Solid inclusion complexes of vanillin with cyclodextrins: Their formation, characterization, and high-temperature stability

Article abstract of DOI:10.1021/jf202915c

This study reports the formation of solid vanillin/cyclodextrin inclusion complexes (vanillin/CD ICs) with the aim to enhance the thermal stability and sustained release of vanillin by inclusion complexation. The solid vanillin/CD ICs with three types of

Full text of DOI:10.1021/jf202915c

ARTICLE
pubs.acs.org/JAFC
Solid Inclusion Complexes of Vanillin with Cyclodextrins: Their
Formation, Characterization, and High-Temperature Stability
Fatma Kayaci and Tamer Uyar*
UNAM-Institute of Materials Science and Nanotechnology, Bilkent University, Ankara 06800, Turkey
ABSTRACT: This study reports the formation of solid vanillin/cyclodextrin inclusion complexes (vanillin/CD ICs) with the aim to
enhance the thermal stability and sustained release of vanillin by inclusion complexation. The solid vanillin/CD ICs with three types
of CDs (α-CD, β-CD, and γ-CD) were prepared using the freeze-drying method; in addition, a coprecipitation method was also
used in the case of γ-CD. The presence of vanillin in CD ICs was confirmed by FTIR and 1H NMR studies. Moreover, 1H NMR
study elucidated that the complexation stoichiometry for both vanillin/β-CD IC and vanillin/γ-CD IC was a 1:1 molar ratio,
whereas it was 0.625:1 for vanillin/α-CD IC. XRD studies have shown channel-type arrangement for CD molecules, and no
diffraction peak for free vanillin was observed for vanillin/β-CD IC and vanillin/γ-CD IC, indicating that complete inclusion
complexation was successfully achieved for these CD ICs. In the case of vanillin/α-CD IC, the sample was mostly amorphous and
some uncomplexed vanillin was present, suggesting that α-CD was not very effective for complexation with vanillin compared to
β-CD and γ-CD. Furthermore, DSC studies for vanillin/β-CD IC and vanillin/γ-CD IC have shown no melting point for vanillin,
elucidating the true complex formation, whereas a melting point for vanillin was recorded for vanillin/α-CD IC, confirming the
presence of some uncomplexed vanillin in this sample. TGA thermograms indicated that thermal evaporation/degradation of
vanillin occurred over a much higher temperature range (150À300 °C) for vanillin/CD ICs samples when compared to pure vanillin
(80À200 °C) or vanillin/CD physical mixtures, signifying that the thermal stability of vanillin was increased due to the inclusion
complexation with CDs. Moreover, headspace GC-MS analyses indicated that the release of vanillin was sustained at higher
temperatures in the case of vanillin/CD ICs due to the inclusion complexation when compared to vanillin/CD physical mixtures.
The amount of vanillin released with increasing temperature was lowest for vanillin/γ-CD IC and highest for vanillin/α-CD IC,
suggesting that the strength of interaction between vanillin and the CD cavity was in the order γ-CD > β-CD > α-CD for solid
vanillin/CD ICs.
KEYWORDS: cyclodextrin, vanillin, inclusion complex, thermal stability, sustained release
’ INTRODUCTION
Vanillin (4-hydroxy-3-methoxybenzaldehyde) (Figure 1e),
which is the major component of natural vanilla, is widely used
as a flavoring additive in the food industry.13À15 Vanillin can also
be used as a food preservative due to its antioxidant and anti-
microbial properties.14,15 Moreover, vanillin is used as a fragrance
constituent in the cosmetics and textile industries.15,16 However,
a short shelf life is a significant problem for vanillin due to its
thermal instability and volatile nature. Here, we studied the forma-
tion ofvanillin/cyclodextrin inclusion complexes (vanillin/CD ICs)
for the enhancement of the thermal stability and sustained release of
vanillin by using three types of CDs, α-CD, β-CD, and γ-CD.
