Enzymatically interesterified fats based on mutton tallow and

walnut oil suitable for cosmetic emulsions

M. Kowalska*, M. Mendrycka*, A. Zbikowska

†

and S. Stawarz*

*Faculty of Material Science, Technology and Design, Kazimierz Pulaski University of Technology and Humanities, Chrobrego Street 27, 26-600
Radom, and

†

Department of Food Technology, Faculty of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), Nowoursynowska

159C, 02-787 Warsaw, Poland

Received 26 August 2014, Accepted 7 October 2014

Keywords:

emulsions, interesterification synthesis, mutton tallow, sensory evaluation, stability of emulsion, walnut oil

Synopsis
OBJECTIVE: Formation of emulsion systems based on interesteri-
fied fats was the objective of the study. Enzymatic interesterifica-
tion was carried out between enzymatic mutton tallow and
walnut oil in the proportions 2 : 3 (w/w) to produce fats not
available in nature. At the beginning of the interesterification
process, the balance between the interesterification and fat
hydrolysis was intentionally disturbed by adding more water to
the catalyst (Lipozyme IR MR) of the reaction to produce more
of the polar fraction monoacylglycerols [MAGs] and diacylglyce-
rols [DAGs]. To obtain a greater quantity of MAGs and DAGs in
the reaction environment via hydrolysis, water was added (11,
13, 14, 16 w-%) to the enzymatic preparation. The obtained fats
were used to form emulsions.
METHODS: The emulsions were evaluated with respect to sensory
and skin moisturizing properties by 83 respondents. Determination
of emulsion stability using temperature and centrifugal tests was
carried out. Morphology and the type of emulsions were deter-
mined.
RESULTS: The respondents described the skin to which the emul-
sions in testing were applied as smooth, pleasant to touch and ade-
quately moisturized.
CONCLUSIONS: The work has demonstrated that interesterifica-
tion of a mutton tallow and walnut oil blend resulted in new fats
with very interesting characteristics of triacylglycerols that are not
present in the environment. The results of the present work indi-
cate the possibility of application of fats with the largest quantity of
MAGs and DAGs as a fat base of emulsions in the cosmetic indus-
tries. The hypothesis assumed in this work of producing additional
quantities of MAGs and DAGs (in the process of enzymatic inter-
esterification) responsible for the stability of the system was con-
firmed. It should be pointed out that the emulsions based on
interesterified fats exhibited a greater level of moisturization of the
skin than the emulsions containing non-interesterified fat. Also, in
the respondents’ opinion, the emulsion containing fat, which was
modified during enzymatic interesterification when 13% of water

was added to the enzymatic preparation, exhibited the best sensory
profile.

R

esum
e

OBJECTIF: La formation des syst

emes d’
emulsion a base de lipides

transest

erifi
es
etait l’objectif de l’
etude. L’est
erification enzymatique

a

et
e r
ealis
ee entre le suif de mouton et l’huile de noix dans les pro-

portions 2: 3 (poids / poids) pour produire des lipides qui ne sont
pas disponibles dans la nature. Au d

ebut du processus de trans-

est

erification, l’
equilibre entre la transest
erification et l’hydrolys
e

des lipides a

et
e volontairement perturb
ee par l’ajout d’eau pour le

catalyseur (Lipozyme IR MR) de la r

eaction en vue de produire

davantage de fractions polaires monoacylglyc

erols [MAG] et diacyl-

glyc

erols (DAG). Pour obtenir une plus grande quantit
e de MAG et

de DAG dans le milieu de r

eaction par hydrolyse, de l’eau a
et
e

ajout

ee (11, 13, 14, 16% en poids) de la pr
eparation enzymatique.

Les corps gras obtenus ont

et
e utilis
es pour former des
emulsions.

M
ETHODES: Les

emulsions ont
et
e
evalu
ees par rapport aux pro-

pri

et
es hydratantes et sensorielles par 83 r
epondants. La d
etermina-

tion de la stabilit

e de l’
emulsion en fonction de la temp
erature et

les essais d’

etalement ont
et
e effectu
es. La morphologie et le type

d’

emulsions ont
et
e d
etermin
es.

RESULTATS: Les volontaires ont d

ecrit la peau sur laquelle les

emulsions des tests ont
et
e appliqu
ees comme lisse, agr
eable au
toucher et hydrat

ee convenablement.

