LWT - Food Science and Technology 140 (2021) 110730

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Thermosonication for peroxidase inactivation in sugarcane juice

Jucelio Kulmann de Medeiros a, Julia Ribeiro Sarkis b, Debora Pez Jaeschke b,
Giovana Domeneghini Mercali c, *
a
Federal Education, Science and Technology Institute of Santa Catarina, 150, Quatorze de Julho St., Florianópolis, SC, Brazil
b
Department of Chemical Engineering, Federal University of Rio Grande do Sul, 2777, Ramiro Barcelos St., Porto Alegre, RS, Brazil
c
Department of Food Science, Institute of Food Science and Technology, Federal University of Rio Grande do Sul, 9500, Bento Gonçalves Av., Porto Alegre, RS, Brazil

A R T I C L E I N F O A B S T R A C T

Keywords: To allow a widespread commercialization of sugarcane juice it is necessary to develop an effective preservation
Enzymatic inactivation method. For that, thermosonication was evaluated as an alternative processing technology. Therefore, ultrasound
Ultrasound (US) and conventional (CH) treatments were compared regarding peroxidase (POD) inactivation. Both treat­
Cavitation
ments were performed for 25 min at 50–80 ◦ C, with matched temperature profiles. POD inactivation kinetics
Thermal treatment
were evaluated under different US parameters: pulse regime, US application time and power intensity. After
Emerging technologies
treatments, sugarcane juice was monitored for 32 days under refrigerated storage to verify juice quality. Results
showed that POD inactivation followed Weibull model for US and CH treatments at 70, 75 and 80 ◦ C. At 70 and
75 ◦ C, US lead to faster POD inactivation when compared to CH. The use of 20s on/10s off pulse regime, US
application for 25 min and 75% power intensity were the best conditions for enzyme inactivation (77.3%).
During refrigerated storage, POD regeneration, phenolic compounds degradation nor sedimentation were
observed in the juice treated by US. For CH, on the other hand, sedimentation and lower phenolic compounds
were observed. Overall, US is a potential technology for sugarcane juice enzymatic inactivation with reduced
processing times and/or temperatures, leading to a better quality juice.

1. Introduction
Sugarcane (Saccharum officinarum L.) is a distinguished crop, being
extensively used for both ethanol and sugar production. Brazil is
currently the main producer, with 7 million hectares of cultivated land,
followed by India, Thailand and Australia (Marin, Pellegrino, Assad,
Pinto, & Zullo Junior, 2009). Sugarcane juice is a sweet and inexpensive
beverage, refreshing and energetic, popular in Brazil and other tropical
countries, sold in restaurants, supermarkets, trailers and kiosks (Frate­
schi et al., 2013). Concerning its appearance, this liquid is opaque with
color varying from grey to dark green (Rezzadori, Petrus, Benedetti,
Carminatti, & Petrus, 2013). Total soluble solids vary between 15 and 25
◦
Brix depending on environmental factors and harvest period; the pH is
moderately acid, ranging from 5 to 6, frequently in the 5.2–5.4 interval.
Sugarcane contains a significative amount of phenolic compounds,
mainly phenolic acids and flavonoids, that have physiological and
morphological functions (Havsteen, 2002). These compounds are found
in several foods and contribute to its sensory characteristics, such as
color, astringency and bitterness, being the most abundant antioxidants
in human diet (Sant’Anna, Gurak, Ferreira Marczak, & Tessaro, 2013).
Sugarcane juice has high water activity, moderate pH and high sugar
content which contribute to the presence of a vast microbial flora,
making it very perishable (Brochier, Mercali, & Marczak, 2016; Garud,
Priyanka, Negi, & Rastogi, 2017). Moreover, the juice contains enzymes,
such as polyphenol oxidase (PPO) and peroxidase (POD), that promote
oxidative reactions, causing undesirable changes in both organoleptic
and nutritional properties. As a result, sugarcane juice marketing and
consumption is usually informal and occur immediately after its pro­
duction (Huang, Chang, & Wang, 2015; Rezzadori, Petrus, Benedetti,
Carminatti, & Petrus, 2013). Thermal treatment can reduce sugarcane
juice microbial count and enzymatic browning. However, it can also
lead to the occurrence of Maillard reaction and accelerate chlorophyll
and phenolic compounds degradation, changing sensory and nutritional
characteristics (Huang, Chang, & Wang, 2015).
POD is an oxidoreductase enzyme that catalyzes several oxidation
reactions of organic and inorganic substrates in plants using mostly

* Corresponding author.
E-mail addresses: jucelio.medeiros@ifsc.edu.br (J. Kulmann de Medeiros), julia@enq.ufrgs.br (J.R. Sarkis), deborapj@enq.ufrgs.br (D.P. Jaeschke), giovana.
mercali@ufrgs.br, gimercali@gmail.com (G.D. Mercali).

