chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592

Contents lists available at ScienceDirect

Chemical Engineering Research and Design

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

Polyphenol extraction from fresh tea leaves by

pulsed electric field: A study of mechanisms

Aleksandra Zderic a , Edwin Zondervan b,∗

a Eindhoven University of Technology, De Zaale, 5612 AJ Eindhoven, The Netherlands
b Bremen University, Leobener Strasse, 21895 Bremen, Germany

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

Article history: The major interest in pulsed electric field treatment of biological tissues is derived from
Received 15 December 2015 its non-thermal application: increasing cell permeability. This application has an important
Received in revised form 2 March implication in extraction of complex organic molecules. In this work, pulsed electric field
2016 treatment is investigated as a mild (non-thermal) processing method for opening the cell
Accepted 5 March 2016 structure in fresh tea leaves. Pulsed electric field utilizes short-duration high-voltage pulses
Available online 12 March 2016 for opening the cell structure by the process called electroporation. Upon the treatment,
subsequent extraction of complex organic molecules, particularly, polyphenols, occurs. The
Keywords: amount of extracted polyphenols (in this case, the extraction yield) has been determined
Pulsed electric field as a function of electric field strength, duration and number of applied pulses, as well as
Polyphenols energy input per unit of mass of the sample. The results indicate that the used conditions
Fresh tea leaves during the treatment increase in temperature did not exceed 10 ◦ C. This limited temperature
Non-thermal method rise provides a valid evidence that pulsed electric field processing is a non-thermal method
applied under used conditions.
© 2016 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1. Introduction (Asavasanti et al., 2011). The term electroporation is used to

Recently, commercial interest in the extraction of intracellu-
lar organic compounds and liquids from cellular plant tissues
using various solid–liquid extraction methods has been grow-
ing. One of the factors influencing the extraction process is the
degree of cell membrane disintegration (Bazhal et al., 2003).
Various physical, chemical or biological treatments damage
the cellular membrane. The pulsed electric field (PEF) tech-
nique is a non-thermal processing method for heterogeneous
food materials. Moreover, PEF is based on cell transforma-
tion or rupture under exposure of an external electric field
resulting in an increase in the electrical conductivity and
permeability of the cell membrane (Lebovka et al., 2000; Soliva-
Fortuny et al., 2009; Haberl et al., 2013). The application of
electric fields for a short duration of a few to several hundred
microseconds is capable of inducing cell membrane perme-
abilization through a phenomenon called “electroporation”
describe phenomena that accompany the exposure of cells to
transmembrane electrical pulses (Weaver, 2000). Applying an
external electric field to the cells results in pore formation in
the membrane. Because pore formation is a dynamic process
depending on the intensity of the PEF treatment, electropo-
ration can be reversible or irreversible. When induced pores
are small in comparison to the membrane area and if they
are generated with PEF treatment of low intensity, then the
electric breakdown is reversible (Soliva-Fortuny et al., 2009).
Increasing the intensity of the treatment by increasing the
electric field strength (E) and/or treatment time (t) (which con-
siders the number of pulses and the pulse width applied in
the system) will result in the formation of large pores, and
reversible permeabilization will turn into irreversible disrup-
tion of the cell membrane. The irreversible permeabilization
of the cell membrane in the plant tissue provides a wide range
of process applications where disruption of the cell membrane

∗
Corresponding author. Tel.: +49 421 218 63301; fax: +49 421 218 63302.
E-mail address: edwin.zondervan@uni-bremen.de (E. Zondervan).
http://dx.doi.org/10.1016/j.cherd.2016.03.010
0263-8762/© 2016 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592 587