CONCLUSIONS: Le travail a d

emontr
e que l’est
erification d’un

m

elange d’huile de suif de mouton et de l’huile de noix aboutit a

de nouvelles lipides avec des caract

eristiques tres int
eressantes de

triglyc

erides qui ne sont pas pr
esents dans l’environnement. Les

r

esultats de la pr
esente
etude indiquent la possibilit
e de l’applica-

tion de lipides avec des plus grandes quantit

es de MAG et DAG

comme base de lipide d’

emulsions dans les industries cosm
etiques.

L’hypoth

ese suppos
ee de ce travail, a savoir de produire des quan-

tit

es suppl
ementaires de DAG et MAG (dans le processus d’inter-

est

erification enzymatique) responsables de la stabilit
e du systeme a

et
e confirm
ee. Il convient de souligner que les
emulsions a base de
graisses inter-est

erifi
ees pr
esentaient un plus grand niveau d’hydra-

tation de la peau que les

emulsions contenant des matieres grasses

non-est

erifi
ees. Aussi, de l’avis des r
epondants, l’
emulsion conten-

ant de la graisse qui a

et
e modifi
ee au cours de transest
erification

enzymatique quand 13% de l’eau a

et
e ajout
ee a la pr
eparation

enzymatique, pr

esentait le meilleur profil sensoriel.

Correspondence: Malgorzata Kowalska, Faculty of Material Science,
Technology and Design, Kazimierz Pulaski University of Technology
and Humanities, Chrobrego Street 27, 26-600 Radom, Poland. Tel.:
[phone]; fax: [phone]; e-mail: [e-mail]

82

© 2014 Society of Cosmetic Scientists and the Soci
et
e Franc
aise de Cosm
etologie

International Journal of Cosmetic Science, 2015, 37, 82–91

doi: 10.1111/ics.12173

Introduction

Emulsions are unquestionably among the most important of all the
colloid and macrodispersed systems, not only from a commercial
point of view, but also due to their great scientific interest [1]. They
are used in the food, pharmaceutical, petrochemical and cosmetic
industries [2

–6]. In the food industry, emulsions have the following

applications: low-calorie products, improved sensory characteristics
and taste masking. In the pharmaceutical field, emulsions are used
as drug delivery systems. In the cosmetic industry, they are used as
easily spreadable creams with encapsulated ingredients in both
water and oil phases [7].

Emulsions are dispersed, multiphase systems consisting of at

least two insoluble liquids. The two immiscible phases are usually
oil and water. The dispersed phase is present in the form of droplets
in a continuous phase. Depending on the emulsification process,
the diameter of the spherical droplets is between 0.1

lm and

0.1 mm. Emulsions of this kind are thermodynamically unstable,
which means that there is a tendency to reduce the interface, caus-
ing the droplets to coalesce and thus reducing the total amount of
interface [7]. There are mainly two types of emulsions: oil-in-water
(o/w), and water-in-oil (w/o). However, recently, there has been
increasing interest in the development of multiple or double emul-
sions, for example water-in-oil-in-water (W/O/W) and oil-in-water-
in-oil (O/W/O). For example, W/O/W emulsions consist of small
water droplets dispersed in larger oil globules, which are them-
selves dispersed in an aqueous continuous phase. Double emulsions
present many interesting possibilities for the controlled release of
chemical substances initially entrapped in the internal droplets,
which can have a significant role in the food industry (low-calorie
products, improved sensory characteristics, taste masking), cos-
metic industry (easily spreadable creams with encapsulated ingredi-
ents in both water and oil phases), pharmaceutical industry (drug
delivery systems) and other fields such as agriculture and the
production of multicompartment microspheres [8, 9].

Kinetically stable emulsions can be formed by adding emulsifiers

and/or thickening agents (proteins and hydrocolloids) to overcome
the activation energy of the system [10

–12]. The conventional

explanation for emulsion stabilization by such agents is their accu-
mulation at the oil

–water interface in the form of a densely packed

layer, which may prevent droplet flocculation and coalescence by a
steric mechanism [12, 13].

Monoacylglycerols and their derivatives have wide application as

emulsifiers in the food, pharmaceutical and cosmetic industries.
Acylglycerol emulsifiers (mainly employed as W/O stabilizers) are
used in food in all countries, because they contain components
which can be metabolized. Their identity with natural lipid struc-
tures results in very favourable ecological properties. These deriva-
tives are obtained by the glycerolysis of triacylglycerols or by the
direct esterification of glycerol with fatty acids [14].

