Journal of Microencapsulation

Micro and Nano Carriers

ISSN: 0265-2048 (Print) 1464-5246 (Online) Journal homepage: https://www.tandfonline.com/loi/imnc20

Characterisation of peppermint (Mentha piperita

L.) essential oil encapsulates

Murat Yilmaztekin, Steva Lević, Ana Kalušević, Mustafa Cam, Branko

Bugarski, Vesna Rakić, Vladimir Pavlović & Viktor Nedović

To cite this article: Murat Yilmaztekin, Steva Lević, Ana Kalušević, Mustafa Cam, Branko
Bugarski, Vesna Rakić, Vladimir Pavlović & Viktor Nedović (2019) Characterisation of peppermint
(Mentha�piperita L.) essential oil encapsulates, Journal of Microencapsulation, 36:2, 109-119, DOI:
10.1080/02652048.2019.1607596

To link to this article: https://doi.org/10.1080/02652048.2019.1607596

Accepted author version posted online: 13

Apr 2019.
Published online: 29 Apr 2019.

Submit your article to this journal

Article views: 147

View related articles

View Crossmark data

Full Terms & Conditions of access and use can be found at

https://www.tandfonline.com/action/journalInformation?journalCode=imnc20
JOURNAL OF MICROENCAPSULATION
2019, VOL. 36, NO. 2, 109–119
https://doi.org/10.1080/02652048.2019.1607596

ORIGINAL ARTICLE

Characterisation of peppermint (Mentha piperita L.) essential oil

encapsulates
Murat Yilmaztekina, Steva Levi
cb, Ana Kalusevi
cb, Mustafa Camc, Branko Bugarskid, Vesna Raki
cb,
Vladimir Pavlovi
cb and Viktor Nedovi
cb
a
Department of Food Engineering, Faculty of Engineering, Inonu University, Malatya, Turkey; bDepartment of Food Technology and
Biochemistry, Faculty of Agriculture, University of Belgrade, Belgrade, Serbia; cDepartment of Food Engineering, Faculty of
Engineering, Erciyes University, Kayseri, Turkey; dDepartment of Chemical Engineering, Faculty of Technology and Metallurgy,
University of Belgrade, Belgrade, Serbia

ABSTRACT ARTICLE HISTORY

Aim: The aim was to choose the optimal encapsulation system and to incorporate encapsulates Received 23 November 2018
loaded with essential oil into the ice cream as a model food product. Accepted 8 April 2019
Methods: Ca-alginate beads were produced by electrostatic extrusion process. Gelatine/alginate
KEYWORDS
coacervates were processed with coacervation. Carnauba wax microparticles were produced
Mentha piperita L. essential
using melt dispersion process. Morphological properties, chemical, and thermal stabilities of oil; encapsulation; ice
encapsulates were tested by SEM, FTIR spectral, and thermogravimetric analysis. cream; sensory evaluation
Results: Alginate provided sufficient emulsion stability over 1 h. Ca-alginate showed higher encap-
sulation efficiency (EE) (98.4 ± 4.3%) compared to carnauba wax (94.2 ± 7.8%) and gelatine/alginate
coacervates (13.2 ± 1.2%). The presence of essential oil in all three types of encapsulates confirmed
with FTIR. The encapsulation process ensured controlled release and thermal stability of the oil.
Conclusions: Ca-alginate matrix as the most suitable for peppermint essential oil encapsulation.
The sensory analysis showed that ice cream incorporating encapsulates is a promising system
for the consumption of health beneficial peppermint essential oil.

1. Introduction the structure of another material (shell) (Gibbs et al.

1999). Numerous encapsulation techniques, such as
Essential oils have a huge historical and practical sig-
spray drying, fluid bed coating, spray-cooling, melt
nificance for human civilization, primarily as therapeu-
tics for the treatment of various diseases, as well as injection, coacervation, and encapsulation in polymer
ingredients in cosmetics and food products (Burt 2004). microsphere have been developed for encapsulation
The peppermint is a rich source of valuable natural of food active compounds (Zuidam and Shimoni
compounds with antimicrobial, antioxidant, and antitu- 2010). One of the promising encapsulation methods is
mor activity (McKay and Blumberg 2006). Peppermint entrapping of the active compound inside the poly-
essential oil is traditionally produced and widely used mer matrix. Among natural polymers, alginate has
in medicine, massage and for personal hygiene (Ody been intensively used for encapsulation of food addi-
1993). Also, peppermint essential oils could be used as tive (Manojlovi
c et al. 2008) and essential oils (Chang
a source of monoterpenes that are suitable for the and Dobashi 2003, Lai et al. 2007). Further, the size of
treatment of some cancers types (Crowell 1997). alginate based encapsulates could be controlled using
Due to exposure to unfavorable environmental con- electrostatic droplet generation, enabling the produc-
ditions, such as high temperatures or oxygen, the tion of spherical calcium alginate particles suitable for
essential oils may lost their valuable ingredients and various biotechnological applications (Nedovi
c et al.
consequently their health benefits. Volatile and chem- 2001, Manojlovi
c et al. 2008, Levi
c et al. 2013).
ically unstable components of essential oils could be Another promising encapsulation technique for pro-
additionally protected by the process commonly tection and controlled delivery of essential oils is coac-
known as encapsulation. Encapsulation could be ervation. Encapsulates produced by coacervation (or
defined as entrapping of active compound (core) into simple coacervates) could be defined as an

