J. of Korean Oil Chemists' Soc.

, 1
Vol. 31, No. 2. June, 2014. 203~209
ISSN 1225-9098 (Print)
ISSN 2288-1069 (Online)
http://dx.doi.org/10.12925/jkocs.2014.31.2.203

Candelilla Wax Nanoemulsions Prepared by Phase Inversion

Composition (PIC) Method

Eun-Hee Kim ․Wan-Goo Cho✝

Basic Medicine Department, College of Medical Sciences, Jeonju University, 303 Cheonjam-ro,
Wansan-gu, Jeonju 560-759, Korea
(Received May 1, 2014; Revised June 24, 2014; Accepted June 25, 2014)

Abstract : Candelilla wax-in-water nanoemulsions stabilized by Span 80/Tween 80 were

prepared by the phase inversion composition (PIC) method. Stable nanoemulsions with droplet
diameters below 50 nm could be formed when the hydrophilic–lipophilic balance (HLB) values
were between 13.5 and 14.5, surfactant concentration was 5.0 wt%, and the surfactant-wax
ratio was 1:1. Increased emulsification temperature and cooling rate were found to improve the
emulsion properties. Process of PIC (adding aqueous phase to the wax phase) produced smaller
droplet size nanoemulsion compared to the process of adding wax phase to the aqueous phase.
The stability of these nanoemulsions was assessed by following the change in droplet diameters
with time of storage at room temperature (∼25 °C). The size remained constant during 2
months storage time.

Keywords : candelilla wax, nanoemulsion, PIC, stability, cosmetics

1. Introduction energy emulsification methods require large

mechanical energy generated by high pressure
The potential benefits of cosmetic homogenizers or ultrasound generators to
nanoemulsions include optical clarity, good produce fine droplets. In contrast, low energy
stability to gravitational separation, flocculation emulsification methods can take advantage of
and coalescence, and improved absorption and the chemical energy stored in the ingredients
bioavailability of functional components [1]. A and produce the nanoemulsions almost
deeper understanding of the basic spontaneously, thus have great attraction both
physiochemical properties of nanoemulsions in theoretical study and practical application.
would, therefore, provide key information to The change in the spontaneous curvature of
better guide formulation and application of surfactants during the emulsifying process has
nanoemulsion to cosmetics. been recognized to be a key factor for the
Nanoemulsions can be prepared by high or formation of nanoemulsions, which can be
low energy emulsification methods [2–4]. High achieved either by changing the temperature
(phase inversion temperature, PIT method) or

✝Corresponding by changing the volume fraction of water or
author
oil (phase inversion composition method, PIC
(E-mail: wgcho@jj.ac.kr)

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Eun-Hee Kim Wan-Goo Cho
method)[2-4].
Although there have been many studies on
nanoemulsions with liquid oil as dispersed
phase, few systematic studies have been done
on emulsions with dispersed crystals, wax, a
crystallizable oil phase. The distribution of the
titanium dioxide inside the carnauba wax
phase of the nanosuspensions was more
effective to reach higher SPFs than that at the
aqueous phase [5]. Traditionally, nanoemulsion
containing carnauba wax was used as fruit
coating agent [6,7]. Paraffin wax has several
advantages, including its relatively large latent
heat, low vapor pressure and chemical
inertness. When paraffin is dispersed in
aqueous medium, a paraffin/water emulsion is
formed which is more suitable for many
applications, due to its greatly accelerated
heat-exchange and negligible thermal resistance
[8]. Leal-Calderon and co-workers have
investigated paraffin wax emulsions stabilized
by different stabilizers [9,10].
In the present work, we prepared candelilla
wax-in-water nanoemulsions using the
low-energy PIC method, and investigated the
effects of HLB value, surfactant concentration,
emulsification temperature and emulsification
route.
2. Materials and Methods
2.1. Materials
The candelilla wax(Kahl GmbH & Co. KG)
had a narrow range of melting point from
68.5-72.5 oC. The component of this
candelilla wax were odd-numbered n-alkanes
(C29 to C33), together with esters of acids
and alcohols with even-numbered carbon
chains (C28 to C34). Water was deionized and
Milli-Q filtered. Sorbitan monooleate (Span
80, c.p.), polyoxyethylene (20) sorbitan
monooleate (Tween 80, c.p.), were obtained
from Ilshin wells Co. Ltd. and Croda Chem.,
respectively. All reagents were used as received
without further purification.
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J. of Korean Oil Chemists' Soc.
2.2. Preparation of emulsions
Emulsions were prepared from a mixture of
candelilla wax and surfactants by slowly
adding EDTA-2Na aqueous solution with
gentle agitation using a magnetic stirrer. The
addition rate of aqueous solution was kept
constant at approximately 1.0 mL/min. The
emulsification temperatures were kept at 65–80
o
C. The concentration of candelilla wax in all
emulsions was kept constant at 5.0 wt%, while
the surfactant concentration was varied from
2.0 to 12.5 wt%.
2.3. Droplet size determination
The size and distribution of nanoemulsion
droplet were determined by dynamic light
scattering (ELS-8000, Otsuka, Japan). A 200
mW green laser (λ = 532 nm) with variable
intensity was used, and measurements were
carried out at room temperature with a
scattering angle of 90o. The droplet size was
measured directly without dilution. The
average radius were calculated from the
intensity autocorrelation data with the
cumulant method. The time-intensity
correlation functions were analyzed by the
CONTIN method [9].
3. Results
3.1. Determination of required HLB for
preparing of candelilla wax emulsions
Stable emulsions, especially where synthetic
surfactants are used, are best formulated with
emulsifiers or a combination of emulsifiers
having HLB values close to that required of
oil phase [11]. To attain a much greater
degree of the droplet stability, a combination
of hydrophilic and lipophilic emulsifiers is
often used, which are thought to align
alongside each other imparting more rigidity
and strength to the emulsifier film [12].
Emulsions were prepared initially with 90 wt%
aqueous phase, 5 wt% candelilla wax and 5
wt% emulsifier blend of Tween 80 and Span
Vol. 31, No. 2 (2014) Candelilla Wax Nanoemulsions Prepared by Phase Inversion Composition (PIC) Method 3