The aim of this work was to assess new formulations (emulsions)

containing interesterified fats. First, the new fats were formed via
enzymatic modification, then those fats were put into emulsions as
a fat base of the emulsions. On the basis of previously conducted
studies on chemical and enzymatic interesterification of fats such
as beef tallow and fractions of vegetable oils, it is known that
fats with a unique structure can be obtained by interesterification
[15

–17]. Using enzymatic interesterification, hard fats can be

enriched in monoenoic fatty acids, or polyenic fatty acids from oils
[16

–18]. For this purpose, vegetable oil (such as walnut oil) rich in

unsaturated fatty acids and mutton tallow as a hard fat were used

as substrates during interesterification. The obtained new fats with
interesting characteristics were used as a fat base in the proposed
cosmetic emulsions. Moisturizing and sensory properties of the syn-
thesized emulsions were studied by the techniques below.

The novelty in this study was the application of interesterified

fats as the fatty base in cosmetic emulsions. Fats of this type have
mainly been used by the food industry before.

Materials and methods

Materials

Walnut oil (WO) was purchased at a local market. The composition
of walnut oil’s fatty acids was as follows: C16:0 (7.3%), C16:1
(0.2%), C 18:0 (2.5%), C 18:1 cis 9 (19.8%), C 18:2 n-6 (58.0%),
C 18:3 n-3 (12.3%).

Mutton tallow (L) was laboratory-refined, bleached and deodor-

ized under a vacuum at 105

°C. Its main fatty acid composition was

as follows: C 14:0 (1.8%), C 16:0 (18.4%), C 16:1 cis 9 (0.8%),
C17:0 (1.9%), C 18:0 (29.2%), C 18:1 cis 9 (31.6%), C 18:2 all cis
(1.7%), C18:2 cis-9, trans-11 (0.8%), C 18:3 all cis (0.6%), C 20:0
(0.3%), C 20:1 cis 11 (0.1%), total saturated FA (SFA) (52.0%), total
monounsaturated FA (MUFA cis) (34.1%), total polyunsaturated FA
(PUFA cis) (2.2%), total n-3 (0.6%) and total n-6 (1.6%).

As a biocatalyst during enzymatic interesterification, Lipozyme

RM IM (Novozymes, Bagsvaerd, Denmark) was used. The commer-
cial Lipozyme RM IM preparation contains immobilized lipase from
Rhizomucor miehei and contains 4 w-% of water. Activity of Lipo-
zyme RM IM is 5

–6 BAUN/g.

Carboxymethylcellulose was obtained from Mikro-Technik GmbH

& Co. KG, B

€urgstadt, Germany. It was used as a thickener for

emulsion solutions.

Sunflower lecithin was obtained from Lasenor Emul, S.L. Barce-

lona, Spain. It was used as an emulsifier.

Methods

Enzymatic interesterification of fats
The process of interesterification was carried out according to our
own specifications and previous experience [2, 15, 16].

Mutton tallow was mixed at 70

°C under nitrogen with walnut

oil (L : WO) in the proportions 2 : 3 (w/w). Flasks containing fat
blends were placed in a thermostated mineral oil shaker bath (type
357, producer Elpin plus, Lubawa, Poland). After thermal equili-
bration of the fat blend at the desired temperature of 60

°C, 8 w-%

of Lipozyme RM IM was added to the blend. To obtain a greater
quantity (MAGs and DAGs

– polar fraction responsible for good sta-

bility of emulsions) in the reaction environment via hydrolysis,
water was added (11, 13, 14 and 16 w-%) to the enzymatic prepa-
ration. That quantity of MAGs and DAGs was desired because
those fats with a higher quantity of the polar fraction are used in
emulsion systems as natural emulsifiers. The interesterification was
performed with continuous shaking for 6 h. After a predetermined
time of interesterification, the samples were filtered to stop the
reaction.

The composition of fatty acids (FA) of walnut oil and mutton tal-

low was determined using gas chromatography (GC) after conver-
sion of the fats to fatty acid methyl esters (FAMEs). They were
analysed using a Hewlett Packard model HP 6890 gas chromato-
graph with HP-88 capillary column (88% cyanopropyl aryl-poly-
siloxane,

100 m

9 0.25 mm 9 0.25 lm film thickness from

© 2014 Society of Cosmetic Scientists and the Soci
et
e Franc
aise de Cosm
etologie

83

International Journal of Cosmetic Science, 37, 82–91

Interestrified fats suitable for cosmetic emulsions

M. Kowalska et al.

Agilent). The temperature of the injector was 280

°C and of the

flame ionization detector (FID) 290

°C. The temperature of the col-

umn was in the range from 60 to 230

°C, at 4°C min

1

. The flow

rate of carrier gas was 1.2 mL min

1

. Helium was used as the car-

rier gas. Identification of the fatty acids was carried out by compar-
ison of the retention times with a standard mixture of fatty acid
methyl esters. GC of the FAMEs was performed according to the
ISO standard.

