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Cistus extract as a valuable component for enriching wheat bread
Journal Pre-proof Cistus extract as a valuable component for enriching wheat bread Anna Mikulec, Stanisław Kowalski, Małgorzata Makarewicz, Łukasz Skoczylas, Małgorzata Tabaszewska PII:
S0023-6438(19)31055-2
DOI:
https://doi.org/10.1016/j.lwt.2019.108713
Reference:
YFSTL 108713
To appear in:
LWT - Food Science and Technology
Received Date: 21 March 2019 Revised Date:
4 October 2019
Accepted Date: 6 October 2019
Please cite this article as: Mikulec, A., Kowalski, Stanisł., Makarewicz, Mał., Skoczylas, Ł., Tabaszewska, Mał., Cistus extract as a valuable component for enriching wheat bread, LWT - Food Science and Technology (2019), doi: https://doi.org/10.1016/j.lwt.2019.108713. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.
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Cistus extract as a valuable component for enriching wheat bread
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Anna Mikulec1, Stanisław Kowalski2, Małgorzata Makarewicz3, Łukasz Skoczylas4,
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MałgorzataTabaszewska4
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1)
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[email protected]
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2)
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Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
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3)
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Technology, University of Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
State Higher Vocational School in Nowy Sacz, ul. Staszica 1, 33-300 Nowy Sacz, Poland,
Department of Carbohydrate Technology, Faculty of Food Technology, University of
Department of Fermentation Technology and Technical Microbiology, Faculty of Food
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4)
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Technology, University of Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
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Corresponding author:
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Anna Mikulec, e-mail: [email protected], ul. Staszica 1, 33-300 Nowy Sacz, Poland
Department of Technology of Fruits, Vegetables and Mushrooms, Faculty of Food
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Keywords: wheat-cistus bread, bread stalling index, polyphenol profile, microbiological
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properties of the crumb, crumb color
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Abstract. The aim of the study was to use cistus extract for the production of wheat bread and
19
determine its impact on selected physicochemical, microbiological, and organoleptic
20
characteristics, the color of the crumb, changes in the crumb texture, polyphenol profile and
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the total polyphenol content. Breads with 5 and 7.5% cistus extract were characterized by
22
lower average scores for taste and smell, compared to wheat bread. During storage (up to 5
23
days) the largest increase of crumb hardness was observed for wheat bread, as much as
24
72.89%, compared to the first day of storage, in contrast to bread with 5 % cistus extract
25
(29.03%). The replacement of water by cistus extract influenced the color of the crumb by
26
increasing its browning index from 30.92 (standard bread) to 66.47 (7.5% cistus extract). The
27
cistus extract contributed to an improvement of the microbiological quality of the bread. The
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addition of the cistus extract influenced the polyphenol content by increasing it from 8.88
29
(wheat bread) to 78.71 mg/100 g (breads with 7.5 % cistus extract) and the total polyphenol
30
content from 62.81 to 105.81 mg GAE per 100 g of product, compared to the wheat bread.
31 32
1. INTRODUCTION
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Bread is a food product of great importance for consumers. It is still popular and consumed
34
every day in many countries. The assortment of bakery products is constantly expanding with
35
new products, often enriched with products of natural origin. For the enrichment of bread, and
36
thus for a significant change not only of its organoleptic characteristics, but also of health-
37
promoting properties, various additives are used, such as non-cereal grains, herbs, oilseeds
38
and dried fruits and vegetables (Ibrahim, Salleh, & Maqsood-ul-Haque, 2015). From a
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nutritional point of view, cistus (Cistus incanus L.) is a valuable source of many antioxidant
40
compounds including ellagic and gallic acids, terpenoids, flavonoids and tannins (Viapiana,
41
Konopacka, Waleron, & Wesolowski, 2017). Cistus has strong anti-inflammatory properties
42
(Benkhnigue, Hachi, Fadli, Douira, & Zidane, 2016; Küpeli, & Yesilada, 2007; Sayah et al.,
43
2017) and, among others, antileukemic, antioxidant, antiperoxidant, antiradicular, antiseptic,
44
antiulcer, astringent, bactericide, candidicide, cardioprotective, dermoprotective, fungicide,
45
gastroprotective, hemostat, myorelaxant, nervine, purgative, revulsive, sedative, and
46
spasmolytic properties (Duke, Duke, & duCellier, 2008). It was also found that the
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polyphenol compounds contained in cistus can inhibit the development of tumors,
48
demonstrating cytotoxic activity (Barrajón-Catalán et al., 2010; Vitali, Pennisi, Attaguile,
49
Savoca, & Tita, 2011). The beneficial effects of various species of cistus against pathogenic
50
microorganisms, bacteria or fungi have also been proved (Tomás-Menor et al., 2013;
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Viapiana et al., 2017). In-vitro studies confirmed that the extract of cistus inhibits
52
multiplication and reduces the population of the bacterium Borrelia burgdorferi, the
53
etiological factor of borreliosis (Hutschenreuther, Birkemeyer, Grötzinger, Straubinger, &
54
Rauwald, 2010) and inhibits human immunodeficiency virus (HIV) infections (Rebensburg et
55
al., 2016). Kuchta et al. (2019) observed that use of Cistus incanus herbal tea on a daily basis
56
can reduces cardiovascular risk factors including dyslipidemia and oxidative stress. Lisiecka,
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Wójtowicz, Dziki, Gawlik-Dziki (2019) enriched wheat pasta with the addition of Cistus
58
incanus leaves and observed an increase in the total phenolic content (TPC) and the
59
antioxidant activity of supplemented pasta. Cacak-Pietrzak et al. (2019) supplemented wheat
60
bread with the addition of 1 up to 5 % of ground Cistus incanus and also observed
61
significantly higher TPC and much higher antioxidant activity of bread with this herb’s
62
leaves. To date, no research has been conducted to determine the effect of cistus extract on
63
bread characteristics. Moreover, other authors used dried Cistus incanus leaves for the
64
production of bread and pasta (Cacak-Pietrzak et al., 2019; Lisiecka et al., 2019), which could
65
be less effective than using it in liquid form.
66
For this reason, appreciating the potential nutritional value of cistus extract in bread
67
production, it was decided to study the possibility of using this extract for enrichment of bread
68
products. The aim of the study was to analyze and compare selected physicochemical,
69
antioxidant, textural and microbiological properties, as well as the technological and
70
organoleptic quality of bread in which water was replaced with 2.5, 5 and 7.5% cistus extract
71
on the baking day and up to 5 days.
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2. MATERIALS AND METHODS
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2.1. Materials
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The study material consisted of wheat bread (WS) obtained from wheat flour type 550 (PZZ
75
Krakow, Poland), the same as used to in previous research (Mikulec et al., 2019), and wheat
76
bread in which water prescribed by the recipe was replaced with cistus extract in the
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concentrations 2.5, 5 and 7.5 % w/v (Bellis Pharma sp. z o.o., Jaroslaw, Poland). The dough
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used for baking of bread consisted of 7.5 kg of wheat flour, 4.60 dm³ of cistus extract, 0.23 kg
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of yeast (Lesaffre, S.A., Wolczyn, Poland) and 0.15 kg of salt (Solino, Inowroclaw, Poland).
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The baking was performed twice, each time obtaining 30 pieces of bread.
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2.1.1. Bread preparation
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The dough was prepared using a single-phase method in a fast rotating spiral mixer (Ibis
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MS 130 Poland). After initial fermentation (60 min, 21 ºC, 55% relative humidity (RH)), the
84
dough was divided into 405 ± 5 g, pieces, formed and placed in steel molds. After the
85
fermentation (in a fermentation cabinet by MIWE GVA (Germany)) (35 min, 32 ºC, 90%
86
RH), the bread was baked in an MIWE IDEAL (Germany) batch oven at 230 °C for 32
87
minutes. After removal from molds, bread was cooled down for two hours at 19 °C and used
88
for further analyses.
89
2.2. Methods
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2.2.1. Analysis of basic quality features
91
Bread volume was measured by the AACC-approved method 10-05.01 (AACC,
92
2000), total baking loss was calculated as the difference between the mass of the raw roll and
93
the mass of the roll one hour after baking (Majzoobi, Farahnaky, & Agah, 2011), and crumb
94
moisture was estimated using the gravimetric method (AOAC 925.10).