https://doi.org/10.1016/j.lwt.2020.110730
Received 14 September 2020; Received in revised form 17 November 2020; Accepted 5 December 2020
Available online 8 December 2020
0023-6438/© 2020 Elsevier Ltd. All rights reserved.
J. Kulmann de Medeiros et al.
Fig. 1. Thermosonication equipment setup: a) heating bath, b) maintenance bath, c) bypass system, d) jacketed glass cell, e) magnetic stirrer, f) data acquisition
system, g) US control unit and h) US probe coupled with temperature sensors.
hydrogen peroxide as the electron acceptor (Chisari, Barbagallo, &
Spagna, 2007). These reactions are responsible for enzymatic browning
which is commonly observed in sugarcane juice (Brochier, Hertz,
Marczak, & Mercali, 2020). POD activity varies mainly with pH and
temperature and the optimum conditions for this enzyme are widely
approached in literature (Chisari, Barbagallo, & Spagna, 2007; Rojas,
Trevilin, Esteves, & Augusto, 2016; Rojas, Trevilin, Funcia, Gut, &
Augusto, 2017); several authors reports, for different food products and
raw materials, a large range of pH (from 4.5 to 6.5) and temperatures
(from 24 ◦ C to 60 ◦ C). Brochier, Mercali, and Marczak (2018) reported
an ideal temperature around 60 ◦ C and an optimum pH between 5.5 and
6.0 for sugarcane juice. POD is heat resistant enzyme, being used as a
quality indicator for thermal processes. It is important to highlight that,
even after thermal treatment, there is a possibility of enzyme regener­
ation; residual enzymatic activity during storage can increase up to the
levels observed prior to treatment (Rojas et al., 2017).
Consumers seek for a sugarcane juice with high quality and micro­
biological safety, rich in antioxidants and without chemical additives
(Huang, Chang, & Wang, 2015). An effective conservation method
should be developed, allowing a wide commercialization by controlling
microbial growth and maintaining the product natural qualities (Huang,
Chang, & Wang, 2015). Thermosonication has been studied as an
alternative for juice thermal processing; this technology combines the
effects of heat and cavitation to inactivate microorganisms and enzymes
using milder temperatures and/or shorter processing times (Anaya-Es­
parza et al., 2017). High-power ultrasound (≥1 W cm− 2) promotes
modifications in food structure and composition manly due to cavita­
tion. The cavitation phenomenon consists on the formation and increase
of microscopic bubbles with fast collapse caused by pressure differences
within the media. The bubbles collapse violently, and, consequently,
localized high temperatures (more than 5000 K), pressures (more than
50,000 kPa) and shear effects are observed (Tiwari, 2015).
Ultrasound (US) effects on enzyme structure is still unclear and many
mechanisms have been proposed to explain the phenomenon. Enzyme
inactivation is mainly attributed to the physical and chemical effects of
cavitation. Shear forces may disrupt hydrogen bonds and van der Waals
interactions in polypeptides chains, causing changes on secondary and
tertiary protein structures, leading to losses in biological activity. In
addition, cavitation may favor free radicals release due to water mole­
cules sonolysis. These free radicals may react with amino acid residues
of enzymes, that play an important role in enzyme stability, substrate
binding or catalytic function, resulting on enzymatic activity changes
(Cao, Cai, Wang, & Zheng, 2018).
In this context, the present study aimed to evaluate the effectiveness
of thermosonication to inactivate peroxidase in sugarcane juice and to
2
LWT 140 (2021) 110730
preserve the product quality parameters (color and total phenolic
compounds); moreover, the stability of the juice during storage was
analyzed.
2. Material and methods
2.1. Sugarcane juice
Sugarcane was donated by a rural property located in Veranópolis
(Rio Grande do Sul, Brazil, 28◦ 56′ 10′′ S, 51◦ 32′ 58′′ W) and was from a 5
years-old cane regrowth regime. The juice was processed in the rural
property, according to the subsequent steps: the sugarcane was har­
vested and mechanically milled, the juice was filtrated, homogenized,
packaged in 150 mL aliquots in polypropylene bags, and stored at
− 18 ◦ C. For the experiments, samples were thawed in water bath, ho­
mogenized and maintained in ice bath until thermal processing. Fresh
sugarcane juice from two different crop seasons were used: harvest I,
from January 2019, and harvest II, from August 2019.
2.2. Sugarcane juice characterization
The fresh sugarcane juice was characterized as follows: pH (DM - 22,
Digimed, São Paulo, Brazil) using a pHmeter; soluble solids (TSS, ◦ Brix
degree) using a digital refractometer (MA871, Milwaukee Instruments,
Rocky Mount, USA); electrical conductivity (EC), measured by a digital
conductivity meter (DM - 32, Digimed, São Paulo, Brazil); and water
activity, measured by an electric hygrometer at 25 ◦ C (Novasina,
Labmaster-aw model, Lachen, Switzerland). Total flavonoids com­
pounds (TFC) were determined according to the methodology described
by Zhuang et al. (1992), using UV–Vis spectrophotometer (UV-1800,
Shimadzu, Kyoto, Japan) at 510 nm. Total phenolic compounds (TPC),
PPO and POD activities were also assayed as described in sections
2.4–2.5. All analyzes were performed in triplicate.
2.3. Color analysis
Color was determined using the CIELAB parameters: L* (lightness or
brightness/darkness), a* (redness) and b* (yellowness). A ColorQuest XE
colorimeter (Hunter Lab, Reston, USA) was used with light source D65,
observation angle of 10◦ and reflectance specular included mode.
2.4. Total phenolic compounds
Total phenolic compounds (TPC) were determined by UV–Vis spec­
trophotometry (UV-1800, Shimadzu, Kyoto, Japan), following the
J. Kulmann de Medeiros et al. LWT 140 (2021) 110730

Fig. 2. Temperature profile over heating time (first 15 min) for US and CH experiments at 80 ◦ C.

methodology described by Waterhouse (2001). Briefly, samples reacted

Table 1
with Folin-Ciocalteau reagent and sodium carbonate prior to absorbance
Parameters evaluated for US optimization at 75 ◦ C.
determination at 765 nm. Results were obtained using a calibration
curve and expressed as mg of galic acid equivalents per liter (mg Treatment US intensity (%) Sonication time (min) Pulses (on/off)
GAE/L). I 75 25 20 s/10 s
II 75 25 40 s/10 s
III 75 25 Continuum
2.5. Polyphenol oxidase and peroxidase enzymatic activity IV 50 25 20 s/10 s
V 100 25 20 s/10 s
POD activity was determined based on protocols described by VI 75 First 5 min 20 s/10 s

Chisari, Barbagallo, & Spagna, 2007, modified by Brochier et al. (2016).

PPO activity followed the method defined by Walker (2001), with
modifications. For both methodologies, the extract was prepared by
mixing (1:5, w:v) sugarcane juice with phosphate-citrate buffer pH 5.4
(McIlvaine buffer) in vortex (Phoenix solutions, AP-56 model) (Walker,
2001b). Afterwards, the mixture was centrifuged (CIENTEC, CT, model
5000 R, Brazil) for 10 min, at 4 ◦ C and 6000 rpm, and filtered through
qualitative filter paper.
For POD activity measurement, the extract was mixed with hydrogen
peroxide 1% (Labsynth, Diadema, Brazil) and guaiacol 0.5% v/v
(Dinâmica, Indaiatuba, Brazil), and the absorbance was read at 460 nm
in a UV–Vis spectrophotometer (UV-1800, Shimadzu, Kyoto, Japan) at
25 ◦ C. For PPO analysis, the extract was added to a catechol solution,
and the absorbance read at 420 nm in the spectrophotometer. For both
assays, a blank was carried out replacing the sample by distilled water.
The enzyme activity (A) was determined measuring the absorbance
variation per minute (Δabs/min) and dividing this value by 0.001. The A
value was obtained by linear regression (determination coefficients
above 0.99), and only the linear part of the absorption curve was taken
into account. Analyses were performed in triplicate.
2.6. Control and ultrasonic treatments
The US apparatus used to conduct the experiments is shown in Fig. 1
and consisted of an US probe equipment (frequency 20 kHz; probe
diameter 1.2 cm; 50 W cm− 2; amplitude 140 μm; Sonics model VC 750,
Newtown, EUA). The experiments were performed with 110 mL of
sample in a 250 mL jacketed glass cell. Temperature was measured by
two pt-100 temperature sensors.
Two thermal treatments were performed for comparison, thermo­
sonication (US) and conventional heating (CH). The experiments
comprised two phases: the come-up and the isothermal phase. The
temperature in the come-up phase was raised by passing hot water
thorough the cell jacket (bath set at 98 ◦ C; Alpha A 6, Lauda, Lauda-
Königshofen, Germany). For US, in the isothermal phase, the ultrasound
equipment was turned on and a second bath (Alpha RA 8, Lauda, Lauda-
Königshofen, Germany) was used to maintain the temperature at the set
point. For CH, samples were heated just using hot water thought the
jacket. A bypass system, consisted of valves and hoses, was used to
alternate water baths. Samples were withdrawn in specific heating
times, cooled and kept in ice bath until further analyses.
Time and temperature profiles of US and CH experiments were
matched to infer the US effects (without the interference of the thermal
effects). Fig. 2 shows temperature over time (for the first 15 min of
heating) during US and CH experiments at 80 ◦ C, indicating similar
profiles. Analogous plots were obtained for all conditions evaluated in
this study.
2.7. Inactivation kinetics at different temperatures
POD and PPO kinetics experiments were carried out at 50, 60, 70, 75
and 80 ◦ C ± 2 ◦ C, using sugarcane juice from harvest I. US and CH
treatments were performed in triplicate following the procedure
described in section 2.6. US was applied with 75% intensity, pulse
regime of 20/10 s on/off. Samples were withdrawn at specific treatment
times: 0 (beginning of the isothermal phase), 2.5, 5, 10, 15, 20 and 25
min. Due to the obtained results, PPO inactivation kinetics was reeval­
uated at 50 ◦ C (US and CH) using a smaller cell (100 mL of volume) with
different samples withdrawn times during the isothermal phase: 0, 1, 3,
5, 7.5, 10 and 15 min.
2.8. US parameters evaluation: intensity, pulse regime and time
Experiments to evaluate the effect of US intensity, pulse regime and
time on POD inactivation were carried out at 75 ◦ C following the same