Polar fraction content was determined using a column chroma-

tography on silica gel, SG 60, 70

–230 mesh, Merck, Darmstadt,

Germany, according to the ISO standard [19].

Emulsion preparation and analysis

Emulsions were prepared according to the specifications shown in
Table I. They were prepared in the same way by adding together
the water with thickener and the fat phase and stirring with the
mixer RW 20 DZM (by Janke & Kunkel, Staufen im Breisgau, Ger-
many) for 4 min with the velocity of 38 332 relative centrifugal
force (RCF). Emulsions were prepared at room temperature (22

°C).

The volume of each mixtures was 100 g.

Determination of skin humidification

Skin humidification was measured by means of a CM 825 Corne-
ometer by Courage

+Khazaka Electronic [20, 21]. Respondents gave

their written consent for taking measurements of functional param-
eters of the skin before starting the study. Testing was carried out
on 83 women aged 30

–45. To have valid calculation, the reading

of skin humidification was taken five times. Some of the women
were educated cosmetology graduates from the University of

Technology and Humanities in Radom, Poland. The remaining
women

represented

the

profession

of

pharmacy

technician.

Research was conducted under the control of a trained person who
helped during the whole period of the research. Research for all
volunteers lasted for 6 weeks (17 February 2014 to 30 March
2014). The effect of the cosmetic emulsions produced on humidifi-
cation of cuticular cornea was evaluated. Testing began on clean,
degreased skin, which constituted the point of reference and zero
time of the cosmetic effect on the skin. Approximately 0.01 g of
the emulsion was then applied to the same 20

9 20 mm fragment

of the forearm skin and left there for 5 min. The probe of the cor-
nea before and after the measurement was always wiped with a
clean cotton cosmetic swab. The region of the skin before and after
each measurement was wiped with a clean, dry cotton cosmetic
swab too. Humidification was measured at 22

 1°C after 5 min

and at intervals of 15 min until 90 min after the moment the skin
was cleaned of the emulsion. Variations of skin humidification were
calculated by a formula, averaging results for all subjects involved
in the test:

ZN

t

¼

N

t

N

PKt

N

PKt

 100%;

where, ZN

t

is change of skin humidification over time ‘t’; N

t

is

average skin humidification after ‘t’ for a place with the tested cos-
metic on; N

PKt

is average skin humidification after ‘t’ for the con-

trol point.

Sensory determination

Consumer testing continued for 2 weeks in the period 02 to
16 March 2014 and was conducted three times a week. Therefore,
the volunteers (83 females) were able to use the emulsions several
times before the final evaluation. Six cosmetic emulsions were
subjected to the sensory analysis. Five emulsions contained inter-
esterified fats, and the reference emulsion was a mix of non-inter-
esterified fats (Table I). The emulsions were assessed for the
following characteristics: consistency (density and cohesion of the
tested cosmetic), homogeneity (behaviour of the preparation when
applied to the skin

– absence of clots or air bubbles), cushion effect

(palpability of the substance when rubbed between fingers), distri-
bution (facility of spreading on the skin surface), smoothing
(smoothing effect when applied to the skin), viscosity (degree of pal-
pable viscosity left on the skin), greasiness (a fat film remaining on
the skin) and absorption (rate of absorption by the skin). Each
characteristic was graded on a scale of 1

–5, with 1 the minimum

and 5 the maximum score. For ease of organoleptic testing, each
survey was enclosed with guidelines for sensory analysis (Table II)
describing the test procedure and the scoring scale [23, 24].
Sensory analyses were conducted in the presence of the person
responsible for sensory assessment (proper behaviour during mea-
surements). In addition, the survey was supplemented with a ques-
tion concerning the selection of the emulsion, which was the most
suitable for the respondents.

Determination of emulsion stability using the centrifugal test

Determination was measured in the centrifugal machine at
1008 RCF. Test tubes were filled with 15 mL of the emulsion and
then centrifuged for 30 min, with the state of emulsion checked
every 10 min. If the emulsion remained homogeneous after
30 min, it was considered to have proper stability.