Therefore, in practice, it is difficult to separate two mech-

Nomenclature anisms (thermal and/or electroporation mechanism) and to
make statements regarding the real contribution of the elec-
c concentration (kg m−3 ) tric field to electroporation followed by transfer of polyphenols
Dd diffusion coefficient (m−2 s) from the cell interior to the surrounding liquid. By mea-
E electric field strength (V m−1 ) suring the total amount of polyphenols (PPs) transferred, it
F0 Fourier number (dimensionless) was possible to monitor the process of pulse-induced mem-
I current (A) brane permeabilization. Therefore, the objective of this work
m mass (kg) is to study the effect of the operational parameters (electric
N number of pulses (dimensionless) field strength, pulse duration and number of pulses) on the
PBP pause between pulses (s) extraction yield of the polyphenols. The total specific energy
PD pulse duration (s) input (kJ/kg leaves) and the temperature increment (◦ C) are
PEF pulsed electric field chosen as parameters to describe treatment intensity as non-
PPs polyphenols thermal.
Ppulse power per pulse (W) The amount of PPs in tea leaves is around 30% on
R radius (m) total amount of solids which is 25% (Graham, 1992; Parsija
T temperature (◦ C) and Anandharamakrishnan, 2015). This makes the con-
t time (s) tent very attractive for extraction and concentration as
U voltage (V) an added-value product. Traditional methods for extrac-
V volume (m3 ) tion of PPs include heating, boiling and Soxhlet extraction.
Wspec specific energy input (J kg−1 ) These methods are limited by low extraction efficiency
wt weight fraction (%) and the degradation of PPs and other components due
to the high temperature or presence of solvent. That is
the reason that extraction, in particular, should be done
is required. Both reversible and irreversible electroporation under mild conditions. PEF has been used as cell opening
have found their applications in different disciplines such as technique under mild conditions and subsequently dif-
biomedicine, biotechnology and environmental science (see, fusion of PPs from the cell matrix into aqueous media
e.g. Haberl et al., 2013; Yarmush et al., 2014). In food processing, occurs. Therefore, at high temperature, neither additional
currently intensive research has been done in non-thermal solvents. From the table below, it can be seen that for
preservation and sterilization by microbial inactivation (Wan mentioned extraction techniques, starting material is dried
et al., 2009; Monfort et al., 2010; Walkling-Ribeiro et al., 2011; leaves. In our study, fresh tea leaves were used for experi-
Evrendilek et al., 2013). ments.

Extraction Starting material Temperature Solvent Efficiency Extraction time

method

Extraction with Dried leaves High/boiling No Low Relatively

hot water short (few
minutes)
Soxhlet method Dried leaves High ∼ 70 ◦ C N-hexane; Relatively high Long, 24 h
(Fui-Seung et al., powder methanol; about 65%
2013) dichloromethane
Direct extraction Dried leaves Room tempera- 20% acetonitrile Relatively high Long, 24 h
(Fui-Seung et al., powder ture about 65%
2013)

