Accepted Manuscript

Enhancement of antioxidant activity and physicochemical

properties of yogurt enriched with concentrated strawberry pulp
obtained by block freeze concentration

Henrique Jaster, Giordana Demaman Arend, Katia Rezzadori,

Vitor Clasen Chaves, Flávio Henrique Reginatto, José Carlos
Cunha Petrus

PII: S0963-9969(17)30691-9
DOI: doi:10.1016/j.foodres.2017.10.006
Reference: FRIN 7054
To appear in: Food Research International
Received date: 17 July 2017
Revised date: 3 October 2017
Accepted date: 6 October 2017

Please cite this article as: Henrique Jaster, Giordana Demaman Arend, Katia Rezzadori,
Vitor Clasen Chaves, Flávio Henrique Reginatto, José Carlos Cunha Petrus , Enhancement
of antioxidant activity and physicochemical properties of yogurt enriched with
concentrated strawberry pulp obtained by block freeze concentration. The address for
the corresponding author was captured as affiliation for all authors. Please check if
appropriate. Frin(2017), doi:10.1016/j.foodres.2017.10.006

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 ACCEPTED MANUSCRIPT
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Enhancement of antioxidant activity and physicochemical properties of yogurt

enriched with concentrated strawberry pulp obtained by block freeze
concentration

Henrique Jaster1, Giordana Demaman Arend1, Katia Rezzadori1*, Vitor Clasen Chaves2,
Flávio Henrique Reginatto2, José Carlos Cunha Petrus1

1
Department of Chemical and Food Engineering, Federal University of Santa Catarina,

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Florianopolis, SC, Brazil
2
Postgraduate Program in Biotechnology and Biosciences, Federal University of Santa
Catarina, Florianopolis, SC, Brazil

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* Corresponding: Tel: +55 48 3721 6443
E-mail address: katia.rezzadori@gmail.com
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Universidade Federal de Santa Catarina - Centro Tecnológico/CTC
Campus Reitor João David Ferreira Lima
CEP: 88040-970 Caixa Postal 476
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Florianópolis – SC – Brasil

Abbreviated Running title: YOGURT PRODUCTION WITH CRYOCONCENTRATE

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STRAWBERRY PULP.
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Abstract

Strawberry juice was concentrated using block freeze concentration process. The
concentrate was used to produce two yogurts with different concentrations of
cryoconcentrated strawberry pulp (15 % and 30 %). Total lactic acid bacteria count,
physicochemical and rheological properties was evaluated during storage (7 days) for
all yogurts. Also, the beverages produced were compared with two commercial
trademarks. It was observed that the total lactic acid bacteria count remained higher than

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108 CFU·mL-1 during the storage time for all beverages studied. The viscosity of the
yogurts decreased when the ratio of strawberry cryoconcentrate was increased. The

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Power Law model was successfully applied to describe the flow of the yogurts, which

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had a thixotropic behaviour. The incorporation of the cryoconcentrated strawberry pulp
in the yogurt resulted in a product with 3-fold more anthocyanins content and
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antioxidant activity. The enrichment of natural yogurt with strawberry cryoconcentrated
pulp proved to be effective in the production of a beverage with higher nutritional
characteristics.
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Keywords: anthocyanins; pelargonidin-3-O-glycoside; phenolic compounds; functional

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yogurt; cryoconcentration; bioactive compounds.

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1 Introduction

Yogurt is a dairy product appreciated worldwide once it presents important

sensorial and nutritional characteristics. The frequent consumption of yogurt with lactic
prebiotic cultures is well known for the reduction of intestinal inflammations and
infections, diarrhoea, irritable bowel and cholesterol levels (de Vrese & Schrezenmeir,
2008). Aiming to improve the nutritional characteristics of yogurt, several authors have
studied the incorporation of natural products to the yogurt, as hazelnuts shells (Bertolino

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et al., 2015), algae extract (O’Sullivan et al., 2015) and black, white and green tea
(Muniandy, Shori, & Baba, 2016).

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The yogurts with natural fruit pulps addition have been increasing their worldwide

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market due to the large demand of products for a healthier diet, rich in compounds with
biological activity. Strawberry yogurt is one of the most consumed because of the
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sensorial properties, like flavour and colour, as well as the nutritional properties such as
the presence of bioactive compounds, e.g. anthocyanins. These natural compounds can
provide better sensorial, nutritional and antioxidant quality when compared to the
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synthetic additives in yogurt (Caleja et al., 2016).