Table I Composition and parameters of emulsions

Component [%]

Type of emulsion

E I

E II

E III

E IV

E V

E VI

Sunflower lecitin

5.0

5.0

0

0

0

0

Content of polar

fraction(MAGs, DAGs, FFA)

3

a

14

b

23

c

26

d

27

d

32

e

*

Carboxymethyl-cellulose

1.0

Fat Blend

30.0

Sodium benzoate

0.20

Water

Up to 100%

Parameter
Mixing speed, [RFC]

38 332

Mixing time, minutes

4

Legendary: E I: Emulsion containing initial blend (L : WO; 2 : 3, w/w)

– non-in-

teresterified blend of fats; E II: Emulsion containing fat blend (L : WO; 2 : 3, w/
w) interesterified by Lipozyme IM containing 4% water in the enzymatic prepara-
tion; E III: Emulsion containing fat blend (L : WO; 2 : 3, w/w) interesterified by
Lipozyme IM containing 11% water in the enzymatic preparation; E IV: Emulsion
containing fat blend (L : WO; 2 : 3, w/w) interesterified by Lipozyme IM contain-
ing 13% water in the enzymatic preparation; E V: Emulsion containing fat blend
(L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 14% water in the
enzymatic preparation; E VI: Emulsion containing fat blend (L : WO; 2 : 3, w/w)
interesterified by Lipozyme IM containing 16% water in the enzymatic prepara-
tion. Different letter in lines indicate mean values that differ statistically signifi-
cantly (p<0.05).

84

© 2014 Society of Cosmetic Scientists and the Soci
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aise de Cosm
etologie

International Journal of Cosmetic Science, 37, 82–91

Interestrified fats suitable for cosmetic emulsions

M. Kowalska et al.

Determination of emulsion stability using the temperature test at
35 and 3

°C

All emulsions were stored in the dryer at 35

°C (0.3°C) and in the

refrigerator at 3

–5°C alternately for 5 days, with a change every

24 h. If the emulsion remained homogeneous in such conditions, it
was considered to have proper stability.

Determination of mean particle size and particle size distribution of
fat emulsions

Determination was carried out in the range 0.12

–704.0 lm by

laser scattering using a Microtrac Particle Size Analyzer (Leeds &
Northrup, Philadelphia, U.S.A.) after preparation. The emulsions
were diluted 1 : 200 with distilled water. Determination was car-

ried out 48 h from the emulsion manufacturing. The measure-
ments were taken three times.

Viscosity examination

The viscosities of the samples were determined at 25

°C using spin-

dle no. 1 and no. 2 in a Brookfield Rheometer DV-I

+ 48 h after

their production.

Determination of type of emulsion

Conductivity of determined emulsions was measured by microcom-
puter pH/conductivity meter CPC-551 ELMETRON with chloride
electrode EClL-305W EUROSENSOR-GLIWICE, Poland. Conductivity
of the all tested emulsions was determined.

Table II Guidelines for sensory analysis of the cosmetic emulsions tested (study based on literature [22, 23] and own experience)

Feature

Description of test procedure

Score (1–5)

C

Position hand at an angle of 60

° and place 5 cm

3

of the

test substance there. Proceed to analyse its consistency by
assessing the ability to keep the cosmetic adhering to the
hand.

5. Cosmetic is easy to apply, not flowing
4. Easy to apply yet flowing can be observed
3. Cosmetic is hard to apply
2. Too thick to apply to the hand
1. Impossible to apply

H

Spread the substance on your hand and assess

smoothness of its layer, presence of clots or air bubbles.

5. Completely homogeneous, no clots or air bubbles, forms a smooth layer on the

skin when applied

4. Homogeneous, no clots and few air bubbles, forms an uneven layer
3. Observable and palpable clots and air bubbles in the substance and on the

skin when applied

2. Heterogeneous
1. Formulation components are not dissolved.

CE

Scoop 0.5 cm

3

of the emulsion and rub between the thumb

and index finger.

5. Imperceptible substance
4. Weakly perceptible substance
3. Somewhat perceptible substance
2. More perceptible substance
1. Highly perceptible substance

D

Spread 0.5 cm

3

of the preparation on the forearm skin

and observe its resistance to spreading.

5. No resistance to spreading
4. Little resistance to spreading
3. Incomplete cover, good spreading
2. Difficult to spread
1. Impossible to spread

SM

Apply 0.5 cm

3

of the emulsion on the cleaned forearm

skin and after an hour appraise the skin’s smoothness
in reference to a standard to which the substance has
not been applied.

5. Very smooth, soft skin surface
4. Smoother and softer skin surface than of the reference standard
3. The skin surface is as smooth as that of the reference standard
2. Rough skin
1. Very rough skin

ST

Apply and spread the emulsion on the cleaned forearm

skin, then press the other hand against this skin section
and assess viscosity.

5. No palpable skin viscosity
4. Low skin viscosity
3. Palpable skin viscosity
2. Increased skin viscosity
1. High skin viscosity

G

Apply 0.5 cm

3

of the substance on the cleaned forearm

skin and assess formation of a greasy film.