Note: starting materials in all extraction methods are dried

Electric field strength is an important factor that con-
trols the efficiency of electroporation of cellular tissue. Bazhal
et al. (2003) presented a classification of the PEF modes
as low (E ≤ 100–200 V/cm), moderate (E = 300–1500 V/cm) and
high (E > 1500 V/cm). With a low electric field strength,
the treatment time should be longer for electroporation
of the cellular membranes. It has been found experimen-
tally that the time needed for electroporation of cellular
membranes of the different biological tissues is inversely pro-
portional to the electric field strength (Bouzrara and Vorobiev,
2003).
PEF processing of foods involves the application of short
pulses (duration of micro- to milliseconds) of high elec-
tric field intensity. The pulsed electric field application
to the tea leaves can result in increase in temperature
(through ohmic heating) and electrically stimulated damage of
cell walls/membranes (through electroporation mechanism).
tea leaves. In our study, fresh tea leaves were used, not more
than 2 days old after plucking.
2. Materials and methods
2.1. Composition of tea leaf
Fresh tea leaves from Kenya (Camellia sinensis variety) were
used. The tea leaf contains 30% of catechins, 2% simple
polyphenols (gallic acid), 3% caffeine and the rest (proteins,
minerals, organic acids and carbohydrates). The composition
is given as weight fraction (% w/w) on dry mater. Catechins,
gallic acid and caffeine concentrations have been measured
by HPLC.
588 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592
2.2. Moisture content analysis
Fresh tea leaves were plucked 1 day before shipment to Europe
(The Netherlands) and they were stored at 5 ◦ C until required.
Before treatment, leaves were allowed to reach the ambient
temperature of approximately 20 ◦ C. To determine the mois-
ture content, fresh leaves were freeze-dried and subsequently
subjected to a hot-air oven at 105 ◦ C. Moisture content was
75–80 wt%. Samples that were used for the moisture content
analysis were separated from the samples used for PEF treat-
ment. For all experiments, thin slices of fresh tea leaves were
used (approximately 1 cm width).
2.3. Natural extraction without electrical treatment:
“control” experiments
For the control experiments without electrical treatment,
slices of fresh tea leaves (untreated, m = 20 g) were placed into
a cylindrical glass beaker. Distilled water (200 g) was added
at 20 ◦ C, and then diffusion was studied. A careful agita-
tion at 250 rpm was provided. The concentrations of the total
polyphenols in water ware measured after 20 min. It is possi-
ble to determine the length of experiments, using the Fourier
number to calculate how much time is needed to reach steady
state. At that point in time, the extracted amount of polyphe-
nols is in equilibrium with the rest of polyphenols in the spent
leaves:
Dd · t
F0 =
R2
where D is the diffusion coefficient (m2 /s), t the time (s) and
R the radius of the particle (m).
When the Fourier number reaches Eq. (1), steady state
is achieved. The time necessary to reach equilibrium is the
time needed for our experiments. The equilibrium can be
represented with the following mass balance. Here d in the
subscript represents the dispersed PPs (in spent leaves) and c
the continuous phase (water) at time t = 0 and t = ∞.
Vd · (cd,0 − cd,∞ ) = Vc · (cc,∞ − cc,0 )
where V is the volume (m3 ) and c the concentration (kg/m3 ).
2.4. Pulsed electric field treatments
Tea leave samples were treated using the pulsed electric field
(PEF) equipment with a batch treatment configuration of the
Nutri-Pulse NP110-60 System (IXL Netherlands B.V.) which
consists of a PEF treatment chamber and a high-voltage gen-
erator. The batch treatment chamber (100 mm length × 70 mm
width × 50 mm height, 350 mL capacity) consists of two par-
allel stainless steel electrodes of 5 mm thickness separated
by a distance of 20 mm (see Fig. 1a). A high-voltage genera-
tor provided rectangular pulses in the range of 0.1 × 10−3 –0.1 s
with a maximum voltage of 2.2 kV and a maximum number
of pulses 50. Samples were placed in the treatment chamber
between the two stainless steel electrodes filled with a ster-
ile salt solution (Fig. 1a). The conductivity of the sterile salt
solution was adjusted to correspond with the conductivity of
the sample ( = 3.5 mS/cm). NaCl as a salt was used for the
preparation of the salt solution. After cutting, the leaves were
subjected to various PEF treatments. All experiments were
carried out using an electric field strength (E) ranging from
0.1 to 1.1 kV/cm, pulse duration (PD) from 0.1 × 10−3 –0.1 s and
number of pulses (N) from 10 to 50. The total treatment time
was defined as the product of the number of pulses and the
pulse width applied to the system (Fig. 1b). Apart from the
treatment time, the pause between two pulses (PBP) is the
interval between two pulses and represents the relaxation
time.
The temperature was measured both at the inlet and outlet
of the treatment chamber. In all experiments, the increase in
the temperature due to the treatment never exceeded 10 ◦ C.
All experiments were duplicated.
2.5. Specific energy input
The pulse shape (square wave bipolar) was monitored on-line
with an oscilloscope (Model RTM 2000) during the treatment.
For a square-wave pulse, the specific energy input for each
PEF-treated sample was calculated on the basis of Eq. (3):