According to Aaby et al. (2012), the anthocyanins are the main phenolic compounds
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in strawberry. These compounds show antioxidant and anti-inflammatory properties, as

well as a large variety of chemotherapeutic effects, avoiding the oxidative stress and
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improving the antioxidant defences (Forbes-Hernández et al., 2014; Gasparrini et al.,

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2017). However, these compounds are often highly thermosensitive, which is their main
disadvantage. In this sense, the block freeze concentration is a low cost and low
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complexity alternative technology that has been successfully employed to concentrate

phenolic compounds extracted from natural products (Adorno et al., 2017), avoiding the
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degradation of these compounds (Aider, De Halleux, & Melnikova, 2009). The block
freeze concentration is based in the application of temperatures below the water freezing
point to form pure water crystals, thus concentrating the solutes. Therewith, the solutes
(anthocyanins) are partially separated from the solvent (water). After 24 hours, the
product is partially thawed and the concentrated solute is first obtained while the water
crystals remains frozen (Aider & Halleux, 2009).
For the best of our knowledge, there are few researches about the application of
cryoconcentrated products in yogurt. Furthermore, the use of dairy products as a natural
source of antioxidant compounds seems to be a suitable form to satisfy the consumers
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interest in health benefits. Therefore, the aim of this study was to evaluate the use of
block freeze concentrated strawberry pulp to enhance the phenolic compounds in
yogurts and to investigate its microbiological, physical, chemical and rheological
properties during storage time.

2 Material and Methods

2.1 Strawberry block freeze concentration

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The block freeze concentration was performed according to Aider et al. (2007)

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and adapted by Adorno et al. (2017) (Figure 1 (a)). Firstly, the in natura juice was

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fractionated in plastic containers of 100 mL and then frozen at -16 ± 2 °C during 24
hours. When the solution was completely frozen, 50% of the initial volume was
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defrosted at controlled atmosphere (17 ± 2 °C), using the scheme showed in Figure 1
(b), and the thawed liquid (cryoconcentrate) was used as feed solution for the next step.
This process was repeated three times for obtaining the final concentrate.
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2.2 Yogurt preparation

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The yogurt was prepared based on the methodology proposed by Bertolino et al.
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(2015). Pasteurized milk, standardized (3% of fat) and homogenized was purchased at
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local market (Tirol®, Brazil). The milk was warmed until 42 °C, inoculated with dairy
culture of Streptococcus thermophiles and Lactobacillus delbruckii subspecies
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Bulgaricus (Chr. Hansen, Danmark) and the fermentation was performed until the pH of
4.8 was reached. After that, the fermentation was interrupted immediately by
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refrigeration at thermostatic bath at 20 °C. The product obtained was used as control
sample (0% of cryoconcentrate).
Concentrated strawberry juice (C3) was added to the yogurt in the proportions of
15% (yogurt 1) and 30% (yogurt 2). The different samples were maintained at 5 °C and
analysed after 1 and 7 days of storage. Yogurt manufacturing was carried out in
duplicate for each group.
Simultaneously, commercial yogurt indicating the presence of strawberry pulp in
their labels, marketed in Brazil, belonging to trademarks from known companies (C1
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and C2) was obtained from local supermarkets with approximately 7 days old when
evaluated and kept under refrigerated storage at 5 °C in order to perform the analysis.

2.3 Physicochemical analysis

The pH determination was measured on pH meter (QUIMISQ 400A). Colour

parameter of luminosity (L*) and red colour intensity (a*) was determined on
colorimeter (Minolta CR400). Total solids were analysed by drying the samples at 105

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°C during 24 h and the solids content obtained by weight difference (AOAC, 1990). The
acidity was determined according to the methodology suggested by Adolfo Lutz

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Institute (IAL, 2008) and the results expressed as grams per 100 millilitres of acid lactic

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(g·100mL-1). All the analyses were performed in triplicate.
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2.4 Microbiological analysis

The counts of acid lactic bacteria (LAB) were taken in triplicate according to the
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methodology described in APHA (2001). The results were expressed as colony-forming

units per millilitre of yogurt (CFU·mL-1).
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2.5 Antioxidant activity