Very few studies have been reported for the formation of
vanillin/CD ICs by using β-CD, and these CD ICs were mostly
studied in solutions.14,17À20 However, to date, the solid vanillin/
CD IC systems have not been studied in detail, and the inclusion
complexation of vanillin with different types of CDs has not been
evaluated in the solid state. In this study, we formed solid
vanillin/CD ICs by using α-CD, β-CD, and γ-CD by a freeze-
drying method and coprecipitation method (only for γ-CD), and
we have carried out extensive characterization of these vanillin/CD
Cyclodextrin inclusion complexation is widely used in the
food, cosmetics, and textile industries to increase thermal stabi-
lity and to control/sustain the release of volatile flavors and
fragrances or other functional additives.1À9 Cyclodextrins (CDs)
are cyclic oligosaccharides that are produced by the enzymatic
degradation of starch.1 The three most common CDs have 6, 7,
and 8 glucopyranose units in the cyclic and are named α-CD,
β-CD, and γ-CD, respectively. Whereas the depth of the cavity
for these CDs is ∼8 Å, the sizes of the cavity are different for α-CD,
β-CD, and γ-CD, being ∼6, 8, and 10 Å, respectively (Figure 1b).1,2
CDs have a truncated cone-shaped molecular structure having a
relatively hydrophobic inner cavity and hydrophilic outer surface
because of their unique chemical structure. The hydrophobic
cavities of CDs have a remarkable ability to form noncovalent
hostÀguestinclusioncomplexes(ICs) withavarietyofmolecules.1À9
The formation and stability of the CD ICs depend on various factors
including the size/shape fit, chemical surroundings, and binding
forces (hydrophobic interactions, van der Waals attractions, hydro-
gen bonding, electrostatic interactions, etc.) between the host CD
and guest molecules.10À12 The physical and chemical properties of
the guest molecules are improved when complexed with CDs; there-
fore, CD ICs are quite applicable in the food, cosmetics, and textile
industries because CDs enhance the stabilization and controlled/
sustained release of volatile or unstable flavors and fragrances.1À9
1
ICs by using FTIR, H NMR, XRD, DSC, TGA, and headspace
Received: July 22, 2011
Revised:
September 17, 2011
Accepted: September 19, 2011
Published: September 20, 2011
r
2011 American Chemical Society
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Figure 1. (a) Chemical structure of α-CD; (b) approximate dimensions of α-CD, β-CD, and γ-CD; schematic representation of packing structures of
(c) cage-type and (d) channel-type CD crystals; (e) chemical structure of vanillin, and (f) schematic representation of vanillin/CD IC.
GC-MS techniques. The physical mixtures of vanillin/CDs and
pure vanillin were also analyzed for comparison.
mortar to obtain a fine powder. For comparison, physical mixtures of
vanillin/CDs having a 1:1 molar ratio were prepared in the solid state by
admixing vanillin and CDs and ground with a pestle in an agar mortar.
Characterization and Measurements. A Fourier transform
infrared (FTIR) spectrometer (VERTEX70, Bruker) was used to obtain
the infrared spectra of the samples. The FTIR measurements were per-
formed by blending the samples with potassium bromide (KBr), and
pellets were formed under high pressure. The spectra were recorded
between 4000 and 400 cmÀ1 with a resolution of 4 cmÀ1 at total scans of
64. The proton nuclear magnetic resonance (1H NMR, Bruker DPX-400)
spectra were recorded at 400 MHz at 25 °C. About 20 g/L vanillin/CD
ICs was dissolved in d6-DMSO to evaluate the stoichiometries of the
complexes. Integration of the chemical shifts (δ) given in parts per
million (ppm) of the samples was calculated by using NMR software.
The X-ray diffraction (XRD, PANalyticalX’Pert Powder diffractometer)
patterns for the samples were collected by using Cu Kα radiation in a
range of 2θ = 5À30°. The thermal properties of the vanillin/CD ICs,
vanillin/CD physical mixtures, and pure vanillin were investigated by
using a differential scanning calorimeter (DSC, Q2000, TA Instruments)
and a thermogravimetric analyzer (TGA, Q500, TA Instruments). For
DSC analyses, the samples were initially equilibrated at 25 °C and then
heated to 250 °C at a 20 °C/min heating rate under N2 as a purge gas.
