LWT - Food Science and Technology 73 (2016) 197e204

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LWT - Food Science and Technology

journal homepage: www.elsevier.com/locate/lwt

Effect of addition of green coffee extract and nanoencapsulated

chlorogenic acids on aroma of different food products
_
Grazyna Budryn*, Donata Zaczyn
 ska, Joanna Oracz
Institute of Food Technology and Analysis, Faculty of Biotechnology and Food Sciences, Lodz University of Technology, 90-924, Lodz, Poland

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

Article history: Between food polyphenols and proteinaceous compounds occur noncovalent interactions. The in-
Received 24 February 2016 teractions can affect some properties of these compounds, such as nutritional value, bioactivity and
Received in revised form organoleptic characteristics. When polyphenols are used as food additive, the changes can by limited by
5 June 2016
their encapsulation. The aim of the study was to determine, how the addition of polyphenols, i.e.
Accepted 6 June 2016
chlorogenic acids, from green coffee in free form or nanoencapsulated by b-cyclodextrin and further the
Available online 7 June 2016
addition of hydrolysate of ovalbumin, whey proteins or soy proteins will affect the content and the
profile of volatile aroma compounds of six food products: bread, cookies, caramel cottage cheese, nutty
Keywords:
Green coffee
filling, and mushroom or meat stuffing. Considering the six food products a general trend of the content
Chlorogenic acids of volatile compounds caused by supplementing with the used additives was observed. The increase of
Volatile substances aroma volatiles content in the headspace of the products increased in the following order: nano-
Aroma encapsulated chlorogenic acids < control < protein hydrolysates < free chlorogenic acids < chlorogenic
GC-MS acids in free form with protein hydrolysates.
© 2016 Elsevier Ltd. All rights reserved.

1. Introduction one serving can strongly interact with proteinaceous components

Polyphenols are often used to enrich food products because of
the high activity to prevent oxidative changes in food and to limit
oxidative stress in human organism (Maat et al., 2005). Coffee is
one of rich sources of polyphenols, especially phenolic acids
(Daglia, Papetti, Gregotti, Berte , & Gazzani, 2007). Green coffee
beans contain mainly hydroxycinnamic acids, such as caffeic and
ferulic acids, as well as their esters with quinic acid, referred to as
chlorogenic acids (CHAs) (Budryn, Zy _ zelewicz,
_ Nebesny, Oracz, &
Krysiak, 2013). They exhibit anti-inflammatory and anti-
mutagenic effects, which prevent chronic disorders, such as tu-
mors, cardiovascular and rheumatologic diseases (Cheng, Dai, Zhou,
Yang, & Liu, 2007). Consumption of coffee can also be helpful in the
fight against obesity and in limiting the effects of type 2 diabetes
(Bassoli, et al., 2008).
To achieve the positive effects of polyphenols consumption,
their average daily intake has to be on a level not lower than 1 g
(Williamson & Holst, 2008). Many studies considering supple-
mented food showed that such a large amount of polyphenols in
* Corresponding author.
E-mail address: grazyna.budryn@p.lodz.pl (G. Budryn).
(Renouf et al., 2010). The interactions could limit the functionality
of both proteins and polyphenols, especially they decrease their
absorption and reduce the antioxidant activity (Rawel & Rohn,
2010). Because of these facts it may be preferable to nano-
encapsulate CHAs by forming inclusion complexes with b-cyclo-
dextrin (b-CD) and adding in such form to foods (Nasirullah, Kumar
& Shariff, 2011; Szejtli & Szente, 2005). The aromatic ring of CHAs
located inside the b-CD can be protected against reactions with
food proteins. Such properties of b-CD-CHAs inclusion complexes
during processing with food proteins and protein hydrolysates
were already confirmed in model studies (Budryn et al., 2014;
Budryn, Zaczyn  ska, Rachwał-Rosiak, & Oracz, 2015; Budryn et al.
2015; Budryn et al., 2016; Budryn, Zaczyn  ska, & Rachwał-Rosiak,
2016). Inclusion of CHAs with b-CD presumably does not limit
their bioavailability and antioxidant activity (Paramera, Konteles, &
Karathanos, 2011).
The interactions of polyphenols with proteinaceous prepara-
tions could change not only pro-health activities but also organo-
leptic properties of foods and forming of some specific volatiles
could take place (Ahmad et al., 2015; Noor-Soffalina, Jinap, Naz-
amid, & Nazimah, 2009). Thus the form of added polyphenols and
their ability to interact with proteins would be significant for aroma
volatiles formation. So far no studies on the effect of food