n 
ttreatment
n
Wspec = U(t) · I(t) · dt (eq.3)
m
0
0
The total specific energy input was calculated by multiply-
ing the energy delivered per pulse with the number of pulses
applied into the system.
Wspec is the specific energy input (in kJ/kg). The energy per
pulse (in kJ) is calculated from the power of the pulse (Ppulse )
multiplied by pulse duration (s). Ppulse is the result of out-
(eq. 1) put voltage U(t) and the total electric current I(t) supplied to
the sample on the basis of Ohm’s law. n is the pulse number
applied to the system (dimensionless) and m is the total weight
of sample (kg) charged to the treatment chamber for the PEF
treatment applied.
2.6. Determination of extraction yield—measurement
of total polyphenols content
The disintegration of the cellular membrane was detected
indirectly by a total polyphenols (PPs) content measurement.
(eq. 2) Based on the fact that the cellular membrane is ruptured,
transport of the cell material together with the polyphenols
occurred and the extraction yield is defined as a fraction
of extracted PPs. After the treatment, samples were stored
at ambient temperature and left in an aqueous solution for
20 min (extraction time). Upon extraction, the leaves were sep-
arated from the aqueous solution. The amount of extracted
PPs was measured in the solution. The total amount of phe-
nols was determined by directly reading the absorbance at
725 nm (SpectraMax 190 Absorbance Microplate Reader, USA)
of diluted samples 1/10 (v/v). The total amount of PPs was
expressed as gallic acid equivalents (GAE) by means of a cor-
responding calibration curve with standard gallic acid. The
extraction yield of PPs was calculated according to Eq. (4):
extraction yield of PPs
polyphenols content (aquoues solution)
= · 100% (eq.4)
polyphenols content (fresh tea leaves)
2.7. Statistical analysis
All experiments and measurements of characteristics were
repeated at least twice. A one-way analysis of variance was
 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592
Fig. 1 – Experimental set up (lab scale). Scheme of PEF treatment chamber (a) and PEF pulse protocol (b).
used for statistical analysis of the data using the Statgraphics
Centurion® XVI. For each analysis, a significance level of 5%
was assumed. The error bars presented in the figures corre-
spond to the standard deviations.
3. Results and discussion
There are several techniques for extraction of PPs from tea
leaves such as microwave-assisted extraction (Nkhilli et al.,
2009; Pan et al., 2003) solvent-based extraction (Meterc et al.,
2007) and (Dong et al., 2011), molecular distillation (Tang
et al., 2011) and ultrasonication (Koiwai and Masuzawa, 2006).
Microwave-assisted extraction uses hot/boiling water and in
addition, irradiation power of 600 W (Nkhilli et al., 2009). PPs,
proteins and other valuable components from fresh tea leaves
are sensitive to high temperatures. Usually, ethanol is used
as solvent for extraction of PPs from tea. In a later stage,
ethanol is removed from the process. However, there could
still be some residue of ethanol in the final product. In the
proposed process, no additional solvent is used and due to
complete absence of ethanol, the proposed process is more
effective compared to the solvent-based extraction method.
In this work, we will focus on the pulsed electric field method.
Fresh tea leaves were subjected to pulsed electric field
(PEF) treatments. Extraction of polyphenols occurred as a
result of pore formation and disintegration of the cellular
2500
2000
1500
Voltage, V
1000
500
0 -100
-2.00E-04 -1.00E-04 0.00E+00 1.00E-04
Fig. 2 – A recording of voltage (blue dotted line) and current (red dotted line) during a PEF experiment.
589
membrane. Therefore, when PEF-treated leaves were in con-
tact with an aqueous solution after 20 min of extraction time,
their color changed in yellowish brown. This phenomenon
occurs due to the transport of cellular material into the sur-
rounding aqueous solution and the reaction of PPs catalyzed
by the enzyme polyphenol oxidase (PPO). PPO is located in the
cytoplasm of the plant cells and PPs are in organelles called
vacuoles. When PPO and PPs are brought into contact, oxi-
dized PPs are produced as result of enzymatic oxidation. The