CONTACT Murat Yilmaztekin [email protected] Department of Food Engineering, Faculty of Engineering, Inonu University, 44280
Malatya, Turkey
ß 2019 Informa UK Limited, trading as Taylor & Francis Group
110 M. YILMAZTEKIN ET AL.

encapsulation system produced by liquid-liquid phase Table 1. Chemical composition of peppermint essential oil.
separation of polymer(s) aqueous solution. By careful Compound %
adjustment of temperature and pH, it is possible to cre- a-pinene 7.55
camphene 0.23
ate polymer layer around dispersed oil droplets. After b-pinene 8.56
formation, coacervates are separated from the solution sabinene 0.24
b-myrcene 0.76
and could be further processed by crosslinking of shell limonene 9.50
material or processing coacervates by drying (Zuidam 1,8-cineole 0.88
b-cymene 0.27
and Shimoni 2010). Coacervation has been applied for 3-octanol 0.64
encapsulation of aromatic compounds using different p-menthone 14.89
b-bourbonene 0.83
natural shell materials. Jun-xia et al. (2011) showed that linalool 0.37
coacervation between soybean protein isolate and gum 1-octanol 0.51
menthyl acetate 8.54
arabic can be used for encapsulation of sweet orange oil isopulegol 4.30
into spherical shaped particles. Also, heat resistance of longifolene 0.19
essential oil could be improved using coacervates based a-ylangene 0.39
neomenthol 6.98
on gelatine/gum arabic (Lv et al. 2014). b-caryophyllene 1.31
Encapsulation of highly hydrophobic active materials, neoisomenthol 1.24
menthol 20.31
such as essential oils could be achieved by applying pulegone 1.83
hydrophobic shell material. This approach has been lavandulol 0.61
a-terpineol 6.09
considered for encapsulation of pharmaceutical com- piperitone 2.98
pounds inside hydrophobic natural compounds, such total 100
as waxes. Usually, preparation of wax particles are
based on the dispersion of molten mixture wax/active composition of essential oil determined by GC-MS is
compound in hot water followed with solidification by given in Table 1. The alginate (ALGOGELTM3001,
cooling and fast solidification of lipid carrier enables Cargill, Minnetonka, MN, USA) was generously
efficient entrapment of hydrophobic compounds inside donated by PALCO (Sabac, Serbia). Bovine gelatine
wax microparticles (Milanovi
c et al. 2010). (food grade) was supplied from the domestic market
The aim of this study was to investigate the possible in Belgrade (Serbia). Carnauba wax was supplied from
solutions for essential oils encapsulation. Peppermint Carl Roth GmbH (Karlsruhe, Germany). Sodium citrate
essential oil was used in this study as a model core and dichloromethane (GC grade) were obtained from
compound, while Ca-alginate, gelatine/alginate, and car- Merck (Darmstadt, Germany). All other chemicals were
nauba wax were carriers for encapsulation. In order to of analytical reagent grade and used without any fur-
eliminate water and provide better storage performance ther purification.
on encapsulates, the samples were freeze-dried. The
morphology, chemical composition and thermal proper-
2.2. Encapsulation of peppermint essential oil in
ties of encapsulates were studied in order to define the
Ca-alginate beads
optimal system for peppermint essential oil encapsula-
tion. Considering the potential health benefits of using The production of Ca-alginate beads was realized in
peppermint essential oil in the food industry, produc- two phases. The first phase was the preparation of
tion of encapsulates were optimized in that respect that liquid systems-emulsions that are used for beads pro-
the final encapsulates could be applied in various food duction. Prior to the preparation of emulsion, Na-algin-
products. Carnauba wax beads loading peppermint ate was dissolved in distilled water (concentration of
essential oil were chosen as optimal encapsulation sys- 0.02 g/ml). Further, different ratios of peppermint
tem and were incorporated into the ice cream as a essential oil to Na-alginate (10 and 20%, w/w oil in
model food product. Sensory analysis was carried out alginate solution) were mixed in order to produce sta-
on ice cream samples enriched with carnauba wax ble emulsions. The emulsions were prepared using
mechanical stirrer Ultra-TurraxV T25 (Janke and Kunkel
R
beads containing peppermint essential oil.
Ika-Labortechnik, Staufen, Germany) at 10 000 rpm for
2. Materials and methods 2 min. Emulsions prepared in this way remain stable
over the 1 h, which is a period of time enough for
2.1. Materials
emulsion extrusion and beads formation. The addition
Peppermint essential oil (Arifoglu brand) was supplied of emulsifiers was not necessary since alginate pro-
from a local market (Kayseri, Turkey). The chemical vided sufficient emulsion stability, which is in
 JOURNAL OF MICROENCAPSULATION 111

accordance with previously reported results (Levi
c washed with distilled water and freeze-dried as
et al. 2015). described above.
In the second phase, the Ca-alginate beads with
encapsulated peppermint essential oil were produced
by the procedure described previously by Nedovi
c
et al. (2001). Droplets were formed by extrusion of the 2.4. Encapsulation of peppermint essential oil in
sodium alginate/essential oil emulsion through a blunt carnauba wax
stainless steel needle using a syringe pump (Pump 11, Encapsulation of peppermint essential oil in carnauba
Harvard Apparatus, Cambridge, MA). The needle was wax was realized by the melt dispersion technique
grounded, while the collecting solution (CaCl2 in (Milanovi
c et al. 2010). Carnauba wax was melted in
water, 0.015 g/ml) was positively charged. Beads pro- distilled water at 95  C in a water bath. Peppermint oil
duction was carried out by applying electrostatic was added to the dispersion of molten wax in the
potentials of 8 kV. Electrostatic encapsulation unit (VAR water while stirring rigorously (10 000 rpm for 2 min)
V1, Nisco Engineering Inc., Zurich, Switzerland) was by a mechanical stirrer Ultra-TurraxV T25 (Janke and
R

