U.S. patent application number 13/824734 was filed with the patent office on 2015-11-19 for method for extracting molecules of interest from grape pomace.
This patent application is currently assigned to UNIVERSITE TECHNOLOGIE DE COMPIEGNE-UTC. The applicant listed for this patent is Nadia Bousseta, Jean-Louis Lanoiselle, Michel Logeat, Sebastien Manteau, Eugene Vorobiev. Invention is credited to Nadia Bousseta, Jean-Louis Lanoiselle, Michel Logeat, Sebastien Manteau, Eugene Vorobiev.
Application Number | 20150327582 13/824734 |
Document ID | / |
Family ID | 44072536 |
Filed Date | 2015-11-19 |
United States Patent
Application |
20150327582 |
Kind Code |
A2 |
Lanoiselle; Jean-Louis ; et
al. |
November 19, 2015 |
METHOD FOR EXTRACTING MOLECULES OF INTEREST FROM GRAPE POMACE
Abstract
The invention relates to a method for extracting molecules of
interest from a plant matrix, the method including the following
steps: electrically processing the plant matrix by means of pulsed
power; diffusing the molecules of interest from the processed plant
matrix in a hydroalcoholic solvent and/or a solvent including ethyl
acetate; and recovering the molecules of interest that were
diffused.
Inventors: |
Lanoiselle; Jean-Louis;
(Salency, FR) ; Vorobiev; Eugene; (Compiegne,
FR) ; Bousseta; Nadia; (Compiegne, FR) ;
Manteau; Sebastien; (Boursault, FR) ; Logeat;
Michel; (Varennes Vauzelles, FR) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Lanoiselle; Jean-Louis
Vorobiev; Eugene
Bousseta; Nadia
Manteau; Sebastien
Logeat; Michel |
Salency
Compiegne
Compiegne
Boursault
Varennes Vauzelles |
|
FR
FR
FR
FR
FR |
|
|
Assignee: |
UNIVERSITE TECHNOLOGIE DE
COMPIEGNE-UTC
Compiegne
FR
SOCIETE FRANCAISE DE LABORATOIRES D'OENOLOGIE SOFRALAB
Magenta
FR
|
Prior
Publication: |
|
Document Identifier |
Publication Date |
|
US 20130323372 A1 |
December 5, 2013 |
|
|
Family ID: |
44072536 |
Appl. No.: |
13/824734 |
Filed: |
November 21, 2011 |
PCT Filed: |
November 21, 2011 |
PCT NO: |
PCT/EP2011/070597 PCKC 00 |
371 Date: |
August 14, 2013 |
Current U.S.
Class: |
426/239; 426/655;
568/717 |
Current CPC
Class: |
A23N 1/006 20130101;
A23L 33/105 20160801; A23L 5/30 20160801; C02F 1/4608 20130101;
A23L 27/11 20160801; B01D 11/0211 20130101 |
International
Class: |
A23L 1/221 20060101
A23L001/221; A23L 1/025 20060101 A23L001/025 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 19, 2010 |
FR |
1059547 |
Claims
1. A method for extracting molecules of interest from a plant
matrix made up of all or part of grape pomace, the method
comprising the following steps: electrical treatment of the plant
matrix by pulsed power; diffusion of the molecules of interest of
the treated plant matrix in a hydroalcoholic solvent or a solvent
composed of ethyl acetate; and recovery of the molecules of
interest having diffused.
2. The extraction method according to claim 1, wherein the
molecules of interest are polyphenols.
3. The method according to one of claims 1 to 2, wherein the
solvent is hydroalcoholic and includes between 25% and 50% of
alcohol.
4. The method according to claim 3, wherein the alcohol is
ethanol.
5. The method according to one of claims 1 and 2, wherein the
solvent includes a mixture of alcohol and ethyl acetate.
6. The method according to claim 5, wherein the mixture includes
between 5% and 30% of ethyl acetate.
7. The method according to claim 5, wherein the solvent further
includes water.
8. The method according to one of claims 1 to 7, wherein the
diffusion temperature is between 40.degree. C. and 70.degree.
C.
9. The method according to one of claims 1 to 8, wherein the
duration of the diffusion step is of at least 10 minutes.
10. The method according to one of claims 1 to 9, wherein the
electrical treatment and diffusion steps are carried out with a
continuous flow of solvent in a treatment tube.
11. The method according to claim 10, wherein the electrical
treatment is applied via coaxial electrodes, the distance between
the electrodes being about the radius of the treatment tube.
12. The method according to one of claims 1 to 10, wherein the
electrical treatment is applied via electrodes spaced apart by
about 5 mm.
13. The method according to one of claims 1 to 12, wherein the
electrical treatment is carried out by the application of
high-voltage electrical discharges.
14. The method according to claim 13, wherein the total specific
energy of treatment of the high-voltage discharges is between 70
and 90 kJkg.sup.-1, preferably 80 kJkg.sup.-1.
15. The method according to one of claims 13 or 14, wherein the
solvent-grape pomace ratio used during the diffusion step is
between 4 and 6, preferably 5.
16. The method according to one of claims 13 to 15, wherein the
diffusion temperature is 60.degree. C.
17. The method according to one of claims 1 to 12, wherein the
electrical treatment is carried out by the application of pulsed
electric fields.
18. The method according to claim 17, wherein the intensity of the
pulsed electric field is between 15 and 25 kVcm.sup.-1, preferably
20 kVcm.sup.-1.
19. The method according to one of claims 17 and 18, wherein the
diffusion temperature is 50.degree. C.
20. The method according to one of claims 1 to 19, further
including a step of purification and/or a step of pulverization of
the molecules obtained following the recovery step.
21. The method according to one of claims 1 to 20, wherein the
plant matrix includes at least one element chosen from the group
consisting of: grape pomace and constituents thereof, lees, must
deposits, tea, cocoa beans, berries, and oilseeds.
22. The polyphenols likely to be obtained by the implementation of
the method described in claims 2 to 21.
23. Use of the polyphenols of claim 22 to improve the gustatory
properties of wine.
Description
TECHNICAL FIELD
[0001] The invention relates to the field of the extraction of
molecules of interest from a plant matrix or the like, and more
particularly from a by-product of wine making such as grape pomace,
lees or must deposits.
[0002] Grape pomace results from the pressing of grapes, and
includes notably seeds, skins, and stems.
[0003] Lees are obtained after fermentation of grape juice with
yeasts. It is the deposit formed after fermentation containing
transformed yeasts or yeast fragments.
[0004] Must deposits are all of the particles that sediment after
pressing.
[0005] The invention is thus applicable to the extraction of
molecules of interest from seeds, skins and stems, taken
individually or in combination, lees or must deposits.