95
Analysis of crumb color according to the CIELab system was performed by the
96
reflectance method using a Konica Minolta CM-3500d spectrophotometer (Konica Minolta
97
Sensing, Osaka, Japan; illuminant D65, observer 10°). The total color difference was
98
calculated according to Fernandez-Artigas, Guerra-Hernandez, & Garcia-Villanova (1999)
99
using the formula (Eq. 1):
100
∆ = √∆
+∆
+∆
(Eq. 1)
101
where:
102
∆E = the total color difference,
103
∆L = brightness difference,
104
∆a = redness difference,
105
∆b = yellowness difference.
106
Color measurements were performed in twelve replicates per loaf. On the basis of L*a*b*
107
parameters, the browning index (BI) was estimated (Buera, Retriella, & Lozano, 1985) as
108
follows (Eq. 2):
109
=
[
.
]
(Eq. 2)
.
110
where:
111
=
.
∗
.
∗
∗
∗
.
∗
112
L = brightness,
113
a = redness,
114
b = yellowness.
115
2.2.2. Microbiological analysis
116
Microbiological analyses of the bread were performed during the 1st and the 5th day of
117
storage. Samples were obtained from a different part of the loaf (including the crust and
118
crumb). To determine the number of aerobic amylolytic bacteria 1 cm3 of each decimal
119
dilution was transferred to three parallel Petri dishes and poured with Waksman medium
120
(Biocorp). The plates were incubated at 37 °C for 48 h. The reading of the results consisted in
121
filling the plate with Lugol's liquid and counting the colonies around which the yellow, clear
122
zone was formed. To determine the number of yeast and mold, from two successive decimal
123
dilutions, 1 cm3 was transferred into three parallel Petri dishes and poured with Sabouraud
124
agar with chloramphenicol (Biocorp). The plates were incubated at 25 °C for 3 to 5 days. The
125
obtained results were counted as cfu/g bread.
126
The determination of the microbiological stability of the bread consisted of thermostating the
127
product sample at 30 ºC and observing it every 24 hours to determine possible organoleptic
128
changes caused by molds or at 37 ºC to determine changes caused by aerobic amylolytic
129
bacteria. Time of occurrence of the first organoleptic changes (mold, malleability,
130
deformability or crumb viscosity) determined the shelf life of the product. Measurements
131
were done in two replicates.
132
2.2.3. Texture analysis
133
Selected mechanical features of the breadcrumb, such as hardness, springiness, resilience,
134
chewiness and gumminess (Szcześniak, 1963), was performed using a single-arm
135
TA.XT2.Plus texture analyzer (Stable Micro System Ltd, Godalming, Surrey, United
136
Kingdom). Breadcrumb samples were sliced mechanically and the slices for analysis were cut
137
from the middle part of the loaf. The tests were done on cylindrical samples (r=30 mm, h=35
138
mm). A P/36R aluminum probe with a diameter of 36 mm was used, with the compression
139
rate 2 mm·s-1 to reach 50% deformation in two cycles with 5 s waiting time between the
140
cycles (Mikulec et al., 2019). Bread for examination of the crumb texture changes during
141
storage were packed in polyethylene film pouches (HDPE) and stored at ambient temperature
142
(19 ± 1 °C, 60% RH). The analysis was done at 1st and the 5th day of storage. Measurements
143
were done in ten repetitions.
144
To assess the changes in texture caused by storage, the percentage changes of hardness,
145
chewiness, gumminess, springiness and resilience were calculated according to Różyło (2014)
146
with modification (Eq. 3): !"# !%#
147
=
148
Where:
149
x 5d = textural property marked after 5 days of storage,
150
x 1d = textural property marked after 1 day of storage.
!%#
∗ 100
(Eq. 3)
151
2.2.4. Sensory analysis
152
Consumer acceptability tests were performed by a panel consisting of 60 untrained consumers
153
(20–51 years old, 43 females and 17 males) who were recruited from the Institute of
154
Technology (State Higher Vocational School in Nowy Sacz) (staff and students) and who
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were habitual consumers of bread. Assessors were asked demographic questions about age
156
and gender and a question about bread consumption. Two hours after baking bread samples
157
were sliced mechanically into 20 mm slices and were cooled until bread reached 19 °C. Then
158
slices were placed on disposable paper trays and coded with two digits random numbers.
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Samples were presented to the assessors simultaneously. The bread was evaluated only on the
160
day of baking using a 9-point hedonic scale (1 = dislike extremely; 2 = dislike very much; 3 =
161
dislike moderately; 4 = dislike slightly; 5 = neither like nor dislike; 6 = like slightly; 7 = like
162
moderately; 8 = like very much; 9 = like extremely), as described by Meilgaard, Civille, &
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Carr (2007). During the assessment the assessors neutralized the taste using pure water at
164
temperature 19 °C. The results were calculated by Microsoft Excel 10.0 software. The
165
assessors followed the ethical requirements of the sensory laboratory approved by the State
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Higher Vocational School in Nowy Sacz and an informed consent form was signed by the
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panelists.
168
2.2.5. Total phenolic content
169
Total phenolic content was estimated using Folin-Ciocalteu reagent (Meda, Lamien, Romito,
170
Millogo, & Nacoulma, 2005). Samples were prepared at 0.05 g/mL concentration using water
171
solution as described by Mikulec et al. (2019). Results were expressed as mg of gallic acid
172
equivalent (GAE) per 100 mg of product. Measurements were done in two replicates.
173
2.2.6. Chromatographic analysis of polyphenol profile
174
Separation and identification of polyphenols were carried out using high-performance liquid
175
chromatography (HPLC) according to the method described by Klimczak, Małecka, Szlachta,
176
& Gliszczyńska-Świgło (2007) with modification as described by Mikulec et al. (2019).
177
Briefly, Dionex Ultimate 3000 HPLC set equipped with DAD detector (Thermo Scientific,
178
Germering, Germany), fitted with Cosmosil 5C18 – MS - II 250 x 4.6 mm ID, 5 µm particle
179
size (Nacalai Tesque, INC. Kyoto, Japan) column was used. Gradient elution was used
180
(mobile phase: A - 2% aqueous solution of acetic acid, and B - 100% methanol); flow rate
181
was 1 mL/min. using the following gradient: Eluent A -0 min 95%, 10 min. 70%; 25 min.
182
50%; 35 min. 30%; 40 min. 95%; to the end of analysis. Measurements were done in two
183
replicates.
184
2.2.7. Statistical analysis
185
Statistical analysis was carried out using Statistica 13.0 (StatSoft, Poland). The significance of
186
differences was verified with the Duncan test at p<0.05. The results were presented as
187
averages ± standard deviation.
188
3. RESULTS AND DISCUSSION
189
3.1. Analysis of basic quality features
190
No influence of cistus extract on baking loss or volume of bread was observed, which ranged
191
from 11.13% and 845 cm3 for bread with 7.5% cistus extract to 11.46% and 865 cm3 for
192
wheat bread respectively. The moisture content of the crumb on the day of baking did not
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differ significantly among all breads and ranged from 46.34 to 46.96% (Table 1). Similar
194
results was observed by Pasrija, Ezhilarasi, Indrani, & Anandharamakrishnan (2015), who
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added a green tea extract and encapsulates to bread and did not observe any change in bread
196
volume. Other results were obtained by Cacak-Pietrzak et al. (2019), who observed a
197
significant increase in moisture of bread (which increased with cistus content), a baking loss
198
from 3 to 5% concentrations of cistus leaves in bread and a decrease in bread volume from
199
2% addition of this herb. A similar trend, to that reported by Cacak-Pietrzak et al. (2019) was
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observed in another study by Ning, Hou, Sun, Wan, & Dubat (2017) where specific volume
201
decreased with higher concentrations of green tea powder.
202
Breads with cistus extract were characterized by significantly lower lightness (brightness) of
203
the crumb, compared to wheat bread (Table 1). Bread with cistus extract differed significantly
204
in terms of the red component (redness) content (a) in the crumb compared to wheat bread.
205
The lowest level was recorded for wheat bread (1.00), and bread with cistus extract ranged
206
from 4.25 to 4.29. In the case of the yellowness component (b), wheat bread showed the
207
lowest value (20.03), and this value significantly increased with the increase of cistus extract
208
content to 27.45 in WCE7.5 (Table 1). The total color difference (△E) increased significantly
209
with the increase of cistus extract (Table 1). According to total color difference values, the
210
color difference between control and cistus extract bread was perceptible with close
211
observation since ∆E values were lower than 3 (Hernández-Carrión et al., 2015). All breads
212
differed significantly in terms of the browning index (BI) value. The lowest value was found
213
for standard bread (30.92), and the highest was found for WCE7.5 (66.47) (Table 1). Cacak-
214
Pietrzak et al. (2019) also observed a significant difference in the color of bread with cistus
215
leaves from the control bread. The differences in color parameters of breads were caused by
216
the pigments present in Cistus incanus.