5. No sense of grease or film formation on the skin after application
4. Weak sense of greasiness, no film on the skin
3. Thin, greasy film on the skin after application
2. Greasy film on the skin directly on application
1. A compact, greasy film after application

A

Apply the substance on cleaned skin and assess the time

of its absorption.

5. Very good absorption below 30 s
4. Good absorption from 30 s to 1 min
3. Average absorption from 1 to 3 min
2. Poor absorption from 3 to 5 min
1. Very poor absorption for more than 5 min

C, Consistency; H, Homogeneity; CE, Cushion effect; D, Distribution; SM, Smoothing; ST, Stickiness; G, Greasiness; A, Absorption.

© 2014 Society of Cosmetic Scientists and the Soci
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e Franc
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etologie

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International Journal of Cosmetic Science, 37, 82–91

Interestrified fats suitable for cosmetic emulsions

M. Kowalska et al.

Morphology of emulsions

Optical microscopic observation of the emulsions was done using a
usual optical microscope (Nikon

– Polska, production PZO, lens

Nikon 4 4/0.10 (160/-WD 25), ocular Nikon China CF WE 15/12,
camera Panasonic type GP-KR 222, software

MICROSCAN

for Win-

dows). An appropriate amount of freshly prepared emulsion was
placed onto the microscopic slide. A cover slip was placed on the sam-
ple, ensuring that no air or bubbles were trapped between the sample
and cover slip, and the samples were tested with a 40

9 objective.

Statistical analysis

The results were subjected to one-way ANOVA. The Duncan test
was used to assess the differences between means. The level of

P

< 0.05 was considered significant.

STATGRAPHICS

plus 4.0 package

(Statistical Graphics Corp., Warrenton, VA, U.S.A.) was used.

Results and discussion

Analysing the interesterification process, it was found that new a-
cylglycerols were formed. Generally, FFA, MAG and DAG contents
increased compared to the starting blend. The highest value for
MAG and DAG contents was observed for the fat blend interesteri-
fied by Lipozyme RM IM with 16% water in the enzymatic prepara-
tion (Table I). Consequently, addition of water to the enzymatic
preparation generated a greater quantity of the polar fraction in
the interesterified blends. Appearance of MAGs and DAGs during
interesterification is very important because they are seen as good
emulsifiers [14].

Figure 1 Changes of the hydration degree of the skin with the passage of time. Legendary: E I: Emulsion containing initial blend (L : WO; 2 : 3, w/w) – non-
interesterified blend of fats; E II: Emulsion containing fat blend (L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 4% water in the enzymatic prep-
aration; E III: Emulsion containing fat blend (L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 11% water in the enzymatic preparation; E IV:
Emulsion containing fat blend (L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 13% water in the enzymatic preparation; E V: Emulsion contain-
ing fat blend (L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 14% water in the enzymatic preparation; E VI: Emulsion containing fat blend
(L : WO; 2 : 3, w/w) interesterified by Lipozyme IM containing 16% water in the enzymatic preparation.

Table III Mean values of sensory assessment obtained as a result of the survey

Feature

Type of emulsion*

E I

E II

E III

E IV

E V

E VI

C

5.0

 0.75

4.8

 0.093

4.3

 0.27

4.5

 0.35

4.3

 0.37

4.5

 0.34

H

5.0

 0.121

5.0

 0.87

5.0

 0.89

4.8

 0.27

5.0

 0.45

5.0

 0.26

CE

5.0

 0.71

4.8

 0.23

4.8

 0.64

4.8

 0.64

4.2

 0.74

4.8

 0.47

D

5.0

 1.17

5.0

 0.28

5.0

 0.26

5.0

 0.1.02

5.0

 0.28

4.7

 0.36

SM

5.0

 1.02

5.0

 0.73

5.0

 0.34

5.0

 0.74

5.0

 0.54

5.0

 0.65

ST

4.5

 1.04

4.0

 0.63

5.0

 1.02

4.8

 0.27

4.7

 0.48

4.5

 0.50

G

4.0

 0.82

4.8

 0.81

4.3

 0.24

4.5

 0.47

4.5

 0.50

4.3

 0.26

A

4.0

 0.76

4.5

 0.27

4.3

 0.54

4.7

 0.38

4.8

 0.93

4.5

 0.34

C, Consistency; H, Homogeneity; CE, Cushion effect; D, Distribution; SM, Smoothing; ST, Stickiness; G, Greasiness; A, Absorption.
*See legends of E I to E VI is given in Table I.