[0006] The invention also relates to the field of the extraction of
molecules of interest from tea (notably green tea), cocoa beans,
berries (notably red berries), oilseeds such as flax, apples,
etc.
PRIOR ART
[0007] In the following, we will describe in more detail the
invention as it applies to the extraction of polyphenols from a
plant matrix, notably grape pomace. This is, however, in no way
restrictive, insofar as these molecules can also be extracted from
seeds, skins, stems, lees and must deposits, taken individually or
in combination.
[0008] The structure of polyphenols, also called phenolic
compounds, includes a benzene nucleus with one or more hydroxyl
groups, free or engaged with a substituent such as an alkyl, ester
or sugar. The molecular weight of polyphenols can vary from less
than 100 g/mol to more than 30,000 g/mol.
[0009] The polyphenols present in grape pomace belong to various
groups: simple phenolic derivatives, flavonoids (flavonols and
anthocyanins) and condensed, more complex phenolic structures.
[0010] A conventional method for recovering polyphenols from grape
pomace is based on solid/liquid extraction by solvent followed by
purification and drying. "Solid/liquid extraction" refers to the
selective dissolution of one or more solutes of a solid matrix in
liquid solvent. It is one of the oldest unit operations. This
operation consists in contacting the liquid solvent with the solid
matrix. In the case of the extraction of polyphenols, this contact
is carried out by total immersion of the solid matrix in the liquid
solvent or by spraying/washing the solid matrix with solvent.
[0011] According to the solvent used, the membranes of the cells of
the seeds, skins or stems are more or less weakened, which
facilitates the release of polyphenols from the cells.
[0012] Much work has been done concerning the influence of the
operational parameters of solid/liquid extraction of polyphenols.
The principal parameters are solvent type, temperature, contact
time, liquid/solid ratio, particle size and pH.
[0013] Regarding solvents, methanol, ethanol, ethyl acetate and
water are used most often in extractions from skins and from grape
pomace in general. Among these, methanol seems to be the solvent
that provides the best extraction rate, followed by ethanol and
then water (Pinelo et al., 2005). Indeed, polyphenols are
solubilized more easily in methanol than in the other two solvents.
However, ethanol and water are preferable when extracting
polyphenols for an application in foods.
[0014] With regard to the liquid/solid ratio, it seems that the
higher the liquid/solid ratio, the better the extraction of
polyphenols. However, from an economic point of view, this variable
must be optimized in order to reduce the method's costs, notably by
reducing the consumption of liquid solvent.
[0015] At the conclusion of the solid/liquid extraction, the
obtained extracts contain a large number of other compounds such as
sugars, proteins, amino acids, mineral salts, etc. As a result,
purification is carried out in order to remove them. Several
purification techniques exist. The principal techniques used are
adsorption-desorption and filtration.
[0016] Following this purification, the purified extracts are dried
in order to obtain a polyphenol powder. In powder form, the product
is stabilized.
[0017] Numerous intensification techniques have also been
developed, such as ultrasound and pulsed power. These
intensification techniques act on the membranes and/or walls of
cells constituting the solid plant matrix in order to facilitate
the extraction of biocompounds such as polyphenols.
[0018] Two main pulsed-power intensification techniques exist. They
use electrical pulses to concentrate, in very short time periods,
electrical energy stored in a condenser. This electrical energy is
then injected into a treatment chamber containing the solid plant
matrix, for example grape pomace. This sudden injection of
electrical energy into the chamber makes it possible to destabilize
the physical, biological and/or chemical properties of the cells of
the solid plant matrix, which can have highly advantageous
characteristics for the extraction of biocompounds such as
polyphenols.
[0019] A first pulsed-power technique uses a pulsed electric field
(PEF technique, or simply PEF hereinafter). The pulsed electric
field induces pores on the membrane of a plant or animal cell: this
is the phenomenon of electroporation.
[0020] The pulsed electric field can also act on the intracellular
contents of the cell (e.g., to detach the cell membrane from the
cell wall as well as to disrupt its intracellular contents).
[0021] The creation of a pulsed electric field requires a
high-voltage generator and a treatment chamber including at least
two electrodes, one being connected to the generator and the other
being connected to ground.
[0022] The plant matrix is placed in the treatment chamber. The
high-voltage generator then transforms the alternating electric
current into pulsating direct current. The energy of each pulse is
temporarily stored in one or more condensers and then discharged by
the electrodes in the treatment chamber.
[0023] The ability to extract compounds of interest from plant
cells with PEF depends on several operational parameters, which
fall in two categories: parameters related to the method (amplitude
of the electric field applied, duration and number of pulses,
temperature) and parameters related to the plant matrix
(conductivity).
[0024] The second pulsed-power technique uses high-voltage
electrical discharges (HVED technique, or simply HVED hereinafter).
This technique was first intended for military and scientific
applications requiring very high energies.
[0025] The creation of high-voltage electrical discharges requires
a treatment chamber and an electric generator designed for high
currents (thyristors, IGBT, GTO, etc.) or high voltages (line and
Tesla transformers, Marx generators, etc.).
[0026] The treatment chamber includes electrodes, whose most
commonly used combinations are tip-planar and tip-tip. The
electrodes are entirely or partially submerged in water.
[0027] The plant matrix is placed in the treatment chamber and
submerged. The electric generator stores electrical energy in a set
of storage condensers or inductors. The presence of high voltage at
the terminals of the electrodes causes a phenomenon of electrical
breakdown and the creation of an electric discharge between the two
electrodes. When an electric discharge is applied in water, as it
is here, it produces shock waves which come into contact with the
plant matrix. The latter then fragments according to the number of
pulses injected, thus releasing biocompounds, including
polyphenols.
[0028] This state of the art made it possible to highlight certain
shortcomings of traditional solid/liquid extraction methods and led
toward research into intensification of the extraction. First, the
traditional solid/liquid extraction method is relatively long
(between 3 hours and 20 hours). A high temperature (above
50.degree. C.) is also required, resulting in a sizeable energy
cost.
[0029] Moreover, the addition of organic solvents or sulfites has
an environmental cost and limits applications of the final product.
Finally, in a general manner, for the extraction of alcohol,
tartaric acid or possibly polyphenols, there is no method the
parameters of which are rationalized and optimized.
[0030] Consequently, it appears necessary to improve the extraction
method by an intensification method.
[0031] Presentation
[0032] The invention thus aims at overcoming these disadvantages of
the prior art, in order to increase the effectiveness of the
extraction of molecules of grape pomace or compounds thereof, while
providing a treatment of lower energy costs and while reducing the
addition of chemicals.