TGA measurements were performed from room temperature to 500 °C
at a heating rate of 20 °C/min under N2 atmosphere.
Headspace gas chromatographyÀmass spectrometry (GC-MS) ana-
lyses were performed on an Agilent Technologies 7890A gas chromato-
graph coupled to an Agilent Technologies 5975C inert MSD with a
triple-axis detector. The headspace GC-MS experiments were carried
out by using a CTCPAL autosampler. Different amounts of CD ICs
(0.0185 g vanillin/α-CD IC, 0.0211 g vanillin/β-CD IC, and 0.238 g
vanillin/γ-CD IC) were placed separately in 20 mL headspace glass vials
to have the same amount of vanillin (0.0025 g) in each vial. For com-
parison, the same amounts of vanillin/CD physical mixtures were placed
separately in 20 mL headspace glass vials for the headspace GC-MS
measurements. The tops of all vials were tightened to prevent the escape
of the vanillin. Every sample was agitated at 250 rpm at different incuba-
tion temperatures in the order 35, 50, 75, and 100 °C for 30 min by using
an agitator and then analyzed by headspace GC-MS. One thousand
’ MATERIALS AND METHODS
Materials. Vanillin (4-hydroxy-3-methoxybenzaldehyde, C8H8O3;
molecular weight, 152.15; melting point, 81À83 °C; 99% purity) was
obtained from Sigma-Aldrich. Cyclodextrins (α-CD, β-CD, and γ-CD)
were purchased from Wacker Chemie AG (Germany). All materials
were used without any purification. Water used for the preparation of the
complexes was from a Millipore Milli-Q ultrapure water system.
Preparation of Vanillin/CD ICs. The inclusion complexes of
vanillin with α-CD, β-CD, and γ-CD were prepared by adding an
equimolar amount of vanillin to an aqueous solution of α-CD, β-CD,
and γ-CD. The amount of the CD used was determined according to the
solubility of CDs in water at 25 °C, that is, 14.5, 1.85, and 23.2 g/100 mL
for α-CD, β-CD, and γ-CD, respectively.1,2 First, 0.25 g of α-CD
dissolved in 1.725 mL of water and, then, 0.039 g of vanillin were added
into the aqueous α-CD solution, and the vanillin/α-CD solution was
mixed overnight at room temperature. The vanillin/α-CD solution
became slightly turbid, but precipitation did not occur. For vanillin/
β-CD IC, 0.25 g of β-CD was dissolved in 14 mL of water, and then
0.034 g of vanillin was added into this clear β-CD aqueous solution. The
vanillin/β-CD solution became clear after mixing overnight at room
temperature, and no precipitation was observed. In the case of vanillin/
γ-CD IC, 0.25 g of γ-CD was dissolved in 1.08 mL of water and 0.029 g
of vanillin was added into this solution. The vanillin/γ-CD solution first
became clear and then highly turbid and precipitation took place after
mixing overnight at room temperature. Thereafter, all three vanillin/CD
solutions were frozen at À80 °C and then lyophilized in a freeze-dryer
(Labconco) to obtain solid vanillin/CD ICs. In addition, the inclusion
complex of vanillin with γ-CD was also prepared by a coprecipitation
method. Similarly, 0.029 g of vanillin was added to an aqueous solution
of γ-CD (0.25 g of γ-CD dissolved in 1.08 mL of water) and stirred
overnight. The resulting suspension was filtered through a borosilicate
filter (por 2), and then the filtrate was washed with water several times to
remove uncomplexed vanillin and γ-CD and then dried overnight under
a hood. After drying, the vanillin/γ-CD IC solid sample was crushed in a
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Figure 2. FTIR spectra of (a) as-received (i) α-CD, (ii) β-CD, and (iii) γ-CD and (b) (i) vanillin/α-CD IC, (ii) vanillin/β-CD IC, (iii) vanillin/γ-CD
IC, (iv) vanillin/γ-CD IC (coprecipitation), and (v) vanillin.