217
3.2. Microbiological analysis
218
The cistus extract contributed to an improvement of the microbiological quality of the bread.
219
On the baking day, breads with a cistus extract were characterized by a lower content of
220
microorganisms, compared to wheat bread (Table 1). In all breads in which the water was
221
replaced by the cistus extract there was observed a decrease in the total number of bacteria
222
and the total number of spores of amylolytic bacteria and a reduction of total number of
223
aerobic amylolytic bacteria on the fifth day of storage. Inhibition of microbiological changes
224
in bread with 7.5% cistus extract was observed, compared to wheat bread (Table 1).
225
The time of occurrence of the first organoleptic changes and the positive thermostat test result
226
determined the shelf life of the product. When analyzing thermostatic stability, there were
227
observed in wheat bread crumb, from the 3rd day of storage, both visible mold (Aspergillus
228
niger) and changes in the crumb indicating the development of Bacillus bacteria (soft, sticky
229
and viscous breadcrumb). In the WCE2.5 and WCE5 bread, no mold-like changes were
230
observed until the 5th day, but from the 3rd day changes in the crumb indicating the
231
development of Bacillus bacteria were visible. In WCE7.5 bread, no mold-like changes were
232
observed and the bacterial-like changes were characterized by slight variation. Based on these
233
results it can be concluded that the addition of cistus extract contributed to improvement of
234
the microbiological quality of the tested bread.
235
The obtained effect may result from the antibacterial properties of the cistus extract observed
236
by Barrajón-Catalán et al. (2010) in their studies. They observed antibacterial properties of
237
extracts from Cistus ladanifer, which inhibits growth of the Gram-positive bacterium
238
Staphylococcus aureus, and extract from Cistus populifolius reveals high growth inhibitory
239
activity against the Gram-negative bacterium Escherichia coli. Tomás-Menor et al. (2013)
240
posited that the cause of the inhibitory activity against bacteria may be related to the presence
241
of galloylated flavanols and specific flavonols and polar compounds and to other flavonols.
242
3.3. Texture analysis
243
The stalling process leads to changes in the texture and reduction of fresh bread
244
characteristics, which results in a short shelf life and best before date (Gray & Bemiller, 2003;
245
Ribotta & Le Bail, 2007). During storage (up to 5 days) an increase in breadcrumb hardness
246
was observed for all breads. The largest increase was characterized by standard wheat bread,
247
as much as 72.89%, compared to the first day of storage, and the smallest for WCE5 bread
248
(29.03%). There was a reduction in the springiness and resilience of all loaves at a similar
249
level. The largest increase in gumminess and chewiness was observed in wheat bread,
250
whereas in bread with cistus extract a reduction of chewiness was observed. The greatest
251
variability of texture parameters was identified for breadcrumb of standard wheat bread
252
(Table 1) and the smallest for bread with cistus extract, which can be considered as an
253
inhibitor of staling of the breadcrumb. Others authors have observed different dynamics of
254
changes in the texture features of bread, both increases and decreases of such parameters as
255
hardness, springiness, cohesiveness, and chewiness depending on the form of the additive.
256
The changes of bread crumb texture are probably a result of change in the nature of starch,
257
dilution of gluten and enrichment of fibre. It can be seen that the addition of herbal extracts
258
had a rather positive effect on the textural properties of the bread (Cacak-Pietrzak et al., 2019;
259
Ning et al., 2017; Pasrija et al., 2015).
260
3.4. Sensory analysis
261
In the organoleptic evaluation, wheat bread and bread with a 2.5% cistus extract did not differ
262
statistically significantly and obtained the highest average marks for specific characteristics,
263
which ranged from 8.8 to 8.9 and 8.7 to 8.8 respectively (Figure 1). Bread with 5% and 7.5%
264
of cistus extract differed significantly in the results of the sensory evaluation compared to WS
265
and WCE2.5. Breads with 5 and 7.5% cistus extract did not differ significantly for taste and
266
smell or crumb porosity. The scores for other features were statistically different. WCE5 and
267
WCE7.5 received the lowest marks for taste and smell (7.5 and 7.3 respectively) (Figure 1).
268
Bread with a 5% cistus extract received the highest marks for appearance and crumb porosity
269
(8.6 and 8.5 respectively) and bread with a 7.5% cistus extract for crumb porosity (8.3) and
270
appearance and crust thickness (8.1.) (Figure 1). Cacak-Pietrzyk et al. (2019) observed
271
deterioration in the organoleptic characteristics of taste, aroma and overall acceptability with
272
a 4% addition of the cistus leaves. Deterioration of the organoleptic properties of bread which
273
were observed in our research can also be seen in bread enriched with herbs such as basil,
274
oregano, and thyme, which can be explained by the content of essential oils that cause its
275
specific taste and smell (Adams, Kruma, Verhé, De Kimpe, & Kreicbergs, 2011).
276
3.5. Total polyphenol content and chromatographic analysis of polyphenol profile
277
The addition of the cistus extract clearly contributed to increasing the antioxidative potential
278
of the obtained bread products (Table 2). Breads with the addition of cistus extract were
279
characterized by a higher total polyphenol content, from 62.81 to 105.81 mg GAE per 100 g
280
of product, compared to the standard (41.23 mg GAE per 100 g). Along with the addition of a
281
more concentrated cistus extract, an increase in the sum of polyphenols determined
282
chromatographically in breads was observed from 34.68 (WCE2.5) to 78.71 mg/100 g
283
(WCE7.5). Cacak-Pietrzak et al. (2019) and Lisiecka et al. (2019), who enriched pasta and
284
bread with dried cistus leaves, observed significantly higher TPC and much higher
285
antioxidant activity of breads and pasta compared to the control sample. The bread enriched
286
with Cistus incanus was characterized by significantly higher TPC already at 1% of the
287
amount of this additive (Caca-Pietrzak et al., 2019). Breads enriched with herbs are
288
characterized by beneficial features in many aspects such as high antioxidant content (Ibrahim
289
et al., 2015). Cistus herb exhibited the highest content of polyphenols (2635.16 mg/100 g),
290
and the dominant polyphenols were (+) catechin (726.65) and ellagic acid (637.18 mg/100 g)
291
(Table 2). Wheat flour and wheat bread were characterized by the lowest polyphenol content
292
(8.50 and 8.88 mg/100 g respectively). No rutin, ellagic acid, 3-hydroxybenzoic acid, (+)
293
catechin, protic acid or t-cinnamic acid was found in the wheat flour or wheat bread in the
294
polyphenol profile. Chlorogenic and t-cinnamic acid were not found in bread with cistus
295
extract despite being present in the cistus herb (93.10 and 0.12 mg/100 g respectively). The
296
content of polyphenols in bread obtained with the addition of cistus extract increased as a
297
result of the substitution of water with this extract. Nevertheless, the polyphenol profile
298
showed quantitative, as well as qualitative, changes. Gallic acid appeared in wheat bread
299
(2.14), but was not present in wheat flour, and there was no phloridzin in wheat bread,
300
although it was detected in wheat flour (0.74 mg/100 g). Some fluctuations were also
301
observed in the content of the other polyphenols. In contrast to the other polyphenols
302
measured by chromatography, more ferulic acid and rutin were determined in bread with the
303
cistus extract in comparison to the amount resulting from the calculation (Table 2). These
304
changes are the result of heat treatment (baking process). Riehle, Vollmer, and Rohn (2013)
305
reported that when preparing C. incanus beverages, decreased amounts of phenolic substances
306
and reduced antioxidant activity are observed if an incorrect selection of brewing process
307
parameters (brewing water, temperature, and duration) is made. Spontaneous changes in
308
polyphenols can occur at elevated temperatures. Such relationships were also observed in the
309
case of grape pomace heating (Chamorro, Goni, & Viveros, 2012). A significant increase in
310
the total polyphenol content was also observed, along with an increase in the cistus extract
311
content in bread (Table 2).