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For the ABTS assay, the method proposed by Rufino et al. (2007a) was
followed. The samples and ABTS solution reacted during 6 min and absorbance was
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measured at 734 nm in a, ethanol was used as the blank. Using the same procedure, the
Gallic acid standard curve was performed and the results were expressed in milligrams
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of Gallic acid equivalent (GAE) per millilitre of yogurt (mg GAE·mL-1). The analyses
were performed in triplicate.
The DPPH assay was performed according to method suggested by Rufino et al.
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(2007 ). Samples and the DPPH solution reacted during 30 min and the absorbance was
measured at 515 nm in a spectrophotometer (UV-Vis mini-1241) using methanol as the
blank. By means of the same procedure, the Gallic acid standard curve was performed
and the results were expressed in milligrams of Gallic acid equivalent (GAE) per
millilitre of yogurt (mg GAE·mL-1). The analyses were performed in triplicate.
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2.6 Anthocyanin content (AC)

The analyses of total anthocyanin content were performed through Liquid

chromatography (LC) (PerkinElmer® Series 200), equipped with diode array detector
(DAD), quaternary pump, degasser and autosampler. For the separation of the different
anthocyanins a C18 column (150 x 4.6 mm id, 5 mm, PerkinElmer®) was used at
constant flow of 1 mL·min-1, volume injection of 20 µL and temperature of 21 ± 2 °C.
As mobile phase, a linear gradient consisting of methanol (A) and an aqueous solution

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of 3 % formic acid (B) was used. An isocratic elution of 10 % A (0-2 min) was followed
by a linear gradient to 20 % A (2-3 min), 20-41 % A (3-19 min), then increasing to 70

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% A (19-30 min). The chromatograms were monitored at 520 nm and the spectra were

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acquired in the 190-700 nm range. The characterisation of the major peaks was
performed by the comparison of retention times with the reference substances
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(pelargonidin-3-O-glucoside and cyanidin-3-O-glucoside) and by co-injection with
reference standards. The results were expressed as pelargonidin-3-O-glycoside·100 mL-
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(mg·100 mL-1).
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2.7 Rheological measurements

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The measurements were carried out using a rotational viscometer (Thermo

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Haake DC 10, model VT 550, Karlsruhe, Germany), with concentric cylinders (NV ST
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807-0713 CE and NV 807-0702) and the data collected by the software program Pro
Rheowin© (version 2.93, Haake). These analyses were performed in triplicate. The
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temperature used was 25 °C and the shear rate varying from 0 to 2000 s-1 (upward
curve) and from 2000 to 0 s-1 (downward curve); each curve was obtained during 3
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minutes. The flow behaviour was described by the Power Law model and the
thixotropic behaviour of the yogurt was evaluated by calculating the hysteresis loop area
between the upward and downward flows curves.

2.8 Statistical analyses

The influence of the addition of cryoconcentrated pulp, as well as the storage

time, and their effect on the yogurt characteristics were subjected to analysis of variance
(ANOVA), and significant differences among mean values from replicates analysis (p <
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0.05) were determined by Tukey’s test with the statistical software Statistica 7.0
(StatSotft Inc., EUA).

3 Results and discussion

3.1 Physicochemical analysis

Table 1 shows the results for pH, titratable acidity, colour parameters (L*, a*)

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and total solids for the yogurts prepared with cryoconcentrated pulp and the commercial
yogurts studied. It can be observed that the yogurt has undergone an acidification when

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the cryoconcentrate (pH around 3.3, data not shown) was added to the control yogurt

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with consequent reduction on the pH values and increase on the titratable acidity values.
Also, after 7 days of storage it can be observed that the acidity of yogurts 1 and 2
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decreased (p < 0.05). The LAB produces β-galactosidase that cleaves the lactose present
in the yogurt and remained active including under refrigeration (0-5°C), resulting in the
accumulation of the acids produced, as metabolic by-products, by the starter
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microorganisms.
The commercial trademarks present lower acidity than the yogurts produced on
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this paper. However, the values obtained for yogurt 1 and 2 are in accordance with
Costa et al. (2013). The same authors affirm that the acidity of milk products must be
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between 0.6 and 1.5 %. Benedetti et al. (2016) reported that consumers prefer fermented
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products with pH on the range of 4.2 and 4.4. Yet, for yogurts 1 and 2, and also the
commercial yogurts, lower values for this parameter were observed. The commercial
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trademarks use acidulants (lactic and citric acid) in their composition, which can be
related with the lower values found.
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Total solids ranged from 11 to 13 % (Table 1) for all the yogurts prepared
(control, yogurt 1 and yogurt 2). The highest value was found to the yogurt 2, which
was expected since they were prepared with higher content of cryoconcentrated
strawberry pulp. Also, Varnam and Sutherland (1994) related that it is common to
increase the solids of commercial yogurts to approximately 15 % to improve the texture
of the final yogurt. Besides that, higher values for total solids on the commercial
trademarks were observed and associated with the different components present in the
commercial yogurts, such as, carboxymethylcellulose, modified starch, whole milk
powder and whey.
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The colour is the first characteristic that determines the quality and the flavour
on strawberry products and it is important in the marketability of these foods and
beverages (Alves, Corrêa, Pinheiro, & Oliveira, 2013; de Moura et al., 2012).
According to the results obtained for the colour parameters (Table 1) significant (p <
0.05) changes were observed with the addition of different proportions of
cryoconcentrate. A decrease in the luminosity (L*, scale ranges from 0 to 100, from
black to white) was observed with the increase of the strawberry pulp proportion. This
behaviour is related with an increase of the red colour of the yogurt, reducing the