http://dx.doi.org/10.1016/j.lwt.2016.06.019
0023-6438/© 2016 Elsevier Ltd. All rights reserved.
198 G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204
supplementation with phenolic compounds in different form on
aroma volatiles profile were conducted. The aim of this study was
to supplement six food products with free or nanoencapsulated
CHAs, and also with protein hydrolysates, to compare the compo-
sition of headspace aroma volatiles of the obtained products
depending on the used additives.
2. Materials and methods
2.1. Chemicals and raw materials
Analytical-grade ethanol, methanol and ethyl acetate were
purchased from Poch (Gliwice, Poland), b-cyclodextrin (b-CD,

98%) from Sigma Aldrich (St. Louis, MO, USA) and nylon filters
from Chromacol (Herts, UK).
Green Robusta coffee beans (Coffea canephora L.) harvested in
Brazil in 2012, hulled by dry method were purchased from Bero
Polska (Gdynia, Poland). Protein hydrolysates: whey protein hy-
drolysate Amino 4500 (WPH) was purchased from Trec Nutrition
(Gdynia, Poland), egg ovalbumin hydrolysate A 6710 (EOH) from
Sigma (St. Luis, MO, USA) and soy protein hydrolysate S 1674 (SPH)
from Fluka (St. Luis, USA). All the used ingredients for obtaining
food products were purchased from the local market.
2.2. Preparation and purification of green coffee extract (GCE)
The aqueous extract from green coffee beans was obtained as
previously described (Budryn et al. 2014). The extract contained
isomers of caffeoylquinic, feruloylquinic and dicaffeolyquinic acids.
The solution was frozen at 80  C, freeze-dried in a DELTA 1-24LSC
Christ freeze drier (Osterode am Harz, Germany) and purified by
centrifugal partition chromatography (CPC) method with SPOT
Prep II 50 chromatograph from Armen Instrument (Saint-Ave ,
France) integrated with UV/VIS detector and a fraction collector.
Briefly, the two-phase system of solvents was prepared from water,
ethanol and ethyl acetate (5:1:4, v/v/v). Elution of CHAs occurred
from 20 to 24 min and from 29 to 32 min of the analysis, detected
by UV absorption at 320 nm. Collected fractions were concentrated
in ScanMaxiVac concentrator Labogene (Lynge, Denmark) and
again freeze-dried. The content of CHAs in the purified GCE was
564.8 mg g1 db. (dry basis) and caffeine was eliminated. The
remainder of the extract were proteins (29.5 mg g1 db.), sugars
(92.8 mg g1 db.), soluble fiber (121.6 mg g1 db.) and minerals
(191.3 mg g1 db.).
2.3. Preparation of inclusion complexes of b-CD with CHAs from
GCE (b-CD-CHAs)
Inclusion complexes of b-CD with CHAs were prepared with
accordance to the previously developed method (Budryn et al.
2014). Briefly 113.5 mg of b-CD and 130.3 mg of GCE, which con-
tained 70.8 mg of CHAs were dissolved in 2 mL of water. The
complexation was conducted for 2 h at 50  C in a Pierce Reacti-
Therm TS-18821 reactor from Thermo Scientific (Palo Alto, CA,
USA). After complexation the solution was left for 24 h at 0  C and
the suspension was centrifuged in MIKRO 22R centrifuge from
Hettich (Kirchlengern, Germany) at 4  C for 20 min at 10 000  g.
The precipitate was washed twice with 5 mL of methanol, sus-
pended in 5 mL of ice-cold water, frozen at 80  C and freeze-dried.
A mixture of b-CD complexes with particular CHAs was obtained
and characterized by ESI-MS/MS method (Budryn et al. 2014). The
concentration of CHAs in complexes amounted 237.7 mg g1.
2.4. Food products formulations and obtaining
Types of food products were chosen due to their characteristic
taste properties harmonizing with those of CHAs. The obtained
products included: bread (BR), cookies (CO), caramel cottage cheese
(CT), nutty filling (NF), mushroom (MU) and meat stuffing (ME),
and each product was prepared in five modifications comprising:
control (C), supplemented with GCE (GCE), supplemented with
protein hydrolysate (PH), supplemented with both GCE and PH
(GCE-PH), and supplemented with CHAs in the form of inclusion
complexes (b-CD-CHAs). Three different protein hydrolysates were
used, each of them was added to two products, taking into account
organoleptic and functional properties accordant with those of the
products. BR and CO were supplemented with egg ovalbumin hy-
drolysate (EOH), CT and NF with whey protein hydrolysate (WPH),
MU and ME with soy protein hydrolysate (SPH).
The formulations of all prepared food products are given in
Table 1 (Budryn et al., 2016).
Cookies: all ingredients were mixed with blender, formed into
round biscuits with diameter of 40 mm and a height of 3 mm and
baked in a baking oven Piccolo PRO II Winkler Wachtel (Düsseldorf,
Germany) at 220  C for 10 min.
Bread: a half of water amount was mixed with yeast, sugar and a
quarter of the flour amount. After 20 min of pre-fermentation at
33  C remaining ingredients were added. Prepared mixture was left
for 30 min to continue fermentation. Afterwards the loaf was
formed and baked in a baking oven Piccolo PRO II Winkler Wachtel
(Düsseldorf, Germany) at 200  C for 55 min.
Caramel cottage cheese: all the ingredients were mixed with
blender.
Nutty filling: fat was liquefied at 40  C and mixed with the
remaining components with blender.
Mushroom and meat stuffing: shredded onion was fried in rape
oil with mushrooms or pork depending on a kind of stuffing. After
cooling spices, CHAs and optionally soy protein hydrolysate were
added.
2.5. HS-SPME-GC-MS analysis of aroma volatile compounds
The aroma volatiles from headspace (HS) layer were absorbed
by solid phase microextraction (SPME) technique. Absorption of
headspace volatiles was conducted using a Divinylbenzene/Car-
boxen/Polydimethylsiloxane (DVB/CAR/PDMS) fiber (50/30,
80 mm). Before each analysis, the SPME fiber was conditioned in the
dispenser of a gas chromatograph at 250  C. A sample of 500 mg of
a product was taken into a glass vial of 2 mL capacity. The vial was
closed with a cap and PTFE-silicone seal and conditioning for
45 min at 40  C. Subsequently the SPME fiber was placed in HS layer
over the product for 40 min at 25  C. After that the fiber was
inserted to the injector system operating in the splitless mode at
220  C. The fiber was left for 1 min to desorb the volatile
substances.
The analysis of the volatile compounds desorbed from the SPME
fiber was performed using a Varian gas chromatograph GC-450
coupled with a mass spectrometer Saturn 220-MS Varian (GC-
MS) (Palo Alto, CA, USA). GC was equipped with a capillary column
ZB-WAX 30 m  0.25 mm  0.25 mm. Helium was used as GC
carrier gas with a flow of 0.7 mL min1. The used temperature of the
operating program included initial 35  C maintained for 5 min, then
increased by 5  C min1 to 50  C and further increased by
5.5  C min1 to 230  C maintained for 10 min. Mass spectra were
obtained under electron ionization (EI) at 70 eV with m/z range of
30e200. The temperature of the ion source and that of the transfer
line was 180  C and 200  C, respectively. The identification of vol-
atiles was performed by comparing the mass spectra of the
 G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204 199