[0033] In a particular application, the invention aims at providing
a method for extracting polyphenols having a better extraction
yield than conventional methods, while maintaining, even improving,
the oxidizing activity of the extracted polyphenols thanks to the
method.
[0034] To that end, the invention provides a method for extracting
molecules of interest from a plant matrix made up of all or part of
grape pomace, the method comprising the following steps: [0035]
electrical treatment of the plant matrix by pulsed power; [0036]
diffusion of the molecules of interest of the treated plant matrix
in a hydroalcoholic solvent or a solvent composed of ethyl acetate;
and [0037] recovery of the molecules of interest having
diffused.
[0038] Certain preferred but nonrestrictive aspects of the method
are as follows: [0039] the molecules of interest are polyphenols;
[0040] the solvent is hydroalcoholic and includes between 25% and
50% alcohol; [0041] the alcohol is ethanol; [0042] the solvent
includes a mixture of alcohol and ethyl acetate; [0043] the mixture
includes between 5% and 30% ethyl acetate; [0044] the solvent
further includes water; [0045] the diffusion temperature is between
40.degree. C. and 70.degree. C.; [0046] the duration of the
diffusion step is at least 10 minutes; [0047] the electrical
treatment and diffusion steps are carried out with a continuous
flow of solvent in a treatment tube; [0048] the electrical
treatment is applied via coaxial electrodes, the distance between
the electrodes being about the radius of the treatment tube; [0049]
the electrical treatment is applied via electrodes spaced apart by
about 5 mm; [0050] the electrical treatment is carried out by the
application of high-voltage electrical discharges; [0051] the total
specific energy of the treatment of the high-voltage discharges is
between 70 and 90 kJkg.sup.-1, preferably 80 kJkg.sup.-1; [0052]
the solvent-grape pomace ratio used during the diffusion step is
between 4 and 6, preferably 5; [0053] the diffusion temperature is
60.degree. C.; [0054] the electrical treatment is carried out by
the application of pulsed electric fields; [0055] the intensity of
the pulsed electric field is between 15 and 25 kVcm.sup.-1,
preferably 20 kVcm.sup.-1; [0056] the diffusion temperature is
50.degree. C.; [0057] it further includes a step of purification
and/or a step of pulverization of the molecules obtained following
the recovery step; and [0058] the plant matrix includes at least
one element chosen from the group consisting of: grape pomace and
constituents thereof, lees, must deposits, tea, cocoa beans,
berries, and oilseeds.
[0059] According to a second aspect, the invention provides
polyphenols likely to be obtained by the implementation of a method
in accordance with the invention.
[0060] According to a final aspect, the invention provides the use
of these polyphenols to improve the gustatory properties of
wine.
DETAILED DESCRIPTION
[0061] We will now describe a method for extracting molecules in
accordance with the invention from grape pomace, illustrated by the
extraction of polyphenols.
[0062] This method can be implemented on each solid element
constituting pomace, namely seeds, skins and stems, on lees or on
must deposits, or a combination thereof. Grape pomace, seeds,
skins, stems, lees, must deposits and combinations thereof are
referred to as "plant matrix" hereinafter.
[0063] The raw material used for the extraction can result directly
from wine making, or can have been preserved beforehand for a
predetermined period (generally up to a year or more, as needed) by
deep freezing or by adding an antioxidant (such as sulfur dioxide).
Indeed, it should be noted that grape pomace is produced only once
a year during the harvest, after pressing, and it breaks down
rapidly. Its use throughout the year thus requires the ability to
store it between two grape harvests.
[0064] Grape pomace can thus notably be preserved in hermetic and
opaque plastic bags, to protect it from photo-oxidation, either at
about 4.degree. C., preferably in the presence of 0.01% sulfur
dioxide, or at -31.degree. C., after deep freezing.
[0065] The polyphenols are then extracted in three principal
steps:
[0066] (1) electrical treatment of the raw material for the purpose
of damaging cell membranes or walls in order to facilitate
extraction;
[0067] (2) solid-liquid diffusion during which the molecules
migrate from the grape pomace toward a hydroalcoholic solvent;
and
[0068] (3) recovery of the diffused molecules.
[0069] The purpose of the electrical treatment is to intensify the
extraction of polyphenols during the subsequent step of
solid-liquid diffusion of polyphenols from the grape pomace into a
solvent. It can notably be selected from treatment with
high-voltage electrical discharges (HVED) or treatment with pulsed
electric fields (PEF).
[0070] The order of steps (1) and (2) is not restrictive. Indeed,
the solid-liquid diffusion can begin before the electrical
treatment.
[0071] As a variant, the electrical treatment can begin during the
diffusion, after a first extraction in a hydroalcoholic solvent or
water. Indeed, the objective of this first extraction is to modify
the electrical conductivity of the solvent by enrichment in ionic
compounds stemming from pomace in order to improve the application
of the electric field.
[0072] The plant matrix can further undergo a pressing intended to
remove part of its water of constitution in order to promote the
application of the electrical treatment and to limit the volumes of
materials and solvents used.
[0073] Polyphenols include simple phenolic derivatives and
flavonoids.
[0074] Simple phenolic derivatives are derivatives of
hydroxybenzoic acid (gallic acid), hydroxycinnamic acid (caffeic
acid, coumaric acid, ferulic acid, stilbenes) or lignins.
[0075] Hydroxycinnamic acid derivatives include caffeic acid,
coumaric acid, ferulic acid and stilbenes (trans-resveratrol,
cis-resveratrol, glucosides of trans- or cis-resveratrol,
trans-piceids, cis-piceids, etc.).
[0076] Flavonoids include anthocyanins, flavanols and
flavanols.
[0077] The principal anthocyanins are glucosylated derivatives of
cyanidin, peonidin, petunidin, delphinidin and malvidin.
[0078] Flavanols include, among others, catechin, epicatechin
gallate, epicatechin, procyanidins, in particular procyanidins
B.sub.1 and B.sub.2, and polymers thereof: poly(catechin),
poly(epicatechin), poly(gallocatechin), poly(epigallocatechin), or
heteropolymers.
[0079] Polyphenols can be used in wine making methods to improve
the gustatory properties of wine.
[0080] Flavanols include, among others, kaempferol and glycosides
thereof, quercetin and glycosides thereof, and isorhamnetin
glycosides.
[0081] In the present application, the term "polyphenol" refers
either to a single particular compound cited above, or to a mixture
of at least two compounds cited above.
[0082] Treatment with HVED
[0083] HVED treatments intensify the extraction of polyphenols by
mechanically breaking down the structure of the raw material of
pomace. More precisely, treatment with HVED uses pressure waves,
cavitation processes and turbulence phenomena, all of which cause
the material to fragment, thus promoting the transfer of compounds
from the cell's interior to its exterior.