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tendency to white visualized on the control yogurt. However, during the storage time,
no significant (p < 0.05) difference was observed for this parameter.

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The red colour parameter (a*) increased with the increase on the proportion of

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cryoconcentrate, which was expected, presenting values of 10 and 18, for yogurts 1 and
2, respectively. The control yogurt presented negative a* values, since it has no addition
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of cryoconcentrate, corroborating with the L* tendency presented above. For a*
parameter, a slightly reduction around 6 and 15 % was seen after 7 days of storage, for
yogurt 1 and 2, respectively. According to Karaaslan et al. (2011) the anthocyanins can
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be easily degradable and affected by the pH, storage temperature, enzymes and
microbial activity and can exhibit colourless or brown-coloured compounds.
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Nevertheless, even with this reduction, the yogurt 2 remained with similar values to
commercial yogurts.
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This is one important feature of this study since, according to the ingredients
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table of commercial trademarks, both of them present artificial dyes as red ponceau,
bright blue and azorubine in their formulation. The red colour observed in the produced
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yogurt with cryoconcentrate is obtained by the addition of natural pulp and, as discussed
in the Item 3.3, attributed to the anthocyanins present in the strawberry, which also
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provides nutritional characteristics to the product.

3.2 Microbiological analysis

Table 2 presents the LAB count obtained for all the samples during the storage
time. In this study, even with the addition of strawberry cryoconcentrate, LAB count
was maintained over 108 CFU·mL-1. This results are in agreement with the Codex
Alimentarius Commission (2011), which stablishes that the counting of lactic acid
bacteria must be over 107 CFU·mL-1.
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No significant (p < 0.05) differences were observed with the increase on the
cryoconcentrate proportion, suggesting that the addition of concentrated pulp did not
influence the development of the starter microorganisms. Sun-Waterhouse et al. (2013)
studied the development of the starter microorganisms with the addition of berry
polyphenols before and after the fermentation and reported the same behaviour.
During the storage time, a significant reduction (p < 0.05) in LAB count was
observed. This reduction can occur due to the decrease of pH and, consequently, the
increase in the acidity. Shah (2006) verified that in pH below 4.2, some starter

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microorganisms can disappear, mainly the Streptococcus, contributing for the decrease
in LAB count. Also, the maintenance of the LAB can be affected by the chemical

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composition of the yogurt that can disturb the growth of the microorganisms (Almeida,

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Tamime, & Oliveira, 2009). Nevertheless, even with some slight variations in the LAB
count in each storage period evaluated (less than one cycle log), the recommended
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counting of lactic acid bacteria was maintained.

3.3 Anthocyanin content and antioxidant activity of the yogurts

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The enrichment of the yogurt with natural fruit pulps rich in bioactive
compounds can improve the health benefits of this product. The major compound with
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biological activity present in strawberries are the anthocyanins, which are characterized
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as unstable and easily degradable compounds (Aaby et al., 2012).