3
J. Kulmann de Medeiros et al. LWT 140 (2021) 110730

Table 2 fitted to the first order and Weibull distribution models, shown in Eqs.
Physicochemical composition of different batches of in natura sugarcane juice. (2) and (3), using non-linear regression (STATISTICA version 12.0,
Parameter Harvest I Harvest II StatSoft, Inc. Tulsa, OK).
Harvest period January 2019 August 2019 A = A0 exp( − k ⋅ t) (2)
TSS (◦ Brix) 19.6 ± 0.7aa 17.6 ± 0.2b
pH 5.38 ± 0.05a 5.68 ± 0.05b A = A0 exp( − b ⋅ tn ) (3)
EC (μS⋅cm− 1, 20 ◦ C) 1955 ± 57a 3647 ± 14b
0.947 ± 0.003a 0.962 ± 0.001b − 1
aw In these equations, t is the time (min), k is the rate constant (min ), b
TPC (mg EAG⋅L− 1) 476 ± 26a 490 ± 39a
is the Weibull scale parameter (min-n), n is the Weibull shape parameter,
APOD (UAE⋅min− 1⋅g− 1) 3044 ± 331a 4813 ± 527b
APPO (UAE⋅min− 1⋅g− 1) 1148 ± 114a 2170 ± 239b A0 and A are the enzymatic activity at time zero and time t, respectively
a
(Weibull, 1951).
Different lower-case letters indicate a significant difference between har­
The goodness of fit was evaluated by three statistical parameters:
vests (p < 0.1).
determination coefficient (R2), chi-square (χ2) and root mean square
error (RMSE). The last two parameters are represented by Eqs. (4) and
procedure defined in section 2.6. Harvest II was used to perform the (5), respectively. In these equations, the experimental and predicted
experiments that are described in Table 1. A control treatment was also values are represented by αexp and αpred, correspondingly, n is the
performed in the same temperature. For all kinetic curves, samples were number of observations and p is the number of parameters.
withdrawn in specific times of the isothermal phase: 0, 2.5, 5, 10, 15, 20 ( )
and 25 min. Σ αexp− αpred
χ2 = (4)
(n − p)
2.9. Enzymatic regeneration during refrigerated storage
1[ ( )2 ]0,5
RMSE = Σ n αexp − αpred (5)
Experiments to evaluate the POD regeneration during refrigeration n
storage were carried out. Samples from harvest II were pasteurized by Data were compared by Student t-test, ANOVA and Tukey test with
US and CH at 80 ◦ C (±2 ◦ C) during the time necessary to reach a decimal 90% confidence level (STATISTICA version 12.0, StatSoft, Inc. Tulsa,
reduction of POD activity. The times for US and CH were determined OK).
according to the kinetic experiments (fitted to the Weibull model) and
were of 23 and 36 min, respectively. The experiments were carried out 3. Results and discussion
in triplicate following the methodology described in section 2.6. US
experiments were accomplished using 75% of intensity and pulse regime 3.1. Sugarcane juice physicochemical characterization
of 20/10s on/off.
Fresh and pasteurized samples were packaged in 15 mL falcon tubes, Table 2 shows the physicochemical characterization of in natura
which were filled up to the top to eliminate air in the head space, closed sugarcane juice for both harvests used in the present work. Differences
and sealed with plastic laboratory film. Sugarcane juice samples were between batches were observed for total soluble solids content (TSS),
stored at the bottom of a domestic refrigerator at 4 ± 2 ◦ C. POD activity, pH, electrical conductivity (EC) and water activity (aw); all values found
TPC and color were measured in different storage times: 0, 1, 2, 4, 8, 14, were in the expected range for sugarcane juice (Brochier et al., 2016;
18, 22 and 32 days. Brochier et al., 2018; Saxena, Makroo, & Srivastava, 2016). Total
phenolic compounds did not show significant differences between
2.10. Mathematical modeling and statistical analysis batches and the results were in accordance with previous work (Brochier
et al., 2020, 2016).
Residual activities (A/A0) were plotted against processing time and

Fig. 3. Sugarcane juice PPO residual enzymatic activity over time for CH and US treatments at 50 ◦ C. The points are the experimental data, and the lines represent
the Weibull model.