[0084] "High voltage" refers to voltages sufficient to produce
electrical discharges, advantageously greater than 20 kV, for
example 50 kV, for tip-planar electrodes spaced apart by 5 mm.
[0085] Treatment with HVED can also be carried out on a laboratory
scale (in a 1 liter treatment enclosure, for example), a semi-pilot
scale (in a 35 liter treatment enclosure, for example), or an
industrial scale using a continuous treatment cell within which the
product to be treated circulates, such as, for example, a tube 1
meter in length and a few centimeters in diameter capable of
treating several tons of plant matrix per hour, indeed up to 40
tons per hour (electrodes thus being placed along the path of
circulation).
[0086] An example of a device for applying HVED includes notably a
high-voltage generator connected to a treatment chamber.
[0087] The treatment chamber includes two electrodes between which
grape pomace diluted in a solvent such as water or a hydroalcoholic
mixture is introduced. The electrodes are made of stainless steel
or aluminum, and include a tip electrode (typically 10 mm in
diameter) connected to the generator and a planar electrode
(typically 120 mm in diameter) connected to the mass. The distance
between the electrodes is between 2 and 10 mm, and is preferably
about 5 mm. Indeed, at this distance, the latency before electric
breakdown is reduced, thus limiting energy losses.
[0088] It will be noted, however, that the optimal distance between
electrodes varies with the shape of the electrodes, the voltage
applied to the electrodes and the dimensions of the treatment
chamber.
[0089] The high-voltage generator includes a condenser designed to
store electrical energy and then to discharge it in the treatment
cell via a spark gap in order to produce electric breakdown in the
water and to generate an electric discharge.
[0090] The electrical treatment thus consists in applying a given
number n of pulses (i.e., electrical discharges) to the solid and
liquid mixture consisting of pomace and water.
[0091] The parameters acting on the effectiveness of the treatment
are notably: [0092] treatment temperature, selected between
40.degree. C. and 70.degree. C.; [0093] duration t of the treatment
(proportional to the number of pulses)
[0093] with t=nxt.sub.i
wherein: n is the number of pulses, and [0094] t.sub.i is the
duration of a pulse (s), [0095] solvent/grape pomace ratio, also
called liquid/solid ratio, preferentially selected to be 5; [0096]
distance between electrodes; [0097] voltage applied; [0098] energy
supplied; and [0099] pulse frequency.
[0100] These parameters indeed make it possible to optimize the
permeabilization of the cell structure of pomace.
[0101] On a laboratory scale, the generator can, for example,
supply a maximum voltage of 40 kV for a maximum current of 10 kA
and generate pulses of a duration of about 10 .mu.s at a frequency
of about 0.5 Hz. The average energy of an electrical pulse supplied
by the generator is thus 160 J per pulse.
[0102] On a semi-pilot scale, the generator can, for example,
supply a maximum power of 40 kV for a maximum current of 30 kA and
generate pulses of a duration of about 100 .mu.s at a frequency of
about 0.5 Hz. The characteristics of the electrical treatment (such
as the average energy of an electrical pulse) are, however, more
flexible than in the case of the laboratory generator. For example,
at low energy the discharge of a 200 nF condenser can supply an
average energy of 160 J per pulse, whereas at high energy the
discharge of a 5 .mu.F condenser can supply an average energy of
4,000 J per pulse.
[0103] The total specific energy of the treatment (in relation to
the weight of treated grape pomace) with HVED is between 70 and 90
kJ/kg, preferably 80 kJ/kg.
[0104] On an industrial scale, the parameters are identical except
for the distance between the electrodes, which can be greater. For
example, when the electrodes are assembled coaxially (wherein a
first electrode extends parallel to the axis of revolution of the
treatment tube (typically along this axis), while the second
electrode extends coaxially to the first electrode, so that the
flow of the product passes between the two electrodes) in the
treatment tube, the distance between electrodes being preferably
about equal to the radius of the tube, for example 1.27 cm. On the
other hand, when the electrodes are assembled collinearly (wherein
the cathode and the anode are substantially aligned and alternate
along the tube), the optimal distance for the batch configuration
can be retained, which is thus about 5 mm between the cathode and
the anode along the treatment tube.
[0105] Treatment with PEF
[0106] In the case of treatment with PEF, the extraction of
polyphenols is principally intensified by electroporation of the
cell membranes of the grape pomace. This treatment can be applied
to relatively small quantities of plant matrix (about 1 to 10 g of
plant matrix) or to greater quantities (about 100 to 500 g).
[0107] It is in particular possible to apply a treatment with PEF
at low intensity (on a laboratory scale, with 0.1-1.3 kV/cm), or at
high intensity (on a semi-pilot or industrial scale, with 0.5-20
kV/cm).
[0108] In the case of a low-intensity treatment, the experimental
device consists of a low-volume (a few cm.sup.3) PEF treatment cell
connected to a PEF generator.
[0109] The total PEF treatment time (t.sub.PEF, s) is defined by
the duration of the pulse (t.sub.i, s) and the total number of
pulses (N.sub.tot). The latter depends on the number of trains (N)
and the number of pulses per train (n).
[0110] All of the generator's treatment parameters can be
controlled by control software. For example, the generator can
supply a maximum power of 400 V for a maximum current of 40 A and
can generate pulses of a duration between about 10 and 10,000
.mu.s. The generator can thus supply between 1 and 1,000 pulse
trains, each comprising 1 to 10,000 pulses with an idle period
between each train of between 1 and 3,600 s.
[0111] Given that the intensity of the electric field (E, V/cm) is
defined by the ratio of the voltage applied (U, V) and the
distances between electrodes (d, cm),
E = U d ##EQU00001##
with this experimental device, for a distance between electrodes d
of 3 mm, it is thus possible to reach a maximum intensity of about
1.3 kV/cm.
[0112] Generally, the intensity selected is greater than 0.5
kV/cm.
[0113] When a high-intensity treatment is used, and according to
the nature of the plant matrix (pomace, seeds, etc.), the intensity
of the electric field required can be rather high, such as, for
example, for uncrushed grape seeds. At the most, for grape seeds,
the intensity of the field is typically selected between 15 and 25
kV/cm, and preferably 20 kV/cm.
[0114] It is possible to use the same generator and the same
treatment chamber as for the low-intensity treatment used to
generate HVED on a laboratory scale, using, for example, two
parallel, planar stainless steel electrodes in the place of tip and
planar electrodes. The distance between the electrodes can also
vary from 2 to 10 mm, i.e., a corresponding PEF intensity of 4 to
20 kV/cm.