used in this work for beads production under electro- Kunkel Ika-Labortechnik, Staufen, Germany). Different
static potential. The distance between the needle tip weight ratios of essential oil to wax (10, 20, 30, and
(id: 0.7 mm) and the collecting solution was 2.5 cm, 50%, w/w) were tested. The solidification of the molten
while the flow rate of the liquid was 70 ml/h. After the wax droplets with oil was performed by adding 150 ml
formation of the beads, they were left in hardening of cold water (5  C) to the resulting dispersion. After
solution without stirring for 60 min in order to assure the addition of cold water, the dispersion was cooled
the finishing of the gelling process. The formed algin- down and solid particles were formed. Further, the
ate beads were removed from the CaCl2 solution by
particles were collected by filtration, washed with dis-
filtration and washed with distilled water. After gel for-
tilled water and freeze-dried. All encapsulates were
mation period, the beads were frozen at
20  C over-
then stored at þ4  C in closed falcon tubes for fur-
night before freeze drying, which was carried out at
ther analyses.

50  C at a pressure of 1.1 Pa for 24 h in a freeze drier
(GAMMA Martin Christ, GmbH, Osterode am
Harz, Germany).
2.5. Analysis of the bead dimensions and shape
2.3. Encapsulation of peppermint essential oil in Dimension and shape of wet and dried Ca-alginate
gelatine/alginate coacervates beads, as well as carnauba wax particles, were eval-
Formation of coacervates loading peppermint essential uated by binocular microscope Leica XTL-3 400 D
oil was performed according to the procedure (Leica, Wetzlar, Germany), equipped with a camera
described by Siow and Ong (2013), with modifications. (DC 300, Leica, Germany) and software for measuring
The main modifications were the usage of Na-alginate (IM 1000, Leica, Wetzlar, Germany). For each measure-
as carrier material and citric acid for pH control ment, a sample of 30 beads was taken and the diame-
instead of gum arabic and acetic acid, respectively. ters of the beads were measured. The diameter for
Gelatine (concentration of 0.02 g/ml) and Na-alginate each particle was calculated as the average of the
solutions (concentration of 0.02 g/ml) were prepared horizontal and perpendicular dimensions (Levi
c
by dissolving these compounds in distilled water. An et al. 2015).
aliquot 25 ml of each gelatine and Na-alginate solu-
tions were mixed with 1, 2, and 4 g of essential oil at
45  C by a mechanical stirrer Ultra-TurraxV T25 (Janke
R

2.6. Scanning electron microscopy (SEM)

and Kunkel Ika-Labortechnik, Staufen, Germany),
respectively. A solution of 10% (w/v) citric acid was Morphological investigations of the surface of the
added to the mixture to adjust pH to 2–3. Such rela- samples have been carried out by JEOL JSM-6390LV
tively low pH was applied in order to ensure fast coac- scanning electron microscope (JEOL, Ltd., Tokyo,
ervates formation and to minimize the loss of Japan). Prior to analysis, samples were sputter-coated
essential oil during preparation steps. The mixture was with gold during 100 s under 30 mA ion current on
cooled down immediately to 10  C in an ice bath. BALTEC SCD 005 sputter coater (Capovani Brothers
Further, the coacervates were collected by filtration, Inc., Scotia, NY) (Levi
c et al. 2015).
112 M. YILMAZTEKIN ET AL.