Table 1 measurements and their standard deviation as well as one-way

Formulation of food products supplemented with hydroxycinnamic acids (HCAs) ANOVA (analysis of variation) and statistical values comparing
from green coffee beans (g 100 g1 of a final product) (Budryn et al. 2016b).
sample groups using the student’s t-test by Statistica 10.0 software
Ingredient CON GCE PRH GCE-PRH b-CD-HCAs at the significance level p < 0.05.
Cookies
Wheat flour 53.3 51.4 48.3 46.4 49.1
Butter 27.0 27.0 27.0 27.0 27.0
Sugar 14.3 14.3 14.3 14.3 14.3
Baking powder 1.3 1.3 1.3 1.3 1.3
Egg yolk 4.1 4.1 4.1 4.1 4.1 3. Results and discussion
Green coffee extract e 1.9 e 1.9 e
Egg ovalbunim hydrolysate (EOH) e e 5.0 5.0 e 3.1. Total aroma volatiles content
b-CD-HCAs inclusion complex e e e e 4.2
Bread
Wheat flour 50.0 48.1 45.0 43.1 45.8
Bread and cookies were obtained at high temperature above
Yeast 5.0 5.0 5.0 5.0 5.0 200  C, comparing to other products, which only contained in-
Sugar 1.0 1.0 1.0 1.0 1.0 gredients derived from heat processing, but after mixing with the
Salt 0.7 0.7 0.7 0.7 0.7 used additives processed at room temperature.
Water 43.3 43.3 43.3 43.3 43.3
From the GC-MS chromatograms (Supplementary Figs. 1e6) it
Green coffee extract e 1.9 e 1.9 e
Egg ovalbunim hydrolysate (EOH) e e 5.0 5.0 e could be observed that the highest concentration of volatile com-
b-CD-HCAs inclusion complex e e e e 4.2 pounds was contained in HS of ME, then CO, NF, CT, BR and MU, and
Caramel cottage on average of five modifications it amounted 329.4, 234.8, 217.6,
Cottage with natural flavor 90.0 88.1 85.0 83.1 85.8 199.0, 196.4 and 163.4 kCounts  min  105, respectively (Fig. 1).
Caramel mass 10.0 10.0 10.0 10.0 10.0
Green coffee extract e 1.9 e 1.9 e
Considering the six food products a general trend of volatiles
Whey protein hydrolysate (WPH) e e 5.0 5.0 e content related to the used different additives was observed. On
b-CD-HCAs inclusion complex e e e e 4.2 average the increase of the concentration of volatiles was induced
Nutty filling by the different additives in the following order: b-CD-
Palm fat 25.0 25.0 25.0 25.0 25.0
CHAs < C < PH < GCE < GCE-PH (Fig. 1, p < 0.05). The primary
Milled wafer 18.0 16.1 18.0 16.1 13.8
Sugar 15.0 15.0 15.0 15.0 15.0 objective of nanoencapsulation of CHAs was to isolate CHAs from
Skimmed milk powder 17.0 17.0 12.0 12.0 17.0 the proteinaceous substances in order to limit their interaction, and
Low-fat cocoa powder 5.0 5.0 5.0 5.0 5.0 consequently to reduce the degradation of the CHAs and to increase
Ground roasted peanuts 20.0 20.0 20.0 20.0 20.0 their bioavailability (Budryn et al., 2016). However, this method
Green coffee extract e 1.9 e 1.9 e
Whey protein hydrolysate (WPH) e e 5.0 5.0 e
also contributed to the reduction of the formation of aroma volatile
b-CD-HCAs inclusion complex e e e e 4.2 substances (GCE vs. b-CD-CHAs, p < 0.05).
Mushroom stuffing The addition of free CHAs in form of GCE to baked products, i.e.
Fried champignons 41.0 39.1 36.0 34.1 36.8 BR and CO caused decrease (p < 0.05) of content of volatile sub-
Fried onion 47.5 47.5 47.5 47.5 47.5
stances comparing to the controls by 7.8 and 12.0%, respectively
Salt 1.0 1.0 1.0 1.0 1.0
Sugar 0.1 0.1 0.1 0.1 0.1 (Figs. 1e3). When CHAs were added in form of b-CD-CHAs the
Pepper 0.2 0.2 0.2 0.2 0.2 content of volatiles decreased by 17.6 and 12.8% for BR and CO,
Rape oil 10.2 10.2 10.2 10.2 10.2 respectively. Both products were supplemented with EOH. Its
Green coffee extract e 1.9 e 1.9 e addition to BR caused an increase of volatiles content by 11.4%,
Soy protein hydrolysate (SPH) e e 5.0 5.0 e
b-CD-HCAs inclusion complex e e e e 4.2
while for cookies opposite e decrease by 9.3% compared to the
Meat stuffing controls. The addition of GCE along with EOH resulted in the for-
Fried minced pork 53.8 51.9 48.8 46.9 49.6 mation of the highest content of volatiles in BR, namely an increase
Fried onion 35.0 35.0 35.0 35.0 35.0 by 22.8% compared to BR-C and in cookies a decrease by 8.9%. Thus
Salt 1.3 1.3 1.3 1.3 1.3
the differences in composition of both products, i.e. lower water
Sugar 0.8 0.8 0.8 0.8 0.8
Pepper 0.3 0.3 0.3 0.3 0.3 activity and higher sugars and fat contents and higher temperature
Dried garlic 0.3 0.3 0.3 0.3 0.3 in cross-section resulted probably in the formation of non-volatile,
Gyro spice 1.0 1.0 1.0 1.0 1.0 more advanced degradation products in cookies.
Herbes de Provence 0.5 0.5 0.5 0.5 0.5 Changes in the content of aroma volatiles in NF and CT despite
Rape oil 7.0 7.0 7.0 7.0 7.0
Green coffee extract e 1.9 e 1.9 e
the use of the same additives did not show the same trends
Soy protein hydrolysate (SPH) e e 5.0 5.0 e (Figs. 1e3). The CT volatiles content increased in the following or-
b-CD-HCAs inclusion complex e e e e 4.2 der: CT-WPH]CT-C]CT-b-CD-CHAs < CT-GCE < CT-GCE-WPH. The
presence of CHAs in their free form, in particular when CT was also
enriched in WPH led to the formation of the highest content of
analyzed compounds with those stored in the NIST 08 MS and volatiles. For NF volatiles content increased in the order: NT-C]NF-
AMDIS 2.6 libraries. For this purpose MS Data Review software was b-CD-CHAs < NF-GCE]NF-GCE-WPH < NF-WPH. The addition of
used. To determine the percent of a particular compound in the WPH caused the highest increase of the content of aroma volatiles,
head space the percent of its peak area in total chromatogram area but the combination of WPH with CHAs in their free form caused
was calculated (Budryn, Nebesny, Kula, Majda, & Krysiak, 2011). lower increase of volatiles (p < 0.05). This observation suggests that
CHAs limited the effect of enhancing the formation of volatile
components by WPH.
2.6. Statistical analysis Both kinds of stuffing revealed similar trends of the effect of
various additives on the content of volatile components, i.e. it
The products were obtained twice. Analyzes were carried out in increased in the order: C < SPH < GCE-SPH < GCE. For MU-b-CD-
triplicate for both replicates. Results were subjected to statistical CHAs the amount of volatiles was higher than in MU-C but the
analysis. It comprised determination of average values of six opposite trend was observed for ME.
200 G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204