microliters of vapor was injected to the GC-MS by using a headspace
injector (MSH 02-00B, volume = 2.5 mL, scale = 60 mm). The syringe
temperature was kept the same as the incubation temperature. The
separation of compounds was performed on an HP-5MS (Hewlett-
Packard, Avondale, PA) capillary column (30 m  0.25 mm i.d., 0.25 μm
film thickness). Column temperature was held at 50 °C for 2 min and
increased to 250 °C at the rate of 20 °C/min and equilibrated at this
temperature for 3 min. Helium was used as a carrier gas at a flow rate of
1.2 mL/min. Thermal desorption was carried out in the splitless mode
during 2 min. The temperatures of the ion source and the transfer line
were 230 and 280 °C, respectively. The GC-MS analyses were carried
out in the complete scanning mode (SCAN) in the 50À550 mass range.
The vanillin peak was identified by comparing its mass spectrum with
that of vanillin in the Flavor 2 and NIST 0.5 libraries. The retention time
of vanillin was 8.76 min, and the major peaks of the vanillin were 151 and
152 mass over charge.
samples. This is very typical for CD ICs, and FTIR peak shifts
were commonly reported for various CD IC systems.22,23
The presence of vanillin in the CD ICs and their stoichiome-
tries were determined by 1H NMR studies (Figure 3). The peaks
for vanillin observed at about 3.8, 6.9, 7.4, and 9.8 ppm correspond
to different protons in vanillin.18,19 The quantity of vanillin in the
CD ICs was calculated by integrating the peak ratio of the
characteristic chemical shifts corresponding to vanillin and CD. The
integration of the CD peak at 4.8 ppm20 and the vanillin peak at
9.8 ppm was taken into account to calculate the stoichiometry
of the host and guest molecules in the CD IC samples. It was
found that vanillin/β-CD IC and vanillin/γ-CD IC have a 1:1
(vanillin/CD) molar ratio, whereas the ratio of vanillin/α-CD
was 0.625:1, suggesting that the 1:1 initial molar ratio used for the
formation of CD ICs was not conserved in the case of vanillin/
α-CDIC. Thissuggeststhatα-CD was not very effectiveinencapsu-
lating vanillin, which is possibly because of the small dimension
of the α-CD cavity. Additionally, the chemical shifts for pure
vanillin and the vanillin/α-CD system were almost the same in
the NMR spectrum, indicating that vanillin and α-CD were
decomplexed in the d6-DMSO solvent system. However, we
observed differences in chemical shifts (about 0.1 ppm shift) for
vanillin peaks in vanillin/β-CD and vanillin/γ-CD samples when
compared to pure vanillin, which is due to the presence of
complexation.
XRD studies were performed to investigate the crystalline
structure of vanillin/CD ICs. The as-received CDs have a cage-
type packing structure in which the cavity of each CD molecule is
blocked by neighboring CD molecules (Figure 1c).24,25 The
inclusion complexation is generally confirmed by the formation
of channel-type arrangement of the CD molecules in which CD
molecules are aligned and stacked on top of each other by
forming cylindrical channels (Figure 1d).24À26 The XRD patterns
of as-received CDs (Figure 4a) and vanillin/CD ICs (Figure 4b)
have characteristic diffraction peaks in the range of 2θ = 5À30°.