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Skin care formulations, that is cosmetic and dermal products,

form a distinctive group among emulsions, because they must meet
a whole series of requirements. They must have a pleasing appear-
ance and retain it during storage, give an agreeable feeling during
application and, most importantly, provide long-term beneficial
effects to the skin [25]. It is important that cosmetic emulsion-type
topical products must provide skin hydration in both healthy and
diseased conditions [26].

Assays of humidification properties of the emulsions found the

greatest average humidification (63%) on application of emulsion

VI (Fig. 1). Emulsions IV and V exhibited somewhat lower values in
the range 50

–60%. Application of emulsion II generated minimum

humidification, whereas skin covered with emulsions I and III dis-
played a relatively high degree of humidification, that is, nearly
60%, five minutes after application. It can be assumed that the
greater amount of natural emulsifiers in the emulsion could influ-
ence the higher degree of skin humidification. It seems that their
presence in the emulsion is desirable due not only to their stabilizing
properties but also their humidification properties. The humidifica-
tion was observed to decline after successive intervals, however, to

Figure 2 Sensory profile of emulsions E I to E VI. See legends of E I to E VI is given in Fig. 1.

© 2014 Society of Cosmetic Scientists and the Soci
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M. Kowalska et al.

reach 30

–40% after 90 min (Fig. 1). The degree of skin humidifica-

tion was clearly at a minimum after application of emulsion II.
Results of the sensory evaluation are presented in Table III. Accord-
ing to the results shown in Table III and Fig. 2, emulsion E IV had
optimal properties. The field formed by its average assessments most
closely approximates ‘the spider web’, whereas the mean evalua-
tions generated for emulsion E1 create a field the most distant from

‘the spider web’. According to presented data, it appears that emul-
sion IV had a sufficient amount of natural emulsifiers which stabi-
lized this emulsion, and, simultaneously, the emulsion was
considered the best in terms of sensory properties. Additional emul-
sifiers present in emulsion VI increased the moisturizing properties
but did not increase the explanatory sensory emulsion.

It can be said that the respondents most highly valued the

following parameters: smoothing, homogeneity and spreading.
Their mean values were 5.0, 4.96 and 4.96 points, respectively.
Greasiness and skin absorption were rated the lowest, on the other
hand (4.37, 4.46 points).

Choice of an emulsion for everyday use was one of the questions

in the survey. Responses are illustrated in Fig. 3. It is clear that
most respondents (33%) chose emulsion IV for everyday skin care.
In the respondents’ opinion, this emulsion displayed characteristics
similar to those of the remaining substances, yet the skin was less
shiny than in the case of other emulsions.

According to the type, all tested emulsions were o/w emulsions.

After immersing the electrode in a drop of emulsion, one observed
deviation of the amperometer indicator, which confirmed the con-
ductivity of these layouts (Fig. 4). It was also confirmed by the
digestion test of emulsion drops in water. All emulsions in a few

Figure 3 Percentage of people reporting the daily use of the emulsion. See
legends of E I to E VI is given in Fig. 1.

Figure 4 Conductivity of studied emulsions 48 h after production. See legends of E I to E VI given is in Fig. 1.

Figure 5 Percentage of particles of given sizes and accumulated distribution in the emulsion: E I to E VI 48 h after production. See legends of E I to E VI is
given in Fig. 1.

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M. Kowalska et al.

minutes were solubilized in the water. It is believed that o/w emul-
sions generally show a fairly high rate of penetration into the skin,
are less greasy and form a protective film on the skin [5].

Generally, the temperature test was passed successfully by all

emulsions; however, emulsion III showed some delamination on
day three. Low quantities of air bubbles and development of a
heterogeneous layer could be observed in emulsions IV, V and VI.
These changes did not result in adverse assessments though. As
regards emulsions I and II, it was noticed that these emulsions
exhibited appropriate homogeneity, a smooth and uniform struc-
ture and absence of clots after each test cycle. Their high and
proper stability was reaffirmed by the centrifugal test. They were
the only dispersion systems to pass the test without altering their
structure. Certain changes suggestive of de-lamination

were

observed in the remaining emulsions after 20 min of the test. It
was particularly apparent in emulsion III. Finally, at the end of
the

test,

there

was

noticed

stratification

of

emulsion

III.

Remaining emulsions (IV, V, VI) retained the proper form to the
end of test.