312
4. CONCLUSION
313
Cistus extract has a high nutritional and pro-health value, so it could be used as a valuable
314
component of bread. To date there are no studies concerning utilization of cistus extract in
315
bread formulations, so the research was
316
microbiological, and organoleptic characteristics, polyphenol profile and the total polyphenol
317
content as nutrients in bread. Cistus extract influenced the color of the crumb by increasing its
318
browning index in comparison to wheat bread. The total color difference (△E) increased
319
significantly with the increase of cistus extract and the total color difference between control
320
and cistus extract breads was perceptible with close observation. The cistus extract positively
321
influenced microbiological properties of bread enhancing its shelf life. Both on the day of
322
baking and after 5 days of storage bread with a cistus extract was characterized by a lower
323
content of microorganisms, compared to wheat bread. During storage (up to 5 days) the
focused on selected physicochemical,
324
greatest variability of texture parameters was identified for wheat breadcrumb and the
325
smallest for bread with cistus extract, which can be considered as an inhibitor of staling of the
326
breadcrumb. The observed effect is advantageous, because crumb hardening is one of the
327
main symptoms of bread staling. The addition of the cistus extract positively influenced
328
antioxidant properties of bread by increasing the total polyphenol content. The results show
329
that preparations, based on cistus extract, may be valuable components of bread significantly
330
improving its nutritional and pro-health value, with almost unchanged sensory attributes and
331
limiting the aging of the bread.
332 333
ACKNOWLEDGEMENTS
334
This Research was finance by the Ministry of Science and Higher Education of the Republic
335
of Poland.
336
Statistical analysis was performed using the skills acquired during the training entitled
337
"Application of statistics and Statistica in planning empirical research and developing its
338
results in natural sciences - basic methods" implemented as part of the project: "Integrated
339
Program of the Agricultural University H. Kołłątaja w Krakowie ”, co-financed from
340
European Union funds.
341
Declarations of interest: none.
342 343
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455
Figure 1. Results of organoleptic analysis of tested breads
456
Explanatory notes:
457
…….
458
with 5 % cistus extract, ⸺ WCE7.5 – bread with 7.5 % cistus extract
WS – wheat bread, - - - WCE2.5 – bread with 2.5 % cistus extract, ⸺⸺ WCE5 – bread
Table 1. Evaluation of selected quality parameters of studied breads Bread type
WS*
WCE2.5
WCE5
WCE7.5
Physico-chemical parameters of bread Total baking loss [%]
11.46**a ± 0.42
11.16a ± 0.26
11.33a ± 0.28
11.13a ± 0.34
Volume [cm³]
865.00a ± 17.84
859.67a ± 15.17
856.67a ± 12.89
845.0a ± 16.46
Moisture of bread crumb [%]
46.81a ± 0.08
46.96a ± 0.19
46.34a ± 0.13
46.48a ± 0.96
Color parameters of bread crumb c
L*** [-]
75.17** ± 0.87
60.86b ± 0.77
58.94a ± 0.78
58.18a ±0.78
a [-]
1.00a ± 0.08
4.29b ± 0.16
4.25b ± 0.22
4.26b ± 0.22
b [-]
20.03a ± 0.74
22.53b ± 0.46
25.41c ± 0.54
27.45d ± 0.40
a
b
∆E [-]
-
14.90 ± 0.73
17.41 ± 0.48
18.83c ± 0.75
BI [-]
30.92a ± 1.68
49.85b ± 1.28
59.53c ± 1.91
66.47d ± 1.94
Microbiological characteristic of bread (cfu / g) I day TNB****
1.9 x 103
3.3 x 101
4.7 x 101
3.0 x 101
NAAB
1.1 x 103
3.0 x 101
2.7 x 101
2.3 x 101
NSAB
6.7 x 102
2.0 x 101
6.8 x 101
0.5 x 100
Microbiological characteristic of bread (cfu / g) V day TNB
8.3 x 105
4.2 x 105
2.2 x 105
1.6 x 104
NAAB
8.1 x 105
0.0
6.0 x 104
0.0
NSAB
4.2 x 105
1.8 x 104
1.5 x 101
4.0 x 103
Changes in texture after five day storage [%] Hardness [N]
72.89c ± 2.31
36.73b ± 0.51
29.03a ± 0.47
29.27a ± 0.33
Springiness [-]
-7.61b ± 0.57
-8.78ab ± 0.66
-8.49ab ± 0.33
-9.09a ± 0.59
Gumminess [N]
19.59d ± 1.01
2.25a ± 0.11
3.04b ± 0.09
5.14c ± 0.25
Chewiness [N x mm]
8.44d ± 0.44
-2.62c ± 0.09
-9.77a ± 0.37
-6.62b ± 0.12
Resilience [-]
-33.33a ± 1.01
-40.00b ± 0.91
-40.00b ±1.11
-40.63b ± 1.29
Explanatory notes: * WS – wheat bread. WCE2.5 – bread with 2.5 % cistus tea. WCE5 - bread with 2.5 % cistus tea. WCE7.5 - bread with 7.5 % cistus tea **values in the same row marked with different letters are statistically significantly different at p≤ 0.05 ± SEM *** L – lightness. a – redness. b – yellowness. △E- total color difference. BI - browning index ****TNB - total number of bacteria. NAAB - total number of aerobic amylolytic bacteria. NSAB- total number of spores amylolytic bacteria
Table 2. Polyphenol profile and the total polyphenol content of bread Sample
chlorogenic acid
p-coumaric acid
ferulic acid
rutin
ellagic acid
phloridzin
3hydroxybenzo ic acid
(+) catechin
protic acid
gallic acid
t-cinnamic acid
total
TPC***
mg GAE* 100g-1
[mg*100g-1] Wheat flour WF550*
0.00**±0.00
0.30±0.02
7.46±0.20
0.00±0.00
0.00±0.00
0.74±0.08
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
8.50±0.23
0.00±0.00
8.88c±0.15
48.12±0.77
Bread d
c
WS
0.00±0.00
0.30 ±0.01
6.43 ±0.13
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
2.14c±0.06
WCE2.5
0.00±0.00
1.07c±0.01
7.34a±0.25
2.85b±0.12
9.10c±0.41
1.17c±0.08
1.78b±0.41
6.94c±0.91
2.50c±0.49
1.93c±0.06
0.00±0.00
34.68b±1.58
WCE5
0.00±0.00
1.81a±0.02
7.01b±0.16
7.36a±0.49
23.40b±2.39
1.56b±0.08
4.11a±0.66
16.40b±1.44
5.93b±0.47
4.80b±0.36
0.00±0.00
72.38a±5.71
WCE7.5
0.00±0.00
2.26a±0.03
6.51c±0.18
7.72a±0.14
26.57a±0.34
1.83a±0.03
3.87a±0.55
18.20a±0.72
6.56a±0.19
5.19a±0.10
0.00±0.00
78.71a±0.89
cistus 93.10±1.77 73.95±10.27 35.48±4.17 231.55±8.44 637.18±53.00 37.59±0.51 192.90±4.74 726.65±118.9 313.69±68.75 herb 4 cistus extract 2.33 1.85 0.89 5.79 15.93 0.94 4.82 18.17 7.84 ****2.5% cistus extract 4.66 3.69 1.77 11.58 31.86 1.88 9.65 36.33 15.68 ****5.0% cistus extract 6.98 5.55 2.66 17.37 47.79 2.82 14.47 54.49 23.53 ****7.5% Explanatory notes: * WF- wheat flour, WS – wheat bread, WCE2.5 – bread with 2.5 % cistus tea, WCE5 - bread with 2.5 % cistus tea, WCE7.5 - bread with 7.5 % cistus tea **values in the same column marked with different letters are statistically significantly different at p≤ 0.05 ± SEM *** TPC –the total phenolic content **** values calculated on the basis of individual compound concentration in cistus herb
292.94±11.07
0.12±0.02
2635.16±243.37
7.32
0.00
65.88
14.65
0.01
131.76
21.97
0.01
197.64
41.23d±0.7 3 62.81c±0.70 82.73b±0.5 1 105.81a±2.1 5
Cistus 930.76 ±9.72 23.27 36.55 69.81
1. Cistus extract enhanced antioxidant properties of bread 2. Cistus extract had a positive effect on textural features of the bread 3. Cistus extract inhibited the growth of microflora improving shelf life of bread 4. Cistus extract influenced the color of the crumb by increasing its browning index
Declaration of interests ☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. ☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:
S0023-6438(19)31055-2
DOI:
https://doi.org/10.1016/j.lwt.2019.108713
Reference:
YFSTL 108713
To appear in:
LWT - Food Science and Technology
Received Date: 21 March 2019 Revised Date:
4 October 2019
Accepted Date: 6 October 2019
Please cite this article as: Mikulec, A., Kowalski, Stanisł., Makarewicz, Mał., Skoczylas, Ł., Tabaszewska, Mał., Cistus extract as a valuable component for enriching wheat bread, LWT - Food Science and Technology (2019), doi: https://doi.org/10.1016/j.lwt.2019.108713. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.