Vanillin is a crystalline material having a distinct diffraction centered
at 2θ = 13° (Figure 4b). The XRD data show that the as-received
CDs (α-CD, β-CD, and γ-CD) have cage-type packing structures,
which is consistent with the literature findings.26À28
’ RESULTS AND DISCUSSION
In this study, we prepared solid inclusion complexes of vanillin
with three types of cyclodextrin (α-CD, β-CD, and γ-CD) by
using a freeze-drying method; in addition, a coprecipitation method
was also used in the case of vanillin/γ-CD IC. Initially, the pre-
sence of the guest molecule (vanillin) in the cyclodextrin inclu-
sion complexes was confirmed by the FTIR studies. The character-
istic absorption bands of CDs are observed at around 1030,1080,
and 1155 cmÀ1 corresponding to the coupled CÀC/CÀO stretch-
ing vibrations and the asymmetric stretching vibration of the
CÀOÀC glycosidic bridge in the FTIR spectra of pure CDs
(Figure 2a). The FTIR spectra of the vanillin/CD ICs are
depicted in Figure 2b, and the FTIR spectrum of pure vanillin
is also given for comparison. The FTIR spectrum of pure vanillin
exhibited characteristic peaks at 1510, 1590, and 1665 cmÀ1 cor-
responding to stretching absorption of benzene ring and stretch-
ing of CdO of the aldehyde group.21 The overlapping of absorp-
tion peaks of CDs and vanillin makes the identification of the
individual components rather complicated in vanillin/CD ICs.
Yet, the absorption bands of vanillin at 1512, 1595, and 1670 cmÀ1
were observed for all vanillin/CD ICs, which indicated that vanillin
is present in the samples. In addition, it was noted that the
characteristic absorption bands of vanillin were shifted slightly
for vanillin/CD ICs when compared to pure vanillin, suggesting
the hostÀguest interactions between CD and vanillin in these
The XRD patterns of vanillin/CD ICs are very different from
those of as-received CDs (Figure 4b). The XRD of the vanillin/
α-CD IC shows two broad halo diffraction patterns, indicat-
ing that the material was mostly amorphous; yet, a distinct peak
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diffraction peak at 2θ = 13° (Figure 4b). The overlapping of
vanillin and the α-CD channel-type diffraction peaks makes the
peak assignment rather complicated. The supplementary thermal
analyses by DSC, which will be discussed in the following section,
indicated that some free vanillin was present in the vanillin/
α-CD IC sample. Therefore, the peak at 2θ = 13° was due to both
the presence of some uncomplexed free vanillin and the α-CD
channel-type packing. In short, the XRD data of vanillin/α-CD
IC suggest that some free vanillin is present in vanillin/α-CD IC
and that the sample was mostly amorphous, having some amount
of channel-type structure.
The XRD pattern of vanillin/β-CD IC shows two major peaks
centered at 2θ = 11.8° and 2θ = 17.8°, indicating that the β-CD
adopts a channel-type packing;28 in addition, no peak at 2θ = 13°
for free vanillin was observed, suggesting that complete com-
plexation of vanillin with β-CD was successful. In the case of
vanillin/γ-CD IC obtained by coprecipitation technique, the
XRD pattern has a major peak at 2θ = 7.5° along with minor
diffractions at 2θ = 14.2°, 15°, 16°, 16.8°, and 22°, confirming
the tetragonal channel-type packing of γ-CD.26,27 The XRD
pattern of vanillin/γ-CD IC obtained by freeze-drying is similar
to that of the vanillin/γ-CD IC obtained by coprecipitation
method; yet, the peaks are relatively broad, indicating that the
material was partially amorphous. In addition, the peak at 2θ =
6.2° corresponding to channel structure with hexagonal packing
was also observed.26 The tetragonal channel packing trans-
forms to hexagonal packing upon removal of water residing in
the interstitial sites when vacuum-dried.26 Hence, in the case of
freeze-drying method, vanillin/γ-CD IC having a mixture of
tetragonal and hexagonal channel-type packing was obtained.
Similar to vanillin/β-CD IC, the vanillin peak at 2θ = 13° was
absent, indicating that uncomplexed vanillin was not present in
both vanillin/γ-CD IC samples. The XRD data suggested that
both coprecipitation and freeze-drying methods were successful
for obtaining solid vanillin/γ-CD ICs.