400

350 a

Total area (kCounts × min) × 105

a
300

250 a a a b b
a a b
200 b
a,b
a a a
150

100

50

0
BR CO CT NF MU ME

Fig. 1. Total aroma volatile substances as total area of GC-MS chromatograms of six food products supplemented with green coffee extract, nanoencapsulated chlorogenic acids and
protein hydrolysates; e C; e GCE; e PH; e GCE-PH; e b-CD-CHAs; ±SD; different letters within one product or lack of them correspond to significant differences
(p < 0.05), n ¼ 6.

45

40
Content in aroma (% of total area)

35

30
b b

25
b b
20 a a a
a

15

10
a a
a a a a
5
a a
0
Acids Furans Alcohols Pyrazines Aldehydes & Phenols Esters Lactones
ketones

Fig. 2. Volatile substances of bread supplemented with green coffee extract, nanoencapsulated chlorogenic acids and egg ovalbumin hydrolysate; e C; e GCE e EOH; e
GCE-EOH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.

30 b
b
b
b b
Content in aroma (% of total area)

25
a a
b
b
20 a
a a a

15

10
a a

b b
5 a a
a a
a b a b
0
Esters Acids Alcohols Aldehydes & Furans Pyrazines Phenols Pyrroles Lactones
ketones

Fig. 3. Volatile substances of cookies supplemented with green coffee extract, nanoencapsulated chlorogenic acids and egg ovalbumin hydrolysate; e C; e GCE; e EOH; e
GCE-EOH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.