1
Cistus extract as a valuable component for enriching wheat bread
2
Anna Mikulec1, Stanisław Kowalski2, Małgorzata Makarewicz3, Łukasz Skoczylas4,
3
MałgorzataTabaszewska4
4
1)
5
[email protected]
6
2)
7
Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
8
3)
9
Technology, University of Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
State Higher Vocational School in Nowy Sacz, ul. Staszica 1, 33-300 Nowy Sacz, Poland,
Department of Carbohydrate Technology, Faculty of Food Technology, University of
Department of Fermentation Technology and Technical Microbiology, Faculty of Food
10
4)
11
Technology, University of Agriculture in Krakow, ul. Balicka 122, 30-149 Krakow, Poland
12
Corresponding author:
13
Anna Mikulec, e-mail: [email protected], ul. Staszica 1, 33-300 Nowy Sacz, Poland
Department of Technology of Fruits, Vegetables and Mushrooms, Faculty of Food
14 15
Keywords: wheat-cistus bread, bread stalling index, polyphenol profile, microbiological
16
properties of the crumb, crumb color
17 18
Abstract. The aim of the study was to use cistus extract for the production of wheat bread and
19
determine its impact on selected physicochemical, microbiological, and organoleptic
20
characteristics, the color of the crumb, changes in the crumb texture, polyphenol profile and
21
the total polyphenol content. Breads with 5 and 7.5% cistus extract were characterized by
22
lower average scores for taste and smell, compared to wheat bread. During storage (up to 5
23
days) the largest increase of crumb hardness was observed for wheat bread, as much as
24
72.89%, compared to the first day of storage, in contrast to bread with 5 % cistus extract
25
(29.03%). The replacement of water by cistus extract influenced the color of the crumb by
26
increasing its browning index from 30.92 (standard bread) to 66.47 (7.5% cistus extract). The
27
cistus extract contributed to an improvement of the microbiological quality of the bread. The
28
addition of the cistus extract influenced the polyphenol content by increasing it from 8.88
29
(wheat bread) to 78.71 mg/100 g (breads with 7.5 % cistus extract) and the total polyphenol
30
content from 62.81 to 105.81 mg GAE per 100 g of product, compared to the wheat bread.
31 32
1. INTRODUCTION
33
Bread is a food product of great importance for consumers. It is still popular and consumed
34
every day in many countries. The assortment of bakery products is constantly expanding with
35
new products, often enriched with products of natural origin. For the enrichment of bread, and
36
thus for a significant change not only of its organoleptic characteristics, but also of health-
37
promoting properties, various additives are used, such as non-cereal grains, herbs, oilseeds
38
and dried fruits and vegetables (Ibrahim, Salleh, & Maqsood-ul-Haque, 2015). From a
39
nutritional point of view, cistus (Cistus incanus L.) is a valuable source of many antioxidant
40
compounds including ellagic and gallic acids, terpenoids, flavonoids and tannins (Viapiana,
41
Konopacka, Waleron, & Wesolowski, 2017). Cistus has strong anti-inflammatory properties
42
(Benkhnigue, Hachi, Fadli, Douira, & Zidane, 2016; Küpeli, & Yesilada, 2007; Sayah et al.,
43
2017) and, among others, antileukemic, antioxidant, antiperoxidant, antiradicular, antiseptic,
44
antiulcer, astringent, bactericide, candidicide, cardioprotective, dermoprotective, fungicide,
45
gastroprotective, hemostat, myorelaxant, nervine, purgative, revulsive, sedative, and
46
spasmolytic properties (Duke, Duke, & duCellier, 2008). It was also found that the
47
polyphenol compounds contained in cistus can inhibit the development of tumors,
48
demonstrating cytotoxic activity (Barrajón-Catalán et al., 2010; Vitali, Pennisi, Attaguile,
49
Savoca, & Tita, 2011). The beneficial effects of various species of cistus against pathogenic
50
microorganisms, bacteria or fungi have also been proved (Tomás-Menor et al., 2013;
51
Viapiana et al., 2017). In-vitro studies confirmed that the extract of cistus inhibits
52
multiplication and reduces the population of the bacterium Borrelia burgdorferi, the
53
etiological factor of borreliosis (Hutschenreuther, Birkemeyer, Grötzinger, Straubinger, &
54
Rauwald, 2010) and inhibits human immunodeficiency virus (HIV) infections (Rebensburg et
55
al., 2016). Kuchta et al. (2019) observed that use of Cistus incanus herbal tea on a daily basis
56
can reduces cardiovascular risk factors including dyslipidemia and oxidative stress. Lisiecka,
57
Wójtowicz, Dziki, Gawlik-Dziki (2019) enriched wheat pasta with the addition of Cistus
58
incanus leaves and observed an increase in the total phenolic content (TPC) and the
59
antioxidant activity of supplemented pasta. Cacak-Pietrzak et al. (2019) supplemented wheat
60
bread with the addition of 1 up to 5 % of ground Cistus incanus and also observed
61
significantly higher TPC and much higher antioxidant activity of bread with this herb’s
62
leaves. To date, no research has been conducted to determine the effect of cistus extract on
63
bread characteristics. Moreover, other authors used dried Cistus incanus leaves for the
64
production of bread and pasta (Cacak-Pietrzak et al., 2019; Lisiecka et al., 2019), which could
65
be less effective than using it in liquid form.
66
For this reason, appreciating the potential nutritional value of cistus extract in bread
67
production, it was decided to study the possibility of using this extract for enrichment of bread
68
products. The aim of the study was to analyze and compare selected physicochemical,
69
antioxidant, textural and microbiological properties, as well as the technological and
70
organoleptic quality of bread in which water was replaced with 2.5, 5 and 7.5% cistus extract
71
on the baking day and up to 5 days.
72
2. MATERIALS AND METHODS
73
2.1. Materials
74
The study material consisted of wheat bread (WS) obtained from wheat flour type 550 (PZZ
75
Krakow, Poland), the same as used to in previous research (Mikulec et al., 2019), and wheat
76
bread in which water prescribed by the recipe was replaced with cistus extract in the
77
concentrations 2.5, 5 and 7.5 % w/v (Bellis Pharma sp. z o.o., Jaroslaw, Poland). The dough
78
used for baking of bread consisted of 7.5 kg of wheat flour, 4.60 dm³ of cistus extract, 0.23 kg
79
of yeast (Lesaffre, S.A., Wolczyn, Poland) and 0.15 kg of salt (Solino, Inowroclaw, Poland).
80
The baking was performed twice, each time obtaining 30 pieces of bread.
81
2.1.1. Bread preparation
82
The dough was prepared using a single-phase method in a fast rotating spiral mixer (Ibis
83
MS 130 Poland). After initial fermentation (60 min, 21 ºC, 55% relative humidity (RH)), the
84
dough was divided into 405 ± 5 g, pieces, formed and placed in steel molds. After the
85
fermentation (in a fermentation cabinet by MIWE GVA (Germany)) (35 min, 32 ºC, 90%
86
RH), the bread was baked in an MIWE IDEAL (Germany) batch oven at 230 °C for 32
87
minutes. After removal from molds, bread was cooled down for two hours at 19 °C and used
88
for further analyses.
89
2.2. Methods
90
2.2.1. Analysis of basic quality features
91
Bread volume was measured by the AACC-approved method 10-05.01 (AACC,
92
2000), total baking loss was calculated as the difference between the mass of the raw roll and
93
the mass of the roll one hour after baking (Majzoobi, Farahnaky, & Agah, 2011), and crumb
94
moisture was estimated using the gravimetric method (AOAC 925.10).