4
J. Kulmann de Medeiros et al.
Fig. 4. Sugarcane juice POD residual enzymatic activity over time for CH and US treatments at 50 and 60 ◦ C (A) and at 70, 75 and 80 ◦ C (B). The points are the
experimental data, and the lines represent the Weibull model.
The initial enzymatic activity evaluation demonstrated that both
peroxidase (POD) and polyphenoloxidase (PPO) are active in sugarcane
juice. Sample comparison highlights the variability among harvests,
which are expected and may be related to the weather differences be­
tween summer (January 2019) and winter (August 2019), as previously
described in literature (Brochier et al., 2018, 2016; Saxena, Makroo, &
Srivastava, 2016). The high perishability of this juice, caused by the
action of microorganisms and enzymes, is strongly related to high values
observed for pH, water activity, soluble solids, and initial enzymatic
activity.
3.2. PPO inactivation kinetics at different temperatures
PPO inactivation in sugarcane juice was evaluated at 50, 60, 70 and
80 ± 2 ◦ C. Results showed that at 60, 70 and 80 ◦ C, inactivation occurred
mostly during the come-up phase. At the end of this stage (time zero of
the isothermal phase), 76 ± 8, 97.3 ± 0.8 and 97.4 ± 0.5% of inacti­
vation was observed at 60, 70 and 80 ◦ C, respectively. Therefore, during
the isothermal phase, PPO inactivation did not present the expected
exponential behavior, and data could not be fitted to an inactivation
5
LWT 140 (2021) 110730
model. At 50 ◦ C, on the other hand, the inactivation rate was slower: at
the end of the come-up phase, 39 ± 7% of inactivation was found. This
inactivation level is still considered high and comparison between
technologies could not be performed.
Similar results were found by Brochier et al. (2016) during conven­
tional and ohmic heating of sugarcane juice; these authors observed high
PPO inactivation rates during the come-up phase at 60–90 ◦ C, and
modeling was not possible. According to these authors, only the most
resistant enzymatic fraction was active during the isothermal phase,
which resulted in an unpredicted inactivation trend. PPO heat sensi­
tivity may be attributed to its complex structure, containing 3 or 4
subunits in higher plants, making this enzyme more susceptible to
inactivation (Illera et al., 2018).
To better evaluate PPO inactivation, a second batch of experiments
was performed at 50 ◦ C. A smaller cell was used for faster heating. These
results are presented in Fig. 3, which shows experimental and predicted
data (Weibull model) for PPO residual activity during CH and US
treatments. Experimental data were fitted to first order and Weibull
models. The statistical analysis showed that the Weibull model was the
most appropriate one, presenting higher determination coefficients
J. Kulmann de Medeiros et al.
Table 3
Statistical indexes ([minimum; maximum]) for first order and Weibull models
describing POD inactivation in sugarcane juice.
Model R2 χ2 RMSE
First order [0.608; 0.976] − 5
[1.99⋅10 ; 7.87⋅10 ]− 3 − 2
[1.16⋅10 ; 3.03⋅10 ]− 3
Weibull [0.892; 0.9990] [6.89⋅10− 5; 1.64⋅10− 3] [6.89⋅10− 5; 1.64⋅10− 3]
(0.999 for CH and US), and lower RMSE (3.04۰10− 4 and 5.90۰10− 4, for
CH and US respectively) and χ 2 (1.61۰10− 4 and 3.12۰10− 4, for CH and
US respectively) values.
The Weibull model coefficients for PPO inactivation, b (0.21 ± 0.02
and 0.15 ± 0.05, for CH and US, respectively) and n (0.77 ± 0.04 and
0.93 ± 0.15, for CH and US, respectively), did not present statistical
difference between CH and US treatments. These results indicate that
PPO activity was not significantly influence by US application, and the
inactivation was a consequence of temperature only. To the best of our
knowledge, this is the first report regarding US effects on sugarcane juice
PPO structure. Different results were reported by other researchers that
applied US in other food matrices; US influenced PPO inactivation in
bayberry juice (Cao et al., 2018), tomato (Ercan & Soysal, 2011) and
melon juice (Fonteles et al., 2012). It is important to notice that the
aforementioned studies did not matched CH and US temperature pro­
files. Facing the results obtained for PPO inactivation kinetics and
considering that POD is more thermoresistant, the later enzyme was the
focus of the present work.
3.3. POD inactivation kinetics at different temperatures
Fig. 4 shows the experimental data for POD inactivation in sugarcane
juice at all analyzed temperatures. The solid and dashed lines represent
the Weibull model fitted to the experimental data for US and CH,
respectively. Fig. 4-A shows the results at the lowest analyzed temper­
atures, 50 and 60 ◦ C. When CH was applied at 50 ◦ C, enzymatic acti­
vation was observed; at 60 ◦ C this activation was observed only after 2.5
min followed by mild inactivation. This behavior may indicate that the
optimum temperature of POD in sugarcane juice may be close to 60 ◦ C.
Brochier et al. (2016) also reported a POD activation in sugarcane juice
at 60 ◦ C. Overall, as the temperature rises enzyme activity increases up
to a maximum value at the optimum temperature. After this point, the
high temperature breaks the enzyme structure, promoting denaturation.
POD behavior as a function of temperature depends on the food matrix;
the optimum activity temperature for this enzyme in sugarcane juice
was not found in the literature. When US was applied, enzyme activation
was not noted, and results were similar for 50 and 60 ◦ C. This indicates
that US may affect the optimum temperature of the POD activity. These
results are different from the behavior observed by Brochier et al.
(2016), that found an increase in enzyme activity at 60 ◦ C for both
conventional and ohmic heating. The differences suggest that 60 ◦ C is
the limit temperature for POD enzymatic activity.
Data from experiments at 50 and 60 ◦ C were not fitted to the models
because they were not adequate to represent enzyme activation and low
inactivation rates (models showed low goodness of fit). According to
Van Boekel (2008), for a proper estimation of the parameters, the
experiment should be conducted such that a considerable extent of re­
action is reached. For higher temperatures, higher inactivation rates
were observed, and the models were suitable to represent experimental
data. Fig. 4-B shows the enzymatic inactivation over time for CH and US
at 70, 75 and 80 ◦ C; in these temperatures a decrease of the enzyme
activity was observed in all scenarios. At 70 ◦ C differences between both
treatments were found: after 25 min of treatment with US sugarcane
presented 52% of the initial enzymatic activity, whereas with CH the
activity was 82% of the initial value. The experimental curves at 60, 70,
75 and 80 ◦ C displayed a non-linear behavior, that indicates faster
enzyme inactivation at the beginning of thermal treatments. This may be
related to the presence of more heat resistant enzyme fractions
6
LWT 140 (2021) 110730
Table 4
Weibull parameters for POD residual activity in sugarcane juice at all analyzed
temperaturesa.
Temperature b (min-n) n
(◦ C)
CH US CH US
70 0.04 ± 0.08 ± 0.6 ± 0.2aA 0.67 ±
0.01aA 0.01bA 0.05aA
75 0.06 ± 0.25 ± 0.91 ± 0.57 ±
0.02aA 0.05bB 0.09bB 0.06aA
80 0.28 ± 0.26 ± 0.59 ± 0.69 ±
0.08aB 0.05aB 0.08aA 0.06aA
a
Different lower-case letters indicate a significant difference between CH and
US, and different capital letters indicate a significant difference between tem­
peratures for each parameter (p < 0.1).
(isoenzymes). Brochier et al. (2020) obtained a zymogram from fresh
sugarcane juice, where three bands with enzymatic activity were
observed; their findings indicate that there are at least three isozymes in
the juice. The research also demonstrated that these fractions have
different thermal resistances, which can be related to the behavior
observed in the present work.
Table 3 shows the parameters used to evaluate the most adequate
model to describe the experimental data. Minimum and maximum
values for the first order and Weibull models in all temperatures and
treatments are shown. The first order model had more variability within
the parameters and presented higher maximums for RMSE and χ 2 ;
moreover, the Weibull model resulted in higher determination co­
efficients. Therefore, this model was selected to fit experimental data.
Although POD inactivation is most commonly described by the first
order kinetic model (Ali, Russly, Jamilah, Azizah, & Mandana, 2011;
Cao et al., 2018; Ercan & Soysal, 2011), a similar inactivation behavior
was reported by other researchers: POD inactivation in sugarcane juice
during ohmic heating treatment (70–80 ◦ C, 25 V) (Brochier et al., 2016)
and in water green coconut during US treatment was well described by
the Weibull model (Rojas et al., 2017). It is known that POD inactivation
rates depend not only on the intensity of the treatment but also on the
type of fruits or cultivars (Cao et al., 2018). For sugarcane juice, the first
order model may have presented inferior results as it does not consider
isoenzymes and is not able to describe the downward concavity
observed in Fig. 4. It is important to point out that the isoforms found in
sugarcane juice may behave differently in the presence of US (due to the
non-thermal effects) and this may not be correctly represented by any
existing mathematical model.
Table 4 shows the Weibull model parameters (b and n) fitted to the
different treatments. As aforementioned, the Weibull model was not
adequate to fit the experimental data at 50 and 60 ◦ C. Analyzing the
presented results for the scale parameter (b), it is possible to conclude
that inactivation rates increased with temperature, as expected.
Comparing US and CH treatments, both parameters presented signifi­
cant differences at 70 and 75 ◦ C. Once temperature profiles were
matched, the decrease of the enzymatic activity promoted by US may be
related to non-thermal effects. On the other hand, at 80 ◦ C no significant
differences were observed, indicating that an increase in temperature
lead to smaller differences between CH and US. This may be related to
the magnitude of both effects observed in this study: thermal and non-
thermal enzymatic inactivation. Increasing temperatures increases
thermal effects on POD which can shadow the non-thermal effects of US.
The US non-thermal effects on enzymes structure are related to the
cavitation phenomenon that may break up hydrogen bonds and van der
Waals interactions, leading to an enzymatic activity decrease. Further­
more, free radicals may be released in the medium and react with spe­
cific enzymes sites and amino acid residues, destabilizing enzyme
conformation structure (Cao et al., 2018).
Concerning the shape parameter (n), it assumed values lower than 1
in all scenarios studied. This behavior is commonly observed for mi­
crobial and enzyme inactivation curves which present a downward
J. Kulmann de Medeiros et al. LWT 140 (2021) 110730