The solid vanillin/CD ICs were further characterized by DSC
(Figure 5a) to verify whether the vanillin was included inside the
CD cavities or not. For instance, thermal transitions such as the
melting point (Tm) for the guest molecule would be observed if
there were any uncomplexed guest molecules present in the CD
ICs.29À31 The DSC thermograms of vanillin/CD physical mix-
tures have shown an endothermic peak at around 77 °C, which
corresponds to the melting point of vanillin (Figure 5b). The
DSC thermogram of pure vanillin is also given for comparison,
showing a melting point of 82 °C. On the other hand, the DSC
thermogram of vanillin/β-CD IC did not show any melting peak
for free vanillin, suggesting that the vanillin was fully complexed
with the β-CD. Similarly, the melting peak of free vanillin was
also absent in the DSC thermograms of vanillin/γ-CD IC
samples obtained by freeze-drying and coprecipitation methods,
confirming that both techniques resulted in pure vanillin/γ-CD
ICs. The absence of a vanillin melting point in DSC and the
absence of a diffraction peak at 2θ = 13° in XRD for solid
vanillin/β-CD IC and vanillin/γ-CD IC samples comprise
strong evidence for the complete inclusion complexation of
vanillin with β-CD and γ-CD. However, in the case of vanillin/
α-CD IC, a small endothermic peak at around 75 °C was
observed in the DSC thermogram due to the presence of some
free vanillin in this sample. The DSC data also correlate with the
XRD data as discussed in the previous section, where the XRD of
vanillin/α-CD IC has a diffraction peak for some uncomplexed
vanillin crystals.
Figure 3. 1H NMR spectra of (a) vanillin, (b) vanillin/α-CD IC, (c)
vanillin/β-CD IC, (d) vanillin/γ-CD IC, and (e) vanillin/γ-CD IC
(coprecipitation).
at 2θ = 13° and a weak peak at 2θ = 20° were present. The α-CD
having channel-type structure has two prominent peaks centered at
2θ=13° and 2θ=20°.27 Asmentionedabove, vanillin has a major
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Figure 4. XRD patterns of (a) as-received (i) α-CD, (ii) β-CD, and (iii) γ-CD and (b) (i) vanillin/α-CD IC, (ii) vanillin/β-CD IC, (iii) vanillin/γ-CD
IC, (iv) vanillin/γ-CD IC (coprecipitation), and (v) vanillin.
Figure 5. DSC thermograms of (a) (i) vanillin/α-CD IC, (ii) vanillin/β-CD IC, (iii) vanillin/γ-CD IC, and (iv) vanillin/γ-CD IC (coprecipitation)
and (b) (i) vanillin/α-CD physical mixture, (ii) vanillin/β-CD physical mixture, (iii) vanillin/γ-CD physical mixture, and (iv) vanillin.
For CD ICs, the thermal stability and the evaporation of the
volatile guest molecules shift to higher temperature upon inclu-
sion complexation due to guestÀhost interactions.8,26 Hence,
TGA was carried out for vanillin/CD ICs to determine the
thermal stability and evaporation of the vanillin included inside
the CD cavities (Figure 6). The TGAs of vanillin/CD physical
mixtures and pure vanillin were also performed for comparison
(Figure 6). The TGA thermogram of pure vanillin has a major
weight loss observed in the range of 80À200 °C, showing that
vanillin has a volatile nature. TGA thermograms of vanillin/CD
ICs showed three weight losses, the initial weight loss below
100 °C is due to water loss, the second weight loss between 150
and 300 °C is due to the evaporation/degradation of vanillin, and
the major weight loss above 300 °C corresponds to the main
thermal degradation of CD. The TGA thermograms of vanillin/
CD ICs showed that thermal degradation of vanillin occurred
over a much higher temperature range (150À300 °C) when
compared to pure vanillin (80À200 °C), confirming that the
thermal stability of vanillin increased due to inclusion complexa-
tion with CDs. For vanillin/CD physical mixtures, three weight
losses were observed: the one below 100 °C is attributed to
removal of water, the second one in the range of 80À200 °C
corresponds to the evaporation of vanillin, and the one above
300 °C is due to the main thermal degradation of CD. The
temperature range for evaporation of vanillin for vanillin/CD
physical mixtures is similar to that of pure vanillin. However, in
the case of vanillin/CD ICs, the thermal stability of vanillin
shifted to a much higher temperature owing to the strong
interactions between vanillin and the CD cavity in the CD ICs.