95
Analysis of crumb color according to the CIELab system was performed by the
96
reflectance method using a Konica Minolta CM-3500d spectrophotometer (Konica Minolta
97
Sensing, Osaka, Japan; illuminant D65, observer 10°). The total color difference was
98
calculated according to Fernandez-Artigas, Guerra-Hernandez, & Garcia-Villanova (1999)
99
using the formula (Eq. 1):
100
∆ = √∆
+∆
+∆
(Eq. 1)
101
where:
102
∆E = the total color difference,
103
∆L = brightness difference,
104
∆a = redness difference,
105
∆b = yellowness difference.
106
Color measurements were performed in twelve replicates per loaf. On the basis of L*a*b*
107
parameters, the browning index (BI) was estimated (Buera, Retriella, & Lozano, 1985) as
108
follows (Eq. 2):
109
=
[
.
]
(Eq. 2)
.
110
where:
111
=
.
∗
.
∗
∗
∗
.
∗
112
L = brightness,
113
a = redness,
114
b = yellowness.
115
2.2.2. Microbiological analysis
116
Microbiological analyses of the bread were performed during the 1st and the 5th day of
117
storage. Samples were obtained from a different part of the loaf (including the crust and
118
crumb). To determine the number of aerobic amylolytic bacteria 1 cm3 of each decimal
119
dilution was transferred to three parallel Petri dishes and poured with Waksman medium
120
(Biocorp). The plates were incubated at 37 °C for 48 h. The reading of the results consisted in
121
filling the plate with Lugol's liquid and counting the colonies around which the yellow, clear
122
zone was formed. To determine the number of yeast and mold, from two successive decimal
123
dilutions, 1 cm3 was transferred into three parallel Petri dishes and poured with Sabouraud
124
agar with chloramphenicol (Biocorp). The plates were incubated at 25 °C for 3 to 5 days. The
125
obtained results were counted as cfu/g bread.
126
The determination of the microbiological stability of the bread consisted of thermostating the
127
product sample at 30 ºC and observing it every 24 hours to determine possible organoleptic
128
changes caused by molds or at 37 ºC to determine changes caused by aerobic amylolytic
129
bacteria. Time of occurrence of the first organoleptic changes (mold, malleability,
130
deformability or crumb viscosity) determined the shelf life of the product. Measurements
131
were done in two replicates.
132
2.2.3. Texture analysis
133
Selected mechanical features of the breadcrumb, such as hardness, springiness, resilience,
134
chewiness and gumminess (Szcześniak, 1963), was performed using a single-arm
135
TA.XT2.Plus texture analyzer (Stable Micro System Ltd, Godalming, Surrey, United
136
Kingdom). Breadcrumb samples were sliced mechanically and the slices for analysis were cut
137
from the middle part of the loaf. The tests were done on cylindrical samples (r=30 mm, h=35
138
mm). A P/36R aluminum probe with a diameter of 36 mm was used, with the compression
139
rate 2 mm·s-1 to reach 50% deformation in two cycles with 5 s waiting time between the
140
cycles (Mikulec et al., 2019). Bread for examination of the crumb texture changes during
141
storage were packed in polyethylene film pouches (HDPE) and stored at ambient temperature
142
(19 ± 1 °C, 60% RH). The analysis was done at 1st and the 5th day of storage. Measurements
143
were done in ten repetitions.
144
To assess the changes in texture caused by storage, the percentage changes of hardness,
145
chewiness, gumminess, springiness and resilience were calculated according to Różyło (2014)
146
with modification (Eq. 3): !"# !%#
147
=
148
Where:
149
x 5d = textural property marked after 5 days of storage,
150
x 1d = textural property marked after 1 day of storage.
!%#
∗ 100
(Eq. 3)
151
2.2.4. Sensory analysis
152
Consumer acceptability tests were performed by a panel consisting of 60 untrained consumers
153
(20–51 years old, 43 females and 17 males) who were recruited from the Institute of
154
Technology (State Higher Vocational School in Nowy Sacz) (staff and students) and who
155
were habitual consumers of bread. Assessors were asked demographic questions about age
156
and gender and a question about bread consumption. Two hours after baking bread samples
157
were sliced mechanically into 20 mm slices and were cooled until bread reached 19 °C. Then
158
slices were placed on disposable paper trays and coded with two digits random numbers.
159
Samples were presented to the assessors simultaneously. The bread was evaluated only on the
160
day of baking using a 9-point hedonic scale (1 = dislike extremely; 2 = dislike very much; 3 =
161
dislike moderately; 4 = dislike slightly; 5 = neither like nor dislike; 6 = like slightly; 7 = like
162
moderately; 8 = like very much; 9 = like extremely), as described by Meilgaard, Civille, &
163
Carr (2007). During the assessment the assessors neutralized the taste using pure water at
164
temperature 19 °C. The results were calculated by Microsoft Excel 10.0 software. The
165
assessors followed the ethical requirements of the sensory laboratory approved by the State
166
Higher Vocational School in Nowy Sacz and an informed consent form was signed by the
167
panelists.
168
2.2.5. Total phenolic content
169
Total phenolic content was estimated using Folin-Ciocalteu reagent (Meda, Lamien, Romito,
170
Millogo, & Nacoulma, 2005). Samples were prepared at 0.05 g/mL concentration using water
171
solution as described by Mikulec et al. (2019). Results were expressed as mg of gallic acid
172
equivalent (GAE) per 100 mg of product. Measurements were done in two replicates.
173
2.2.6. Chromatographic analysis of polyphenol profile
174
Separation and identification of polyphenols were carried out using high-performance liquid
175
chromatography (HPLC) according to the method described by Klimczak, Małecka, Szlachta,
176
& Gliszczyńska-Świgło (2007) with modification as described by Mikulec et al. (2019).
177
Briefly, Dionex Ultimate 3000 HPLC set equipped with DAD detector (Thermo Scientific,
178
Germering, Germany), fitted with Cosmosil 5C18 – MS - II 250 x 4.6 mm ID, 5 µm particle
179
size (Nacalai Tesque, INC. Kyoto, Japan) column was used. Gradient elution was used
180
(mobile phase: A - 2% aqueous solution of acetic acid, and B - 100% methanol); flow rate
181
was 1 mL/min. using the following gradient: Eluent A -0 min 95%, 10 min. 70%; 25 min.
182
50%; 35 min. 30%; 40 min. 95%; to the end of analysis. Measurements were done in two
183
replicates.
184
2.2.7. Statistical analysis
185
Statistical analysis was carried out using Statistica 13.0 (StatSoft, Poland). The significance of
186
differences was verified with the Duncan test at p<0.05. The results were presented as
187
averages ± standard deviation.
188
3. RESULTS AND DISCUSSION
189
3.1. Analysis of basic quality features
190
No influence of cistus extract on baking loss or volume of bread was observed, which ranged
191
from 11.13% and 845 cm3 for bread with 7.5% cistus extract to 11.46% and 865 cm3 for
192
wheat bread respectively. The moisture content of the crumb on the day of baking did not
193
differ significantly among all breads and ranged from 46.34 to 46.96% (Table 1). Similar
194
results was observed by Pasrija, Ezhilarasi, Indrani, & Anandharamakrishnan (2015), who
195
added a green tea extract and encapsulates to bread and did not observe any change in bread
196
volume. Other results were obtained by Cacak-Pietrzak et al. (2019), who observed a
197
significant increase in moisture of bread (which increased with cistus content), a baking loss
198
from 3 to 5% concentrations of cistus leaves in bread and a decrease in bread volume from
199
2% addition of this herb. A similar trend, to that reported by Cacak-Pietrzak et al. (2019) was
200
observed in another study by Ning, Hou, Sun, Wan, & Dubat (2017) where specific volume
201
decreased with higher concentrations of green tea powder.
202
Breads with cistus extract were characterized by significantly lower lightness (brightness) of
203
the crumb, compared to wheat bread (Table 1). Bread with cistus extract differed significantly
204
in terms of the red component (redness) content (a) in the crumb compared to wheat bread.
205
The lowest level was recorded for wheat bread (1.00), and bread with cistus extract ranged
206
from 4.25 to 4.29. In the case of the yellowness component (b), wheat bread showed the
207
lowest value (20.03), and this value significantly increased with the increase of cistus extract
208
content to 27.45 in WCE7.5 (Table 1). The total color difference (△E) increased significantly
209
with the increase of cistus extract (Table 1). According to total color difference values, the
210
color difference between control and cistus extract bread was perceptible with close
211
observation since ∆E values were lower than 3 (Hernández-Carrión et al., 2015). All breads
212
differed significantly in terms of the browning index (BI) value. The lowest value was found
213
for standard bread (30.92), and the highest was found for WCE7.5 (66.47) (Table 1). Cacak-
214
Pietrzak et al. (2019) also observed a significant difference in the color of bread with cistus
215
leaves from the control bread. The differences in color parameters of breads were caused by
216
the pigments present in Cistus incanus.