formation of oxidized PPs was identified by visual observa-
tion: a change in color of the solution during the oxidation
reaction occurs. Furthermore, UV spectrophotometer analy-
sis was used to determine the total amount of PPs by reading
the absorbance at 725 nm.
3.1. Characterization of pulsed electric field process
and electric measurements
In the experiments performed, electrical measurements, i.e.,
voltage and current, were recorded for each PEF experiment at
regular time intervals. Fig. 2 shows an example of a recording
of voltage (V) and current (mA) using an oscilloscope.
In addition, in the experiments carried out in this study
next to the electrical measurements (voltage and current),
temperature and resistance were also monitored and recorded
at regular intervals during each experiment. Fig. 3 shows a
600
500
400
Current, mA
300
200
100
0
2.00E-04 3.00E-04 4.00E-04 5.00E-04 6.00E-04
Time, s
 590 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592
10.00
8.00
Resistance, Ω
6.00
4.00
2.00
0.00
-1 1 3 5 7 9 11 13 15
Time, s
Fig. 3 – Recording of resistance (blue dotted line) and
temperature (red dotted line) during a PEF experiment.
resistance and temperature time history during a PEF experi-
ment.
Consequently, it was necessary to record these parameters
(temperature and resistance) during each PEF experiment to
obtain a better insight for a proper evaluation of the results.
Although PEF was intended to be a non-thermal technique,
a temperature rise was present due to the electric current
through the material (ohmic heating). The average tempera-
ture rise (T) in the sample can roughly be estimated, Table 1,
collects the PEF treatment conditions for tea samples as well
as the treatment impact on the changes in voltage, current
and temperature.
When tea leaves were exposed to different intensities of
the pulsed electrical energy, the temperature was measured
and calculated to evaluate the degree of cell permeabiliza-
tion and the effect of the pulsed electric field (PEF) treatment.
On average, the temperature of untreated leaves before being
subjected to PEF treatment was 20 ◦ C. As shown in Table 1,
the temperature increased with the increase in applied pulsed
electrical energy at the same level of the applied electric field
strength. This increase in temperature is particularly signif-
icant at 0.6 and 1.1 kV/cm for the number of pulses ranging
from 30 to 50. The temperature of the sample with tea leaves
after PEF treatment increased when the applied pulsed electri-
cal energy was intensified. The highest treatment temperature
increase observed in this study was 7.1 ◦ C after PEF treatment,
using pulsed electrical energy of 29.7 kJ/kg at an electric field
strength level of 1.1 kV/cm.
3.2. Effect of electric field strength on extraction yield
of polyphenols
Sale and Hamilton (1967) identified the electric field strength
E and the total treatment time (which considers the number
of pulses and the pulse duration applied in the system) as the
main variables determining the efficiency of the PEF damage of
the plant tissue. Higher electric field strengths lead to a better
damage efficiency (Canatella et al., 2001; Toepfl et al., 2007).
However, it was noticed that an optimal value of the electric
field strength for many vegetables and fruit tissue is within
300–500 V/cm.
Fig. 4 presents experimental results for the extraction
yield of PPs from fresh tea leaves as a function of the
electric field strength for various pulse protocols: N = 30,
PD = 0.05 s and PBP(EXA) = 0.5 s (protocol I); N = 30, PD = 0.05 s
and PBP(EXB) = 3 s (protocol II). The only difference between
protocols I and II is the interval time between two pulse series
(see Fig. 1b).
21.90
21.70
Temperature, C
21.50
21.30
21.10
Fig. 4 – Experimental results for extraction yield of
polyphenols from fresh tea leaves vs electric field strength
at two pulse protocols: N = 30, PD = 0.05 s and PBP = 0.5 s
(protocol I) and N = 30, PD = 0.05 s and PBP = 3 s (protocol II).
Displayed error bars represent extraction yield
values ± standard experimental error. In all cases, a sample
was taken 20 min after the end of the PEF treatment and
stored at −30 ◦ C until the extracted amount of total PPs was
analyzed. All experiments were duplicated.
The extraction yield (EY) values of the samples subjected to