3.2. Aroma volatiles of baked products have a sharp, fatty odor. They may be formed by the Maillard re-
action (MR) (Pico, Bernal, & Gomez, 2015; Makhoul, et al., 2015)
Among the 48 volatiles identified in BR headspace volatile acids (Fig. 2, Supplementary Fig. 1AeE, Supplementary Table 1). BR-C was
dominated, and among them propionic and octanoic acids, which characterized by the highest content of these compounds.
 G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204
Supplementation with the used additives caused a decrease of
volatile acids content. This decline was the most evident for BR-
EOH. The second group of volatiles in terms of their participation
in BR HS aroma were furans. These group of compounds was
formed by caramelization of sugars or MR, and the addition of EOH
resulted in an increase of their content. CHAs, regardless of their
form, caused a decrease in furans content. Among furans 2-acetyl-
3-hydroxy furan (caramel), 2-furfural (spicy, sweet) and 2-
methylfuran (chocolate) were the dominant representatives.
Another important group of volatiles were alcohols, and among
them in particular 2-ethyl-1-decanol (pungent) and ethanol
(spirit). The last one can arise as a result of ethanolic fermentation
carried by yeast. They dominated the in BR enriched with CHAs in
their both forms. Addition of EOH causes decrease in their contents.
The next group of aroma volatiles in terms of high content were
pyrazines formed by MR, particularly pyrazine (nutty), 2-
methylpyrazine (also nutty) and 2-ethyl-5-methylpyrazine (cof-
fee). The enrichment with CHAs even if they were included in b-CD,
contributed to the maximum formation of pyrazines. The addition
of EOH also resulted in an increase of their content. For samples
supplemented with EOH the highest increase of aldehydes & ke-
tones was observed. These were primarily 3-methylbutanal (pun-
gent) belonging to Strecker aldehydes, resulting mainly from
leucine degraded during MR, (Mohsen, Fadel, Bekhit, Edris, &
Ahmed, 2009), nonanal (floral) and hexanal (fresh, green), the
last formed by the oxidation of fatty acids. BR enriched with EOH
was characterized by the highest content of phenols, including 2-
methoxyphenol (wood) and 2-methoxy-4-vinylphenol (smoked
wood), probably formed as a result of thermic degradation of
polyphenols (Nebesny, Budryn, Kula, & Majda, 2007). Lactones
were represented by g-butyrolactone (creamy, BR-EOH). The
addition of EOH contributed to the lack of esters, including ethyl
acetate (sweet), significant in aroma of other BR variants. These
results showed that addition of EOH to BR contributed to the higher
efficacy of MR and forming volatiles with a pleasant aroma (furans,
pyrazines, aldehydes, lactones), and reduction of those with un-
pleasant smell (acids and alcohols). The addition of free CHAs also
influenced favorably formation of volatiles with pleasant aroma.
CHAs were probably oxidized to quinones and interacted with
amino acids forming volatiles similar to MR products that enriched
the flavor of the bread. Hence, the best properties of aroma had BR-
GCE-EOH and this variant contained the largest amounts of
volatiles.
In CO 53 volatiles were identified. The highest contribution to
the aroma of CO had esters, especially ethyl acetate, and acids,
including propionic acid, that was present predominantly in COeC,
as well as octanoic acid, characteristic for all supplemented variants
(Fig. 3, Supplementary Fig. 2AeE, Supplementary Table 2), (Mohsen
et al. 2009). Addition of EOH affected the decrease of esters amount,
and the content of acids was statistically lower in COeC (not
identified) and CO-b-CD-CHAs (p < 0.05). Another important
groups of volatiles identified in the CO were alcohols, and alde-
hydes & ketones, mainly butanal (sharp), characteristic for food
containing butter, and hexanal, formed during decomposition of
fatty acids. In CO enriched in EOH also 3-methylbutanal occurred.
Furans, including 2-acetyl-3-hydroxyfuran and 2-furfural, found in
highest quantities in CO-EOH were less abundant group of volatiles
in the aroma of CO. Pyrazines e the products of MR, including 2-