2.7. Determination of encapsulation efficiency indice, I) of each compound. The identification was
made by comparison of the obtained mass spectrums
Encapsulation efficiency (EE) of encapsulates were cal-
of the relevant chromatographic peaks with spectrums
culated by the following equation as the percentage
of the National Institute of Standards and Technology
of the total amount of encapsulated essential oil (mE)
(NIST, Gaithersburg, MD) and Wiley libraries. In add-
in the total amount of initial input of essential oil (mI)
ition, tentative identification was carried out by com-
(Siow and Ong 2013, Levi
c et al. 2015):
  paring the experimental retention indices with the
mE
EEð%Þ ¼  100 (1) theoretical ones obtained from the NIST Standard
mI Reference Database (NIST 2013). Quantitative analysis
Quantification of essential oil content in encapsu- of essential oil components (expressed as area per-
lates was performed after extraction with dichlorome- centage) was carried out by peak area normalization
thane. Briefly, 0.5 g of lyophilized alginate encapsulate measurement. Menthol was identified by comparing
was dissolved in 40 ml of sodium citrate water solution the retention times and spectral data with those of
(1.5%, w/v) in a sealed glass bottle under vigorous the appropriate standard. The amount of essential oil
mixing using a vortex mixer. When encapsulates are in beads was calculated by the calibration curve
dissolved, 5 ml of dichloromethane was added and obtained from dichloromethane extracts of known
essential oil was extracted for 15–20 min with intermit- amounts of essential oil.
tent mixing. For carnauba wax and coacervates based
on gelatine, 0.5 g of encapsulate was extracted three 2.9. FTIR spectral analysis
times with 10 ml portions of dichloromethane. After
extraction, the organic phase was separated and sub- FTIR spectra of the freeze-dried encapsulates was
jected to GC-MS analysis. studied using IRAffinity-1 (Shimadzu, Kyoto, Japan)
FTIR spectrometer. Spectra were collected using KBr
pellets (1 mg of sample in 200 mg of KBr) in the spec-
2.8. GC-MS analysis of peppermint essential oil tral range 4000–600 cm
1. Spectra were recorded as
Gas chromatography-mass spectrometry (GC-MS) ana- average curves from 20 acquisitions, with the reso-
lysis was performed in order to determine the com- lution of 4 cm
1 (Levi
c et al. 2013). FTIR spectra of free
position of peppermint essential oil and to measure peppermint essential oil (2 ll) were recorded using
the concentration of the most represented compound, two blank KBr tablets that were used to create a thin
menthol, in dichloromethane extracts of encapsulates liquid sample film.
which is used for calculation of EEs. The analyses
were carried out on a Shimadzu QP 2010 Plus
2.10. Thermal analysis of the samples
(Shimadzu, Kyoto, Japan) GC equipped with an AOC-
20i/20s autosampler and a MS-QP 2010 series mass- Thermal analyses of freeze-dried samples were carried
selective detector. The conditions of GC-MS were out in a Setaram’s system TG/DSC111. The measure-
adapted from Yilmaztekin (2014). Sample (1 ll) was ments were realized under dynamic oxygen of a flow
injected with a split ratio of 1:50. Volatile compounds rate of 30 ml/min (pressure 1 atm) using a heating rate
were separated with a TRB-Wax (Teknokroma, of 10  C/min from 50 to 300  C.
Barcelona, Spain) fused silica capillary column
(60 m  0.25 mm i.d. and 0.25 lm film thickness).
2.11. Production of ice cream incorporated with
Helium was used as the carrier gas at a flow rate of
carnauba wax oil beads
1 ml/min. The column was maintained at 40  C for
5 min after injection, and then programmed at 3  C/ Due to the small size of the particles and relatively
min to 240  C, which was maintained for 15 min. The satisfactory EE, carnauba wax based encapsulates
total run time, including oven cooling, was 86 min. were chosen for incorporation into ice cream.
Injector, transfer line, and ion-source temperatures Although Ca-alginate capsules exhibited better EE (see
were maintained at 250  C. Mass spectrums were below), their size could negatively affect ice cream
acquired in electron-impact (EI) mode; the ionisation textural properties and overall sensorial acceptance.
voltage was 70 eV; the mass range was 35–450 m/z; Four types of ice cream were produced with the fol-
scanning rate was 1 scan/s. A mixture of n-alkanes lowing combinations according to Cam et al. (2013):
(C8–C28) was injected under the above conditions to Control ice cream (IC1), enriched with peppermint oils
calculate the linear retention indexes (as Kovats’ at 0.1% (w/w) (IC2), 0.2% (w/w) (IC3), and 0.3% (w/w)
 JOURNAL OF MICROENCAPSULATION 113

(IC4). The amount of peppermint oil in the samples strong presence of related attributes. The definitions
was based on dry matter content. As an example, ice of the descriptors and sample references are given in
cream ingredients for IC2 were mixed in the following Table 2.
combinations: A glass beaker containing 180 ml of
skim milk was placed on a thermostatically controlled
mechanical stirrer. Ingredients were added into the 2.13. Statistical analysis
beaker at 50  C with regular stirring at 1000 rpm. All chemical and instrumental analyses were done
Cream (40 g), skim milk powder (25 g), sucrose (32 g), minimum in triplicate. Ice cream formulations were
mono-diglyceride mix (0.5 g), sahlep as stabiliser prepared minimum in duplicate. Analysis of variance
(0.5 g) and encapsulated peppermint oil (0.1 g). The was performed to determine significant differences
mixture was pasteurised at 80  C for 10 min followed between the shell materials for bead sizes and EEs at
by cooled to þ4  C. After keeping the mixture at p  0.05 significant level using the SPSS version 17.0.1
þ4  C for 24 h the aged mixture was whipped at 0  C statistical package for Windows (SPSS Inc., Chicago, IL).
for 30 min with a laboratory type ice cream machine
(Gelato, Bologna, Italy). The resulting ice cream was
hardened at
18  C for 24 h in a deep freezer. All 3. Results and discussion
other formulations (IC1, IC3, and IC4) were prepared 3.1. Bead size, encapsulation loading, and
in the same way by changing the peppermint oil encapsulation efficiency of peppermint essential
content but keeping the amount of other ingre- oil encapsulates
dient’s constant.
Ca-alginate, gelatine/alginate coacervates, and car-
nauba wax microparticles were used as carriers for
2.12. Quantitative descriptive sensory analysis of encapsulation of peppermint essential oil. Table 3
ice cream incorporated with carnauba wax
oil beads Table 3. Average bead size and encapsulation efficiency of
peppermint essential oil encapsulates.
Six non-smoking panelists had previous experience in
Encapsulation
testing ice cream (3 female and 3 male, ages between Preparation Bead size (lm) efficiency (%)
26 and 39) from Department of Food Engineering in Calcium alginate beads
Erciyes University were selected for the evaluation of A1 693.5 ± 133.6a 98.4 ± 4.3a
A2 827.2 ± 121.5a 96.5 ± 5.5a
the sensory attributes of the ice creams. The panelists Gelatine and alginate coacervates
were trained until they fully understood the sensory C1 – 13.2 ± 1.2c
C2 – 11.8 ± 1.0c
terms of ice creams and showed consistent replicates. C4 – 10.5 ± 0.8c
Reference sample preparation and administration of Carnauba wax beads
CW1 27.6 ± 15.4b 66.8 ± 4.7b
the panel were conducted according to relevant litera- CW2 37.1 ± 18.7b 72.9 ± 5.2b
ture (Hightower and Edgar 2009, Soukoulis and Tzia CW3 21.7 ± 10.9b 69.6 ± 5.8b
CW4 30.4 ± 14.5b 94.2 ± 7.8a
2010, Cam et al. 2013).
A1: 2 g oil þ 18 g sodium alginate, A2: 4 g oil þ 16 g sodium alginate,
Flavour descriptors of ice creams were selected as: C1: 1 g oil þ 50 g polymer, C2: 2 g oil þ 50 g polymer, C4: 4 g oil þ 50 g
palatable, sweet, sour, astringent, and peppermint fla- polymer, CW1: 1 g oil þ 9 g carnauba wax, CW2: 2 g oil þ 8 g carnauba
wax, CW3: 3 g oil þ 7 g carnauba wax, and CW4: 5 g oil þ 5 g carnauba
vour. A 10-point scale was used for evaluations, where wax. In the same column, means with different letters significantly dif-
0 for non-existence, 5 for imperceptible, and 10 for fered (p < 0.05).