both protocols increase with increasing electric field strength.
The PEF treatment accelerates the rate of the extraction of
PPs from fresh tea leaves to the surrounding liquid. This is
in agreement with the behavior observed for different fruit
and vegetable tissues (Lebovka, 2009). Protocol I resulted in
a maximum value for the extraction yield of 27% when the
electric field strength is 0.9 kV/cm. However, when the interval
between pulses is equal to 3 s, an electric field of 1.1 kV/cm is
needed to obtain the same extraction yield (protocol II).
For both protocols, the same extraction yield was obtained,
but at different electric field strengths. When the interval
between two pulse series (pause between pulses) is short (pro-
tocol I), a moderate electric field seems to be sufficient to cause
cell membrane rupture. However, protocol II allows for a longer
relaxation time and a higher electric field is needed to achieve
an extraction yield of 27%. These results point to the rate of
extraction that is significantly dependent on the time interval
between two pulse series. Also, a longer relation time asks for
higher electric fields to achieve the same extraction yield.
The variation in the electric field strength to obtain the
same extraction yield between protocols I and II can also be
related to the ionic transport between the electrodes and elec-
trolysis and the tissue surface. Depending on the details of
contact (geometry and size of the samples, orientation of the
leaf slices, etc.) as well as the composition of the samples
(bud leaf, 1st, 2nd and 3rd open leaf), electrolysis may give rise
to different amounts of stable ionic compounds which would
result in an increase in conductivity and tissue disintegration.
3.3. Effect of the total treatment time on the extraction
yield of polyphenols
The degree of disintegration strongly depends on the treat-
ment time and the electric field strength (Lebovka, 2009). For
extensive times of PEF treatment, a smaller electric field is
required. Fig. 5 presents experimental results for the extrac-
tion yield at different treatment times for two electric field
strengths 0.4 and 0.9 kV/cm, respectively.
 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592
Table 1 – Summary of PEF treatment conditions for each PEF experiment and the treatment impact on the changes in
electrical parameters and temperature.
Pulse duration, (10−3 , s) Number of pulses (–)
Low electric field strength = 0.1 kV/cm
0.1 30
50 10
50 30
100 30
Moderate electric field strength = 0.6 kV/cm
0.1 10
0.1 50
50 30
100 10
100 50
High electric field strength = 1.1 kV/cm
0.1 30
50 10
50 50
100 30
Fig. 5 – Experimental results for extraction yield of
polyphenols from fresh tea leaves vs different treatment
times at different values of electric field strength 0.4 (blue
dotted bars) and 0.9 dark dotted bars) kV/cm, respectively,
for the fixed value of PBP = 0.5 s. Displayed error bars
represent extraction yield values ± standard experimental
error (95% confidence interval). In all cases, sample was
taken 20 min after the end of PEF treatment and stored at
−30 ◦ C until extracted amount of total PPs was analyzed.
All experiments were duplicated.
Lebovka et al. (2007) reported the effect of pulse duration
on the efficiency of PEF treatment on sugar beet. Experiments
showed that a longer pulse duration was more effective. This
influence of pulse duration was particularly pronounced at
moderate electric field strength (E = 0.3 kV/cm). This is par-
tially in agreement with the observation because the highest
extraction yield of 27% is obtained for moderate electric
field strength (E = 0.4 kV/cm) and a treatment time of 2.5 s
(Fig. 5). However, almost the same extraction yield (EY = 26.6%)
is obtained for a higher electric field (E = 0.9 kV/cm) and a
shorter treatment time of 1.5 s. When unfortunately the total
treatment time was 5 s (for both electric field strengths), the
experiment could not be performed due to problems with PEF
unit itself (low conductivity was detected in the PEF chamber).
A possible explanation lies in Ohm’s law. Electrical resistance
is the ratio of voltage over current by Ohm’s law and conduc-
tivity is inversely proportional to resistance. Therefore, the