Fig. 1. Photograph of nanoemulsions as a function of HLB values with 5.0 wt%

candelilla wax and 5.0 wt% surfactants prepared by PIC method at 80 oC.

80 at HLB values ranging from 11.8 to 14.8

to determine the‘required HLB’of the candelilla
wax. The mixed HLB values were calculated
by the following equation:

HLBmix = HLBT × T + HLBS × S (1)

where HLBT, HLBS and HLBmix are the HLB

values of Tween 80 (15.0), Span 80 (4.3) and
Fig. 2. Droplet diameter as a function of HLB
the mixed surfactants, and T and S are the
values for nanoemulsions with 5.0 wt%
mass percentages of Tween 80 and Span 80 in
candelilla wax and 5.0 wt% surfactants
the mixed surfactants, respectively. All the
prepared by PIC method at 80 oC.
HLB values used were obtained at 25 oC. The
photograph of nanoemulsions as a function of
3.2. The influence of surfactant
HLB is shown in Fig. 1. HLB values between
concentration for preparing of
13.5 and 14.5 were found to be suitable for
candelilla wax nanoemulsions
small droplet size of nanoemulsions.
For the nanoemulsions prepared with a
Nanoemulsion stability is often estimated by
constant HLB (14.0, with the weight ratio of
the average size of the droplets, so the
Span 80/Tween 80 = 0.1/0.9), Fig. 3 shows
droplets size was also measured (Fig. 2). The
the size of nanoemulsion as a function of the
droplets size of emulsions with smallest size
surfactant concentration. The size of droplets
below 40 nm was attained at HLB 14.0,
was rather constant with the surfactant
beyond which the droplet size increased. So
concentration, because the HLB of surfactant
we chose HLB 14.0 as the candelilla wax's
determines the size of droplet of emulsions.
required HLB as stable candelilla wax O/W
nanoemulsions with droplet size below 40 nm
3.3. The influence of temperature for
were obtained within this region. The
preparing of candelilla wax
combination of Tween 80 and Span 80 within
nanoemulsions
the required HLB region may improve the
Candelilla wax, the dispersed phase used in
rigidity and strength of the emulsifier film
this study, is solid at room temperature.
around the wax droplets.
During the emulsification process, the
temperature is increased to melt the candelilla
wax for easier emulsification. Temperature can

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Eun-Hee Kim Wan-Goo Cho J. of Korean Oil Chemists' Soc.