methylpyrazine (nutty) and 2-isobutyl-3-methylpyrazine
(caramel), showed a similar trend. Among phenols 2-
metoxyphenol and 2-methoxy-4-vinylphenol, in particular in CO-
EOH occurred. The only identified g-butyrolactone was character-
istic also mainly for CO-EOH. For CO small amounts of pyrroles: 2-
acetylpyrrole (musty) and 1-(2-furanylmethyl)-1H-pyrrole (vege-
table) were identified, mainly in CO enriched in both forms of the
201
CHAs. As for BR, also for cookies, an increase of the content of
pyrazines and lactones contributing to the pleasure aroma was
observed along with the enrichment in EOH, particularly when GCE
containing free CHAs was also added. Similar observations for
biscuits enriched with soy protein isolate made Mohsen et al.
(2009).
3.3. Aroma volatiles of desserts
Two groups of volatiles (of total 38 identified) dominated in the
aroma of CT: acids, including propionic and octanoic acids (CT-C)
and esters, represented by ethyl acetate, not identified in CT-C,
however forming from 37 to 43% of the aroma of the other vari-
ants, except of CT-b-CD-CHAs where their content was about half
lower (Fig. 4, Supplementary Fig. 3AeE, Supplementary Table 3).
Among the alcohols dominated the 2-ethyl-1-decanol and 1,2-
propanediol, in the largest quantities contained in CT-C and CT-b-
CD-CHAs. Among ketones the largest share in the flavor accounted
for 2-pentanone (fruity), dominant in the CT-C and CT-b-CD-CHAs.
In the group furans 2-furanmethanol (faint burning) and 2-acetyl-
3-hydroxyfuran were the main volatiles. The other four classes of
the identified volatile, i.e. phenol derivatives (benzyl alcohol);
pyrazines (2-ethyl-6-methylpyrazine) coming probably, as in case
of furans, from the caramel used as a flavoring ingredient
(Paravisini et al., 2015); terpenes (limonene) and lactones (g-
butyrolactone) dominated in CT-C, while their lowest contents
characterized the variants supplemented with WPH. CT was not
processed at high temperature and was characterized by far
different composition of aroma volatiles comparing to baked
products. The most beneficial aroma profile was reported for CT-
GCE (high content of esters and low of acid) and CT-b-CD-CHAs.
CT-C, despite the high content of furans was however characterized
by particularly high content of volatile acids.
The most abundant volatiles from 48 identified in NF were acids,
including propionic and octanoic (Fig. 5, Supplementary Fig. 4AeE,
Supplementary Table 4). The addition of WPH caused a decrease of
their content, in contrast to CHAs, but only in their free form.
Second important class of NF aroma compounds were furans, in
particular 2-furfural and 2-acetyl-5-metylfuran (nutty). Statisti-
cally, the addition of b-CD-CHAs caused an increase in their con-
tents, while the addition of WPH resulted in limiting of their
formation (p < 0.05). Esters (ethyl acetate) dominated in NFeC.
They could derive from cocoa powder, obtained from fermented
and roasted cocoa beans (Pino, Ceballos, & Quijano, 2010). The
lowest concentration of alcohols, including 2-ethyl-1-decanol,
ethanol and 3-methylbutanol (spirit), occurred in NT-GCE-WPH.
For others variants, an increase of their content in comparison
with NFeC was observed. Enrichment with CHAs contributed to a
decrease in headspace of pyrazines content (3-ethyl-2,5-
dimethylpyrazine e earthy, roasted) characteristic for roasted in-
gredients, for example roasted peanuts, which were used to pre-
pare NT (Kaneko, Sakai, Kumazawa, Usuki, & Nashimura, 2013), but
resulted in an increase of aldehydes (hexanal). Phenols (2-
methoxyphenol) present in the largest amounts in NFeC were
also significant for NF aroma. The most preferred aroma com-
pounds were characteristic for NF-b-CD-CHAs (high contents of
furans and aldehydes) and NT-C (high content of esters and
pyrazines).
3.4. Aroma volatiles of fried stuffing
Interesting composition of 46 identified volatiles was observed
in MU. The dominant class of volatile compounds were furans, and
among them 2-butyl-furan (spicy), 2-acetyl-5-methylfuran, 2-(2-
propenyl) furan and furfuryl acetate (fruity) (Fig. 6,
202 G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204