Fig. 5. Sugarcane juice POD residual enzymatic activity over time for CH and US treatments: A) different pulse regimes at 75% of power intensity; B) different
process times at 75% of power intensity; and C) different power intensities at 20/10 s on/off pulse regime. The points are the experimental data, and the lines
represent the Weibull model.

concavity. No differences between treatments and temperatures were
found for this parameter, except for the CH treatment at 75 ◦ C; this
suggests that future research could be performed using an average value
for n and determining only the scale factor.
To the best of our knowledge, the present work is the first report on
POD inactivation by thermosonication in sugarcane juice. Although food
matrix may influence enzyme behavior during thermal and US treat­
ments, similar results were reported by other researchers in other
matrices, such as pear and apple juice. POD inactivation was evaluated
after conventional (65 ◦ C, 10 min and 95 ◦ C, 2 min) and thermosoni­
cation (25, 45 and 65 ◦ C, 10 min, 20 KHz, 750 W) treatments in pear
juice. The authors reported that enzyme inactivation by US can be
achieved at lower temperatures, if compared to the conventional pro­
cess. Moreover, analyzing both process at the same temperature (65 ◦ C),
POD residual activity was 4.3 and 66% after thermosonication and
conventional treatment, respectively (Saeeduddin et al., 2015). The
combined effect of US and temperature on POD inactivation was also
reported for apple juice. POD residual activity after thermosonication at
60 ◦ C (10 min, 20 kHz, 70% of power, 750 W) was 9.0% whereas at 40
and 20 ◦ C was 57.0 and 98.1% of its initial value (Abid et al., 2014). The
results found by these authors and in the present work highlight the
importance of using US combined with temperature to achieve satis­
factory enzyme reductions.
3.4. US parameters evaluation: pulse regime, processing time and
intensity
To evaluate the influence of operational variables on POD inactiva­
tion kinetics at 75 ◦ C, different combinations of pulse regime, processing
time and power intensity were evaluated (Table 1). Fig. 5 shows
experimental and predicted data during CH and US treatments under: A)
different pulse regimes at 75% of power intensity; B) different process­
ing times at 75% of power intensity; and C) different power intensities at
20/10 s on/off pulse regime. For a proper comparison among

Table 5
Weibull parameters for POD residual activity in sugarcane juice for CH different US treatments.
Treatment US Intensity (%) Sonication time (min) Pulses b (min-n)a na R2
(on/off)

CH – – – 0.06 ± 0.02c 0.91 ± 0.09a 0.992

I 75 25 20 s/10 s 0.25 ± 0.05a 0.57 ± 0.06c 0.998
II 75 25 40 s/10 s 0.19 ± 0.04ab 0.68 ± 0.07bc 0.984
III 75 25 Continuum 0.18 ± 0.06ab 0.68 ± 0.07bc 0.996
IV 50 25 20 s/10 s 0.07 ± 0.02c 0.89 ± 0.11a 0.990
V 100 25 20 s/10 s 0.13 ± 0.01bc 0.79 ± 0.04ab 0.993
VI 75 First 5 min 20 s/10 s 0.19 ± 0.04ab 0.5 ± 0.07c 0.994
a
Different letters indicate a significant difference between the treatments (p < 0.1).

7
J. Kulmann de Medeiros et al.
Fig. 6. POD residual activity in sugarcane juice during refrigerated storage time: conventional (CH) and ultrasonic treatments (US) at 80 ◦ C.
treatments, POD residual activity over time was fitted to the Weibull
model (R2 varied from 0.984 to 0.998), and b and n parameters are
presented in Table 5.
Pulse regime influence was analyzed by the comparison between CH
and US treatments I, II and III (Fig. 5-A). Results showed that pulse
regime did not affect POD inactivation kinetics, once Weibull model
scale and shape parameters (b and n, respectively) did not significantly
vary among US treatments (p > 0.1). Moreover, higher POD inactivation
rates were achieved by all US treatments, if compared to CH, that pre­
sented lower b value. Differences in POD residual activity for US and CH
treatments were more intense in the beginning of the treatments. These
results may be related to n parameter, as US treatments resulted in lower
n values, indicating a more pronounced non-linear behavior. Therefore,
20/10 s on/off pulse regime was considered the best condition due to the
lower required energy.
Few authors have explored the effects of pulse regime on enzymes
inactivation kinetics and the results are ambiguous. POD inactivation in
green beans assisted by US (20 kHz, 1.5 kW, 45–225 s, 0.2–0.8 s on/1 s
off, 50–90 ◦ C) was higher with pulse regime of 0.65 s on/1 s off;
moreover, an increase in duty cycle over 0.7 s on/1 s off lead to an in­
crease of the POD residual activity (Yolmeh & Najafzadeh, 2014). Other
researchers, on the other hand, reported that pulse mode was less effi­
cient than continuous mode (20 kHz, 15 min, 79 μm of amplitude, 5 s on
and 5 s off or continuous) for PPO inactivation in apple juice (Illera et al.,
2018). Further studies are needed to better explain the influence of this
parameter on enzymatic inactivation.
Comparison of treatments I, VI and CH is presented in Fig. 5-B. In
treatment I sonication was kept during the entire treatment (25 min) and
in treatment VI, only in the first 5 min. These experiments were per­
formed to verify the hypothesis that an initial damage to the enzymes
would be enough for inactivation. Results showed that higher b values
were obtained for US treatments (I and VI) when compared to CH,
indicating higher POD inactivation rates. It is believed that US promoted
modifications in thermostable isoenzymes structure, that may lead to
higher exposure of active bonds sites. Comparing treatment VI and CH,
higher inactivation rates were obtained for treatment VI just in the first
15 min. These results may be attributed to the US effect which was
responsible for the inactivation of less tolerant isoenzymes in the first 5
min. Facing the obtained results, 25 min was chosen as the ideal pro­
cessing time.
Different US power intensities were evaluated: 50% (treatment IV),
75% (treatment I) and 100% (treatment V) (Fig. 5-C). Results showed
8
LWT 140 (2021) 110730
that higher POD inactivation rates were achieved at higher power in­
tensities (treatments I and V), once b values were higher for these
treatments. These results were expected once cavitation is favored at
high power intensities. Similar behavior was reported for PPO inacti­
vation in apple juice treated by US at power intensities of 25–100%
(amplitude of 19–76 μm) (Illera et al., 2018). Weibull model parameters
for CH and treatment IV did not present significant differences, indi­
cating that 50% of power intensity is not enough to promote US
non-thermal effects on POD structure. Therefore, as 75 and 100% of
power intensity had a similar effect on enzyme inactivation, 75% of
power intensity was considered the best condition due to the lower
energy demand.
3.5. Sugarcane juice stability during refrigerated storage
Thermosonication and conventional pasteurization treatments were
performed to evaluate sugarcane juice stability under refrigerated
storage for 32 days. During this period, samples were analyzed
regarding POD activity, total phenolic content and color changes. The
treatments were designed to inactivate 90% of the initial POD activity.
For this, Weibull model was used to predict process time at 80 ◦ C: 36 min
for CH and 23 min for US treatment.
3.5.1. POD residual activity during refrigerated storage
Right after each pasteurization treatment, samples were evaluated
regarding POD activity (time zero), resulting in 6.0 ± 1.0% and 4.4 ±
0.8% for CH and US treatments, respectively. Fig. 6 shows POD residual
activity over time for sugarcane juice ultrasonically and conventionally
treated. Results show that POD residual activity remained constant
during storage for both heating methods once values at the beginning
and at the end of the storage period did not present significant differ­
ences (p > 0.1). It is important to highlight that the standard deviations
were slightly high; nonetheless, the data indicate that enzyme regener­
ation did not occur during this period. Similar results were observed by
Kohli et al. (2019) that evaluated PPO activity during refrigerated
storage in sugarcane juice after HPH treatment; the researchers did not
observe changes in PPO activity during 28 days. Few authors have
evaluated enzyme stability after US treatment. POD activity was moni­
tored in tomato juice after thermosonication (15–75% of power in­
tensities, 20–150 s, up to 75 ◦ C) and during refrigerated storage (- 4 ◦ C).
Inactivation right after the treatments varied from 35.8% (15% of power
intensity, 150 s) to 100% (50–75% of power intensity, 90–150 s) and
J. Kulmann de Medeiros et al. LWT 140 (2021) 110730