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As expected, the anthocyanins content increased (p < 0.05) for the higher
proportion of cryoconcentrate used (Table 3), evidencing a direct influence in the total
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anthocyanins content. Control and commercial yogurts studied presented no content of

anthocyanins and consequently none or almost no antioxidant activity.
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Regarding the storage time, it was observed a significant reduction (p < 0.05)
of the anthocyanins content after 7 days of storage. However, the values remained very
close, which is an evidence of the maintenance of anthocyanins compounds after the
storage time studied. This behaviour is supported by the colour analyses (data shown on
Item 3.1) where only a small reduction in the parameter of red colour (a*) was
observed. This reduction can be explained by the ease of degradation of the
anthocyanins. On the other hand, the yogurt polymer network and gel matrix can protect
the anthocyanins, reducing the degradation rates (Sun-Waterhouse et al., 2013).
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Moreover, the presence of anthocyanins in strawberries is connected to the

higher antioxidant activity in this fruit. The analysis of the correlation of the antioxidant
activity with the anthocyanins on strawberries is well known. Correlation coefficient
between the antioxidant activity (ABTS) and the anthocyanins content for control
yogurt, yogurt 1 and 2 was R2=0.99, for both, the first and the 7th days of storage. In the
same way, for DPPH method, the correlation between the antioxidant activity and the
anthocyanins content for control yogurt, yogurt 1 and yogurt 2, the values obtained were
R²=0,94 and R²=0,96, for 1 and 7 days, respectively.

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For both methods used - ABTS and DPPH - the antioxidant activity increased
when higher quantities of cryoconcentrate were added in the yogurt. Also, for both

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methods, no significant difference (p>0.05) was observed during the storage time. The

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maintenance of the anthocyanins and antioxidant activity during the storage time must
be highlighted as one of the most important features of this study, since it shows the
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possibility to provide a product with higher nutritional value to the consumer.

3.4 Rheological measurements

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The yogurts rheological behaviour is shown in Figure 2. A reduction in the

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apparent viscosity with the increase of the shear rate can be observed. This is
characteristic of a non-Newtonian behaviour and indicates that these beverages have
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shear thinning characteristics. According to Lucey (2002), the shear forces destroy the
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weak bonds and reduce the electrostatic repulsion and hydrophobic interaction between
the molecules. This disruption is greater in the first minutes of the process due to the
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intense hydrodynamics forces. After that, an alignment of the particles with the flow
occurs, reducing the viscosity (Castro, 2003; Karazhiyan et al., 2009).
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Still regarding Figure 2, it can be observed that the apparent viscosity decreases
with the increase of the cryoconcentrated pulp concentration, i.e. the beverage with 30
% of strawberry pulp presented the smallest viscosity compared to the control yogurt
(without strawberry pulp). According to Sun-Waterhouse et al. (2013) the viscosity can
be affected by the formulation composition, type of starter cultures, heat treatment and
processing methods. This same viscosity reduction behaviour was reported by Benedetti
et al. (2016) and İçier et al. (2015) in their studies on the addition of concentrated tofu
whey and apple juice, respectively, on different beverages. The same authors justified
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this behaviour due to the addition of an aqueous extract, which decreases the viscosity
of this products with consequent reduction in the capacity of the proteins to water-bind.
After 7 days of storage, for the control yogurt, the structure was completely
regained but the regeneration of the three-dimensional structure was inversely
proportional to the strawberry cryoconcentrate proportion. In other words, after 7 days,
the beverages with more cryoconcentrated, showed a decrease in the apparent viscosity.
This behaviour can be related with the reduction of the capacity of water-binding of the
proteins caused by the addition of the strawberry cryoconcentrate, consequently

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reducing the regeneration of the three-dimensional structure (Oliveira, Sivieri, Alegro,
& Saad, 2002; Ramaswamy & Basak, 1992).

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The commercial trademarks presented similar trend, but with higher viscosity

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values. This behaviour can be related to the ingredients table of these products. It is
possible to point out several thickeners as starch and carboxymethylcellulose. These
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components can increase the water retention capacity, aiding on the formation of
viscous gels and consequently promoting higher viscosities.
Rheological constants were predicted to describe the yogurts behaviour by
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means of fitting the rheological model of Power law (Table 4). The fitted parameters
presented correlation coefficient (R²) higher than 96 % and can be considered suitable to
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describe the rheological behaviour of the yogurts for the different storage times.
Power Law model was effective in describing the rheological behaviour of all
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the samples studied. During the storage time, for all the samples, the flow behaviour
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index (n) showed values lower than 1, confirming the non-Newtonian behaviour.
Furthermore, an increase in the n values was verified with the increase in the proportion
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of cryoconcentrate. Sun-Waterhouse et al. (2013) also reported that the addition of