50

45

40

Content in aroma (% of total area) 35

30

25

20

15
b
a a,b
10

5 a a a a b a a b b a a,b
a a b a a,b
a a a
0
Esters Acids Furans Alcohols Phenols Aldehydes & Pyrazines Terpenes Lactones
ketones

Fig. 4. Volatile substances of caramel cottage cheese supplemented with green coffee extract, nanoencapsulated chlorogenic acids and whey proteins hydrolysate; e C; e GCE;
e WPH; e GCE-WPH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.

40

35
Content in aroma (% of total area)

30

25

20
a
a
15

10 a a a
b b
5 a a
a a a
a a a
0
Acids Furans Esters Alcohols Pyrazines Aldehydes & Phenols Lactones Pyrroles Terpenes
ketones

Fig. 5. Volatile substances of nutty filling supplemented with green coffee extract, nanoencapsulated chlorogenic acids and whey proteins hydrolysate; e C; e GCE; e WPH;
e GCE-WPH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.

35
a
a
30
Content in aroma (% of total area)

25

20
b b
15
a a a
a
10 a a a a
a
a
5 a a a a a b b a

0
Furans Aldehydes & Pyrazines Acids Alcohols Phenols Terpenes Sulfuric Esters
ketones compounds

Fig. 6. Volatile substances of mushroom stuffing supplemented with green coffee extract, nanoencapsulated chlorogenic acids and soy proteins hydrolysate; e C; e GCE; e
SPH; e GCE-SPH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.