Table 2. Descriptive terms, definition, and reference material used for the training of the panelists .a
Sensory attribute Definition Reference material
Overall acceptability Ice cream has no defects. Sensorial properties are well balanced. A commercial ice cream (Algida Maraş Usul€u,
Istanbul, Turkey)
Sweet The intensity of flavour caused by sucrose is clearly perceivable 2.0% (w/v) sucrose solution
Sour It is characterised by a refreshing and fruity flavour. A commercial sorbet (Algida, Istanbul, Turkey).
Astringent Astringency sensation is dry and gives a puckery feeling in 0.5% (w/v) punicalagin solution.
the mouth.
Oxidised Off-flavour is characterised by a cardboard, papery, and Cream was oxidised at 105  C for 2 h and an ice
tallow flavour and smell. cream mix was prepared with oxidised cream.
Overall peppermint flavour Sweet, green, earth, sharp, mentholic aromatics associated lu Baharat, Istanbul, Turkey)
Peppermint oil (Arifog
with peppermint oil.
a
Sensory attributes, definitions, and reference samples were based on the literature (Hightower and Edgar 2009, Soukoulis and Tzia 2010, Cam
et al. 2013).
114 M. YILMAZTEKIN ET AL.

shows the average size and EEs of peppermint drying process. It seems that solid fillers (i.e. active
essential oil encapsulates. compounds) are more suitable for shape stabilisation
As can be seen, Ca-alginate as carrier material of encapsulate compared to liquid active compounds.
showed higher EE (98.4 ± 4.3%), followed by carnauba In some cases, preservation spherical shape of encap-
wax microparticle (94.2 ± 7.8%), while gelatine/alginate sulate is favorable (e.g. positive visual effects on con-
coacervates exhibited the lowest EE (13.2 ± 1.2%). sumers). Introduction of additional filer in formulation
These results are in accordance with the results from could be solutions for the problem of shape mainten-
other authors who also reported high EE of Ca-algin- ance during drying of alginate based spherical beads.
ate and carnauba wax as carriers for encapsulation of Chan et al. (2011) proposed using starch as a filler
hydrophobic compounds (Milanovi
c et al. 2010, Levi
c substance for shape preservation of lyophilised Ca-
et al. 2015). Evidently, gelatine/alginate coacervates alginate beads loading cells.
are far less suitable for encapsulation of peppermint As we pointed out above, freeze-dried coacervates
essential oil regarding EE. This is in contrast to results (Figure 1(c,d)) show significant changes in morpho-
reported in other studies that usually showed high EE logical properties during the drying process. Namely,
of coacervates loading essential oils and aromatic com- after encapsulation, coacervates remained free-flowing
pounds (Manaf et al. 2018). Low EE of the gelatine/ in working solutions and were easily collected. During
alginate coacervates prepared in this study could be the drying process, the structure of coacervates
explained by repulsive forces between hydrophilic car- changes towards compact-flakes like structure. On the
riers and essential oil as well as the complex structure other hand, microparticles based on carnauba wax
of peppermint essential oil. Also, McClements and Rao (Figure 1(e)) show more regular spherical shape com-
(2011) pointed out that the formation of a stable emul- pared to other formulations, although their sizes were
sion of essential oil is a difficult process since smaller not uniform. According to Milanovi
c et al. (2010), the
oil droplets tend to coalescence into larger droplets, size of carnauba wax microparticles produced by melt-
leading to emulsion breakdown. Furthermore, add- ing dispersion technique could be controlled by the
itional processing of coacervates, such as drying may variation of process conditions. Compared to other
cause further loss of essential oil. This was observed by formulations (i.e. Ca-alginate and gelatine/alginate
Shinde and Nagarsenker (2011) for the gelatine/algin- coacervates) carnauba wax particles exhibit significant
ate coacervates loading eugenol. The same authors surface porosity (Figure 1(f)). The porosity of wax
reported similar EE as we observed in this study, and microparticles is most probably a consequence of
as the main reasons for low retention of eugenol, they essential oil addition that causes changes in particles
suggested the dehydration of coacervates which structure during encapsulation. The irregular surface of
caused structural changes of particles causing carnauba wax particles was also observed by
eugenol leakage. Milanovi
c et al. (2010).
The particles of Ca-alginate are larger compared to
wax particles, while gelatine/alginate coacervates after
3.3. The results of FTIR analysis
drying were in the form of aggregated mass that
could not be studied for their size. The FTIR spectra of the free peppermint essential oil
and encapsulate loading oil are presented in Figure 2.
In the spectrum of free essential oil, the strong band at
3.2. Morphological properties of peppermint
3470 cm
1 is assigned to the vibrations of the
OH
essential oil encapsulates
groups, while bands at 2924 and 2854 cm
1 appeared
Morphological properties of encapsulates loading due to the presence of the
CH3 and
CH2 groups.
peppermint essential oil were studied by scanning The bands in the spectral range 1700–1750 cm
1 are
electron microscopy and results are presented in most probably related to C¼O vibrations. Other bands
Figure 1. After encapsulation and gelling process, wet such as those at 1460 and 1377 cm
1 originate from
Ca-alginate beads exhibited spherical shape. However, various organic compounds, which presence in the
during the freeze-drying, the water was removed from peppermint essential oil was verified by GC/MS analysis
the beads and the alginate matrix shrank significantly (see Table 1). Prakash and Yunus (2013) showed that
(Figure 1(a,b)). vibrations assigned to menthol and menthone (in
According to Levi
c et al. (2013) addition of solid the spectral range 2849–2954 cm
1), dominate in the
flavour into the alginate matrix provide better preser- FTIR spectrum of essential oil produced by hydro-distil-
vation of beads shape and sphericity during the lation of Mentha arvensis. We also observed similar
 JOURNAL OF MICROENCAPSULATION 115