resistance of the treatment chamber is an important param-
eter since the maximum allowed pulse current by the power
switch is 600 mA. This means that at 4 kV, the minimum resis-
tance is 6 . If the resistance is lower than this, the maximum
pulse current of 600 mA will be exceeded and if this situation
591
Specific energy input (kJ/kg) Change in temperature (◦ C)
0.9 0.1
1.4 0.3
4.0 0.9
4.0 1.0
1.7 0.4
7.8 1.9
7.7 1.8
2.6 0.6
12.5 3.0
12.0 2.9
5.9 1.4
29.7 7.1
17.5 4.2
continues for more than five pulses, the system will automat-
ically shut down to avoid damage to the high-voltage switch.
In this particular case, treatment time was 5 s, the total num-
ber of pulses, N = 50 and the pulse duration, PD = 0.1 s. For the
applied electric field strengths of 0.4 and 0.9 kV/cm, respec-
tively, the critical situation mentioned above exceeds and PEF
equipment will shut down.
Existing work mainly discusses the effects of pulse
duration in the PEF-induced inactivation of different microor-
ganisms. Some authors have demonstrated that inactivation
was more efficient at longer pulse duration (Aguiló-Aguayo
et al., 2010). However, others reported little effect of the pulse
duration on inactivation (Monfort et al., 2010). The effect of
pulse duration seems to vary depending on the electric field
strength and a general relationship between PEF-treatment
protocols, type and quality of the soft tissue, contact parame-
ters (geometry and size of the samples and orientation of the
leaf slices) and the resulting degree of material disintegration
requires more thorough study. A quantitative description of
the performance of PEF for extraction of key components from
cellular material (e.g., PPs from fresh tea leaves and proteins
from sugar beet leaves) from first principle of chemistry and
physics cannot be given at the moment.
4. Conclusions
This study provides valid evidence that pulsed electric field
(PEF) processing is a non-thermal method for the extraction of
polyphenols from fresh tea leaves. The electric field strength
as well as treatment time play an important role in polyphe-
nols extraction. Moreover, the efficiency of the PEF treatment
was indirectly connected to electric field effect, i.e., the relax-
ation time after a series of pulses. Different modes of PEF
treatment (electric field strengths and total treatment times)
were applied to investigate the effect on the extraction of
polyphenols from fresh tea leaves. The amount of extracted
polyphenols from the leaves into the aqueous media strongly
depends on the setting of PEF treatment. Protocol I (EXA)
resulted in a maximum extraction yield of 27% when the elec-
tric field strength is 0.9 kV/cm. However, when the interval
between pulses is longer or equal to 3 s, an electric field of
1.1 kV/cm is needed to obtain the same extraction yield (pro-
tocol II-EXB).
592 chemical engineering research and design 1 0 9 ( 2 0 1 6 ) 586–592
The total treatment time is equal to the product of the num-
ber of pulses and pulse width (duration). Experimental results
showed that longer pulses which means a longer treatment
time of 2.5 s were more effective for moderate electric field
(E = 0.4 kV/cm). Moreover, to achieve the same extraction yield
of 27% but with a shorter total treatment time of 1.5 s, a higher
electric field (E = 0.9 kV/cm) is required. Also a total treatment
time of 5 s (for both electric field strengths), the experiment
was unfortunately not possible due to limitation of the PEF
unit.
ACKNOWLEDGEMENTS
The work was financially supported by the Institute for
Sustainable Process Technology under the project FO-10-06
Selective Opening and Fractionation. The authors would like
to acknowledge Biobased Research Wageningen University,
especially to Dr. Hennie Mastwijk and ir. Ariette Matser for
valuable input and feedback.
References
Aguiló-Aguayo, I., Soliva-Fortuny, R., Martín-Belloso, O., 2010.
High-intensity pulsed electric fields processing parameters
affecting polyphenoloxidase activity of strawberry juice. J.
Food Sci. 75 (7), 641–646.
Asavasanti, S., Ristenpart, W., Stroeve, P., Barrett, D.M., 2011.
Permeabilization of plant tissues by monopolar pulsed
electric fields: effect of frequency. J. Food Sci. 76 (1), 98–111.
Bazhal, M.I., Ngadi, M.O., Raghavan, V.G.S., 2003. Influence of
