresis loop of 20%
Polymorphs lipids NLC enclosed a large area than one of the 15% lipids sample,
ˇ, ˇ indicating a more distinct thixotropy of 20% lipids NLC. Thixotropic
ˇ, ˇ property might well contribute to a better subjective impression of
ˇ, ˇ consistency and spreadability on food surface.
 M. Zheng et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 430 (2013) 76–84
Fig. 7. Shear stress (Pa) of NLC dispersions as function of shear rate (s−1 ) to investi-
gate the thixotropy of different NLC samples. The area of the thixotropic loop showed
the capability of thixotropy.
3.6. Drug incorporation and release profile
Conjugated linoleic acid (CLA) referred to a class of positional
and geometric conjugated dienoic isomers of linoleic acid; cis-
9, trans-11 (c9-t11) and trans-10, cis-12 (t10-c12) are the main
isomers [29]. Various positive health effects were ascribed to
CLA consumption, including anticarcinogenic, antiatherogenic, and
antidiabetic activity [30,31]. In this study, CLA was considered as a
model bioactive compound and the application test was conducted
for the NLC system.
Furthermore, the release process of the CLA loaded NLC
was investigated compared with pure CLA solution in water
ethanol (1/1, v/v) (Fig. 8). In the same dialysis solution
(water/ethanol = 50/50), pure CLA (which means 1 mg/mL CLA
water/ethanol solution) was quickly released during the first sev-
eral hours and the cumulative released amount was almost up to
100% after 10 h. Comparatively, CLA was released slowly from NLC.
After 4 days, the cumulative released from 2% loaded NLC was 46%,
Fig. 8. Release profile of different CLA related suspensions in water ethanol (50/50,
v/v) outside solution. (A) 2 wt% CLA loaded NLC suspension; (B) 1 wt% CLA loaded
NLC suspension; (C) pure CLA solution as a control experiment.
83
and released from 1% loaded NLC was about 73%. Both two CLA
loaded NLC reached equilibrium after 6–7 days. Consequently, the
NLC system was proved to have well controlled release profile for
CLA.
Due to its liquid state, CLA was always introduced as a capsula-
tion formula. According to previous researches, it was encapsulated
by many matrices like whey protein concentrate, gum arabic, mal-
todextrin, etc. [32,33]. Few reports were made about NLC delivery
for CLA. Our attempt provided a lipid-based system for CLA encap-
sulation which had well controlled release profile.
4. Conclusion
From the present study, it could be concluded that the lipids
core influenced the particle size and size distribution of the final
NLC products enormously. Liquid oil in the lipids core, decreased
the particle size, onset and melting points, as well as broaden the
width of melting peak linearly. Meanwhile, liquid oil in our NLC sys-
tems produced more imperfect crystallinity, which could promote
the drug incorporation and release control. Based on rheological
measurements, the NLC samples had greater elastic modulus over
viscous modulus with thixotropic property at higher lipid content
(20 wt%).
Acknowledgment
Wilmar (Shanghai) Biotechnology Research & Development
Center Co., Ltd is thanked for the permission to publish the work as
well as partial support for the second author.
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