The amount of vanillin was calculated for solid vanillin/CD IC
samples from the TGA data. For vanillin/β-CD IC, the amount
of vanillin was ∼11% (w/w, with respect to β-CD), which
correlates with the initial amount of vanillin used for the com-
plexation. This corresponds to a 1:1 molar ratio complexation
between the β-CD and vanillin, correlating with the NMR data;
the initial amount of vanillin was preserved, and no loss of guest
molecule occurred during the formation and storage of vanillin/
β-CD IC. The amount of vanillin was calculated as ∼10% (w/w,
with respect to γ-CD) for both vanillin/γ-CD IC samples
obtained by freeze-drying and coprecipitation method. Similar
to vanillin/β-CD IC, vanillin/γ-CD IC samples have also a 1:1
molar ratio complexation between the γ-CD and vanillin as
observed in the NMR study, and the initial amount of vanillin was
preserved during the formation and storage of these samples. In
the case of vanillin/α-CD IC, the amount of vanillin was
calculated as ∼8% (w/w, with respect to CD), which is lower
than the initial weight percent (∼13%, w/w). As DSC and XRD
data confirmed, some uncomplexed vanillin was present in the
sample, and it is likely that free vanillin could not be preserved
during the storage of the vanillin/α-CD IC sample.
The temperature stability and release characteristics of
vanillin/CD ICs were also studied by headspace GC-MS.
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Table 1. Corrected Peak Areasa Calculated at 8.76 min from
Headspace GC-MS Chromatograms for Vanillin/CD ICs and
Vanillin/CD Physical Mixtures Agitated at Different Incuba-
tion Temperatures
sample
35 °C 50 °C 75 °C
100 °C
vanillin/α-CD IC
 1
 1
 52  736  3002
 75  981  4898
vanillin/α-CD physical mixture
vanillin/β-CD IC
 1
 1
 11  108  1119
 41  519  2083
vanillin/β-CD physical mixture
vanillin/γ-CD IC
 1
 1
 1
 10
 10
 51
 63
 912
 283
vanillin/γ-CD IC (coprecipitation)
vanillin/γ-CD physical mixture
a All of the peak area values were divided by the peak area obtained at
 45  385  1486
35 °C for each sample.
was observed that as the temperature was increased, the relative
amount of vanillin released was in the order vanillin/α-CD >
vanillin/β-CD > vanillin/γ-CD for CD IC samples, indicating
that the strength of interaction between vanillin and the CD
cavity was in the order γ-CD > β-CD > α-CD. Moreover, in the
case of vanillin/γ-CD IC, it was noted that the coprecipitation
method was more effective for the stabilization and sustained
release of vanillin at high temperature (100 °C) when compared
to the freeze-drying method. In the coprecipitation method, the
vanillin/γ-CD IC sample was washed several times to remove
uncomplexed and/or loosely complexed vanillin; however, the
vanillin/γ-CD IC sample obtained by the freeze-drying method
may contain some amount of loosely complexed vanillin, and this
may be why CD ICs prepared by the coprecipitation method
were more effective for the stabilization of vanillin at high
temperature. In short, the headspace GC-MS studies showed
that the temperature release profile of vanillin was different for
each CD IC sample and highly dependent on the type of CD.
The reason for the formation of a more stable inclusion complex
between vanillin and γ-CD is possibly because of the bigger
cavity size of γ-CD, resulting in a better fit and size match
between the vanillin molecule and the γ-CD cavity.