Fig. 7. Normalized phenolic compounds concentration (TPC/TPC0) in sugarcane juice during refrigerated storage time: conventional (CH) and ultrasonic treatments
(US) at 80 ◦ C.

residual activity reached 59% during storage (Ercan & Soysal, 2011). In
Table 6
the present work, conventional and thermosonication treatments were
CIELab color parameters for ultrasonic and conventionally treated samples
more severe, with higher processing times and temperature, that may
stored under refrigerationa.
have lead to irreversible enzymatic inactivation.
Distinct results were observed by other authors that evaluated Treatment Time (days) L* a* b*

enzyme regeneration after different thermal treatments in food matrices: CH IN 29.8 ± 1.1b,c 0.91 ± 0.07a 0.92 ± 0.04b
POD activity in chili powder decreased during 6 months of storage at 0 28.7 ± 0.7b 0.56 ± 0.10a 1.25 ± 0.27b,c
8 32.3 ± 0.9a 3.16 ± 0.03b,c 4.80 ± 1.08a
ambient temperature after conventional blanching (90–100 ◦ C, 5–10
14 32.5 ± 0.7a 3.37 ± 0.39c 4.80 ± 1.25a
min) (Schweiggert, Schieber, & Carle, 2006); pectinmethylesterase 32 31.6 ± 0.6a,c 2.59 ± 0.36b 3.53 ± 0.84a,c
(PME) activity did not change during storage after microwave treat­ US IN 27.5 ± 1.5B 1.03 ± 0.15B 1.26 ± 0.01C
ment, whereas after conventional treatment (90 ◦ C, 3.1–12.5 s), enzy­ 0 31.9 ± 1.7A 0.29 ± 0.11A 1.73 ± 0.14A,B
matic activity increased after 12 days of storage (Salazar-González, San 8 33.1 ± 0.1A 0.38 ± 0.03A 1.82 ± 0.10B
14 33.4 ± 0.1A 0.33 ± 0.01A 1.57 ± 0.06A
Martín-González, Vergara-Balderas, López-Malo, & Sosa-Morales, 32 33.4 ± 0.2A 0.18 ± 0.02A 1.60 ± 0.08A,B
2014). This emphasizes that enzyme regeneration depends on the in­
a
tensity of the treatment and on the food matrix. Different letters indicate a significant difference between the treatments (p <
0.1).

3.5.2. Phenolic compounds stability during refrigerated storage

Phenolic compounds concentration in sugarcane juice did not pre­
sent significant changes when comparing fresh and treated samples just
after CH and US treatments. The relative stability of sugarcane juice
phenolic compounds to heat was reported in previous studies (Brochier
et al., 2016, 2018). Phenolic compounds concentration was evaluated
before and after conventional and ohmic heating at 60–80 ◦ C for 6 and
12 min (Brochier et al., 2016). According to the researchers, degradation
varied from 11 to 23% and started after 6 min of treatment. In other
study, 7–12% of degradation was observed during thermal treatment
come-up phase (approximately 2 min) but after 25 min of conventional
and ohmic heating (75 ◦ C) no further decrease on concentration was
found (Brochier et al., 2018).
Normalized phenolic compounds concentration (TPC/TPC0) in sug­
arcane juice during storage is presented in Fig. 7. Results showed that for
CH treatment, phenolic compounds concentration decreased in day 1,
without further significant changes (p > 0.1) up to day 32. For US
treatment, on the other hand, concentration did not change significantly
during storage. These results may be attributed to the different times of
each thermal treatments: CH was performed for 36 min while US, for 23
min. It is believed that after CH treatment phenolic compounds degra­
dation reactions may have been favored.
To the best of our knowledge, the present work is the first report on
phenolic compounds stability in sugarcane juice during storage. How­
ever, similar results were reported after thermal treatment in other food
matrices. Phenolic compounds concentration in pindo palm did not vary
significantly right after pasteurization (85 ◦ C, 20 min) and during
refrigerated storage (Jachna, Hermes, Flôres, & Rios, 2016). Similarly,
pasteurization process did not affect phenolic compounds concentration
in orange passion fruit. After 4 days of refrigerated storage, a decrease in
concentration was observed and the same content kept constant for 15
days (dos Reis, Facco, Flôres, & Rios, 2018). In this regard, the results
obtained in the present work showed that US pasteurization may be used
to obtain sugarcane juice with high phenolic compounds concentration.
3.5.3. Color variation during refrigerated storage
Possible color changes during sugarcane juice storage were evalu­
ated and the results are presented in Table 6. For each color parameter a
comparison over storage time was performed. For CH and US treat­
ments, L* and b × increased over time, indicating luminosity and yellow
color increase, respectively. Parameter a*, on the other hand, increased
during CH treatment, indicating a greener juice, and decreased during
US treatment, indicating a redder juice. The increase in L* may be
attributed to enzymatic inactivation and the decrease of enzymatic
browning reactions. Although Maillard reactions may have occurred,
their effects on color were probably lower than those promoted by
enzymatic inactivation. a × and b × parameters may be related to
phenolic compounds and chlorophyll concentration in sugarcane juice.
However, a correlation with phenolic compounds is difficult to be