affect the viscosity and the cohesive force of

the candelilla wax, so we prepared emulsions
at different temperatures to investigate the
effect of temperature. In Fig. 4, the
photograph of emulsions produced by varying
the emusification temperature is shown. The
transparency of emulsions is increased as the Fig. 4. Photograph as a function of emulsifying
emulsification temperature is increased. Fig. 5 temperature for emulsions with 5.0
shows clearly that the size of emulsion wt% candelilla wax and 5.0 wt%
droplets is decreased with temperature surfactants prepared at an HLB value
increasing from 65 to 80 oC. This is because of 14.0.
the viscosity and cohesive force decreases with
increasing temperature, which is beneficial to
emulsification. Rousseau [13,14] showed that
the crystallization regimes of the paraffin wax
could affect the emulsion's properties. Hence
we changed the cooling rate to study the
effect of the crystallization. For the emulsion
prepared at 80 oC, one part of it was cooled
at ambient temperature, and the other part
was cooled in an ice-bath. The diameter of
emulsion droplets and photograph are shown
in Fig. 6. As expected, the emulsion cooled in Fig. 5. Droplet diameter as a function of
the ice-bath had smaller particle size, 38.4 emulsifying temperature for emulsions
nm. It is thought that wax droplets may with 5.0 wt% candelilla wax and 5.0
partially coalesce when the temperature is wt% surfactants prepared at an HLB
higher than its melting point, the ice-bath value of 14.0.
method can shorten the time when the
emulsion is partially melted, and so it
facilitates formation of emulsions with smaller
droplet size.

Fig. 6. Droplet diameter as a function of

cooling rate for emulsions with 5.0
wt% candelilla wax and 5.0 wt%
surfactants prepared at an HLB value
Fig. 3. Droplet diameter as a function of
of 14.0.
surfactant concentration for emulsions
with 5.0 wt% candelilla wax
prepared at 80 oC with an HLB
value of 14.0.

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Vol. 31, No. 2 (2014) Candelilla Wax Nanoemulsions Prepared by Phase Inversion Composition (PIC) Method 5

3.4. The influence of emulsification route

for preparing of candelilla wax
nanoemulsions
Two different emulsification methods were
compared in this study: addition of aqueous
phase to a surfactant solution in oil (PIC
method, A); addition of oil to an aqueous
phase (method B). As shown in Figure 7,
stable candelilla wax emulsions cannot be
obtained by method B, while the PIC method
Fig. 7. Droplet diameter as a function of
can produce emulsions with small droplet size.
emulsification route for emulsions with
This is consistent with previous studies [3,4].
5.0 wt% candelilla wax and 5.0 wt%
The reason for this is that during the PIC
surfactants prepared at an HLB value
process the emulsion system undergoes a
of 14.0.
change from a W/O emulsion to an O/W
emulsion, and multiple emulsion, bicontinuous
or lamellar structure may be formed during

Fig. 8. Droplet size distribution and polydispersity index: (A) slow cooling, (B) fast
cooling, (c) method A, (D) method B determined by dynamic light scattering
(ELS-8000).

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Eun-Hee Kim Wan-Goo Cho
this process [4]. The PIC method is a
catastrophic phase inversion process, which is
induced by a change in the water-to-oil ratio.
We showed droplet size distribution and
polydispersity index determined by dynamic
light scattering Figure 8.
3.5. Emulsion stability
The stability of these nanoemulsions was
assessed by following the change in droplet
diameters with time of storage at room
temperature (∼25 °C). In Figure 9, we
presented the change of droplet size of
different HLB. The size remained constant
during the same storage time. It is deduced
that the components with crystal in room
temperature could help enhance the stability
against Ostwald ripening.
Fig. 9. Droplet diameter as a function of time
for samples with different HLB and
stored at 25 oC(O/S=1:1), ■: 11.8,
●: 12.9, ◇: 13.4, ◆: 14.0, ○:14.4,
□: 14.8.
4. Conclusion
Phase inversion composition method was
used to prepare candelilla wax-in-water
emulsions stabilized by Span 80/Tween 80.
The formation of emulsions was evaluated in
relation to HLB, surfactant concentrations,
temperature and processing parameters. Stable
nanoemulsions with droplet diameters below
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J. of Korean Oil Chemists' Soc.
50 nm could be formed when the hydrophilic–
lipophilic balance (HLB) values were between
13.5 and 14.5 and the surfactant-wax ratio
was 1:1. Increased emulsification temperature
and cooling rate were found to improve the
emulsion properties. The emulsion prepared
with optimum emulsification parameters was
quite stable and with an average droplet
diameter below 50 nm. We also found that
there was big difference of diameter of
emulsion droplets between addition of aqueous
phase to wax phase(A) and addition of wax
to aqueous phase(B). Candelilla wax used as
the oil phase of the nanoemulsion was a
crystallizable oil phase, which was better for
emulsion stability.
Acknowledgements
This study was supported by a grant of the
Korea Healthcare technology R&D Project,
Ministry of Health & Welfare, Republic of
Korea. (Grant No. HN12C0056)
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