According to authors [3], macroemulsions are emulsion systems

with droplet sizes ranging from 0.1 to 100

lm. In presented work

analysis of emulsion, particle sizes helped to observe that, except
emulsion III, the average particle size was in a relatively low range
of 3.3

–6.5 lm. This means that the emulsions stabilized with natu-

ral emulsifiers (IV, V, VI) displayed a stability comparable to that of
the emulsion containing lecithin. The accumulated distribution of
the dispersed phase particle sizes for emulsions IV, V and VI shows
that the diameter of 90% of droplets was below 5

lm. Their distri-

bution was quite narrow and monodisperse (Fig. 5). The diameter
of 90% of droplets was below 5

lm and the distribution of the dis-

persed phase particle sizes was similar to that of emulsions IV, V
and VI in the case of the emulsion stabilized with lecithin as well.
A marked system instability (presence of three fractions, high aver-
age particle size and a wide distribution of the dispersed phase

Figure 6 Average partice sie and viscosity of emulsion: E I to E VI 48 h after production. *Values of viscosity 910

2

. a,b,c: Different letters indicate mean val-

ues that differ statistically significantly (P

< 0.05). See legends of E I to E VI is given in Fig. 1.

Figure 7 Optical microscopic image of fresh emulsions E I – E VI. See legends of E I to E VI is given in Fig. 1.

© 2014 Society of Cosmetic Scientists and the Soci
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M. Kowalska et al.

particles) could be observed in the case of emulsion III (Figs 5 and
6). A small particle size could also be noted in emulsion II, with
a two-fraction particle distribution. Appearance of an additional
fraction could be presumed to be a symptom of non-adsorbed parti-
cles of an additional emulsifier in the continuous phase. That emul-
sion was stabilized with lecithin but also contained low quantities
of partial acylglycerols (MAGs and DADs) that arose in the process
of interesterification. Taking all these results into consideration, it
could be concluded that synergies between these two types of emul-
sifiers were absent. According to Dickenson [10], the presence of
such particles can affect the stability of an emulsion system.

Emulsions stabilized only with natural emulsifiers exhibited viscos-

ity of 1000 mPs

*s, whereas emulsions I and II displayed far greater

viscosity values. Emulsion III showed the minimal viscosity (Fig. 6).

Morphology of emulsions I and II exhibited the greatest systemic

homogeneity (Fig. 7). It seemed that their droplet size was compa-
rable and their arrangement suggested an orderly, homogeneous
nature. Larger isolated drops were observed in the remaining
systems of E IV, V, and VI. Perhaps, the presence of natural
emulsifiers

in

the

emulsion

requires

other

homogenization

conditions (a different magnitude of RCF or other homogenization
time). Therefore, further study is needed to learn how to avoid lar-
ger isolated drops before application of such emulsions as a com-
mercial product.

Such a merger into larger drops may initiate coalescence [27].

Generally, emulsion coalescence, in which some neighbouring
droplets form larger droplets, damages an original emulsion and is
indicative of emulsion instability [28]. Morphology of emulsion III
pointed to a distinctly larger particle, possibly responsible for
reduced stability of the system (Fig. 7).

Conclusions

In the results of the presented work on the stability of emulsions
I-VI,

an

unsatisfactory

polar

fraction

in

emulsion

III

was

observed. It was the only emulsion that did not completely pass
the tests for stability, and its morphology indicated larger parti-
cles than in the other emulsions. Their average particle size had
the greatest value, and the distribution was very wide. It can be
claimed that the content of the polar fraction in emulsion III was
insufficient to stabilize that dispersion system. Remaining emulsions
showed proper stability. Their average particle size and distribution
resemble those of a monodispersive system. Respondents character-
ized emulsions as easy to scoop, not flowing or pouring around the
skin. Emulsions IV, V and VI displayed cohesion and their consis-
tency was that of cosmetic milk. Only emulsions I and II showed
thick consistency; therefore, the respondents classified them as
creams. In general, all the emulsions were absorbed well and fast,
and the skin after 1 h of application displayed a palpable greasy
deposit of the substance. Ninety percent of respondents assessed the
skin to which the emulsions had been applied as smooth and pleas-
ant to touch.

In summary, the work has demonstrated the possibility of apply-

ing interesterified fats as the fatty base in cosmetic emulsions.
Emulsions formed on the basis of interesterified fats without any
additional emulsifiers (sunflower lecithin) had properties compara-
ble to emulsions containing mixed non-interesterified fat. It should
be pointed out that the emulsions based on interesterified fats
exhibited an even greater level of moisturization of the skin than
those containing non-interesterified fat. The natural emulsifiers
formed as part of interesterification allowed us to obtain stable
emulsion systems while giving the respondents the satisfaction of a
new product not yet available on the market.

Acknowledgements

The authors wish to acknowledge Kazimierz Pulaski University of
Technology and Humanities in Radom, Poland for financial
support.

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