Figure 1. SEM images: A2 (a-low magnification; b-high magnification), C4 (c-low magnification; d-high magnification), and CW4
(e-low magnification; f-high magnification).

prominent bands in the same spectral region of both intensities might be due to carrier/oil bands overlapping.
free and encapsulated peppermint essential oil. Also, These findings clearly show that essential oil and carrier
the results of GC/MS analysis of Mentha piperita L. materials most probably made a simple mixture in the
essential oil conducted in this study suggest that these encapsulates, since there is no strong evidence of chem-
two compounds are present in the high concentration. ical interactions between peppermint essential oil com-
The results of FTIR analysis show the presence of pounds and carrier materials.
essential oil in all three types of encapsulates produced
in this study. Moreover, the spectra of carriers (i.e. Ca-
3.4. The results of thermogravimetric analysis
alginate, gelatine/alginate coacervates, and carnauba
wax) are generally overlapped by the spectrum of essen- Thermal stability of free and encapsulated pepper-
tial oil. Some relatively small variations in the spectra of mint essential oil was investigated by thermogravim-
encapsulate, such as differences in bands shapes or etry and results are presented in Figure 3. Free
116 M. YILMAZTEKIN ET AL.

peppermint essential oil shows first mass loss in the two steps in the TGA thermogram are most probably
temperature region 90–200  C (50% of mass loss). result of different boiling points of essential oil com-
In the second temperature region, between 200 and pounds. Also, it could be expected that some essen-
300  C up to 60% of essential oil evaporates. The tial oil compounds are thermally decomposed to the
new compounds. According to Neuenschwander
et al. (2010), a-pinene autoxidation under elevated
temperature leads to the formation of different types
of peroxyl radicals. Further, peroxyl radicals’ reaction
leads to the production of hydroperoxide or epoxide
and alkoxyl radicals.
In the case of Ca-alginate/essential oil, thermogram
shows that up to 150  C the mass loss of beads is
<10%. Further, mass loss is most probably connected
with oil compounds evaporation/thermal decompos-
ition as well as Ca-alginate thermal decomposition.
The release of encapsulated peppermint essential oil
and thermal decomposition of Ca-alginate matrix most
probably occurred in the close temperature regions.
According to Levi
c et al. (2013), the main mass losses
of empty Ca-alginate beads correspond with two tem-
perature regions: (i) 50–150  C and (ii) 150–300  C.
Figure 2. FTIR spectrum of the free peppermint essential oil Laurienzo et al. (2005) pointed out that mass loss in
(a), A2 (b), C4 (c), and CW4 (d). the first temperature region is connected with the loss

Figure 3. Thermogravimetric analyses of essential oil (a), A2 (b), C4 (c), and CW4 (d). Black line-thermogravimetry curve; red line-
differential thermal gravimetry curve.
 JOURNAL OF MICROENCAPSULATION 117