9
J. Kulmann de Medeiros et al.
Fig. 8. Sugarcane juice after conventional (CH) and ultrasonic (US) treatments
at 80 ◦ C and 32 days of refrigerated storage.
performed due to variations on samples composition (Brochier et al.,
2018; Duarte-Almeida, Salatino, Genovese, & Lajolo, 2011).
Fig. 8 presents pictures of the juice after 32 days of storage. As can be
seen, sedimentation occurred in CH treated juice while US treated
samples were more turbid. Sedimentation problem is often observed in
fruit juices and was recently reported for sugarcane juice after
pasteurization (80 ◦ C, 10 min) and 28 days of storage (Kohli et al.,
2019). These researchers also reported that sedimentation was pre­
vented after a non-thermal hurdle process using high-pressure homog­
enization and the preservatives nisin and polylisine. The result was
attributed to the homogenization effects promoted by the high-pressure
treatment that reduced large particles. Similarly, the increase in
turbidity observed after US treatment may be related to US effects on
non-soluble particles; due to cavitation, large particles may be down­
sized, remaining suspended in the juice rather than settled. These phe­
nomena may also be related to changes in a × and b × parameters, as US
juice became browner (dun) and yellower due to suspended particles
(decrease in a × to almost zero and a small increase in b*). CH juice, on
the other hand, was less turbid and yellower (increase in b*) than US
treated juice.
4. Conclusions
This study evaluated the thermosonication effect on POD inactiva­
tion in sugarcane juice. POD inactivation by US and CH treatments
followed the Weibull model distribution for all the evaluated conditions.
At 70 and 75 ◦ C, higher inactivation rates were obtained by US when
compared to CH. These results were attributed to a combined effect of
cavitation and temperature. An evaluation of juice stability during
refrigerated storage showed that juice sedimentation may be prevented
by US application. Moreover, phenolic compounds and POD residual
activity did not vary during 32 days of storage. Overall, results showed
that thermosonication is an interesting alternative to conventional
thermal processing for sugarcane juice. Thermosonication may be used
to obtain a safe product at milder temperature conditions and/or with
shorter processing time, which can enhance product quality and process
efficiency. The data from the present study contributes for the correct
understanding of the US non-thermal effects on enzyme inactivation,
allowing the prediction of adequate processing conditions.
10
LWT 140 (2021) 110730
CRediT authorship contribution statement
Jucelio Kulmann de Medeiros: experiments, Validation, Formal
analysis, Investigation, Resources, Writing - original draft, Visualization.
Julia Ribeiro Sarkis: Conceptualization, Methodology, Validation,
Formal analysis, Writing - review & editing, Supervision. Debora Pez
Jaeschke: analysis of the results, statistics, Writing - original draft,
Visualization. Giovana Domeneghini Mercali: Conceptualization,
Methodology, Validation, Formal analysis, Resources, Writing - review
& editing, Supervision, Project administration.
Acknowledgements
The authors greatfully acknowledge the financial support received
from FAPERGS (Fundação de Amparo à Pesquisa do Estado do Rio
Grande do Sul) and CNPq (Conselho Nacional de Desenvolvimento
Científico e Tecnológico).
References
Abid, M., Jabbar, S., Hu, B., Hashim, M. M., Wu, T., Lei, S., et al. (2014).
Thermosonication as a potential quality enhancement technique of apple juice.
Ultrasonics Sonochemistry, 21(3), 984–990. https://doi.org/10.1016/j.
ultsonch.2013.12.003
Ali, G., Russly, A. R., Jamilah, A., & Mandana. (2011). Effect of heat and
thermosonication on kinetics of peroxidase inactivation and vitamin C degradation
in seedless guava (Psidium guajava L.). International Food Research Journal, 18.
Anaya-Esparza, L. M., Velázquez-Estrada, R. M., Roig, A. X., García-Galindo, H. S.,
Sayago-Ayerdi, S. G., & Montalvo-González, E. (2017, March 1). Thermosonication:
An alternative processing for fruit and vegetable juices. Trends in Food Science &
Technology, 61, 26–37. https://doi.org/10.1016/j.tifs.2016.11.020. Elsevier Ltd.
Brochier, B., Hertz, P. F., Marczak, L. D. F., & Mercali, G. D. (2020). Influence of ohmic
heating on commercial peroxidase and sugarcane juice peroxidase inactivation.
Journal of Food Engineering, 284, 110066. https://doi.org/10.1016/j.
jfoodeng.2020.110066
Brochier, B., Mercali, G. D., & Marczak, L. D. F. (2016). Influence of moderate electric
field on inactivation kinetics of peroxidase and polyphenol oxidase and on phenolic
compounds of sugarcane juice treated by ohmic heating. Lebensmittel-Wissenschaft &
Technologie, 74, 396–403. https://doi.org/10.1016/J.LWT.2016.08.001
Brochier, B., Mercali, G. D., & Marczak, L. D. F. (2018). Effect of ohmic heating
parameters on peroxidase inactivation, phenolic compounds degradation and color
changes of sugarcane juice. Food and Bioproducts Processing, 111, 62–71. https://doi.
org/10.1016/j.fbp.2018.07.003
Cao, X., Cai, C., Wang, Y., & Zheng, X. (2018). The inactivation kinetics of polyphenol
oxidase and peroxidase in bayberry juice during thermal and ultrasound treatments.
45, 169–178. https://doi.org/10.1016/j.ifset.2017.09.018
Chisari, M., Barbagallo, R. N., & Spagna, G. (2007). Characterization of polyphenol
oxidase and peroxidase and influence on browning of cold stored strawberry fruit.
Journal of Agricultural and Food Chemistry, 55, 3469–3476. https://doi.org/10.1021/
jf063402k
Duarte-Almeida, J. M., Salatino, A., Genovese, M. I., & Lajolo, F. M. (2011). Phenolic
composition and antioxidant activity of culms and sugarcane (Saccharum
officinarum L.) products. Food Chemistry, 125(2), 660–664. https://doi.org/
10.1016/j.foodchem.2010.09.059
Ercan, S.Ş., & Soysal, Ç. (2011). Effect of ultrasound and temperature on tomato
peroxidase. Ultrasonics Sonochemistry, 18(2), 689–695. https://doi.org/10.1016/j.
ultsonch.2010.09.014
Fonteles, T. V., Costa, M. G. M., de Jesus, A. L. T., de Miranda, M. R. A.,
Fernandes, F. A. N., & Rodrigues, S. (2012). Power ultrasound processing of
cantaloupe melon juice: Effects on quality parameters. Food Research International,
48(1), 41–48. https://doi.org/10.1016/j.foodres.2012.02.013