In conclusion, the formation of solid CD ICs was successfully
achieved between vanillin and three types of CDs (α-CD, β-CD,
and γ-CD). The inclusion complexation of vanillin with β-CD
and γ-CD having a 1:1 molar ratio was quite successful, whereas
α-CD has shown poor complexation ability. Vanillin has a
volatile nature; however, higher thermal stability and sustained
release of vanillin at high temperatures were attained by forming
vanillin/CD ICs. The thermal evaporation/degradation of
vanillin shifted to a much higher temperature range (150À300 °C)
for vanillin/CD IC samples when compared to pure vanillin
(80À200 °C) or vanillin/CD physical mixtures. Additionally,
headspace GC-MS analyses showed that the relative amount of
vanillin released as a function of temperature was lowest for
vanillin/γ-CD IC and highest for vanillin/α-CD IC, suggesting
that the strength of interaction between vanillin and the CD
cavity was in the order γ-CD > β-CD > α-CD. In brief, CD ICs of
a widely used flavor/fragrance, vanillin, were achieved with
improved thermal stability, and such solid vanillin/CD ICs can
be quite applicable in the food industry, active food packaging,
the textile industry, etc.
Figure 6. TGA thermograms of (a) (i) α-CD, (ii) vanillin/α-CD IC,
(iii) vanillin/α-CD physical mixture, and (iv) vanillin; (b) (i) β-CD, (ii)
vanillin/β-CD IC, (iii) vanillin/β-CD physical mixture, and (iv) vanillin;
and (c) (i) γ-CD, (ii) vanillin/γ-CD IC, (iii) vanillin/γ-CD IC
(coprecipitation), (iv) vanillin/γ-CD physical mixture, and (v) vanillin.
The vanillin/CD physical mixtures were also analyzed for com-
parison. The release of the vanillin from each sample was quantified
from the corrected peak area calculated at 8.76 min from each
chromatogram. The relative release of vanillin at different
incubation temperatures (35, 50, 75, and 100 °C) was calculated
by dividing the corrected peak areas obtained at different
temperatures with the one obtained at 35 °C for each sample.
The relative release of the vanillin with increasing temperature is
summarized in Table 1. It is evident that the release of vanillin
was much slower for vanillin/CD ICs at all temperatures when
compared to the vanillin/CD physical mixtures. In addition, it
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dx.doi.org/10.1021/jf202915c |J. Agric. Food Chem. 2011, 59, 11772–11778

Journal of Agricultural and Food Chemistry
ARTICLE
’ AUTHOR INFORMATION
(21) Peng, H.; Xiong, H.; Li, J.; Xie, M.; Liu, Y.; Bai, C.; Chen, L.
Vanillin cross-linked chitosan microspheres for controlled release of
resveratrol. Food Chem. 2010, 121, 23–28.
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of cyclodextrin inclusion complexes À a review. J. Pharm. Sci. Technol.
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(23) Lin, S. Y.; Hsu, C. H.; Sheu, M. T. Curve-fitting FTIR studies of
loratadine/hydroxypropyl-β-cyclodextrin inclusion complex induced by
co-grinding process. J. Pharm. Biomed. Anal. 2010, 53, 799–803.
(24) Saenger, W.; Jacob, J.; Gessler, K.; Steiner, T.; Hoffmann, D.;
Sanbe, H.; Koizumi, K.; Smith, S.; Takaha, T. Structures of the common
cyclodextrins and their larger analogues beyond the doughnut. Chem.
Rev. 1998, 98, 1787–1802.
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solid state. Chem. Rev. 1998, 98, 1803–1828.
(26) Uyar, T.; Hunt, M.; Gracz, H.; Tonelli, A. Crystalline cyclodex-
trin inclusion compounds formed with aromatic guests: guest-dependent
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(27) Rusa, C.; Bullions, T.; Fox, J.; Porbeni, F.; Wang, X.; Tonelli, A.
Inclusion compound formation with a new columnar cyclodextrin host.
Langmuir 2002, 18, 10016–10023.
Corresponding Author
*E-mail: tamer@unam.bilkent.edu.tr. Phone: (+90)3122903571.
Fax: (+90)3122664365.
Funding Sources
The State Planning Organization (DPT) of Turkey is acknowl-
edged for the support of UNAM-Institute of Materials Science and
Nanotechnology. T.U. acknowledges EU FP7-PEOPLE-2009-RG
Marie Curie-IRG for funding the NANOWEB (PIRG06-GA-2009-
256428) project. F.K. acknowledges TUBITAK-BIDEB for the
national graduate study scholarship.
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