[0115] The principal operational parameters that can act on the
effectiveness of the treatment are as follows: [0116] extraction
solvent (water alone, water/alcohol mixture, alcohol alone); [0117]
treatment temperature, selected between 40.degree. C. and
70.degree. C., preferably 50.degree. C.; [0118] treatment period
(proportional to the number of pulses), and [0119] electric field
intensity (defined by the distance between electrodes); [0120]
pulse frequency; [0121] pulse shape; and [0122] pulse polarity
(unipolar or bipolar).
[0123] Hydroalcoholic Diffusion
[0124] In order to optimize the extraction of molecules,
preferentially polyphenols, the treated mixture undergoes the
diffusion step in combination with electrical treatment with HVED
or PEF. The diffusion step can begin before the electrical
treatment and end after the electrical treatment, or can begin
immediately after the electrical treatment.
[0125] During the diffusion, it is preferable not to exceed a
temperature of 60.degree. C. in order to limit the thermal
degradation of the polyphenols.
[0126] Moreover, at a low temperature (20.degree. C.) the diffusion
can be rather long (up to 4 hours), whereas at a higher temperature
(between 40.degree. C. and 60.degree. C.) the diffusion is between
1 hour and 1.5 hours. Typically, increasing the diffusion
temperature from 20.degree. C. to 60.degree. C. makes it possible
to increase the extracted polyphenols content of the solvent by 33%
and antioxidant activity by 48% after 60 minutes of extraction.
[0127] On an industrial scale, the diffusion temperature can be
between 50.degree. C. and 60.degree. C., for example.
[0128] The diffusion is carried out in a hydroalcoholic solvent or
a solvent composed of ethyl acetate. If during the electrical
treatment (with HVED or PEF) the grape pomace is submerged in a
hydroalcoholic solvent or a solvent composed of ethyl acetate, this
solvent can be the same as used for the electrical treatment and
the diffusion.
[0129] The hydroalcoholic solvent is a mixture of water and alcohol
with an alcohol content varying from 25% to 50% by weight. The
water can be distilled water or tap water. The alcohol can be
methanol or ethanol. Nevertheless, ethanol is preferable for a
subsequent use of the extracted molecules in the field of
foods.
[0130] If ethyl acetate is used, it can be used in a mixture with
ethanol or methanol. The ethyl acetate/alcohol mixture includes
between 5% and 30% ethyl acetate. It is also possible to use a
ternary ethyl acetate/alcohol/water mixture, preferably a ternary
ethyl acetate/ethanol/water mixture, in volume proportions notably
between 4/1/4 and 10/1/10.
[0131] The alcohol or ethyl acetate is introduced into the water
before, during or after the electrical pretreatments, but before
the diffusion.
[0132] If the solvent is a hydroalcoholic solvent, the quantity of
extracted polyphenols increases with the alcohol content, since
ethanol, which is a polar solvent, promotes the extraction of
polyphenols due to their greater solubility in this solvent than in
water alone. In addition, it disrupts the external structure of
cell membranes, thus enabling the extraction of polyphenols located
within membranes or inside cells.
[0133] The quantity of solvent in relation to grape pomace is
adjusted by respecting a liquid/solid weight ratio between 1 and
20, preferably between 4 and 10 in the case of diffusion carried
out in a 1 hour batch. Indeed, the greater the ratio, the greater
the quantity of extracted polyphenols. However, a plateau is
reached from a liquid/solid ratio of about 5, the saturation of the
solvent (water) occurring at lower ratios.
[0134] For example, following one or the other of the electrical
pretreatments, diffusion in a solvent composed of 30% ethanol and
70% water makes it possible to obtain the best extraction yield,
increasing it by a factor of 3 in comparison with a solvent
composed only of water (2.8.+-.0.4 g gallic acid equivalents (GAE)
per 100 g of dry matter for treatment with HVED and 7.5.+-.0.4 g
GAE for treatment with PEF) and the best antioxidant activity of
the extracts (66.8.+-.3.1 g TEAC per kilogram of solid mass for
treatment with HVED).
[0135] According to a preferred embodiment, during the diffusion
step, the pomace pretreated electrically is placed in a solvent
composed of 30% ethanol and 70% water with a liquid/solid ratio of
about 5. After 1 hour of diffusion at 30.degree. C. or 50.degree.
C., the solvent then includes polyphenols in dissolved form or in
colloidal suspension.
[0136] The water used can be distilled water, purified water or
simply tap water.
[0137] It will be noted, however, that the polyphenols migrate
progressively from the pomace toward the solvent during the first
30 minutes of extraction, and then reach a plateau between 30 and
60 minutes during which the extraction kinetics slow
considerably.
[0138] The pH of the solution is advantageously acidic. A pH below
6 makes it possible to limit degradation of the polyphenols. A pH
of 4, which is the natural pH of grape pomace, makes it possible to
protect anthocyanins. If need be, the pH can be modified by the
addition of an acid, preferably a food acid.
[0139] After diffusion, the solvent containing the polyphenols in
dissolved form or in colloidal suspension is separated from the
grape pomace, for example by filtering.
[0140] The liquid polyphenol extracts are then separated from the
solvent and other undesired extracts by centrifugation and then
transformed into powder. This pulverization step makes it possible
on the one hand to increase the stability of the polyphenols and,
on the other hand, to provide a product in a marketable form.
[0141] Since the extracts can contain sugars and proteins,
according to the subsequent application selected for the
polyphenols (pharmaceutical, cosmetic and/or agri-food), it can
further be necessary to purify them before drying, for example by
solid-phase extraction. This purification technique is notably
founded on the distribution of compounds between a solid phase
(adsorbent) and a liquid phase (sample) in accordance with
conventional techniques.
[0142] Scaling
[0143] Below, we will detail the operational parameters of the
electrical treatment before being applied in order to intensify the
extraction of polyphenols by electrical treatments on a semi-pilot
scale. To that end, we will more particularly describe the case of
treatment with HVED. Nevertheless, this is in no way restrictive
and the person skilled in the art will be able to apply the
teachings that follow to the case of treatment with PEF.
[0144] Treatment with HVED on a laboratory scale can be carried out
in an enclosure containing 50 g of grape pomace and 250 g of water
(for a total mass of plant matrix of 300 g), i.e., according to a
liquid/solid ratio of 5. For the semi-pilot tests, by maintaining
the liquid/solid ratio of 5, a total mass of plant matrix of 7,500
g is introduced into a second enclosure.
[0145] For scaling, the three important operational parameters of
the electrical treatment are the electrical energy of a pulse
(kJ/pulse), the energy of the electrical pulse per mass of treated
plant matrix (kJ/kg/pulse) and the total specific energy of the
treatment (kJ/kg).