phenolic compounds post fermentation resulted on the interaction of these with the milk
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proteins, reducing aggregation of the casein network and, consequently, reducing the
resistance of yogurt to flow.
The rheograms in Figure 3 show the hysteresis curves present in all yogurts
studied, including the commercial ones. This behaviour is related to products with shear
thinning characteristics, which exhibits thixotropic behaviour. Hernández (1996)
reported that high viscosity is related with greater hysteresis areas, demonstrating a
greater correlation with the fluid thixotropy. In the present study, hysteresis areas
decreased with the increase of strawberry pulp proportion (Figure 4), corroborating with
the viscosity results (Table 4). The largest hysteresis areas were found for the control,
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yogurt 1 and yogurt 2, respectively, and can be associated to the changes in the
interactions between the milk proteins promoted by the addition of strawberry
cryoconcentrate that alter the three-dimensional structures. Assuming that the area of
hysteresis is an index of the energy required to destroy the structure responsible for
time-dependence, during the 7 days of storage, the addition of strawberry concentrate
presented a positive effect.

4. Conclusions

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The use of the strawberry cryoconcentrate in the production of yogurts did not

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affect the survival of lactic acid bacteria during 7 days of storage. However, the

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cryoconcentrate addition promoted a reduction in the pH and consequently an increase
in the acidity. The use of 30% strawberry cryoconcentrate promoted a product with high
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anthocyanins content and antioxidant activity. Besides that, the product presented great
colour parameters, similar to two commercial trademarks which use artificial dyes. All
the yogurts showed shear thinning and thixotropic properties, confirming non-
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Newtonian behaviour and the addition of concentrated pulp promoted a reduction in the
hysteresis areas which is positive in terms of yogurt quality standards. Finally, the use
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of strawberry cryoconcentrate showed to be an excellent alternative for the production

of a dairy beverage with high nutritional quality. Nevertheless, shelf life study of the
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enriched yogurt and its sensorial characteristics are of great importance on this matter.
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In this sense, to the next step of this research, the evaluation of the consumers
acceptance, product shelf life and other important factors - as the in vitro digestibility –
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are proposed, in order to achieve a complete screening of the nutritional effects of the
cryoconcentrated strawberry pulp addition in yogurts.
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Acknowledgments

The authors are thankful to the CNPq - Brazil and CAPES - Brazil for the financial
support and scholarships.
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Table 1: pH, titratable acidity, total solids and colour parameters L*and a* obtained for control yogurt,
yogurt enriched with of cryoconcentrated strawberry pulp (yogurt 1 and 2), and commercial yogurts (C1
and C1), during 7 days of storage.
1 day 7 days
aA
Control 4.6 ± 0.03 4.3 aA ± 0.02
pH Yogurt 1 4.0 aB ± 0.01 3.9 aB ± 0.02
Yogurt 2 3.8 aB ± 0.05 3.7 aC ± 0.01
C1 - 3.5 D ± 0.03
C2 - 3.3 E ± 0.02

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1 day 7 days
aC
Control 0.41 ± 0.00 0.44 aC ± 0.03
Acidity

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Yogurt 1 0.71 aB ± 0.06 0.69 aB ± 0.01
(g.100mL )
Yogurt 2 1.05 aA ± 0.01 1.00 aA ± 0.01

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C1 - 0.42 C ± 0.04
C2 - 0.30 D ± 0.01
1 day 7 days
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aC
Control 11.02 ± 0.093 11.23 aE ± 0.08
Total solids (%) Yogurt 1 11.90 aB ± 0.002 12.02 bD ± 0.01
12.61 aA ± 0.29
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Yogurt 2 12.22 bC ± 0.01

C1 - 20.07 A ± 0.04
C2 - 18.39 B ± 0.039
1 day 7 days
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aA
Control 82.12 ± 0.30 82.52 aA ± 0.26
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L* Yogurt 1 73.55 aB ± 0.66 73.40 aB ± 0.28

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Yogurt 2 66.70 aC ± 0.36 67.96 aC ± 0.60

C1 - 66.54 D ± 0.15
C2 - 72.93 B ± 0.11
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1 day 7 days
aC
Control -2.03 ± 0.01 -2,21 bE ± 0,01
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a* Yogurt 1 10.05 aB ± 0.08 9,45 bD ± 0,08