pulsed electroplasmolysis on the porous structure of apple
tissue. Biosyst. Eng. 86 (1), 51–57.
Bouzrara, H., Vorobiev, E., 2003. Solid–liquid expression of cellular
materials enhanced by pulsed electric field. Chem. Eng.
Process. 42 (4), 249–257.
Canatella, P.J., Karr, J.F., Petros, J.A., Prausnitz, M.R., 2001.
Quantitative study of electroporation-mediated molecular
uptake and cell viability. Biophys. J. 80 (2), 755–764.
Dong, J.J., Ye, J.H., Lu, J.L., Zheng, X.Q., Liang, Y.R., 2011. Isolation
of antioxidant catechins from green tea and its
decaffeination. Food Bioproducts Process. 89 (1), 62–66.
Evrendilek, G.A., Altuntas, J., Sangun, M.K., Zhang, H.Q., 2013.
Apricot nectar processing by pulsed electric fields. Int. J. Food
Properties 16 (1), 216–227.
Fui-Seung, C., Khim-Phin, C., Atong, M., Nyet Kui, W., 2013. Tea
polyphenols and alkaloids content using Soxhlet and direct
extraction method. World J. Agric. Sci. 9 (3), 266–270.
Graham, H.N., 1992. Green tea composition, consumption, and
polyphenol chemistry. Prev. Med. J. 21 (3), 334–350.
Haberl, S., Miklavcic, D., Sersa, G., Frey, W., Rubinsky, B., 2013. Cell
membrane electroporation-Part 2: the applications. IEEE
Electr. Insul. Mag. 29 (1), 29–37.
Koiwai, H., Masuzawa, N., 2006. Extraction of catechins using
ultrasound. Proc. Symp. Ultrason. Electron. 27, 483–484.
Lebovka, N., 2009. Electrotechnologies for Extraction from Food
Plants and Biomaterials. Springer, New York.
Lebovka, N.I., Bazhal, M.I., Vorobiev, E., 2000. Simulation and
experimental investigation of food material breakage using
pulsed electric field treatment. J. Food Eng. 44 (4), 213–223.
Lebovka, N.I., Shynkaryk, M.V., El-Belghiti, K., Benjelloun, H.,
Vorobiev, E., 2007. Plasmolysis of sugarbeet: pulsed electric
fields and thermal treatment. J. Food Eng. 80 (2), 639–644.
Meterc, D., Petermann, M., Weidner, E., 2007. Extraction of green
tea and drying with a high pressure spray process. Hemijska
Ind. J. 61 (5), 222–228.
Monfort, S., Gayán, E., Saldaña, G., Puértolas, E., Condón, S., Raso,
J., Álvarez, I., 2010. Inactivation of Salmonella typhimurium and
Staphylococcus aureus by pulsed electric fields in liquid whole
egg. Innov. Food Sci. Emerg. Technol. 11 (2), 306–313.
Nkhilli, E., Tomao, V., El Hajji, H., El Boustani, S., Chemat, F.,
Dangles, O., 2009. Microwave-assisted water extraction of
green tea polyphenols. Phytochem. Anal. 20, 408–415.
Pan, X., Niu, G., Liu, H., 2003. Microwave-assisted extraction of tea
polyphenols and tea caffeine from green tea leaves. Chem.
Eng. Process Intensification 42 (2), 129–133.
Parsija, D., Anandharamakrishnan, C., 2015. Techniques for
extraction of green tea polyphenols: a review. Food Bioprocess
Technol. 8, 935–950.
Sale, A., Hamilton, W., 1967. Effects of high electric fields on
microorganisms. I. Killing of bacteria and yeasts. Biochim.
Biophys. Acta 148 (3), 781–788.
Soliva-Fortuny, R., Balasa, A., Knorr, D., Martín-Belloso, O., 2009.
Effects of pulsed electric fields on bioactive compounds in
foods: a review. Trends Food Sci. Technol. 20 (11–12), 544–556.
Tang, W.Q., Li, D.C., Lv, Y.X., Jiang, J.G., 2011. Concentration and
drying of tea polyphenols extracted from green tea using
molecular distillation and spray drying. Dry. Technol. Int. J. 29
(5), 584–590.
Toepfl, S., Heinz, V., Knorr, D., 2007. High intensity pulsed electric
fields applied for food preservation. Chem. Eng. Process. 46
(6), 537–546.
Walkling-Ribeiro, M., Rodríguez-González, O., Jayaram, S.,
Griffiths, M.W., 2011. Microbial inactivation and shelf life
comparison of cold hurdle processing with pulsed electric
fields and microfiltration, and conventional thermal
pasteurisation in skim milk. Int. J. Food Microbiol. 144 (3),
379–386.
Wan, J., Coventry, J., Swiergon, P., Sanguansri, P., Versteeg, C.,
2009. Advances in innovative processing technologies for
microbial inactivation and enhancement of food
safety—pulsed electric field and low-temperature plasma.
Trends Food Sci. Technol. 20 (9), 414–424.
Weaver, J.C., 2000. Electroporation of cells and tissues. IEEE Trans.
Plasma Sci. 28 (1), 24–33.
Yarmush, M.L., Golberg, A., Serša, G., Kotnik, T., Miklavčič, D.,
2014. Electroporation-based technologies for medicine:
principles, applications, and challenges. Annu. Rev. Biomed.
Eng. 16, 295–320.