[0146] The electrical energy of a pulse is limited by the condenser
that composes the generator.
[0147] The energy of the electrical pulse per mass of treated plant
matrix takes account of the quantity of raw material. For the
laboratory tests of the preceding examples, the generator makes it
possible to deliver an electrical pulse of 0.16 kJ. In the case of
semi-pilot tests, it is possible to use, for example, two different
condensers in a pilot generator in order to supply an electrical
pulse of 0.16 kJ or 4 kJ. Once the pulse energy is set, it is then
possible to determine the effect of the treatment period applied to
the plant matrix by varying the number of pulses.
[0148] Finally, the total treatment energy takes into account both
the number of pulses and the total quantity of plant matrix.
[0149] Following treatment with HVED, a discontinuous aqueous
diffusion is carried out, for example with a 1 hour batch. On an
industrial scale, up to 14 batches can be treated with a continuous
flow of solvent in the treatment tube, which makes it possible to
approach a continuous aqueous diffusion. The extraction yields
obtained on a laboratory scale and a semi-pilot scale can thus be
compared in terms of polyphenols extraction rate and oxidizing
activity (whose measurement makes it possible to verify that the
polyphenols extracted after HVED remain functional, notably in
relation to their ability to trap oxidants).
[0150] When an electrical energy of a pulse of 0.16 kJ/pulse is
applied, no improvement in polyphenols extraction in relation to
simple diffusion is observed on the semi-pilot scale.
[0151] On the other hand, this is not the case when the energy per
mass of treated plant matrix between the two scales is preserved.
In the case of laboratory scale, the relationship between the
energy supplied and the total mass of plant matrix is 0.53
kJ/kg/pulse.
[0152] When a treatment of total specific energy of 53 kJ/kg is
applied (with 100 pulses of 0.53 kJ/kg/pulse, for example) on a
laboratory scale and a semi-pilot scale, the extracted polyphenols
content on a semi-pilot scale represents only 38% of that obtained
on a laboratory scale. Nevertheless, retaining the energy per mass
of treated plant matrix makes it possible to improve the extraction
of polyphenols compared to simple diffusion (control). The same
tendency is found concerning the antioxidant activity of the liquid
extracts of polyphenols.
[0153] In addition, it is also preferable to take account of the
geometry (dimensions and shape) of the treatment chambers when the
scale of the tests is modified (notably when moving from laboratory
scale to semi-pilot scale or industrial scale). Indeed, according
to the shape of the treatment chamber, the electrical discharges
are distributed differently within the treatment chamber on the
same treated plant matrix insofar as they produce high-pressure
shock waves (up to 1,000 MPa) which are responsible for turbulence
and agitation of the liquid within the chamber.
[0154] Typically, if one looks at the height/diameter ratios of
treatment chambers, dead zones within treatment chambers with a
smaller height/diameter ratio can be larger, whereas agitation due
to shock wave propagation can be reduced in treatment chambers with
larger height/diameter ratios. It is, in fact, the pressure field
within the treatment chamber, which results from the energy of the
discharge per unit volume and the distance between the electric
discharge and the walls of the chamber, which is a determining
factor. Pressure levels greater than 100 bars are required to
significantly increase the extraction of polyphenols.
[0155] Consequently, the energy required for HVED to have an effect
on extraction yields depends on the geometry of the treatment
chamber, and a minimal energy function of the configuration of the
treatment chamber is necessary. Below that minimum, electrical
discharges seem to have only little effect on the extraction of
polyphenols.
[0156] It is also preferable to vary the total treatment energy by
adjusting the number of pulses sent on the plant matrix (pomace,
seeds, lees, etc.). Indeed, the larger the total specific energy,
the better the polyphenols extraction yield. More precisely,
antioxidant activity and extraction yield increase linearly with
number of pulses discharged in the plant matrix. However, an
optimal extraction exists: the polyphenols yield increases and then
decreases beyond a certain treatment energy value. For laboratory
tests, the optimal total specific energy is 100-160 kJ/kg. For
semi-pilot tests, it is 400-550 kJ/kg. A greater treatment energy
is thus necessary on a semi-pilot scale in order to obtain results
equivalent to those obtained on a laboratory scale.
[0157] Typically, excellent results are obtained with 1,000 pulses,
a treatment of 533 kJ/kg total energy: the polyphenols
concentration is thus 7 times greater in relation to a control test
without electrical treatment, whereas the antioxidant activity of
the extracts is increased by a factor of 5, and the extraction
rates (.apprxeq.200 mg GAE/l) obtained on a semi-pilot scale with
160 kJ/kg are thus similar to those determined on a laboratory
scale with 53 kJ/kg by preserving specific energy (kJ/kg) and by
varying energy per pulse (J).
[0158] In addition, according to the type of plant matrix, the
energy necessary is more or less high. For example, the stems, the
branched ligneous part, seem to be the most resistant to discharges
because the maximum polyphenols contents are obtained with the
highest energy values (400 kJ/kg on a semi-pilot scale, 213 kJ/kg
on a laboratory scale). On the contrary, skins seem to be more
sensitive; an energy of 133 kJ/kg is sufficient on a semi-pilot
scale to extract about 400 mg GAE/l. A similar quantity of
polyphenols is obtained on a laboratory scale after a treatment of
53 kJ/kg.
[0159] Consequently, even if the tendencies are similar, extraction
on a semi-pilot scale requires a total treatment energy that is,
overall, greater than that applied on a laboratory scale. The
treatment conditions thus do not seem equivalent in the treatment
chambers. The pressure field generated by the electric discharge is
different in the two treatment systems. The physical
characterization of the electric discharge thus makes it possible
to study the role of the pressure field on cell breakdown and
consequently on the extraction of polyphenols.
[0160] Results
[0161] We will now compare the general performance of the
extraction method in accordance with the present invention with the
conventional methods which do not include a preliminary step of
intensification of the extraction and/or use a diffusion solvent
containing only water or only alcohol.
[0162] It is first reminded that pretreatments with HVED and PEF
act differently on the treated plant matrix, require different
operational parameters (generally according to the type of plant
matrix, namely pomace, seeds, etc.) and in the end produce
different yields. Adaptation of the parameters to the type of
treated plant matrix (pomace, seeds, etc.) is, however, left to the
abilities of the person skilled in the art and will not be
systematically detailed further in this description.
[0163] First, we will detail the performance in the case of the
implementation of a treatment with HVED. Nevertheless, comparable
performance was also obtained during the implementation of the
treatment with PEF.