Yogurt 2 18.59 aA ± 0.11 15,93 bB ± 0,08
C1 - 18,64 A ± 0,05
C2 - 12,95 C ± 0,04
Means followed by the same lower-case letter in the line did not differ statistically at 5% significance.
Means followed by the same upper-case letter in the column did not differ statistically at 5% significance.
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Table 2: Results of Lactic Acid Bacteria (LAB) count of control yogurt, yogurt enriched
with 15% (yogurt 1) and 30 % (yogurt 2) of cryoconcentrated strawberry pulp and
commercial yogurts (C1 and C2) during storage time.
1 day 7 days
Control 8.0·108 aA ± 0.03 3.2 ·108 bC ± 0.02
LAB Yogurt 1 5.5·108 aB ± 0.05 3.2 ·108 bC ± 0.02
(UFC.g-1) Yogurt 2 4.9 ·108 aC ± 0.01 3.0 ·108 bD ±0.04
C1 3.6·108 B ± 0.01

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C2 4.5·108 A ± 0.03

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Means followed by the same lower-case letter in the line did not differ statistically at
5% significance. Means followed by the same upper-case letter in the column did not

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differ statistically at 5% significance.
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Table 3: Total anthocyanins content (AC) and antioxidant activity determined by ABTS
for the different yogurts analysed.
1 day 7 days
Control 0.00aC 0.00aC
Yogurt 1 1.86±0.018aB 1.60±0.016bB
AC* (mg.100g-1) Yogurt 2 2.99±0.032aA 2.54±0.029bA
C1 - 0.00C
C2 - 0.00C

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Control 0.01 aB ± 0.02 0.00 bC ± 0.01
0.26 aB ± 0.02 0.25 aB ± 0.00

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Yogurt 1
ABTS
-1
Yogurt 2 0.38 aA ± 0.03 0.37 aA ± 0.02
(mgGAE.mL )

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C1 - 0.00 D ± 0.01
C2 - 0.01 D ± 0.01
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Control 0.00 aC ± 0.00 0.00 bC ± 0.00
Yogurt 1 8.86 aB ± 0.02 8.97 aB ± 0.18
DPPH
Yogurt 2 9.19 aA ± 0.05 9.92 aA ± 0.18
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-1
(mgGAE.mL )
C1 - 0.01 C ± 0.00
C2 - 0.02 C ± 0.00
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5% significance. Means followed by the same upper-case letter in the column did not
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differ statistically at 5% significance. * Expressed as mg of pelargonidin-3-O-glucoside

per 100 mL of yogurt.
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Table 4: Rheological parameters obtained using Power Law model ( ) for

the control, yogurt 1 (15% of strawberry cryoconcentered) and yogurt 2 (30% of
strawberry cryoconcentered) and commercial yogurts during the storage time.
Power Law Model
Day
K (Pa.sn) n R²
1 1.692 0.30 0.965
Control
7 1.275 0.35 0.982

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1 0.562 0.46 0.989
Yogurt 1
7 0.435 0.46 0.982

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1 0.313 0.54 0.987
Yogurt 2
7 0.197 0.55 0.979

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C1 - 1.217 0.50 0.997
C2 - 1.577 0.44 0.989
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K, consistency index; n, flow behaviour index; R², correlation coefficient.
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Figure 1: (a) Fluxogram of block freeze concentration procedure where V0 represent the
initial volume of strawberry juice, C1, C2 and C3 represents the concentrated fractions
and G1, G2 and G3 represents the ice fractions; (b) Schematic representation of the
equipment used for block freeze concentration.

Figure 2: Apparent viscosity versus shear rate on (a) control sample (b) 15% of
strawberry cryoconcentrate, (c) 30% of strawberry cryoconcentrate for the different
times of storage and (d) commercial trademarks

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Figure 3: Apparent viscosity versus shear rate for the: (a) control; (b) yogurt 1 (15% of

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strawberry cryoconcentrate), (c) yogurt 2 (30% of strawberry cryoconcentrate) during

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the storage time and (d) commercial trademarks.
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Figure 4: Hysteresis area for the control sample, yogurt 1, yogurt 2 and commercial
trademarks (C1 and C2) during the storage time.
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Fig. 1
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Fig. 2
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Fig. 4
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Graphical abstract
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Highlights
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- Freeze concentration raised the content of bioactive compounds in strawberry pulp

- Production of yogurt with high phenolic content using a cryoconcentrated pulp
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- Effect of cryoconcentrate pulp addition in the yogurt physicochemical properties

were investigated
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- High quality and nutritional enhanced yogurt was produced with natural additives