Supplementary Fig. 5AeE, Supplementary Table 5). Their highest amino acids. Other important classes of MU volatiles were alde-
content was observed in variants enriched with SPH. Similar trends hydes & ketones [hexanal, 5-methyl-(E)-2-hepten-4-one (nutty),
of an increase of content of furans were observed by Misharina et al. 2,3-pentanedione (creamy) and 2,5-hexanedione (buttery)] espe-
(2010), who supplemented dried champignons with a mixture of cially in MU-C and MU-GCE. Other important classes were

60
Content in aroma (% of total area)
50
40 a a
30
20
10
0
Acids
Fig. 7. Volatile substances of meat stuffing supplemented with green coffee extract, nanoencapsulated chlorogenic acids and soy proteins hydrolysate; e C; e GCE;
GCE-SPH; e b-CD-CHAs; ±SD; different letters within one class of chemical compounds or lack of them correspond to significant differences (p < 0.05), n ¼ 6.
pyrazines (pyrazine, 2-methylpyrazine and 2,3,5-trimethylpyrazine
(nutty) with the highest content in MU-C), acids (propionic acid, in
MU-SPH and MU-b-CD-CHAs), alcohols (2-ethyl-1-decanol and 2-
ethyl hexanol (earthy), also in MU-SPH and MU-b-CD-CHAs) and
esters (ethyl acetate, in variants with the addition of CHAs
regardless of the form and the presence of SPH, while absent in
MU-C). The addition of GCE caused an increase in the content of
terpenes (a-pinene e resin) and sulfuric compounds (methyl-
sulphanylmethane). The most preferred composition of the aroma
was characteristic for MU-C and MU-GCE (high ketones content,
including diketones), and MU-GCE had additionally a high content
of esters, and low of acids.
From 37 of the identified volatiles of ME the dominant were
volatile acids (propionic and octanoic acids), especially in ME-C
(Fig. 7, Supplementary Fig. 6AeE, Supplementary Table 6).
Another important classes of compounds identified in ME were
furans (furfuryl acetate and 2-furfural, with the highest content in
ME-SPH), pyrazines (2-ethyl-5-methylpyrazine, 2-isobutyl-3-
methylpyrazine, 2-methylpyrazine, pyrazine, also in ME-SPH), al-
dehydes & ketones (5-methyl-(E)-2-hepten-4-one, 3-
methylbutanal and nonanal, present in low amounts in ME-C), al-
cohols (2-ethyl-1-decanol and 2-ethyl hexanol, with the highest
content in ME-GCE) and esters (ethyl acetate, in variants with the
addition of CHAs). A lower share of ME aroma was accounted for
lactones (g-butyrolactone e important for the pork smell regard-
less of its low content (Benet et al., 2016), in ME-SPH), phenols (4-
ethyl-2-metoxyphenol; in ME-b-CD-CHAs) and terpenes (limonene
in variants with the addition CHAs). Due to the low acids content
and high of furans and pyrazines, the preferred composition of the
aroma was characteristic for ME-SPH.
4. Conclusions
The supplemented food products were characterized by diverse
composition of headspace volatile compounds. The richest source
of aroma components was ME, which contained among others fried
meat but MU of a similar formulation but obtained with fried
mushrooms was characterized by the lowest content of volatiles.
The overall trend showed that the addition of protein hydrolysates
together with free CHAs contributed to the highest increase in the
content of volatile components, particularly of furans and pyr-
azines, which favorably affect the aroma properties. Protein hy-
drolysates containing amino acids and short peptides enhanced
formation of volatiles as a result of Maillard reaction, thus this trend
G. Budryn et al. / LWT - Food Science and Technology 73 (2016) 197e204 203
b a,b
a
a a a a
b b
a a
a a a a
a
Alcohols Furans Pyrazines Aldehydes & Lactones Terpenes Esters Phenols
ketones
e SPH; e
was especially observed in baked products. Supplementation the
products only with free CHAs also resulted in an increase of the
aroma volatiles concentration, but not in case of CHAs included into
b-cyclodextrin. The main aim of nanoencapsulation of CHAs with b-
cyclodextrin was to separate them from the proteinaceous com-
pounds and to reduce interactions between them, and conse-
quently to limit degradation of CHAs. However, inclusion of CHAs
contributed to the limited formation of preferred aroma volatiles,
especially in baked products. It indicates a significant contribution
of not encapsulated CHAs aromatic ring in aroma formation.
Therefore the products supplemented with CHAs in a more
bioavailable inclusion form are in turn characterized worse
organoleptic quality.
Acknowledgment
Authors are grateful for the financial support provided by Nar-
odowe Centrum Nauki (project No. UMO-2011/03/B/NZ9/00745).
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.lwt.2016.06.019.
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