[0164] Here, an electrical pretreatment with HVED during which 80
pulses of a total effective duration (i.e., the cumulative duration
during which the solution was subjected to a discharge, without
counting the pauses between two pulses or diffusion time) of 0.8 ms
followed by a discontinuous diffusion per batch (with a batch of a
duration of 1 hour) was applied to grape pomace.
[0165] After 1 hour of extraction at 20.degree. C., the yields in
total solutes are about 70% with electrical pretreatment with HVED,
compared with 22.+-.2% in the absence of HVED (but with diffusion
in a hydroalcoholic solvent). It should be noted in addition that
after only 1 hour of extraction with HVED, the polyphenols content
is 30% greater than that obtained after 4 hours of diffusion
without HVED.
[0166] The extraction of polyphenols is also improved by increasing
the temperature to 60.degree. C. The increase in temperature makes
it possible to increase the fluidity of the plasma membrane and to
promote the creation of pores. Thus, the performance of HVED
diffusion at 20.degree. C. is similar to that of simple diffusion
at 40.degree. C. without HVED. The same tendency is observed for
HVED diffusion at 40.degree. C. and simple diffusion at 60.degree.
C.
[0167] The effect of the electrical pretreatment, whether with HVED
or PEF, is thus to reduce the duration and temperature of the
diffusion step (and thus energy cost) while improving the
extraction performance of the diffusion step.
[0168] On a laboratory scale, to treat grape skins with PEF, an
electric field intensity of 1,300 V/cm applied to the skins for an
effective treatment period of 1 second makes it possible to obtain
a maximum cell membrane permeabilization rate. For a treatment with
HVED, the application of 60 pulses (of an effective duration of 0.6
ms) is sufficient to reach a total solutes extraction plateau.
[0169] Thus, the number of pulses necessary to fragment cells of
skins is fewer because the skins are a more fragile plant matrix
than whole pomaces.
[0170] On a semi-pilot scale, the effective treatment time must, on
the other hand, be increased to obtain the same results. On an
industrial scale, the reaction time is, on the other hand, close to
that of a semi-pilot scale, or about 1 ms.
[0171] Pretreatments of the skins with PEF or HVED also have a
positive effect on the extraction of polyphenols and total solutes.
Indeed, the quantity of polyphenols extracted is significantly
greater immediately after HVED (increase by a factor of 4 in
relation to simple diffusion) and then reaches an extraction
plateau, whereas after PEF the extraction of polyphenols is
increased by a factor of 2.
[0172] In addition, the initial extraction rates are different in
the case of conventional diffusion (without intensification
pretreatment) and in the case of PEF- and HVED-assisted diffusions.
The final quantities of polyphenols in the solvent, however, remain
equivalent after about 3 hours of extraction (the final values for
the assisted diffusions being slightly greater, however).
[0173] The principal compounds of the polyphenols obtained,
identifiable for example by high-performance liquid chromatography
(HPLC), are flavanols (catechin and epicatechin) and flavonols
(quercetin-3-O-glucoside and kaempferol-3-O-glucoside).
HVED-assisted diffusion produces an extraction of catechin and
epicatechin that is more effective than that from simple diffusion
or PEF assisted-diffusion. This difference can be attributed to
tissue fragmentation caused with HVED, as PEF does not break down
plant cells.
[0174] The diffusion temperature can also have an impact on the
performance of extractions from skins. Indeed, the damage caused to
the cells (and thus the extraction of polyphenols) induced by the
electrical treatments are all the more pronounced as temperature
increases.
[0175] The highest polyphenols content is obtained for
HVED-assisted diffusion at 60.degree. C. (C=32 .mu.mol GAE/g DM),
whereas HVED-assisted extraction at 20.degree. C. is as effective
as simple diffusion (without pretreatment) at 40.degree. C.
[0176] There is an optimal total specific energy for the extraction
of polyphenols with an electrical treatment with HVED of 80-100
kJ/kg with the application of 160 J pulses for 10 microseconds. The
total polyphenols extraction rate is thus 1.37.+-.0.11 g GAE per
100 g of dry matter with a corresponding antioxidant activity of
23.02.+-.3.06 g Trolox equivalent antioxidant capacity (TEAC) per
kilogram of dry matter. The same tendency was observed for
individual phenolic compounds (catechin, epicatechin,
quercetin-3-O-glucoside and kaempferol-3-O-glucoside). Beyond this
energy dissipated in the plant matrix, the formation of free
radicals and ozone during HVED contributes to the degradation of
the extracted polyphenols.
[0177] On a semi-pilot scale, this optimal total specific energy is
about 400 kJ/kg.
[0178] In the case of electrical treatment with PEF, optimal
extraction occurs when an electric field intensity of 20 kV/cm is
applied to the pomace (or to any other component, such as seeds,
etc.) for 6 ms (i.e., a treatment of 318 kJ/kg), followed by
diffusion at a treatment temperature of 50.degree. C. in an
extraction solvent containing 30% ethanol and 70% water. The
maximum total polyphenols extraction rate is thus 9 g GAE per 100 g
of dry matter and is reached after about 19 minutes. In comparison,
in the case of treatment with HVED (carried out for 1 ms at 40 kV
in the same solvent), the same polyphenols content is reached after
a diffusion time of 14 minutes.
[0179] In all cases, the electrical pretreatment combined with the
use of a hydroalcoholic extraction solvent thus makes it possible
to reduce the duration of the diffusion step.
[0180] The implementation of PEF has the advantage over the
implementation of HVED of preserving the structure of the solid
plant matrix: PEF acts by the electroporation of cell membranes
without fragmentation of the plant matrix, while HVED damages
membranes and cell walls of the plant matrix (the shock waves and
cavitation bubbles produced during the treatment collide with the
plant matrix and fragment it more or less according to the
intensity of the treatment).
[0181] It is also important to note that the combination of ethanol
in the extraction solvent with electrical pretreatment with PEF
further improves the electroporation of the cell membranes. Indeed,
ethanol weakens the cell membranes, thus improving the PEF
treatment which then forms membrane pores that themselves improve
the penetration of ethanol into the membranes and thus the
extraction of polyphenols.
[0182] If the plant matrix consists of seeds, the diameters of the
PEF-treated seeds are similar to those of untreated seeds (about
4,000 .mu.m), and of fine particles (dust located on the seed
surface) of about 10-20 .mu.m in the suspension after simple
diffusion and PEF-assisted diffusion. In comparison, treatment with
HVED reduces seed size by a factor of 20 (up to about 200 .mu.m in
diameter).
[0183] In addition, solid-liquid separation by centrifugation is
faster for suspensions treated with PEF than those treated with
HVED, so that the presence of seed debris makes such a separation
longer in the case of HVED pretreatment.