of water from the alginate matrix. The second tem- wax particles. Based on thermal analysis, the alginate
perature region corresponds with mass loss of encap- and coacervates encapsulates of peppermint essential
sulated oil (and most probably with oil thermal oil are more suitable for applications in the high-tem-
degradation) and with thermal decomposition of Ca- perature food processes (e.g. baker’s products). Also,
alginate. However, the thermal degradation of alginate smaller in size, carnauba microparticles are more suit-
is a complex process including several steps. able regarding ice cream textural properties.
The TGA thermogram of encapsulated peppermint Thermal analyses showed that encapsulated essen-
essential oil in gelatine/alginate coacervates shows tial oil was released at elevated temperatures in the
two main mass losses. Up to 175  C the mass loss is broader temperature regions and in a more control-
10–15% and most probably is connected with water lable manner compared to free peppermint essential
loss from matrix material as well as essential oil oil. These results suggest that encapsulated pepper-
release. In the temperature region from 50 to 175  C, mint essential oil is most probably more suitable for
the mass loss of coacervates with encapsulated oil is thermal food processes, enabling protection and con-
very similar to those based on Ca-alginate. However, trolled release of the aromatic compound into the
at the end of the analysis (at 300  C), the coacervates product. In this study, we tested applications of car-
show higher mass loss (70% mass loss) than systems nauba wax microparticles loading peppermint essen-
based on Ca-alginate (i.e. 50% mass loss). tial oil in ice cream. However, further studies are
Coacervates showed lower EE compared to encapsu- necessary in order to evaluate applications of encapsu-
late produced by other encapsulation methods used in lated peppermint essential oil in other thermal proc-
this study (see Table 2). As a result, the kinetic of mass essed food products.
loss of coacervates at high temperatures is most prob-
ably affected by low essential oil content. Also, the rea- 3.5. Quantitative descriptive sensory evaluation of
son for the difference in the thermal properties of ice cream incorporated with carnauba wax
alginate beads and coacervates may be the fact that oil beads
Ca-alginate beads are produced by gelling of sodium
alginate using Ca2þ ions as a crosslinker. Thus, Ca-algin- Ice cream samples, control and enriched with pepper-
ate is most probably more thermal stable matrix com- mint oil, showed no statistically significant differences in
pared to gelatine/alginate coacervates that are not terms of textural properties and colour in quantitative
crosslinked. Devi et al. (2012) suggested that additional descriptive sensory analysis. There were no differences
crosslinking using glutaraldehyde as crosslinker provides among 4 ice cream types in terms of textural properties,
better thermal properties of gelatine/alginate microcap- e.g. hardness, coarseness, gumminess, iciness,
sules produced by complex coacervation. However, the
usage of glutaraldehyde is limited in the food industry.
This problem could be for example solved by using
enzymatic crosslinking of gelatine (Prata et al. 2008).
The thermogram of wax-based encapsulates exhibit
two mass losses: up to 250  C (max. at 130  C, prob-
ably wax decomposition and oil release); the second
mass loss occurs above 250  C and it is most probably
connected with the decomposition of carnauba wax.
The release of essential oil is more pronounced at
higher temperatures (above 80–85  C) as the melting
point of carnauba wax corresponding to this tempera-
ture range. Milanovi
c et al. (2010) noticed that thermal
processes above 250  C are results of carnauba wax
transformation under the high temperatures. Since wax
particles under elevated temperature (i.e. 80–85  C, car-
nauba wax melting point) are melted, such wax-based
encapsulates are more suitable for applications in the
low-temperature food processes. We have selected ice Figure 4. Sensory scores of ice cream samples in a radar plot
cream as model food for incorporation and sensory ana- (IC1: Control ice cream, IC2, IC3, and IC4: Ice cream enriched
lysis of peppermint oil encapsulated in the carnauba with peppermint oil at 0.1, 0.2, and 0.3%, w/w, respectively).
118 M. YILMAZTEKIN ET AL.

wateriness, and creaminess. The same situation was Disclosure statement

observed for colour as well. Therefore, the texture and No potential conflict of interest was reported by the authors.
appearance of ice cream samples were not
included here.
The increase in the amount of peppermint oil in ice Funding
creams imparted peppermint oil flavour to the samples This study was carried out in the framework of COST Action
(Figure 4). Sourness and astringency of ice creams were FA0907 BIOFLAVOUR under the EU’s Seventh Framework
slightly perceived by the panelists; however, the degree Programme for Research (FP7). This work was supported by
Ministry of Science and Technological Development,
of sourness and astringency were lower than the value
Republic of Serbia (Project nos. III46010 and III46001).
of 4. Refreshing peppermint oil flavour was noted after-
taste by the panelists. Panelists rated the overall accept-
ability of all peppermint oil enriched ice creams higher References
than the value of 7. Therefore, peppermint oil enrich- Burt, S., 2004. Essential oils: their antibacterial properties and
ment might be a suitable option to produce new ice potential applications in foods-a review. International jour-
cream types. We have also tested the possibility of nal of food microbiology, 94 (3), 223–253.
peppermint oil enrichment higher than 0.3% (w/w) in Cam, M., et al., 2013. Enrichment of functional properties of
ice cream with pomegranate by-products. Journal of food
ice cream; however, oral burn and a dramatic decrease
science, 78 (10), C1543–C1550.
in the overall acceptability were noted by panelists. Chan, E.S., et al., 2011. Effects of starch filler on the physical
properties of lyophilized calcium-alginate beads and the
viability of encapsulated cells. Carbohydrate polymers, 83
4. Conclusions (1), 225–232.
Ca-alginate encapsulates showed higher EEs compared Chang, P.C. and Dobashi, T., 2003. Preparation of alginate
complex capsules containing eucalyptus essential oil and
to other shell materials. On the other hand, after the
its controlled release. Colloids and surfaces B biointerfaces,
drying process, carnauba wax microparticles were
32 (3), 257–262.
more regular spherical in shape than the other formu- Crowell, P., 1997. Monoterpenes in breast cancer chemopre-
lations. The essential oil and carrier materials formed vention. Breast cancer research and treatment, 46 (2–3),
simple mixtures inside the encapsulates, without 191–197.
strong chemical interactions. The results of the ther- Devi, N., et al., 2012. Study of complex coacervation of gel-
atin A and sodium alginate for microencapsulation of
mal analysis showed that encapsulation of peppermint
olive oil. Journal of macromolecular science, part A: pure
essential oil ensured controlled release of oil in the
and applied chemistry, 49 (11), 936–945.
broader temperature regions compared to free oil. Gibbs, F.B., et al., 1999. Encapsulation in the food industry: a
According to sensory analysis, incorporation pepper- review. International journal of food sciences & nutrition,
mint oil into ice cream up to 0.3% (w/w) might be a 50, 213–224.
suitable option to add functional properties of pepper- Hightower, C.A. and Edgar, I.V.C., 2009. Descriptive sensory
mint into ice cream without damaging textural proper- analysis of toothpaste flavor and texture using two sam-
pling methods: brushing versus spoon tasting. Journal of
ties. Since tested encapsulation systems are different
sensory studies, 24 (3), 301–316.
in many aspects, our goal was to establish different Jun-Xia, X., Hai-Yan, Y., and Jian, Y., 2011. Microencapsulation
procedures for essential oils encapsulation and to of sweet orange oil by complex coacervation with soybean
open the possibility for their application in encapsu- protein isolate/gum Arabic. Food chemistry, 125 (4),
lated forms in food products. Regarding EE, Ca-algin- 1267–1272.
ate is the most promising system for peppermint Lai, F., et al., 2007. Artemisia arborescens l essential oil
loaded beads: preparation and characterization. AAPS
essential oil encapsulation. On the other hand, car-
PharmSciTech, 8 (3), E126–E132.
nauba wax microparticles are suitable in the applica- Laurienzo, P., et al., 2005. Synthesis and characterization of a
tions where small particles are required due to food novel alginate-poly(ethylene glycol) graft polymer.
textural properties (i.e. here tested ice cream with Carbohydrate polymers, 62 (3), 274–282.
added carnauba wax microparticles loading pepper- Levi
c, S., et al., 2013. Entrapment of ethyl vanillin in calcium
mint essential oil). Coacervates showed the lowest EE. alginate and calcium alginate/poly(vinyl alcohol) beads.
Chemical papers, 67 (2), 221–228.
However, due to its simple preparation procedure,
Levi
c, S., et al., 2015. Characterization of sodium alginate/D-
encapsulation of essential oils by coacervation could limonene emulsions and respective calcium alginate/D-
be improved by additional steps like a crosslinking of limonene beads produced by electrostatic extrusion. Food
carrier material in order to increase EE. hydrocolloids, 45, 111–123.
 JOURNAL OF MICROENCAPSULATION 119