217
3.2. Microbiological analysis
218
The cistus extract contributed to an improvement of the microbiological quality of the bread.
219
On the baking day, breads with a cistus extract were characterized by a lower content of
220
microorganisms, compared to wheat bread (Table 1). In all breads in which the water was
221
replaced by the cistus extract there was observed a decrease in the total number of bacteria
222
and the total number of spores of amylolytic bacteria and a reduction of total number of
223
aerobic amylolytic bacteria on the fifth day of storage. Inhibition of microbiological changes
224
in bread with 7.5% cistus extract was observed, compared to wheat bread (Table 1).
225
The time of occurrence of the first organoleptic changes and the positive thermostat test result
226
determined the shelf life of the product. When analyzing thermostatic stability, there were
227
observed in wheat bread crumb, from the 3rd day of storage, both visible mold (Aspergillus
228
niger) and changes in the crumb indicating the development of Bacillus bacteria (soft, sticky
229
and viscous breadcrumb). In the WCE2.5 and WCE5 bread, no mold-like changes were
230
observed until the 5th day, but from the 3rd day changes in the crumb indicating the
231
development of Bacillus bacteria were visible. In WCE7.5 bread, no mold-like changes were
232
observed and the bacterial-like changes were characterized by slight variation. Based on these
233
results it can be concluded that the addition of cistus extract contributed to improvement of
234
the microbiological quality of the tested bread.
235
The obtained effect may result from the antibacterial properties of the cistus extract observed
236
by Barrajón-Catalán et al. (2010) in their studies. They observed antibacterial properties of
237
extracts from Cistus ladanifer, which inhibits growth of the Gram-positive bacterium
238
Staphylococcus aureus, and extract from Cistus populifolius reveals high growth inhibitory
239
activity against the Gram-negative bacterium Escherichia coli. Tomás-Menor et al. (2013)
240
posited that the cause of the inhibitory activity against bacteria may be related to the presence
241
of galloylated flavanols and specific flavonols and polar compounds and to other flavonols.
242
3.3. Texture analysis
243
The stalling process leads to changes in the texture and reduction of fresh bread
244
characteristics, which results in a short shelf life and best before date (Gray & Bemiller, 2003;
245
Ribotta & Le Bail, 2007). During storage (up to 5 days) an increase in breadcrumb hardness
246
was observed for all breads. The largest increase was characterized by standard wheat bread,
247
as much as 72.89%, compared to the first day of storage, and the smallest for WCE5 bread
248
(29.03%). There was a reduction in the springiness and resilience of all loaves at a similar
249
level. The largest increase in gumminess and chewiness was observed in wheat bread,
250
whereas in bread with cistus extract a reduction of chewiness was observed. The greatest
251
variability of texture parameters was identified for breadcrumb of standard wheat bread
252
(Table 1) and the smallest for bread with cistus extract, which can be considered as an
253
inhibitor of staling of the breadcrumb. Others authors have observed different dynamics of
254
changes in the texture features of bread, both increases and decreases of such parameters as
255
hardness, springiness, cohesiveness, and chewiness depending on the form of the additive.
256
The changes of bread crumb texture are probably a result of change in the nature of starch,
257
dilution of gluten and enrichment of fibre. It can be seen that the addition of herbal extracts
258
had a rather positive effect on the textural properties of the bread (Cacak-Pietrzak et al., 2019;
259
Ning et al., 2017; Pasrija et al., 2015).
260
3.4. Sensory analysis
261
In the organoleptic evaluation, wheat bread and bread with a 2.5% cistus extract did not differ
262
statistically significantly and obtained the highest average marks for specific characteristics,
263
which ranged from 8.8 to 8.9 and 8.7 to 8.8 respectively (Figure 1). Bread with 5% and 7.5%
264
of cistus extract differed significantly in the results of the sensory evaluation compared to WS
265
and WCE2.5. Breads with 5 and 7.5% cistus extract did not differ significantly for taste and
266
smell or crumb porosity. The scores for other features were statistically different. WCE5 and
267
WCE7.5 received the lowest marks for taste and smell (7.5 and 7.3 respectively) (Figure 1).
268
Bread with a 5% cistus extract received the highest marks for appearance and crumb porosity
269
(8.6 and 8.5 respectively) and bread with a 7.5% cistus extract for crumb porosity (8.3) and
270
appearance and crust thickness (8.1.) (Figure 1). Cacak-Pietrzyk et al. (2019) observed
271
deterioration in the organoleptic characteristics of taste, aroma and overall acceptability with
272
a 4% addition of the cistus leaves. Deterioration of the organoleptic properties of bread which
273
were observed in our research can also be seen in bread enriched with herbs such as basil,
274
oregano, and thyme, which can be explained by the content of essential oils that cause its
275
specific taste and smell (Adams, Kruma, Verhé, De Kimpe, & Kreicbergs, 2011).
276
3.5. Total polyphenol content and chromatographic analysis of polyphenol profile
277
The addition of the cistus extract clearly contributed to increasing the antioxidative potential
278
of the obtained bread products (Table 2). Breads with the addition of cistus extract were
279
characterized by a higher total polyphenol content, from 62.81 to 105.81 mg GAE per 100 g
280
of product, compared to the standard (41.23 mg GAE per 100 g). Along with the addition of a
281
more concentrated cistus extract, an increase in the sum of polyphenols determined
282
chromatographically in breads was observed from 34.68 (WCE2.5) to 78.71 mg/100 g
283
(WCE7.5). Cacak-Pietrzak et al. (2019) and Lisiecka et al. (2019), who enriched pasta and
284
bread with dried cistus leaves, observed significantly higher TPC and much higher
285
antioxidant activity of breads and pasta compared to the control sample. The bread enriched
286
with Cistus incanus was characterized by significantly higher TPC already at 1% of the
287
amount of this additive (Caca-Pietrzak et al., 2019). Breads enriched with herbs are
288
characterized by beneficial features in many aspects such as high antioxidant content (Ibrahim
289
et al., 2015). Cistus herb exhibited the highest content of polyphenols (2635.16 mg/100 g),
290
and the dominant polyphenols were (+) catechin (726.65) and ellagic acid (637.18 mg/100 g)
291
(Table 2). Wheat flour and wheat bread were characterized by the lowest polyphenol content
292
(8.50 and 8.88 mg/100 g respectively). No rutin, ellagic acid, 3-hydroxybenzoic acid, (+)
293
catechin, protic acid or t-cinnamic acid was found in the wheat flour or wheat bread in the
294
polyphenol profile. Chlorogenic and t-cinnamic acid were not found in bread with cistus
295
extract despite being present in the cistus herb (93.10 and 0.12 mg/100 g respectively). The
296
content of polyphenols in bread obtained with the addition of cistus extract increased as a
297
result of the substitution of water with this extract. Nevertheless, the polyphenol profile
298
showed quantitative, as well as qualitative, changes. Gallic acid appeared in wheat bread
299
(2.14), but was not present in wheat flour, and there was no phloridzin in wheat bread,
300
although it was detected in wheat flour (0.74 mg/100 g). Some fluctuations were also
301
observed in the content of the other polyphenols. In contrast to the other polyphenols
302
measured by chromatography, more ferulic acid and rutin were determined in bread with the
303
cistus extract in comparison to the amount resulting from the calculation (Table 2). These
304
changes are the result of heat treatment (baking process). Riehle, Vollmer, and Rohn (2013)
305
reported that when preparing C. incanus beverages, decreased amounts of phenolic substances
306
and reduced antioxidant activity are observed if an incorrect selection of brewing process
307
parameters (brewing water, temperature, and duration) is made. Spontaneous changes in
308
polyphenols can occur at elevated temperatures. Such relationships were also observed in the
309
case of grape pomace heating (Chamorro, Goni, & Viveros, 2012). A significant increase in
310
the total polyphenol content was also observed, along with an increase in the cistus extract
311
content in bread (Table 2).