[0184] In particular for treatment with HVED, the application of
HVED on grape pomaces makes it possible to:
[0185] (1) increase by up to 3.5 times the yields in total solutes,
i.e., all the compounds having passed from the skins to the
solvent, and up to 2.5 times the level of polyphenols;
[0186] (2) reduce the diffusion temperature (yields in solutes
after extraction at 40.degree. C. without HVED being the same as
after extraction at 20.degree. C. with HVED);
[0187] (3) reduce the duration of the diffusion (after an effective
HVED time of 0.8 ms, the yield in solutes is 50% whereas it is only
25% after 60 minutes of extraction without HVED).
[0188] Pretreatment with electrical discharges improves the
extraction of polyphenols, on both a laboratory scale and a
semi-pilot scale (which then makes it possible to envisage a
possible application of HVED on an industrial scale).
[0189] Preservation Method
[0190] Concerning the method for preserving pomace, the addition of
sulfur dioxide has no visible influence on the extraction of total
solutes during diffusion with or without HVED. On the contrary,
when the pomaces are frozen and then unfrozen, the final yield in
solutes increases from 27.+-.2% (fresh pomace) to 68.+-.4% (frozen
pomace) for simple diffusion. With HVED pretreatment, the
extraction yield reaches a maximum of 88.+-.4% after four hours of
diffusion for frozen pomaces. With regard to polyphenols, these two
preservation methods influence their extraction with a maximum rate
of 0.69.+-.0.07% after four hours of extraction with HVED from
frozen pomaces.
[0191] Thus, as a method of preservation, the addition of sulfur
dioxide is preferable because the results of the extraction of
solutes and polyphenols are closest to those obtained with fresh
pomaces. In order to increase extraction yields, freezing is very
effective and acts by affecting the structure of cells through the
formation of ice crystals. In addition, freezing and treatment with
HVED seem to act on different cellular levels, explaining the
synergy observed between these two treatments. However, generally,
electrical pretreatments (such as PEF) are effective only on intact
plant cells, and thus require fresh plant material.
[0192] The method described above is not limited to the extraction
of polyphenols. It can also be used to extract other molecules of
interest such as polysaccharides, sugars, proteins, peptides,
organic acids (malic acid, tartaric acid, etc.), amino acids, fatty
acids, lipids, aromatic compounds, berry defense compounds,
etc.
[0193] The method is not limited to the extraction of molecules of
interest from grape pomace, as was seen above, but can also be
applied to lees, must deposits, tea, cocoa beans, berries, oilseeds
such as flax, apple, having undergone processing (pressing,
fermentation, etc.) or not.
[0194] Furthermore, the polyphenols obtained by a method in
accordance with the invention are powerful antioxidants (in
particular flavonols and anthocyanins).
[0195] The content in percentages of dry matter obtained from the
extracted polyphenols is thus greater than 60 for catechin, greater
than 30 for epicatechin, less than 5 for quercetin-3-O-glucoside,
and less than 1 for kaempferol-3-O-glucoside.
Example 1
Electrical Treatment with HVED
[0196] The grape pomace used in this example is residue of pressed,
unfermented Vitis vinifera var. Pinot Meunier grapes. The grape
pomace is composed of seeds, stems and skins. The dry matter
content of the grape pomace is 22.0.+-.0.1% by weight.
[0197] The 1 liter treatment chamber used includes two stainless
steel electrodes, one a tip 10 mm in diameter and the other a plate
35 mm in diameter, the two separated by 5 mm.
[0198] Treatment with HVED was applied to the grape pomace with the
following parameters: [0199] total specific energy: 80 kJ/kg;
[0200] solvent/grape pomace ratio: 5; [0201] temperature:
20.degree. C.; [0202] total diffusion time: 60 min.
[0203] The diffusion step is also carried out at a temperature of
20.degree. C. with stirring at 160 rpm.
[0204] The extraction yields results for the grape pomace are
summarized in the following table:
TABLE-US-00001 (g GAE/100 g 0% 10% 20% 30% dry matter) ethanol
ethanol ethanol ethanol Without HVED 0.15 Not 0.2 0.3 determined
With HVED 1.3 1.8 2.2 2.8
[0205] The antioxidant activities results for the grape pomace are
summarized in the following table:
TABLE-US-00002 (g TEAC/kg 0% 10% 20% 30% dry matter) ethanol
ethanol ethanol ethanol Without HVED 2 4 3 3.5 With HVED 25 35 40
68
Example 2
Electrical Treatment with PEF
[0206] Grape seeds were obtained beforehand from industrial grape
pomace resulting from the pressing of Vitis vinifera var. Pinot
Meunier grapes. These grape seeds were separated from the grape
pomace, dried industrially by treatment with hot air for a period
of 15 to 20 min and then recovered. The average diameter of the
grape seeds is 4 mm. The dry matter content of the seeds is
93.+-.1%.
[0207] The 1 liter treatment chamber used includes two planar
electrodes with a 95 cm surface area mounted in parallel and
separated by 5 mm. For the PEF treatment, 50.0 g of grape seeds was
placed between the two electrodes. The treatment chamber is then
filled with solvent composed of 30% ethanol and 70% distilled water
by weight.
[0208] The PEF treatment was applied to the plant matrix with the
following parameters: [0209] electric field intensity: 20 kV/cm;
[0210] duration: 6 ms; [0211] temperature: 50.degree. C.
[0212] The diffusion step is carried out at a temperature of
50.degree. C. in the same solvent.
[0213] The results of the extraction yields are summarized in the
following table:
TABLE-US-00003 (g GAE/100 g dry matter) 30% ethanol Without PEF 5.5
With PEF 7.5
Example 3
Profile of Polyphenols with an Electrical Treatment with HVED
TABLE-US-00004 [0214] Minimum concentration in a solution obtained
after diffusion Polyphenol (clarified supernatant) Gallic acid 3
mg/l Tryptophan 15 mg/l Catechin 100 mg/l Epicatechin 70 mg/l
Quercetin-3-O-glucoside + 25 mg/l glucuronide
Kaempferol-3-O-glucoside 5 mg/l Peonidin-3-O-glucoside 20 mg/l
Malvidin-3-O-glucoside 100 mg/l Flavanols 500 mg/l
Example 4
Profile of Polyphenols with an Electrical Treatment with PEF
TABLE-US-00005 [0215] Minimum concentration in a solution obtained
after diffusion Polyphenol (clarified supernatant) Gallic acid 0.5
mg/l Tryptophan 2 mg/l Catechin 6.5 mg/l Epicatechin 5.5 mg/l
Malvidin-3-O-glucoside 20 mg/l Flavanols 80 mg/l
* * * * *