Lv, Y., et al., 2014. Formation of heat-resistant nanocapsules Prakash, N. and Yunus, M., 2013. Fourier transform infrared
of jasmine essential oil via gelatin/gum arabic based com- spectroscopy analysis of oil of Mentha arvensis grown at
plex coacervation. Food hydrocolloids, 35, 305–314. sites varying with vehicular traffic loads in Lucknow city,
Manaf, M.A., et al., 2018. Encapsulation of volatile citronella India. International journal of environment, 2 (1), 16–25.
essential oil by coacervation: efficiency and release study. Prata, S.A., et al., 2008. Release properties of chemical and
IOP conference series: materials science and engineering, enzymatic crosslinked gelatin-gum Arabic microparticles
358, 012072. containing a fluorescent probe plus vetiver essential oil.
Manojlovi
c, V., et al., 2008. Application of electrostatic extru- Colloids and surfaces B: biointerfaces, 67 (2), 171–178.
sion-flavour encapsulation and controlled release. Sensors, Shinde, U. and Nagarsenker, M., 2011. Microencapsulation of
8, 1488–1496. eugenol by gelatin-sodium alginate complex coacervation.
McClements, D.J. and Rao, J., 2011. Food-grade nanoemul- Indian journal of pharmaceutical sciences, 73, 311–315.
Siow, L.F. and Ong, C.S., 2013. Effect of pH on garlic oil
sions: formulation, fabrication, properties, performance,
encapsulation by complex coacervation. Journal of food
biological fate, and potential toxicity. Critical reviews in
processing and technology, 4, 199.
food science and nutrition, 51 (4), 285–330.
Soukoulis, C. and Tzia, C., 2010. Response surface mapping
McKay, L.D. and Blumberg, B.J., 2006. A review of the bio-
of the sensory characteristics and acceptability of choc-
activity and potential health benefits of peppermint tea olate ice cream containing alternate sweetening agents.
(Mentha piperita L.). Phytotherapy research, 20 (8), 619–633. Journal of sensory studies, 25 (1), 50–75.
Milanovi
c, J., et al., 2010. Microencapsulation of flavors in Yilmaztekin, M., 2014. Characterization of potent aroma com-
carnauba wax. Sensors, 10 (1), 901–912. pounds of cape gooseberry (Physalis peruviana L.) fruits
Nedovi
c, V.A., et al., 2001. Electrostatic generation of alginate grown in antalya through the determination of odor activ-
microbeads loaded with brewing yeast. Process biochemis- ity values. International journal of food properties, 17 (3),
try, 37 (1), 17–22. 469–480.
Neuenschwander, U., Guignard, F., and Hermans, I., 2010. Zuidam, N. J. and Shimoni, E., 2010. Overview of microencap-
Mechanism of the aerobic oxidation of alpha-pinene. sulates for use in food products or processes and methods
ChemSusChem, 3 (1), 75–84. to make them. In: N.J. Zuidam and V.A. Nedovic, eds.
Ody, P., 1993. The complete medicinal herbal. London: Encapsulation technologies for food active ingredients and
Dorling Kindersley Limited. food processing. Dordrecht, The Netherlands: Springer, 3–31.