Frateschi, C. S., Durigan, J. F., Marques, M. O., Hojo E, T. D., Santos, L. O., Cunha
Júnior, L. C., et al. (2013). Storage of sugarcane stalks (Saccharum officinarum cv.
SP 79-1011) in low oxygen atmospheres and the effects on enzymatic browning.
Postharvest Biology and Technology, 86, 154–158. https://doi.org/10.1016/j.
postharvbio.2013.06.032
Garud, S. R., Priyanka, B. S., Negi, P. S., & Rastogi, N. K. (2017). Effect of
thermosonication on bacterial count in artificially inoculated model system and
natural microflora of sugarcane juice. Journal of Food Processing and Preservation, 41
(2). https://doi.org/10.1111/jfpp.12813
Havsteen, B. H. (2002). He biochemistry and medical significance of the flavonoids.
Pharmacology and Therapeutics. T. https://doi.org/10.1016/S0163-7258(02)00298-X
Huang, H. W., Chang, Y. H., & Wang, C. Y. (2015). High pressure pasteurization of
sugarcane juice: Evaluation of microbiological shelf life and quality evolution during
refrigerated storage. Food and Bioprocess Technology, 8(12), 2483–2494. https://doi.
org/10.1007/s11947-015-1600-2
Illera, A. E., Sanz, M. T., Benito-Román, O., Varona, S., Beltrán, S., Melgosa, R., et al.
(2018). Effect of thermosonication batch treatment on enzyme inactivation kinetics
and other quality parameters of cloudy apple juice. Innovative Food Science &
Emerging Technologies, 47, 71–80. https://doi.org/10.1016/j.ifset.2018.02.001
J. Kulmann de Medeiros et al.
Jachna, T. J., Hermes, V. S., Flôres, S. H., & Rios, A. O. (2016). Bioactive compounds in
pindo palm (Butia capitata) juice and in pomace resulting of the extraction process.
Journal of the Science of Food and Agriculture, 96(4), 1216–1222. https://doi.org/
10.1002/jsfa.7209
Kohli, G., Jain, G., Bisht, A., Upadhyay, A., Kumar, A., & Dabir, S. (2019). Effect of non-
thermal hurdles in shelf life enhancement of sugarcane juice. Lebensmittel-
Wissenschaft & Technologie, 112, 108233. https://doi.org/10.1016/j.
lwt.2019.05.131
Marin, F. R., Pellegrino, G. Q., Assad, E. D., Pinto, H. S., & Zullo Junior, J. (2009). Cana-
de-açúcar. In J. E. B. A. Monteiro (Ed.), Agrometeorologia dos cultivos: O fator
meteorológico na produção agrícola (pp. 109–130). Brasília: INMET.
dos Reis, L. C. R., Facco, E. M. P., Flôres, S. H., & A de O, Rios (2018). Stability of
functional compounds and antioxidant activity of fresh and pasteurized orange
passion fruit (Passiflora caerulea) during cold storage. Food Research International,
106, 481–486. https://doi.org/10.1016/j.foodres.2018.01.019
Rezzadori, K., Petrus, R. R., Benedetti, S., Carminatti, C. A., & Petrus, J. C. C. (2013).
Effects of tangential microfiltration and pasteurisation on the rheological,
microbiological, physico-chemical and sensory characteristics of sugar cane juice.
International Journal of Food Science and Technology, 48(1), 1–9. https://doi.org/
10.1111/j.1365-2621.2012.03124.x
Rojas, M. L., Trevilin, J. H., Esteves, P., & Augusto, D. (2016). The ultrasound technology
for modifying enzyme activity. Scientia Agropecuaria, 7(2), 145–150. https://doi.org/
10.17268/sci.agropecu.2016.02.07
Rojas, M. L., Trevilin, J. H., Funcia, E., dos, S., Gut, J. A. W., & Augusto, P. E. D. (2017).
Using ultrasound technology for the inactivation and thermal sensitization of
peroxidase in green coconut water. Ultrasonics Sonochemistry, 36, 173–181. https://
doi.org/10.1016/j.ultsonch.2016.11.028
Saeeduddin, M., Abid, M., Jabbar, S., Wu, T., Hashim, M. M., Awad, F. N., et al. (2015).
Quality assessment of pear juice under ultrasound and commercial pasteurization
processing conditions. Lebensmittel-Wissenschaft und -Technologie- Food Science and
Technology, 64(1), 452–458. https://doi.org/10.1016/j.lwt.2015.05.005
Salazar-González, C., San Martín-González, M. F., Vergara-Balderas, F. T., López-
Malo, A., & Sosa-Morales, M. E. (2014). Physical-chemical and microbiological
11
LWT 140 (2021) 110730
stability during refrigerated storage of microwave-pasteurized guava nectar. Focusing
on Modern Food Industry, 3, 43. https://doi.org/10.14355/fmfi.2014.03.006, 0030.
Sant’Anna, V., Gurak, P. D., Ferreira Marczak, L. D., & Tessaro, I. C. (2013). Tracking
bioactive compounds with colour changes in foods – a review. Dyes and Pigments, 98
(3), 601–608. https://doi.org/10.1016/j.dyepig.2013.04.011
Saxena, J., Makroo, H. A., & Srivastava, B. (2016). Optimization of time-electric field
combination for PPO inactivation in sugarcane juice by ohmic heating and its shelf
life assessment. Lebensmittel-Wissenschaft und -Technologie- Food Science and
Technology, 71, 329–338. https://doi.org/10.1016/j.lwt.2016.04.015
Schweiggert, U., Schieber, A., & Carle, R. (2006). Effects of blanching and storage on
capsaicinoid stability and peroxidase activity of hot chili peppers (Capsicum
frutescens L.). Innovative Food Science & Emerging Technologies, 7(3), 217–224.
https://doi.org/10.1016/j.ifset.2006.03.003
Tiwari, B. K. (2015). Ultrasound: A clean, green extraction technology. TRAC Trends in
Analytical Chemistry, 71, 100–109. https://doi.org/10.1016/J.TRAC.2015.04.013
Van Boekel, M. A. J. S. (2008). Kinetic modeling of food quality: A critical review.
Comprehensive Reviews in Food Science and Food Safety, 7(1), 144–158. https://doi.
org/10.1111/j.1541-4337.2007.00036.x
Walker, J. R. L. (2001a). Expression and measurement of enzyme activity. In Current
protocols in food analytical chemistry (pp. C4.1.1–C4.1.15). John Wiley & Sons, Inc.
Walker, J. R. L. (2001b). Polarographic and spectrophotometric assay of diphenol
oxidases. In R. E. Worlstad, T. E. Acree, A. Haejung, E. A. Decker, M. H. Penner,
D. R. Reid, et al. (Eds.), Current protocols in food analytical chemistry. New York: John
Willey and Sons (p. C4.1.7-C4.1.8).
Waterhouse, A. L. (2001). Determination of total phenolics. In Current protocols in food
analytical chemistry.. John Wiley & Sons, Inc. https://doi.org/10.1002/0471142913.
faa0101s06.
Weibull, W. (1951). Statistical distribution function of wide applicability. Journal of
Applied Mechanics, 103, 293–297.
Yolmeh, M., & Najafzadeh, M. (2014). Optimisation and modelling green bean’s
ultrasound blanching. International Journal of Food Science and Technology, 49(12),
2678–2684. https://doi.org/10.1111/ijfs.12605
Zhuang, X. P., & Lu, Y. Y. (1992). Extraction and determination of flavonoid in ginkgo.
Chinese Herbal Medicine, (23), 122–124.