312
4. CONCLUSION
313
Cistus extract has a high nutritional and pro-health value, so it could be used as a valuable
314
component of bread. To date there are no studies concerning utilization of cistus extract in
315
bread formulations, so the research was
316
microbiological, and organoleptic characteristics, polyphenol profile and the total polyphenol
317
content as nutrients in bread. Cistus extract influenced the color of the crumb by increasing its
318
browning index in comparison to wheat bread. The total color difference (△E) increased
319
significantly with the increase of cistus extract and the total color difference between control
320
and cistus extract breads was perceptible with close observation. The cistus extract positively
321
influenced microbiological properties of bread enhancing its shelf life. Both on the day of
322
baking and after 5 days of storage bread with a cistus extract was characterized by a lower
323
content of microorganisms, compared to wheat bread. During storage (up to 5 days) the
focused on selected physicochemical,
324
greatest variability of texture parameters was identified for wheat breadcrumb and the
325
smallest for bread with cistus extract, which can be considered as an inhibitor of staling of the
326
breadcrumb. The observed effect is advantageous, because crumb hardening is one of the
327
main symptoms of bread staling. The addition of the cistus extract positively influenced
328
antioxidant properties of bread by increasing the total polyphenol content. The results show
329
that preparations, based on cistus extract, may be valuable components of bread significantly
330
improving its nutritional and pro-health value, with almost unchanged sensory attributes and
331
limiting the aging of the bread.
332 333
ACKNOWLEDGEMENTS
334
This Research was finance by the Ministry of Science and Higher Education of the Republic
335
of Poland.
336
Statistical analysis was performed using the skills acquired during the training entitled
337
"Application of statistics and Statistica in planning empirical research and developing its
338
results in natural sciences - basic methods" implemented as part of the project: "Integrated
339
Program of the Agricultural University H. Kołłątaja w Krakowie ”, co-financed from
340
European Union funds.
341
Declarations of interest: none.
342 343
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344 345
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Figure 1. Results of organoleptic analysis of tested breads
456
Explanatory notes:
457
…….
458
with 5 % cistus extract, ⸺ WCE7.5 – bread with 7.5 % cistus extract
WS – wheat bread, - - - WCE2.5 – bread with 2.5 % cistus extract, ⸺⸺ WCE5 – bread
Table 1. Evaluation of selected quality parameters of studied breads Bread type
WS*
WCE2.5
WCE5
WCE7.5
Physico-chemical parameters of bread Total baking loss [%]
11.46**a ± 0.42
11.16a ± 0.26
11.33a ± 0.28
11.13a ± 0.34
Volume [cm³]
865.00a ± 17.84
859.67a ± 15.17
856.67a ± 12.89
845.0a ± 16.46
Moisture of bread crumb [%]
46.81a ± 0.08
46.96a ± 0.19
46.34a ± 0.13
46.48a ± 0.96
Color parameters of bread crumb c
L*** [-]
75.17** ± 0.87
60.86b ± 0.77
58.94a ± 0.78
58.18a ±0.78
a [-]
1.00a ± 0.08
4.29b ± 0.16
4.25b ± 0.22
4.26b ± 0.22
b [-]
20.03a ± 0.74
22.53b ± 0.46
25.41c ± 0.54
27.45d ± 0.40
a
b
∆E [-]
-
14.90 ± 0.73
17.41 ± 0.48
18.83c ± 0.75
BI [-]
30.92a ± 1.68
49.85b ± 1.28
59.53c ± 1.91
66.47d ± 1.94
Microbiological characteristic of bread (cfu / g) I day TNB****
1.9 x 103
3.3 x 101
4.7 x 101
3.0 x 101
NAAB
1.1 x 103
3.0 x 101
2.7 x 101
2.3 x 101
NSAB
6.7 x 102
2.0 x 101
6.8 x 101
0.5 x 100
Microbiological characteristic of bread (cfu / g) V day TNB
8.3 x 105
4.2 x 105
2.2 x 105
1.6 x 104
NAAB
8.1 x 105
0.0
6.0 x 104
0.0
NSAB
4.2 x 105
1.8 x 104
1.5 x 101
4.0 x 103
Changes in texture after five day storage [%] Hardness [N]
72.89c ± 2.31
36.73b ± 0.51
29.03a ± 0.47
29.27a ± 0.33
Springiness [-]
-7.61b ± 0.57
-8.78ab ± 0.66
-8.49ab ± 0.33
-9.09a ± 0.59
Gumminess [N]
19.59d ± 1.01
2.25a ± 0.11
3.04b ± 0.09
5.14c ± 0.25
Chewiness [N x mm]
8.44d ± 0.44
-2.62c ± 0.09
-9.77a ± 0.37
-6.62b ± 0.12
Resilience [-]
-33.33a ± 1.01
-40.00b ± 0.91
-40.00b ±1.11
-40.63b ± 1.29
Explanatory notes: * WS – wheat bread. WCE2.5 – bread with 2.5 % cistus tea. WCE5 - bread with 2.5 % cistus tea. WCE7.5 - bread with 7.5 % cistus tea **values in the same row marked with different letters are statistically significantly different at p≤ 0.05 ± SEM *** L – lightness. a – redness. b – yellowness. △E- total color difference. BI - browning index ****TNB - total number of bacteria. NAAB - total number of aerobic amylolytic bacteria. NSAB- total number of spores amylolytic bacteria
Table 2. Polyphenol profile and the total polyphenol content of bread Sample
chlorogenic acid
p-coumaric acid
ferulic acid
rutin
ellagic acid
phloridzin
3hydroxybenzo ic acid
(+) catechin
protic acid
gallic acid
t-cinnamic acid
total
TPC***
mg GAE* 100g-1
[mg*100g-1] Wheat flour WF550*
0.00**±0.00
0.30±0.02
7.46±0.20
0.00±0.00
0.00±0.00
0.74±0.08
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
8.50±0.23
0.00±0.00
8.88c±0.15
48.12±0.77
Bread d
c
WS
0.00±0.00
0.30 ±0.01
6.43 ±0.13
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
0.00±0.00
2.14c±0.06
WCE2.5
0.00±0.00
1.07c±0.01
7.34a±0.25
2.85b±0.12
9.10c±0.41
1.17c±0.08
1.78b±0.41
6.94c±0.91
2.50c±0.49
1.93c±0.06
0.00±0.00
34.68b±1.58
WCE5
0.00±0.00
1.81a±0.02
7.01b±0.16
7.36a±0.49
23.40b±2.39
1.56b±0.08
4.11a±0.66
16.40b±1.44
5.93b±0.47
4.80b±0.36
0.00±0.00
72.38a±5.71
WCE7.5
0.00±0.00
2.26a±0.03
6.51c±0.18
7.72a±0.14
26.57a±0.34
1.83a±0.03
3.87a±0.55
18.20a±0.72
6.56a±0.19
5.19a±0.10
0.00±0.00
78.71a±0.89
cistus 93.10±1.77 73.95±10.27 35.48±4.17 231.55±8.44 637.18±53.00 37.59±0.51 192.90±4.74 726.65±118.9 313.69±68.75 herb 4 cistus extract 2.33 1.85 0.89 5.79 15.93 0.94 4.82 18.17 7.84 ****2.5% cistus extract 4.66 3.69 1.77 11.58 31.86 1.88 9.65 36.33 15.68 ****5.0% cistus extract 6.98 5.55 2.66 17.37 47.79 2.82 14.47 54.49 23.53 ****7.5% Explanatory notes: * WF- wheat flour, WS – wheat bread, WCE2.5 – bread with 2.5 % cistus tea, WCE5 - bread with 2.5 % cistus tea, WCE7.5 - bread with 7.5 % cistus tea **values in the same column marked with different letters are statistically significantly different at p≤ 0.05 ± SEM *** TPC –the total phenolic content **** values calculated on the basis of individual compound concentration in cistus herb
292.94±11.07
0.12±0.02
2635.16±243.37
7.32
0.00
65.88
14.65
0.01
131.76
21.97
0.01
197.64
41.23d±0.7 3 62.81c±0.70 82.73b±0.5 1 105.81a±2.1 5
Cistus 930.76 ±9.72 23.27 36.55 69.81
1. Cistus extract enhanced antioxidant properties of bread 2. Cistus extract had a positive effect on textural features of the bread 3. Cistus extract inhibited the growth of microflora improving shelf life of bread 4. Cistus extract influenced the color of the crumb by increasing its browning index
Declaration of interests ☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. ☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: