U.S. patent application number 15/092667 was filed with the patent office on 2016-11-10 for production of soluble protein solutions from soy ("s701" cip).
This patent application is currently assigned to BURCON NUTRASCIENCE (MB) CORP.. The applicant listed for this patent is BURCON NUTRASCIENCE (MB) CORP.. Invention is credited to Brandy Gosnell, Brent E. Green, James Logie, Sarah Medina, Martin Schweizer, Kevin I. Segall.
Application Number | 20160324203 15/092667 |
Document ID | / |
Family ID | 45937790 |
Filed Date | 2016-11-10 |
United States Patent
Application |
20160324203 |
Kind Code |
A1 |
Schweizer; Martin ; et
al. |
November 10, 2016 |
PRODUCTION OF SOLUBLE PROTEIN SOLUTIONS FROM SOY ("S701" CIP)
Abstract
A soy protein product, which may be an isolate, produces
transparent heat-stable solutions at low pH values and is useful
for the fortification of soft drinks and sports drinks without
precipitation of protein. The soy protein product is obtained by
extracting a soy protein source material with an aqueous calcium
salt solution to form an aqueous soy protein solution, separating
the aqueous soy protein solution from residual soy protein source,
adjusting the pH of the aqueous soy protein solution to a pH of
about 1.5 to about 4.4 to produce an acidified clear soy protein
solution, which may be dried, following optional concentration and
diafiltration, to provide the soy protein product.
Inventors: |
Schweizer; Martin;
(Winnipeg, CA) ; Segall; Kevin I.; (Winnipeg,
CA) ; Green; Brent E.; (Warren, CA) ; Medina;
Sarah; (Winnipeg, CA) ; Logie; James;
(Winnipeg, CA) ; Gosnell; Brandy; (Winnipeg,
CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
BURCON NUTRASCIENCE (MB) CORP. |
Winnipeg |
|
CA |
|
|
Assignee: |
BURCON NUTRASCIENCE (MB)
CORP.
Winnipeg
CA
|
Family ID: |
45937790 |
Appl. No.: |
15/092667 |
Filed: |
April 7, 2016 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13879418 |
Aug 1, 2013 |
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15092667 |
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12603087 |
Oct 21, 2009 |
8691318 |
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13879418 |
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61107112 |
Oct 21, 2008 |
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61193457 |
Dec 2, 2008 |
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61202070 |
Jan 26, 2009 |
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61202553 |
Mar 12, 2009 |
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61213717 |
Jul 7, 2009 |
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61272241 |
Sep 3, 2009 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A23L 2/66 20130101; A23L
11/32 20160801; A23J 3/16 20130101; A23V 2300/34 20130101; A23J
1/14 20130101; A23L 33/185 20160801; A23V 2200/238 20130101; A23V
2200/238 20130101; A23V 2002/00 20130101; A23V 2002/00 20130101;
A23V 2300/34 20130101; A23V 2250/5488 20130101; A23V 2300/14
20130101; C11B 1/10 20130101 |
International
Class: |
A23L 33/185 20060101
A23L033/185; A23L 2/66 20060101 A23L002/66; A23J 3/16 20060101
A23J003/16; A23J 1/14 20060101 A23J001/14 |
Claims
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17. A soy protein product having a protein content of at least
about 60 wt % (N.times.6.25) d.b. which is substantially completely
soluble in an aqueous medium at a pH of about 7 to about 8.
18. An aqueous solution with a near neutral pH, preferably in the
range of about 6 to about 8, having dissolved therein the soy
product of claim 17.
19. The aqueous solution of claim 18 which is a beverage.
20. The aqueous solution of claim 18 which is incorporated in the
manufacture of a dairy analogue or a product that is a dairy/soy
blend.
21. The aqueous solution of claim 18 which is heat stable.
22. The aqueous solution of claim 18 which is transparent at a pH
of about 7.5 to about 8.
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34. A soy protein product having a protein content of at least
about 60 wt % (N.times.6.25) d.b. which has colorimeter readings
for a solution thereof in water, prepared by dissolving sufficient
soy protein product to supply 3.2 g of protein per 100 ml of water
used, which are a combination of L*=about 82 to about 100, a*=about
-2 to about 5 and b*=about 0 to about 30.
35. The soy protein product of claim 34 wherein said colorimeter
readings are a combination of L*=about 94 to about 100, a*=about -1
to about 1 and b*=about 0 to about 10.
36. The soy protein product of claim 34 which has a protein content
of at least about 90 wt % (N.times.6.25) d.b., preferably at least
about 100 wt % (N.times.6.25) d.b.
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Description
REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 13/879,418 filed Aug. 1, 2013, which itself is
a continuation-in-part of U.S. patent application Ser. No.
12/603,087 filed Oct. 21, 2009 (US Patent Publication No.
2010-0098818 now U.S. Pat. No. 8,691,318 issued Apr. 8, 2014;
WO2010/045727), which itself claims priority under 35 USC 119(e)
from U.S. Patent Applications No. 61/107,112 filed Oct. 21, 2008;
61/193,457 filed Dec. 2, 2008; 61/202,070 filed Jan. 26, 2009;
61/202,553 filed Mar. 12, 2009; 61/213,717 filed Jul. 7, 2009 and
61/272,241 filed Sep. 3, 2009.
FIELD OF INVENTION
[0002] The present invention is directed to the production of
protein solutions from soy and to novel soy protein products.
BACKGROUND TO THE INVENTION
[0003] A protein product, particularly an isolate, that is highly
soluble and produces transparent solutions at low pH would be
greatly valued in the food industry for use in various products,
particularly beverages, such as soft drinks and sports drinks. The
above properties combined with heat stability would further
increase the value of the product. Proteins for food use may be
derived from plant or animal sources but plant proteins are often
less expensive. Soy is a very common source of plant proteins for
food use. Soy proteins are recognized for their excellent
nutritional properties and health benefits.
[0004] Soy protein isolates conventionally are formed by an
isoelectric precipitation procedure in which meal from the
separation of soy oil from soybeans is processed by an initial
extraction under alkaline conditions, before the alkaline extract
is acidified to the isoelectric point of soybean protein to result
in protein precipitation. The precipitated soy protein may be
washed and/or neutralized, then is dried to provide the soy protein
isolate. Soy protein isolates have a protein content of at least
about 90 wt % (N.times.6.25) on a dry weight basis (d.b.).
[0005] Although a range of soy protein products is available, with
a variety of functional properties, to our knowledge, there does
not exist a soluble soy protein isolate or product that produces
transparent and heat stable solutions under low pH conditions.
SUMMARY OF INVENTION
[0006] It has now been found that it is possible to provide a soy
protein product having a protein content of at least about 60 wt %
(N.times.6.25) d.b. that produces transparent and heat stable
solutions at low pH values and, therefore, which may be used for
protein fortification of, in particular, soft drinks and sports
drinks, as well as other aqueous systems, without precipitation of
protein.
[0007] The novel soy protein product provided herein has a unique
combination of parameters not found in other soy protein products.
The product is completely soluble in aqueous solution at acid pH
values less than about 4.4 and is heat stable in this pH range
permitting thermal processing of the aqueous solution of the
product, such as hot fill applications. Given the complete
solubility of the product, no stabilizers or other additives are
necessary to maintain the protein in solution or suspension. The
soy protein product has been described as having no "beany" flavour
and no off odours. The product is low in phytic acid, generally
less than about 1.5 wt %, preferably less than about 0.5 wt %. No
enzymes are required in the production of the soy protein product.
The soy protein product is preferably an isolate having a protein
content of at least about 90 wt %, preferably at least about 100 wt
% (N.times.6.25).
[0008] In accordance with one aspect of the present invention,
there is provided a method of producing a soy protein product
having a soy protein content of at least about 60 wt %
(N.times.6.25) on a dry weight basis, which comprises: [0009] (a)
extracting a soy protein source with an aqueous calcium chloride
solution to cause solubilization of soy protein from the protein
source and to form an aqueous soy protein solution, [0010] (b) at
least partially separating the aqueous soy protein solution from
residual soy protein source, [0011] (c) optionally diluting the
aqueous soy protein solution, [0012] (d) adjusting the pH of the
aqueous soy protein solution to a pH of about 1.5 to about 4.4,
preferably about 2 to about 4, to produce an acidified clear soy
protein solution, [0013] (e) optionally polishing the acidified
clear soy protein solution to remove residual particulates, [0014]
(f) optionally concentrating the aqueous clear soy protein solution
while maintaining the ionic strength substantially constant by
using a selective membrane technique, [0015] (g) optionally
diafiltering the concentrated soy protein solution, and [0016] (h)
optionally drying the concentrated soy protein solution.
[0017] The soy protein product preferably is an isolate having a
protein content of at least about 90 wt %, preferably at least
about 100 wt %, (N.times.6.25) d.b.
[0018] The present invention further provides a novel soy protein
isolate which is water soluble and forms heat stable transparent
solutions at acid pH values of less than about 4.4 and is useful
for the protein fortification of aqueous systems, including soft
drinks and sports drinks, without leading to protein precipitation.
The soy protein isolate is also low in phytic acid content,
generally less than about 1.5% by weight, preferably less than
about 0.5% by weight. The soy protein in the product is not
hydrolyzed.
[0019] Thus, in another aspect to the present invention, there is
provided a soy protein isolate having a protein content of at least
about 90 wt % (N.times.6.25) d.b., preferably at least about 100 wt
% (N.times.6.25) d.b., which is substantially completely soluble in
an aqueous medium at a pH of less than about 4.4, preferably about
1.5 to about 4.4.
[0020] The soy protein isolate provided herein may be provided as
an aqueous solution thereof having a high degree of clarity at acid
pH values, generally from less than about 4.4, preferably about 1.5
to about 4.4, and which is heat stable at these pH values.
[0021] The novel soy protein product of the invention can be
blended with powdered drinks for the formation of aqueous soft
drinks or sports drinks by dissolving the same in water. Such blend
may be a powdered beverage.
[0022] While the present invention refers mainly to the production
of soy protein isolate, it is contemplated that soy protein
products of lesser purity may be provided having similar properties
to the soy protein isolate. Such lesser purity products may have a
protein concentration of at least about 60% by weight
(N.times.6.25) d.b.
[0023] In another aspect of the present invention, there is
provided an aqueous solution of the soy product provided herein
which is heat stable at a pH of less than about 4.4. The aqueous
solution may be a beverage, which may be a clear beverage in which
the soy protein product is completely soluble and transparent or an
opaque beverage in which the soy protein product does not increase
the opacity.
[0024] The present invention also provides a soy protein product
having a protein content of at least about 60 wt % (N.times.6.25)
d.b., preferably at least about 90 wt % and more preferably at
least about 100 wt %, which is substantially completely soluble at
a pH of about 7 to about 8. An aqueous solution of the soy protein
product prepared at a near neutral pH, such as a pH of about 6 to
about 8, may be a beverage. The aqueous solution of the soy protein
product prepared at near neutral pH may also be utilized in the
production of any food application that makes use of a protein
product soluble at near neutral pH, such as a plant based dairy
analogue, such as soy milk and soy ice cream, or a dairy-type
product containing a mix of dairy and plant ingredients. The
aqueous solutions are heat stable and solutions at a pH of about
7.5 to about 8 are completely transparent.
[0025] In a further aspect of the present invention, there is
provided a soy protein product having a protein content of at least
about 60 wt % (N.times.6.25) d.b., preferably at least about 90 wt
%, more preferably at least about 100 wt %, which has a solubility
at 1% protein w/v in water at a pH of about 2 to about 4 greater
than about 95%, as determined by the methods described in Example
14 below.
[0026] Additionally, the present invention provides a soy protein
product having a protein content of at least about 60 wt %
(N.times.6.25) d.b., preferably at least about 90 wt %, more
preferably at least about 100 wt %, which has an absorbance of
visible light at 600 nm (A600) for a 1% protein w/v aqueous
solution at a pH of about 2 to about 4 of less than 0.150,
preferably less than about 0.100, more preferably less than 0.050,
as determined by the method described in Example 15 below.
[0027] In accordance with a further embodiment of the invention,
there is provided a soy protein product having a protein content of
at least about 60 wt % (N.times.6.25) d.b., preferably at least
about 90 wt %, more preferably at least about 100 wt %, which has a
haze reading for a 1% protein w/v aqueous solution at a pH of about
2 to about 4, of less than about 15%, preferably less than about
10% and more preferably less than about 5%, as determined by the
method described in Example 15 below.
[0028] In accordance with a yet further embodiment of the
invention, there is provided a soy protein product having a protein
content of at least about 60 wt % (N.times.6.25) d.b., preferably
at least about 90 wt %, more preferably at least about 100 wt %,
which has a haze reading for a solution thereof in water, prepared
by dissolving 2 g of protein per 100 ml of water used, after heat
treatment at 95.degree. C. for 30 seconds of less than 15%,
preferably less than about 10% and more preferably less than 5%, as
determined by the method described in Example 16 below.
[0029] As may be seen from the data presented below, the aqueous
solutions of the soy protein product provided herein are
practically colourless, unlike aqueous solutions provided from
typical commercial soy protein products. It will be appreciated
that a completely clear, colourless solution would provide
colorimeter readings of L*=100, a*=0 and b*=0. Based on the data
generated herein, in accordance with another aspect of the present
invention, there is provided a novel protein product having a
protein content of at least 60 wt % (N.times.6.25) d.b., preferably
at least about 90 wt %, more preferably at least about 100 wt %,
which has colorimeter readings for a solution thereof in water,
prepared by dissolving sufficient soy protein product to supply 3.2
g of protein per 100 ml of water used, which are a combination of
L*=about 82 to about 100, preferably about 94 to about 100,
a*=about -2 to about 5, preferably about -1 to about 1, and
b*=about 0 to about 30, preferably about 0 to about 10.
[0030] The soy protein product produced according to the process
herein lacks the characteristic beany flavour of soy protein
products and is suitable, not only for protein fortification of
acid media, but may be used in a wide variety of conventional
applications of protein products, including but not limited to
protein fortification of processed foods and beverages,
emulsification of oils, as a body former in baked goods and foaming
agent in products which entrap gases. In addition, the soy protein
product may be formed into protein fibers, useful in meat analogs
and may be used as an egg white substitute or extender in food
products where egg white is used as a binder. The soy protein
product may also be used in nutritional supplements. Other uses of
the soy protein product are in pet foods, animal feed and in
industrial and cosmetic applications and in personal care
products.
GENERAL DESCRIPTION OF INVENTION
[0031] The initial step of the process of providing the soy protein
product involves solubilizing soy protein from a soy protein
source. The soy protein source may be soybeans or any soy product
or by-product derived from the processing of soybeans, including
but not limited to soy meal, soy flakes, soy grits and soy flour.
The soy protein source may be used in the full fat form, partially
defatted form or fully defatted form. Where the soy protein source
contains an appreciable amount of fat, an oil-removal step
generally is required during the process. The soy protein recovered
from the soy protein source may be the protein naturally occurring
in soybean or the proteinaceous material may be a protein modified
by genetic manipulation but possessing characteristic hydrophobic
and polar properties of the natural protein.
[0032] Protein solubilization from the soy protein source material
is effected most conveniently using calcium chloride solution,
although solutions of other calcium salts, may be used. In
addition, other alkaline earth metal compounds may be used, such as
magnesium salts. Further, extraction of the soy protein from the
soy protein source may be effected using calcium salt solution in
combination with another salt solution, such as sodium chloride.
Additionally, extraction of the soy protein from the soy protein
source may be effected using water or other salt solution, such as
sodium chloride, with calcium salt subsequently being added to the
aqueous soy protein solution produced in the extraction step.
Precipitate formed upon addition of the calcium salt is removed
prior to subsequent processing.
[0033] As the concentration of the calcium salt solution increases,
the degree of solubilization of protein from the soy protein source
initially increases until a maximum value is achieved. Any
subsequent increase in salt concentration does not increase the
total protein solubilized. The concentration of calcium salt
solution which causes maximum protein solubilization varies
depending on the salt concerned. It is usually preferred to utilize
a concentration value less than about 1.0 M, and more preferably a
value of about 0.10 to about 0.15 M.
[0034] In a batch process, the salt solubilization of the protein
is effected at a temperature of from about 1.degree. C. to about
100.degree. C., preferably about 15.degree. to about 60.degree. C.,
more preferably about 15.degree. C. to about 35.degree. C.,
preferably accompanied by agitation to decrease the solubilization
time, which is usually about 1 to about 60 minutes. It is preferred
to effect the solubilization to extract substantially as much
protein from the soy protein source as is practicable, so as to
provide an overall high product yield.
[0035] In a continuous process, the extraction of the soy protein
from the soy protein source is carried out in any manner consistent
with effecting a continuous extraction of soy protein from the soy
protein source. In one embodiment, the soy protein source is
continuously mixed with the calcium salt solution and the mixture
is conveyed through a pipe or conduit having a length and at a flow
rate for a residence time sufficient to effect the desired
extraction in accordance with the parameters described herein. In
such a continuous procedure, the salt solubilization step is
effected rapidly, in a time of up to about 10 minutes, preferably
to effect solubilization to extract substantially as much protein
from the soy protein source as is practicable. The solubilization
in the continuous procedure is effected at temperatures between
about 1.degree. C. and about 100.degree. C., preferably about
15.degree. to about 60.degree. C., more preferably between about
15.degree. C. and about 35.degree. C.
[0036] The extraction is generally conducted at a pH of about 5 to
about 11, preferably about 5 to about 7. The pH of the extraction
system (soy protein source and calcium salt solution) may be
adjusted to any desired value within the range of about 5 to about
11 for use in the extraction step by the use of any convenient food
grade acid, usually hydrochloric acid or phosphoric acid, or food
grade alkali, usually sodium hydroxide, as required.
[0037] The concentration of soy protein source in the calcium salt
solution during the solubilization step may vary widely. Typical
concentration values are about 5 to about 15% w/v.
[0038] The protein extraction step with the aqueous salt solution
has the additional effect of solubilizing fats which may be present
in the soy protein source, which then results in the fats being
present in the aqueous phase.
[0039] The protein solution resulting from the extraction step
generally has a protein concentration of about 5 to about 50 g/L,
preferably about 10 to about 50 g/L.
[0040] The aqueous calcium salt solution may contain an
antioxidant. The antioxidant may be any convenient antioxidant,
such as sodium sulfite or ascorbic acid. The quantity of
antioxidant employed may vary from about 0.01 to about 1 wt % of
the solution, preferably about 0.05 wt %. The antioxidant serves to
inhibit oxidation of any phenolics in the protein solution.
[0041] The aqueous phase resulting from the extraction step then
may be separated from the residual soy protein source, in any
convenient manner, such as by employing a decanter centrifuge or
any suitable sieve, followed by disc centrifugation and/or
filtration, to remove residual soy protein source material. The
separated residual soy protein source may be dried for disposal.
Alternatively, the separated residual soy protein source may be
processed to recover some residual protein. The separated residual
soy protein source may be re-extracted with fresh calcium salt
solution and the protein solution yielded upon clarification
combined with the initial protein solution for further processing
as described below. Alternatively, the separated residual soy
protein source may be processed by a conventional isoelectric
precipitation procedure or any other convenient procedure to
recover residual protein.
[0042] Where the soy protein source contains significant quantities
of fat, as described in U.S. Pat. Nos. 5,844,086 and 6,005,076,
assigned to the assignee hereof and the disclosures of which are
incorporated herein by reference, then the defatting steps
described therein may be effected on the separated aqueous protein
solution. Alternatively, defatting of the separated aqueous protein
solution may be achieved by any other convenient procedure.
[0043] The aqueous soy protein solution may be treated with an
adsorbent, such as powdered activated carbon or granulated
activated carbon, to remove colour and/or odour compounds. Such
adsorbent treatment may be carried out under any convenient
conditions, generally at the ambient temperature of the separated
aqueous protein solution. For powdered activated carbon, an amount
of about 0.025% to about 5% w/v, preferably about 0.05% to about 2%
w/v, is employed. The adsorbing agent may be removed from the soy
solution by any convenient means, such as by filtration.
[0044] The resulting aqueous soy protein solution may be diluted
generally with about 0.5 to about 10 volumes, preferably about 0.5
to about 2 volumes, of aqueous diluent in order to decrease the
conductivity of the aqueous soy protein solution to a value of
generally below about 90 mS, preferably about 4 to about 18 mS.
Such dilution is usually effected using water, although dilute salt
solution, such as sodium chloride or calcium chloride, having a
conductivity of up to about 3 mS, may be used.
[0045] The diluent with which the soy protein solution is mixed may
have a temperature of about 2.degree. to about 70.degree. C.,
preferably about 10.degree. to about 50.degree. C., more preferably
about 20.degree. to about 30.degree. C.
[0046] The diluted soy protein solution then is adjusted in pH to a
value of about 1.5 to about 4.4, preferably about 2 to about 4, by
the addition of any suitable food grade acid, to result in a clear
acidified aqueous soy protein solution. The clear acidified aqueous
soy protein solution has a conductivity of generally below about 95
mS, preferably about 4 to about 23 mS.
[0047] The clear acidified aqueous soy protein solution may be
subjected to a heat treatment to inactivate heat labile
anti-nutritional factors, such as trypsin inhibitors, present in
such solution as a result of extraction from the soy protein source
material during the extraction step. Such a heating step also
provides the additional benefit of reducing the microbial load.
Generally, the protein solution is heated to a temperature of about
70.degree. to about 160.degree. C., for about 10 seconds to about
60 minutes, preferably about 80.degree. to about 120.degree. C. for
about 10 seconds to about 5 minutes, more preferably about
85.degree. to about 95.degree. C., for about 30 seconds to about 5
minutes. The heat treated acidified soy protein solution then may
be cooled for further processing as described below, to a
temperature of about 2.degree. to about 60.degree. C., preferably
about 20.degree. C. to about 35.degree. C.
[0048] The optionally diluted, acidified and optionally heat
treated protein solution may optionally be polished by any
convenient means, such as by filtering, to remove any residual
particulates.
[0049] The resulting clear acidified aqueous soy protein solution
may be directly dried to produce a soy protein product. In order to
provide a soy protein product having a decreased impurities content
and a reduced salt content, such as a soy protein isolate, the
clear acidified aqueous soy protein solution may be processed prior
to drying.
[0050] The clear acidified aqueous soy protein solution may be
concentrated to increase the protein concentration thereof while
maintaining the ionic strength thereof substantially constant. Such
concentration generally is effected to provide a concentrated soy
protein solution having a protein concentration of about 50 to
about 300 g/L, preferably about 100 to about 200 g/L.
[0051] The concentration step may be effected in any convenient
manner consistent with batch or continuous operation, such as by
employing any convenient selective membrane technique, such as
ultrafiltration or diafiltration, using membranes, such as
hollow-fibre membranes or spiral-wound membranes, with a suitable
molecular weight cutoff, such as about 3,000 to about 1,000,000
Daltons, preferably about 5,000 to about 100,000 Daltons, having
regard to differing membrane materials and configurations, and, for
continuous operation, dimensioned to permit the desired degree of
concentration as the aqueous protein solution passes through the
membranes.
[0052] As is well known, ultrafiltration and similar selective
membrane techniques permit low molecular weight species to pass
therethrough while preventing higher molecular weight species from
so doing. The low molecular weight species include not only the
ionic species of the food grade salt but also low molecular weight
materials extracted from the source material, such as
carbohydrates, pigments, low molecular weight proteins and
anti-nutritional factors, such as trypsin inhibitors, which are
themselves low molecular weight proteins. The molecular weight
cut-off of the membrane is usually chosen to ensure retention of a
significant proportion of the protein in the solution, while
permitting contaminants to pass through having regard to the
different membrane materials and configurations.
[0053] The concentrated soy protein solution then may be subjected
to a diafiltration step using water or a dilute saline solution.
The diafiltration solution may be at its natural pH or at a pH
equal to that of the protein solution being diafiltered or at any
pH value in between. Such diafiltration may be effected using from
about 2 to about 40 volumes of diafiltration solution, preferably
about 5 to about 25 volumes of diafiltration solution. In the
diafiltration operation, further quantities of contaminants are
removed from the clear aqueous soy protein solution by passage
through the membrane with the permeate. This purifies the clear
aqueous protein solution and may also reduce its viscosity. The
diafiltration operation may be effected until no significant
further quantities of contaminants or visible colour are present in
the permeate or until the retentate has been sufficiently purified
so as, when dried, to provide a soy protein isolate with a protein
content of at least about 90 wt % (N.times.6.25) d.b. Such
diafiltration may be effected using the same membrane as for the
concentration step. However, if desired, the diafiltration step may
be effected using a separate membrane with a different molecular
weight cut-off, such as a membrane having a molecular weight
cut-off in the range of about 3,000 to about 1,000,000 Daltons,
preferably about 5,000 to about 100,000 Daltons, having regard to
different membrane materials and configuration.
[0054] Alternatively, the diafiltration step may be applied to the
clear acidified aqueous protein solution prior to concentration or
to the partially concentrated clear acidified aqueous protein
solution. Diafiltration may also be applied at multiple points
during the concentration process. When diafiltration is applied
prior to concentration or to the partially concentrated solution,
the resulting diafiltered solution may then be additionally
concentrated. The viscosity reduction achieved by diafiltering
multiple times as the protein solution is concentrated may allow a
higher final, fully concentrated protein concentration to be
achieved. This reduces the volume of material to be dried.
[0055] The concentration step and the diafiltration step may be
effected herein in such a manner that the soy protein product
subsequently recovered contains less than about 90 wt % protein
(N.times.6.25) d.b., such as at least about 60 wt % protein
(N.times.6.25) d.b. By partially concentrating and/or partially
diafiltering the clear aqueous soy protein solution, it is possible
to only partially remove contaminants. This protein solution may
then be dried to provide a soy protein product with lower levels of
purity. The soy protein product is still able to produce clear
protein solutions under acidic conditions.
[0056] An antioxidant may be present in the diafiltration medium
during at least part of the diafiltration step. The antioxidant may
be any convenient antioxidant, such as sodium sulfite or ascorbic
acid. The quantity of antioxidant employed in the diafiltration
medium depends on the materials employed and may vary from about
0.01 to about 1 wt %, preferably about 0.05 wt %. The antioxidant
serves to inhibit the oxidation of any phenolics present in the
concentrated soy protein solution.
[0057] The concentration step and the optional diafiltration step
may be effected at any convenient temperature, generally about
2.degree. to about 60.degree. C., preferably about 20.degree. to
about 35.degree. C., and for the period of time to effect the
desired degree of concentration and diafiltration. The temperature
and other conditions used to some degree depend upon the membrane
equipment used to effect the membrane processing, the desired
protein concentration of the solution and the efficiency of the
removal of contaminants to the permeate.
[0058] There are two main trypsin inhibitors in soy, namely the
Kunitz inhibitor, which is a heat-labile molecule with a molecular
weight of approximately 21,000 Daltons, and the Bowman-Birk
inhibitor, a more heat-stable molecule with a molecular weight of
about 8,000 Daltons. The level of trypsin inhibitor activity in the
final soy protein product can be controlled by manipulation of
various process variables.
[0059] As noted above, heat treatment of the clear acidified
aqueous soy protein solution may be used to inactivate heat-labile
trypsin inhibitors. The partially concentrated or fully
concentrated acidified soy protein solution may also be heat
treated to inactivate heat labile trypsin inhibitors. When the heat
treatment is applied to the partially concentrated acidified soy
protein solution, the resulting heat treated solution may then be
additionally concentrated.
[0060] In addition, the concentration and/or diafiltration steps
may be operated in a manner favorable for removal of trypsin
inhibitors in the permeate along with the other contaminants.
Removal of the trypsin inhibitors is promoted by using a membrane
of larger pore size, such as about 30,000 to about 1,000,000 Da,
operating the membrane at elevated temperatures, such as about
30.degree. to about 60.degree. C., and employing greater volumes of
diafiltration medium, such as about 20 to about 40 volumes.
[0061] Acidifying and membrane processing the diluted protein
solution at a lower pH of about 1.5 to about 3 may reduce the
trypsin inhibitor activity relative to processing the solution at
higher pH of about 3 to about 4.4. When the protein solution is
concentrated and diafiltered at the low end of the pH range, it may
be desired to raise the pH of the retentate prior to drying. The pH
of the concentrated and diafiltered protein solution may be raised
to the desired value, for example pH 3, by the addition of any
convenient food grade alkali such as sodium hydroxide.
[0062] Further, a reduction in trypsin inhibitor activity may be
achieved by exposing soy materials to reducing agents that disrupt
or rearrange the disulfide bonds of the inhibitors. Suitable
reducing agents include sodium sulfite, cysteine and
N-acetylcysteine.
[0063] The addition of such reducing agents may be effected at
various stages of the overall process. The reducing agent may be
added with the soy protein source material in the extraction step,
may be added to the clarified aqueous soy protein solution
following removal of residual soy protein source material, may be
added to the concentrated protein solution before or after
diafiltration or may be dry blended with the dried soy protein
product. The addition of the reducing agent may be combined with a
heat treatment step and the membrane processing steps, as described
above.
[0064] If it is desired to retain active trypsin inhibitors in the
concentrated protein solution, this can be achieved by eliminating
or reducing the intensity of the heat treatment step, not utilizing
reducing agents, operating the concentration and diafiltration
steps at the higher end of the pH range, such as pH 3 to about 4.4,
utilizing a concentration and diafiltration membrane with a smaller
pore size, operating the membrane at lower temperatures and
employing fewer volumes of diafiltration medium.
[0065] The concentrated and optionally diafiltered protein solution
may be subject to a further defatting operation, if required, as
described in U.S. Pat. Nos. 5,844,086 and 6,005,076. Alternatively,
defatting of the concentrated and optionally diafiltered protein
solution may be achieved by any other convenient procedure.
[0066] The concentrated and optionally diafiltered clear aqueous
protein solution may be treated with an adsorbent, such as powdered
activated carbon or granulated activated carbon, to remove colour
and/or odour compounds. Such adsorbent treatment may be carried out
under any convenient conditions, generally at the ambient
temperature of the concentrated protein solution. For powdered
activated carbon, an amount of about 0.025% to about 5% w/v,
preferably about 0.05% to about 2% w/v, is employed. The adsorbent
may be removed from the soy protein solution by any convenient
means, such as by filtration.
[0067] The concentrated and optionally diafiltered clear aqueous
soy protein solution may be dried by any convenient technique, such
as spray drying or freeze drying. A pasteurization step may be
effected on the soy protein solution prior to drying. Such
pasteurization may be effected under any desired pasteurization
conditions. Generally, the concentrated and optionally diafiltered
soy protein solution is heated to a temperature of about 55.degree.
to about 70.degree. C., preferably about 60.degree. to about
65.degree. C., for about 30 seconds to about 60 minutes, preferably
about 10 minutes to about 15 minutes. The pasteurized concentrated
soy protein solution then may be cooled for drying, preferably to a
temperature of about 25.degree. to about 40.degree. C.
[0068] The dry soy protein product has a protein content in excess
of about 60 wt % (N.times.6.25) d.b. Preferably, the dry soy
protein product is an isolate with a high protein content, in
excess of about 90 wt % protein, preferably at least about 100 wt %
(N.times.6.25) d.b.
[0069] The soy protein product produced herein is soluble in an
acidic aqueous environment, making the product ideal for
incorporation into beverages, both carbonated and uncarbonated, to
provide protein fortification thereto. Such beverages have a wide
range of acidic pH values, ranging from about 2.5 to about 5. The
soy protein product provided herein may be added to such beverages
in any convenient quantity to provide protein fortification to such
beverages, for example, at least about 5 g of the soy protein per
serving. The added soy protein product dissolves in the beverage
and does not impair the clarity of the beverage, even after thermal
processing. The soy protein product may be blended with dried
beverage prior to reconstitution of the beverage by dissolution in
water. In some cases, modification to the normal formulation of the
beverages to tolerate the composition of the invention may be
necessary where components present in the beverage may adversely
affect the ability of the composition of the invention to remain
dissolved in the beverage.
EXAMPLES
[0070] A series of trial experiments (Examples 1 to 3) were carried
out to ascertain if exposure to calcium could be used to generate a
soluble soy protein that produced transparent, heat stable
solutions at low pH.
Example 1
[0071] Dry soybeans (30 g) were combined with either water, 0.01 M
CaCl.sub.2 or 0.15 M NaCl (300 ml) in a kitchen blender and
processed for 5 minutes at top speed. The samples were then
centrifuged at 7,100 g for 10 minutes to separate the extract from
the fat and residual solids. The samples prepared with water and
calcium chloride solutions had poor separation and so were
centrifuged again at 10,200 g for 10 minutes. The pH of the
extracts was measured and then aliquots filtered with a 0.45 .mu.m
pore size syringe filter and the protein content determined using a
Leco FP528 Nitrogen Determinator. The clarity of the filtered
extracts (NaCl and CaCl.sub.2 trials) was measured as the
absorbance at 600 nm (A600) and then a portion of the sample was
acidified to pH 3 with diluted HCl and the A600 measured again.
Aliquots of the clarified extracts (all trials) were also diluted
1:10 in room temperature water and the A600 and pH measured then
the samples acidified to pH 3 with diluted HCl and the A600
measured again. Another aliquot of the NaCl extract was filtered
with a 25 .mu.m pore size filter paper. The conductivity of this
sample was measured and then raised to 19 mS by the addition of
calcium chloride. This sample was syringe filtered (0.45 .mu.m) and
then the effect of adjustment to pH 3 on sample clarity was
assessed for the undiluted sample and a sample diluted 1:10 with
room temperature water.
[0072] When the three extract samples were centrifuged at 7,100 g
for 10 minutes, only the sodium chloride extract sample separated
well. The fat was still highly dispersed in the aqueous layer for
the water and calcium chloride samples. Centrifuging again at
10,200 g for 10 minutes did not much improve the separation.
Perhaps this poor separation was an effect of aqueous phase density
as the sodium chloride sample had much more dissolved salt than the
calcium chloride sample. The post centrifugation extracts were
further clarified by syringe filtering through a 0.45 .mu.m pore
size filter. The water extract plugged the filter rapidly and the
calcium chloride extract did not come out entirely clear.
[0073] Surprisingly, water was found to extract more protein than
the salt solutions used (Table 1, below). Addition of calcium
chloride to the sodium chloride extract, which raised the
conductivity from 16.70 mS to 19.99 mS was observed to introduce
precipitate and protein must have been lost along with any other
species removed.
TABLE-US-00001 TABLE 1 Protein content of various clarified extract
samples sample % protein 0.15M NaCl 1.28 0.15M NaCl plus CaCl.sub.2
1.03 0.01M CaCl.sub.2 0.74 water 1.98
[0074] Although not clear, the undiluted extract samples acidified
in the presence of calcium were clearer than the acidified sodium
chloride extract, which was very cloudy (Table 2, below).
TABLE-US-00002 TABLE 2 Clarity of various clarified then acidified
extract samples sample initial pH final pH final A600 0.15M NaCl
6.19 3.01 >3.000 0.15M NaCl plus CaCl.sub.2 5.38 3.00 1.220
0.01M CaCl.sub.2 6.11 2.84 1.066
[0075] Diluting the extract samples with water prior to
acidification resulted in excellent clarity for the samples exposed
to calcium chloride, particularly with calcium chloride in the
extraction (Table 3, below). Acidification of the diluted sodium
chloride and water extracts resulted in cloudy samples.
Interestingly, after dilution and before acidification, notable
precipitation was observed in the sodium chloride plus calcium
chloride and water samples but as mentioned, after acidification
only the samples with calcium were clear.
TABLE-US-00003 TABLE 3 Clarity of various diluted (1:10) then
acidified extract samples sample initial pH final pH final A600
0.15M NaCl 6.31 2.81 0.789 0.15M NaCl plus CaCl.sub.2 5.62 3.00
0.094 0.01M CaCl.sub.2 6.39 2.76 0.024 water 6.86 3.01 0.679
Example 2
[0076] Dry soybeans (30 g) were combined with either 0.05 M
CaCl.sub.2, 0.10 M CaCl.sub.2 or 0.15 M CaCl.sub.2 (300 ml) in a
kitchen blender and processed for 5 minutes at top speed. The
samples were then centrifuged at 7,100 g for 10 minutes to separate
the extract from the fat and residual solids. The extracts were
filtered with 0.45 .mu.m pore size syringe filters and the protein
content determined by Leco analysis and the clarity of the samples
measured by A600. Clarified extract samples were then either
acidified directly to pH 3 with diluted HCl and the A600 measured
or diluted 1:10 with room temperature water and the resulting
solution adjusted to pH 3 with diluted HCl and the A600
measured.
[0077] When the various calcium chloride extraction samples were
centrifuged the 0.05 M and 0.10 M CaCl.sub.2 samples separated
quite well but the 0.15 M CaCl.sub.2 sample did not. When the
centrifuged extracts were syringe filtered the 0.05 M sample was
crystal clear, the 0.10 M sample was slightly hazy and the 0.15 M
sample was very hazy, almost milky (Table 4, below). It is thought
that fat was responsible for the cloud. Working with defatted soy
meal as the starting material should eliminate the problem.
However, as the 0.15 M CaCl.sub.2 extract could not be clarified it
was excluded from the testing.
TABLE-US-00004 TABLE 4 Results for clarified soybean extracts with
different CaCl.sub.2 concentrations sample % protein A600 0.05M
CaCl.sub.2 0.84 0.017 0.10M CaCl.sub.2 1.42 0.085 0.15M CaCl.sub.2
2.03 1.900
[0078] Dilution of the extracts into water appeared to produce some
precipitate, with the amount appearing to increase at the higher
salt concentration. Directly acidifying the 0.05 M and 0.10 M
CaCl.sub.2 extracts gave fairly clear solutions but excellent
clarity was achieved by diluting these samples with water before
acidifying (Table 5, below).
TABLE-US-00005 TABLE 5 Clarity of acidified soybean extracts with
and without dilution sample initial pH final pH final A600 0.05M
CaCl.sub.2 5.55 3.06 0.079 0.05M CaCl.sub.2 (diluted 1:10) 5.60
3.02 0.007 0.10M CaCl.sub.2 5.41 3.07 0.101 0.10M CaCl.sub.2
(diluted 1:10) 5.43 3.07 0.014
Example 3
[0079] Toasted soy meal (10 g) was extracted with either 0.15 M
NaCl, 0.15 M CaCl.sub.2 or water (100 ml) for 30 minutes at room
temperature on an orbital shaker platform. The samples were then
centrifuged at 10,200 g for 10 minutes to separate extract from the
spent meal. The supernatant was then further clarified by
filtration through 25 .mu.m pore size filter paper and the pH and
conductivity of the samples measured. Small samples were then
further clarified with a 0.45 .mu.m pore size syringe filter and
then analyzed for clarity (A600) and protein content (Leco). A
clarified sample of each extract was diluted into 4 parts room
temperature water and the A600 measured again. Diluted and
undiluted extract samples were acidified to pH 3 with diluted HCl
and the clarity measured again. A sample of the sodium chloride
extract was also made up to a conductivity of 19 mS with calcium
chloride and the clarity of full strength and 1:5 diluted samples
assessed at natural pH and pH 3.
[0080] The water and calcium chloride solutions appeared to extract
more protein than the sodium chloride solution (Table 6, below).
Overall extractability was quite low as the meal was toasted and so
exposed to relatively severe heat treatment.
TABLE-US-00006 TABLE 6 Properties of various extracts of toasted
soy meal sample pH cond. (mS) % protein water 6.63 3.47 0.43 0.15M
NaCl 6.47 16.34 0.33 0.15M CaCl.sub.2 5.70 22.60 0.44
[0081] All three extracts were of relatively similar clarity after
filtering (Table 7, below). Dilution of the water extract and the
sodium chloride extract with four parts of water did not result in
any protein precipitation. However, precipitate formed when the
calcium chloride extract was diluted. This precipitate dissolved
completely when the pH was lowered to 3 giving a crystal clear
sample. The undiluted calcium chloride extract also stayed quite
clear when acidified. The water and sodium chloride extracts became
highly cloudy when acidified regardless of whether the sample was
diluted with water.
TABLE-US-00007 TABLE 7 Clarity of extracts before and after
acidification sample A600 natural pH A600 pH 3 water 0.261 2.786
water (diluted 1:5) 0.051 1.493 0.15M NaCl 0.154 2.733 0.15M NaCl
(diluted 1:5) 0.033 1.302 0.15M CaCl.sub.2 0.133 0.100 0.15M
CaCl.sub.2 (diluted 1:5) 2.058 0.017
[0082] Adding calcium chloride to the sodium chloride extract
sample to achieve a conductivity of 19 mS resulted in the
development of a cloud in the sample. This calcium induced
precipitate did not appear to resolubilize with the addition of
acid. As such, both the solutions tested contained significant
cloud (Table 8, below). The precipitate should have been removed by
centrifugation or filtration prior to acidification of the
samples.
TABLE-US-00008 TABLE 8 Clarity of NaCl extract with added
CaCl.sub.2 before and after acidification sample A600 natural pH
A600 pH 3 0.15M NaCl plus CaCl.sub.2 2.536 0.986 0.15M NaCl plus
CaCl.sub.2 (diluted 1:5) 1.261 1.296
Example 4
[0083] This Example was conducted to determine if a transparent,
acidified calcium chloride extract of soy stays clear when
concentrated and desalted.
[0084] `a` g of toasted soy meal was added to `b` ml of 0.15 M
CaCl.sub.2 solution at ambient temperature and agitated for 30
minutes to provide an aqueous protein solution. The residual soy
meal was removed and the resulting protein solution was clarified
by centrifugation and filtration to produce `c` ml of filtered
protein solution having a protein content of `d` % by weight.
[0085] The filtered protein solution was then added to e' ml of
water and the pH of the sample lowered to 3 with diluted HCl.
[0086] The diluted and acidified protein extract solution was
reduced in volume to `f` ml by concentration on a `g` membrane
having a molecular weight cutoff of `h` Daltons and then an aliquot
of `i` ml of concentrated, acidified protein solution was
diafiltered with ml of reverse osmosis purified water. The
resulting acidified, diafiltered, concentrated protein solution had
a protein content of `k` % by weight and represented a yield of `l`
wt % of the initial filtered protein solution. The acidified,
diafiltered, concentrated protein solution was dried to yield a
product found to have a protein content of `m`% (N.times.6.25) w.b.
The product was termed S701 soy protein isolate (SPI).
[0087] The parameters `a` to `m` are set forth in the following
Table 9:
TABLE-US-00009 TABLE 9 a 240 b 1,000 c 480 d 1.13 e 960 f 28 g
Hydrosart h 10,000 i 26 j 260 k 11.24 l 68.27 m 93.61
[0088] 3.125 g of S701 SPI product was produced that dissolved well
in water. The colour, clarity and pH of a solution of the S701
sample provided herein and in others of the Examples were assessed
by the following procedure. Sufficient S701 product to supply 0.48
g of protein was dissolved in 15 ml of reverse osmosis purified
water and the solution colour and haze level determined using a
HunterLab Color Quest XE instrument operated in transmission mode.
The pH of the solution was then measured. The resulting
transparent, low pH (3.29) solution of S701 SPI was very light in
colour and had excellent clarity (Table 10).
TABLE-US-00010 TABLE 10 Colour and haze readings for solution of
S701 SPI from toasted soy meal in water sample L* a* b* haze (%)
S701 SPI 96.98 -0.97 9.69 3.1
Example 5
[0089] `a` g of dry soybeans were added to `b` ml of 0.10 M
CaCl.sub.2 solution at ambient temperature and processed for 5
minutes at the top speed of a kitchen blender to provide an aqueous
protein solution. The residual solids and the extracted fat were
removed and the resulting protein solution was clarified by
centrifugation and filtration to produce `c` ml of filtered protein
solution having a protein content of `d` % by weight.
[0090] The filtered protein solution was then added to e' ml of
water and the pH of the sample lowered to 3 with diluted HCl.
[0091] The diluted and acidified protein extract solution was
reduced in volume to `f` ml by concentration on a `g` membrane
having a molecular weight cutoff of `h` Daltons and then an aliquot
of `i` ml of concentrated, acidified protein solution was
diafiltered with ml of reverse osmosis purified water. The
resulting acidified, diafiltered, concentrated protein solution had
a protein content of `k` % by weight and represented a yield of `l`
wt % of the initial filtered protein solution. The acidified,
diafiltered, concentrated protein solution was dried to yield a
product found to have a protein content of `m`% (N.times.6.25) w.b.
The product was termed S701 soy protein isolate (SPI).
[0092] The parameters `a` to `m` are set forth in the following
Table 11:
TABLE-US-00011 TABLE 11 a 150 b 1,000 c 610 d 1.3 e 1,220 f 35 g
Hydrosart h 10,000 i 32 j 384 k 11.85 l 53.59 m 95.34
[0093] The process yielded 3.69 g of S701 SPI product that
dissolved well in water and produced a slightly hazy, low pH (3.19)
solution that was very light in colour (Table 12, below). For some
reason, the sample prepared from soybeans did not clear quite as
nicely as the sample prepared from meal when the pH was lowered to
3 and the sample concentrated. Perhaps this was the influence of
some residual oil that somehow escaped the filter press or else a
protein species that was not extractable from the toasted meal or
an effect of the differing strengths of calcium chloride used. Note
that the clarity of the initial diluted and acidified extract in
this Example was in line with the results achieved with soybeans in
Example 2. An additional pass through the filter press after pH
adjustment and prior to starting ultrafiltration or at some other
point later in the process likely would have yielded a product with
better clarity
TABLE-US-00012 TABLE 12 Colour and haze readings for solution of
S701 SPI from soybeans in water sample L* a* b* haze (%) S701 SPI
96.12 -0.57 8.87 21.3
Example 6
[0094] The procedure of Example 4 was scaled-up from bench top
laboratory scale to pilot plant scale.
[0095] `a` kg of toasted soy meal was added to `b` L of 0.15 M
CaCl.sub.2 solution at ambient temperature and agitated for 30
minutes to provide an aqueous protein solution. The residual soy
meal was removed and the resulting protein solution was clarified
by centrifugation and filtration to produce `c` L of filtered
protein solution having a protein content of `d` % by weight.
[0096] The filtered protein solution was then added to `e` L of
water and the pH of the sample lowered to 3.03 with diluted
HCl.
[0097] The diluted and acidified protein extract solution was
reduced in volume to `f` L by concentration on a `g` membrane
having a molecular weight cutoff of `h` Daltons. An aliquot of `i`
L of concentrated, acidified protein solution was diafiltered with
`j` L of reverse osmosis purified water then pasteurized at
60.degree. C. for 1 minute and filtered. The resulting acidified,
diafiltered, concentrated protein solution had a protein content of
`k` % by weight and represented a yield of `l` wt % of the initial
filtered protein solution. The acidified, diafiltered, concentrated
protein solution was dried to yield a product found to have a
protein content of `m`% (N.times.6.25) d.b. The product was termed
S001-H05-08A S701.
[0098] The parameters `a` to `m` are set forth in the following
Table 13:
TABLE-US-00013 TABLE 13 a 20 b 100 c 80 d 0.66 e 160 f 5 g
Polyethersulfone (PES) h 10,000 i 5 j 25 k 6.87 l 59.46 m
100.24
[0099] The phytic acid content was determined by the procedure of
Latta and Eskin (J. Agric. Food Chem., 28: 1313-1315) for the S701
sample provided herein and in others of the Examples. The phytic
acid content of the S001-H05-08A S701 was 0.00 wt % d.b.
[0100] 187 g of S701 was produced that dissolved well in water and
yielded a transparent, low pH (3.35) solution that was very light
in colour (Table 14, below).
TABLE-US-00014 TABLE 14 Colour and haze readings for solution of
S001-H05-08A S701 in water sample L* a* b* haze (%) S001-H05-08A
S701 95.65 -0.31 9.38 5.6
[0101] The dry powder was also very light in colour (Table 15,
below).
TABLE-US-00015 TABLE 15 Colour readings for dry S001-H05-08A S701
sample L* a* b* S001-H05-08A S701 87.59 +0.43 8.49
Example 7
[0102] This Example assesses the heat stability of soy protein
isolate prepared according to the procedure of Example 6.
[0103] A solution of S001-H05-08A S701, prepared as described in
Example 6, was prepared in water by dissolving sufficient
S001-H05-08A S701 to supply 1 g of protein in 50 ml of RO water.
The clarity of the sample was assessed by measuring the A600 and
the colour was measured using a HunterLab Colour Quest XE
instrument in transmission mode. The protein solution was then
heated to 95.degree. C. and held at this temperature for 30 seconds
before being cooled to room temperature rapidly in ice water. The
colour and clarity of the solution was then assessed again.
[0104] Heat treatment of the protein solution actually slightly
improved the clarity and had little effect on the colour of the
sample (Table 16, below). The maintenance of clarity under heat
treatment conditions is particularly beneficial for the use of the
protein in beverage systems, many of which are heated as part of
their processing.
TABLE-US-00016 TABLE 16 Clarity and colour results for solution of
S001-H05-08A S701 in water heat treated at 95.degree. C. for 30
seconds sample A600 L* a* b* haze (%) before heating 0.048 97.25
-0.24 6.25 8.1 after heating 0.038 97.25 -0.16 6.16 5.0
Example 8
[0105] This Example was performed to evaluate the effect of
extraction solution properties on the yield of protein extracted
from toasted soy meal and to determine if a high pH calcium
chloride extraction would generate a transparent solution at pH
3.
[0106] Samples of toasted soy meal (10 g) were combined with 100 ml
of the following solvents: [0107] Water [0108] Water plus
sufficient diluted NaOH to raise the pH of the extraction to 8.50
[0109] 0.05 M CaCl.sub.2 [0110] 0.10 M CaCl.sub.2 [0111] 0.15 M
CaCl.sub.2 [0112] 0.15 M CaCl.sub.2 plus sufficient diluted NaOH to
raise the pH of the extraction to 8.79 [0113] 0.05 M NaCl [0114]
0.10 MNaCl [0115] 0.15 MNaCl [0116] 0.15 M NaCl plus sufficient
diluted NaOH to raise the pH of the extraction to 8.64
[0117] The samples were mixed for 30 minutes using an orbital
shaker platform. Small samples were then clarified using a 0.45
.mu.m pore size syringe filter and the protein content of the
filtered solutions determined by Leco analysis. Small samples of
the clarified, high pH sodium chloride and calcium chloride
extracts were diluted with 2 parts water and the pH of the sample
adjusted to 3 with diluted HCl. The clarity of the samples was then
assessed visually.
[0118] Increasing the calcium chloride concentration appeared to
increase the amount of protein extracted from the meal (Table 17,
below). The number recorded for the 0.05M extract was extremely
low, likely due to experimental error in the measurement.
Increasing the concentration of sodium chloride in the extraction
had less of an impact in increasing the amount of protein
extracted. Conducting the extraction at high pH appeared to result
in a significant increase in the amount of protein extracted
regardless of the solvent type. The highest yield obtained was for
the 0.15M CaCl.sub.2 extraction at high pH. When this sample was
diluted with water a precipitate formed but this material
re-solubilized and the sample was completely clear when the pH was
lowered to 3. This suggested that the calcium treatment process to
produce a soy protein isolate, soluble and transparent at low pH
can be combined with an alkaline extraction to improve the yield if
desired. It was observed that the higher pH extraction samples were
a darker yellow colour than the natural pH extractions although
this may just be a function of the higher protein concentrations.
Dilution of the pH 8.64 sodium chloride extract with water did not
result in the formation of any haze or precipitate. Lowering the pH
of the sample to 3 did result in the formation of haze and
precipitation.
TABLE-US-00017 TABLE 17 Protein content of various clarified
extracts extraction solution % protein water 0.37 water, pH 8.50
0.48 0.05M CaCl.sub.2 0.03 0.10M CaCl.sub.2 0.38 0.15M CaCl.sub.2
0.47 0.15M CaCl.sub.2, pH 8.79 0.81 0.05M NaCl 0.25 0.10M NaCl 0.24
0.15M NaCl 0.32 0.15M NaCl, pH 8.64 0.68
Example 9
[0119] This Example illustrates extraction of another soy source
with water, sodium chloride and calcium chloride and the effect of
acidification on clarity.
[0120] Defatted, minimally heat processed soy flour (10 g) was
extracted with either water, 0.15 M NaCl or 0.15 M CaCl.sub.2 (100
ml) using a stir bar/stir plate for 30 minutes at room temperature.
The samples were then centrifuged at 10,200 g for 10 minutes to
separate extract from the residual solids. The supernatant was then
further clarified by filtration through 25 .mu.m pore size filter
paper and a 0.45 .mu.m pore size syringe filter then analyzed for
pH, conductivity, clarity (A600) and protein content (Leco). A
clarified sample of each extract was diluted into 4 parts room
temperature water and the A600 measured again. Diluted and
undiluted extract samples were acidified to pH 3 with diluted HCl
and the clarity measured again. Small amounts of CaCl.sub.2 were
also added to samples of the post 25 .mu.m water and sodium
chloride extracts and the conductivity measured. The mixtures were
then centrifuged at 7,800 g for 10 minutes and the supernatants
filtered with a 0.45 .mu.m pore size syringe filter. The pH,
protein content and A600 of these supernatants were measured and
then the pH was lowered to 3 and the A600 measured again.
[0121] After centrifugation, the supernatant from the calcium
chloride extraction appeared to be the clearest sample while the
supernatant from the sodium chloride extraction was a little bit
cloudy and the supernatant from the water extraction was very
cloudy. Even after filtering the sample, the water extract was
still hazy (Table 18, below). The extractability of the soy flour
was very good for all the extraction solutions, particularly the
water.
TABLE-US-00018 TABLE 18 Properties of initial extracts sample A600
% protein cond. (mS) water 0.285 3.53 4.25 0.15M NaCl 0.028 2.84
17.20 0.15M CaCl.sub.2 0.058 2.90 23.80
[0122] When the undiluted samples were acidified to pH 3, only the
calcium chloride extract stayed clear (Table 19, below). This
result indicates that the dilution step may not be necessary for
the generation of the soy protein product that is soluble and
produces transparent solutions at low pH.
TABLE-US-00019 TABLE 19 Effect of acidification on clarity of full
strength extracts sample initial pH final pH final A600 water 6.59
3.04 >3.0 0.15M NaCl 6.44 3.01 >3.0 0.15M CaCl.sub.2 5.44
3.04 0.060
[0123] When a dilution step was applied, again the calcium chloride
extract was the only sample that was clear at pH 3 (Table 20,
below).
TABLE-US-00020 TABLE 20 Effect of acidification on clarity of
diluted extracts sample initial pH initial A600 final pH final A600
water 6.72 0.028 2.90 0.860 0.15M NaCl 6.75 0.443 3.03 2.765 0.15M
CaCl.sub.2 5.66 2.827 2.96 0.032
[0124] Addition of calcium chloride to the water extract raised the
conductivity of the sample to 7.76 mS. The conductivity of the
sodium chloride extract was raised to 22.10 mS with calcium
chloride. Both samples contained significant amounts of precipitate
after the calcium chloride was added but were clarified by the
centrifugation and filtration steps. Significant amounts of protein
were lost in the clarification process, with the clarified
water/CaCl.sub.2 extract testing at 1.19% protein and the
NaCl/CaCl.sub.2 extract testing at 2.27% protein. The water extract
with the added calcium stayed clear upon acidification to pH 3
while the NaCl/CaCl.sub.2 extract went cloudy (Table 21, below).
Interestingly, both of these extract samples, if diluted with water
before the acidification, gave clear solutions at pH 3 (data not
shown). The water/CaCl.sub.2 sample was clear upon both dilution
and acid addition. The NaCl/CaCl.sub.2 sample precipitated upon
dilution but went clear when the pH was lowered to 3.
TABLE-US-00021 TABLE 21 Effect of acidification on clarity of
extracts with added calcium chloride sample initial pH initial A600
final pH final A600 water/CaCl.sub.2 5.69 0.014 3.04 0.062
NaCl/CaCl.sub.2 5.48 0.044 2.96 1.889
Example 10
[0125] This Example illustrates the production of soy protein
isolate on a pilot plant scale using organic soy flour purchased at
a bulk food store.
[0126] `a` kg of soy flour was added to `b` L of 0.15 M CaCl.sub.2
solution at ambient temperature and agitated for 30 minutes to
provide an aqueous protein solution. The residual soy flour and fat
phase was removed and the resulting protein solution was clarified
by centrifugation and filtration to produce `c` L of filtered
protein solution having a protein content of `d` % by weight.
[0127] The filtered protein solution was then added to `e` L of
water and the pH of the sample lowered to 3.05 with dilute HCl.
[0128] The diluted and acidified protein extract solution was
reduced in volume to `f` L by concentration on a `g` membrane
having a molecular weight cutoff of `h` Daltons. The concentrated,
acidified protein solution was diafiltered with `i` L of reverse
osmosis purified water. The resulting acidified, diafiltered,
concentrated protein solution had a protein content of `j` % by
weight and represented a yield of `k` wt % of the initial filtered
protein solution. The acidified, diafiltered, concentrated protein
solution was diluted with an equal volume of water and filtered.
The protein solution was then dried to yield a product found to
have a protein content of `l` wt % (N.times.6.25) d.b. and a phytic
acid content of `m` wt %. The product was termed S003-I18-08A
S701.
[0129] Parameters `a` to `m` are shown in the following Table
22:
TABLE-US-00022 TABLE 22 a 8.12 b 81 c 76 d 1.10 e 76 f 5 g PES h
10,000 i 25 j 12.73 k 73.1 l 103.01 m 0.01
[0130] When the S003-I18-08A S701 was dissolved in water, the
resulting solution (pH 3.33) was transparent and very light in
colour, as seen in the following Table 23.
TABLE-US-00023 TABLE 23 Colour and haze readings for solution of
S003-I18-08A S701 in water sample L* a* b* haze (%) S003-I18-08A
S701 96.74 -0.23 6.67 4.7%
[0131] The dry powder was also very light in colour as seen in the
following Table 24:
TABLE-US-00024 TABLE 24 Colour readings for dry S003-I18-08A S701
sample L* a* b* S003-I18-08A S701 88.03 +0.35 5.90
Example 11
[0132] This Example illustrates the production of soy protein
isolate on a pilot plant scale using defatted, minimally heat
processed soy flour.
[0133] `a` kg of soy flour was added to `b` L of 0.15 M CaCl.sub.2
solution at ambient temperature and agitated for 30 minutes to
provide an aqueous protein solution. The residual soy flour was
removed and the resulting protein solution was clarified by
centrifugation and filtration to produce `c` L of filtered protein
solution having a protein content of `d` % by weight.
[0134] The filtered protein solution was then added to `e` L of
water and the pH of the sample lowered to 3.01 with dilute HCl.
[0135] The diluted and acidified protein extract solution was
reduced in volume to `f` L by concentration on a `g` membrane
having a molecular weight cutoff of `h` Daltons. The concentrated,
acidified protein solution was diafiltered with `i` L of reverse
osmosis purified water. The resulting acidified, diafiltered,
concentrated protein solution had a protein content of `j` % by
weight and represented a yield of `k` wt % of the initial filtered
protein solution. The acidified, diafiltered, concentrated protein
solution was then dried to yield a product found to have a protein
content of `l` wt % (N.times.6.25) d.b. and a phytic acid content
of `m` wt % d.b. The product was termed S004-J02-08A S701.
[0136] Parameters `a` to `m` are shown in the following Table
25:
TABLE-US-00025 TABLE 25 a 10 b 100 c 94 d 1.26 e 94 f 7 g PES h
10,000 i 28 j 12.66 k 74.02 l 101.22 m 0.16
[0137] When the S004-J02-08A S701 isolate was dissolved in water,
the resulting solution (pH 3.09) was transparent and very light in
colour, as seen in the following Table 26:
TABLE-US-00026 TABLE 26 Colour and haze readings for solution of
S004-J02-08A S701 in water sample L* a* b* haze (%) S004-J02-08A
S701 97.92 -1.21 7.72 1.2
[0138] The dry powder was also very light in colour as seen in the
following Table 27:
TABLE-US-00027 TABLE 27 Colour readings for dry S004-J02-08A S701
sample L* a* b* S004-J02-08A S701 87.02 -0.82 10.32
Example 12
[0139] This Example illustrates the production of the novel, acid
soluble soy protein isolate (S701).
[0140] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 60 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0141] The filtered protein solution was then added to `e` L of
reverse osmosis purified water and the pH of the sample lowered to
`f` with diluted HCl.
[0142] The diluted and acidified protein extract solution was
reduced in volume to `g` L by concentration on a `h` membrane
having a molecular weight cutoff of `i` Daltons. The concentrated,
acidified protein solution was diafiltered with `j` L of reverse
osmosis purified water. The resulting acidified, diafiltered,
concentrated protein solution had a protein content of `k` % by
weight and represented a yield of `l` wt % of the initial filtered
protein solution. The acidified, diafiltered, concentrated protein
solution was then dried to yield a product found to have a protein
content of `m`% (N.times.6.25) d.b. The product was given
designation `n` S701.
[0143] The parameters `a` to `n` for three runs are set forth in
the following Table 28:
TABLE-US-00028 TABLE 28 Parameters for the runs to produce S701 n
S005-K18-08A S005-K24-08A S005-L08-08A a 60 60 20 b 600 600 200 c
410 360 170 d 2.63 2.53 2.03 e 410 360 170 f 3.07 3.07 3.06 g 70 81
49 h PES PES PES i 10,000 10,000 10,000 j 350 405 250 k 13.34 13.52
N/A l 89.6 91.1 N/A m 102.71 103.19 105.54 N/A = not available
[0144] When the S701 isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the following Table 28A.
TABLE-US-00029 TABLE 28A pH, colour and haze readings for solutions
of S701 in water Sample pH L* a* b* haze (%) S005-K18-08A S701 3.31
97.95 -1.33 8.27 1.4 S005-K24-08A S701 3.26 98.06 -1.35 8.10 0.3
S005-L08-08A S701 3.35 97.33 -1.00 8.03 1.0
Example 13
[0145] This Example illustrates the production of a soy protein
isolate by the protein micellar mass method.
[0146] 10 kg of defatted, minimally heat processed soy flour was
added to 200 L of 0.5 M NaCl solution at ambient temperature and
agitated for 60 minutes to provide an aqueous protein solution. The
residual soy flour was removed and the resulting protein solution
was clarified by centrifugation and filtration to produce 165 L of
filtered protein solution having a protein content of 1.34% by
weight.
[0147] The protein extract solution was reduced to 12.06 kg by
concentration on a PES membrane having a molecular weight cutoff of
100,000 Daltons, producing a concentrated protein solution with a
protein content of 17.51% by weight.
[0148] The concentrated solution at 30.degree. C. was diluted 1:5
into cold RO water having a temperature of 4.degree. C. A white
cloud formed immediately and was allowed to settle. The upper
diluting water was removed and the precipitated, viscous, sticky
mass (PMM) was recovered from the bottom of the vessel in a yield
of 20.8 wt % of the filtered protein solution. The dried PMM
derived protein was found to have a protein content of 99.66%
(N.times.6.25) d.b. The product was given a designation
S005-K19-08A S300.
Example 14
[0149] This Example contains an evaluation of the solubility in
water of the soy protein isolate produced by the method of Example
12 (S701), soy protein isolate produced by the PMM method of
Example 13 (S300) and two commercial soy protein isolates, namely
Pro Fam 825 and Pro Fam 873 (ADM, Decatur, Ill.), products
indicated by the manufacturer as being highly soluble. Solubility
was tested based on protein solubility (termed protein method, a
modified version of the procedure of Mon et al., J. Food Sci.
50:1715-1718) and total product solubility (termed pellet
method).
[0150] Sufficient protein powder to supply 0.5 g of protein was
weighed into a beaker and then a small amount of reverse osmosis
(RO) purified water was added and the mixture stirred until a
smooth paste formed. Additional water was then added to bring the
volume to approximately 45 ml. The contents of the beaker were then
slowly stirred for 60 minutes using a magnetic stirrer. The pH was
determined immediately after dispersing the protein and was
adjusted to the appropriate level (2, 3, 4, 5, 6 or 7) with diluted
NaOH or HCl. A sample was also prepared at natural pH. For the pH
adjusted samples, the pH was measured and corrected two times
during the 60 minutes stirring. After the 60 minutes of stirring,
the samples were made up to 50 ml total volume with RO water,
yielding a 1% protein w/v dispersion. The protein content of the
dispersions was measured using a Leco FP528 Nitrogen Determinator.
Aliquots (20 ml) of the dispersions were then transferred to
pre-weighed centrifuge tubes that had been dried overnight in a
100.degree. C. oven then cooled in a desiccator and the tubes
capped. The samples were centrifuged at 7,800 g for 10 minutes,
which sedimented insoluble material and yielded a clear
supernatant. The protein content of the supernatant was measured by
Leco analysis and then the supernatant and the tube lids were
discarded and the pellet material dried overnight in an oven set at
100.degree. C. The next morning the tubes were transferred to a
desiccator and allowed to cool. The weight of dry pellet material
was recorded. The dry weight of the initial protein powder was
calculated by multiplying the weight of powder used by a factor of
((100-moisture content of the powder (%))/100). Solubility of the
product was then calculated two different ways:
[0151] 1) Solubility (protein method) (%)=(% protein in
supernatant/% protein in initial dispersion).times.100
[0152] 2) Solubility (pellet method) (%)=(1-(weight dry insoluble
pellet material/((weight of 20 ml of dispersion/weight of 50 ml of
dispersion).times.initial weight dry protein
powder))).times.100
[0153] The natural pH values of the protein isolates produced in
Examples 12 and 13 and the commercial isolates in water are shown
in the following Table 29:
TABLE-US-00030 TABLE 29 Natural pH of dispersions prepared in water
at 1% protein w/v Batch Product Natural pH S005-K18-08A S701 3.21
S005-K24-08A S701 3.36 S005-L08-08A S701 3.35 S005-K19-08A S300
6.76 Pro Fam 825 7.23 Pro Fam 873 7.19
[0154] The solubility results obtained are set forth in the
following Tables 30 and 31:
TABLE-US-00031 TABLE 30 Solubility of products at different pH
values based on protein method Solubility (protein method) (%) Nat.
Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 pH S005- S701 97.1 99.1
100.0 1.0 26.2 94.4 98.0 K18-08A S005- S701 97.8 99.0 95.2 15.2
27.6 100.0 100.0 K24-08A S005- S701 100.0 100.0 100.0 4.2 28.6
100.0 100.0 L08-08A S005- S300 100.0 100.0 85.3 8.1 23.7 100.0 94.7
K19-08A Pro Fam 50.0 32.6 12.1 8.3 56.1 49.5 58.4 825 Pro Fam 57.4
31.1 23.2 13.5 29.9 42.9 45.2 873
TABLE-US-00032 TABLE 31 Solubility of products at different pH
values based on pellet method Solubility (pellet method) (%) Nat.
Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 pH S005- S701 100.0
100.0 100.0 24.3 37.5 99.0 97.1 K18-08A S005- S701 99.8 100.0 99.9
20.2 40.4 91.5 98.7 K24-08A S005- S701 100.0 100.0 100.0 66.8 72.4
99.7 100.0 L08-08A S005- S300 96.5 96.1 76.3 5.7 29.1 93.1 86.8
K19-08A Pro Fam 48.5 30.1 15.3 17.5 50.6 53.7 54.1 825 Pro Fam 49.7
30.9 18.4 18.0 36.6 42.7 43.1 873
[0155] As can be seen from the results of Tables 30 and 31, the
S701 products were far more soluble than the commercial isolates in
the range of pH 2 to 4 and also at pH 7 regardless of the
solubility testing method used. The excellent solubility in the low
pH range is a key factor in the applicability of the S701 product
for use in acidic beverages. The solubility of the S300 product
followed a similar pattern to the S701, except that the solubility
at pH 4 was not quite as good, although still better than the
commercial products.
Example 15
[0156] This Example contains an evaluation of the clarity in water
of the soy protein isolate produced by the method of Example 12
(S701), soy protein isolate produced by the PMM method of Example
13 (S300) and the commercial soy protein isolates Pro Fam 825 and
Pro Fam 873.
[0157] The clarity of the 1% protein w/v dispersions prepared as
described in Example 14 was assessed by measuring the absorbance of
visible light at 600 nm (A600) with water used to blank the
spectrophotometer. Analysis of the samples on a HunterLab
ColorQuest XE instrument in transmission mode also provided a
percentage haze reading, another measure of clarity. For both
tests, a lower score indicated greater clarity.
[0158] The clarity results are set forth in the following Tables 32
and 33:
TABLE-US-00033 TABLE 32 Clarity of solutions at different pH values
as assessed by A600 A600 Nat. Batch Product pH 2 pH 3 pH 4 pH 5 pH
6 pH 7 pH S005- S701 0.007 0.009 0.023 >3.0 >3.0 0.225 0.013
K18- 08A S005- S701 0.013 0.014 0.028 >3.0 >3.0 0.355 0.014
K24- 08A S005- S701 0.014 0.018 0.028 >3.0 >3.0 0.174 0.026
L08- 08A S005- S300 0.059 0.117 1.995 >3.0 >3.0 0.319 0.468
K19- 08A Pro 2.842 >3.0 >3.0 >3.0 2.944 2.891 2.879 Fam
825 Pro 2.765 2.907 >3.0 >3.0 2.875 2.824 2.806 Fam 873
TABLE-US-00034 TABLE 33 Clarity of solutions at different pH values
as assessed by Hunter Lab analysis Hunter Lab haze reading (%) Nat.
Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 pH S005- S701 N/A N/A
N/A N/A N/A N/A N/A K18-08A S005- S701 0.0 0.0 0.2 94.5 94.4 47.0
0.0 K24-08A S005- S701 0.0 0.0 0.0 93.5 93.3 20.2 0.0 L08-08A S005-
S300 5.8 16.9 92.4 93.4 93.4 40.2 54.1 K19-08A Pro Fam N/A N/A N/A
N/A N/A N/A N/A 825 Pro Fam 95.1 95.4 95.6 95.7 95.6 95.3 95.3 873
N/A = not available
[0159] As can be seen from the results of Tables 32 and 33,
solutions of S701 prepared in the pH range 2 to 4 were extremely
clear, regardless of the method used to assess clarity. The
commercial isolates were extremely cloudy at all pH values tested.
The S300 was fairly clear at pH 2 to 3 but not as sharp as the S701
solutions. At pH 4, the S300 solution was quite cloudy. Solutions
of S701 and S300 were clearer than the commercial isolates at pH 7,
but the solutions at this pH were not nearly as clear as the acidic
solutions.
Example 16
[0160] This Example contains an evaluation of the heat stability in
water of the soy protein isolates produced by the method of Example
12 (S701). Solutions of S701 were produced by dissolving sufficient
S701 to supply 0.8 g of protein in 40 ml of water. The pH was then
adjusted to 3 if necessary. The clarity of these solutions was
assessed by haze measurement with the HunterLab Color Quest XE
instrument. The solutions were then heated to 95.degree. C., held
at this temperature for 30 seconds and then immediately cooled to
room temperature in an ice bath. The clarity of the heat treated
solutions was then measured again.
[0161] The clarity of the protein solutions before and after
heating is set forth in the following Table 34:
TABLE-US-00035 TABLE 34 Effect of heat treatment on clarity of
solutions haze (%) before haze (%) after batch product heating
heating S005-K18-08A S701 0.0 0.0 S005-K24-08A S701 0.0 0.0
S005-L08-08A S701 0.0 0.0
[0162] As can be seen from the results in Table 34, the solutions
of S701 were initially completely clear and remained so after heat
treatment.
Example 17
[0163] This Example contains an evaluation of the solubility in a
soft drink (Sprite) and sports drink (Orange Gatorade) of the soy
protein isolate produced by the method of Example 12 (S701), soy
protein isolate produced by the PMM method of Example 13 (S300) and
the commercial soy protein isolates Pro Fam 825 and Pro Fam 873.
The solubility was determined with the protein added to the
beverages with no pH correction and again with the pH of the
protein fortified beverages adjusted to the level of the original
beverages.
[0164] When the solubility was assessed with no pH correction, a
sufficient amount of protein powder to supply 1 g of protein was
weighed into a beaker and a small amount of beverage was added and
stirred until a smooth paste formed. Additional beverage was added
to bring the volume to 50 ml, and then the solutions were stirred
slowly on a magnetic stirrer for 60 minutes to yield a 2% protein
w/v dispersion. The protein content of the samples was analyzed
using a LECO FP528 Nitrogen Determinator then an aliquot of the
protein containing beverages was centrifuged at 7,800 g for 10
minutes and the protein content of the supernatant measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0165] When the solubility was assessed with pH correction, the pH
of the soft drink (Sprite) (3.39) and sports drink (Orange
Gatorade) (3.19) without protein was measured. A sufficient amount
of protein powder to supply 1 g of protein was weighed into a
beaker and a small amount of beverage was added and stirred until a
smooth paste formed. Additional beverage was added to bring the
volume to approximately 45 ml, and then the solutions were stirred
slowly on a magnetic stirrer for 60 minutes. The pH of the protein
containing beverages was measured and then adjusted to the original
no-protein pH with HCl or NaOH as necessary. The total volume of
each solution was then brought to 50 ml with additional beverage,
yielding a 2% protein w/v dispersion. The protein content of the
samples was analyzed using a LECO FP528 Nitrogen Determinator then
an aliquot of the protein containing beverages was centrifuged at
7,800 g for 10 minutes and the protein content of the supernatant
measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0166] The results obtained are set forth in the following Table
35:
TABLE-US-00036 TABLE 35 Solubility of products in Sprite and Orange
Gatorade no pH correction pH correction Solubility (%) Solubility
(%) Solubility (%) in Orange Solubility (%) in Orange Batch Product
in Sprite Gatorade in Sprite Gatorade S005-K18-08A S701 100.0 98.0
100.0 91.7 S005-K24-08A S701 98.9 100 97.1 100 S005-L08-08A S701
100.0 93.4 100.0 100 S005-K19-08A S300 4.8 71.0 95.3 85.2 Pro Fam
825 5.5 19.0 26.6 33.0 Pro Fam 873 12.1 16.4 23.2 26.5
[0167] As can be seen from the results of Table 35, the S701
product was extremely soluble in the Sprite and the Orange
Gatorade. The S701 is an acidified product and had little effect on
the pH of the beverages. The S300 and the commercial isolates were
not acidified products. The solubility of these products was
somewhat improved by correcting the pH of the beverages. However,
even after pH correction the commercial isolates were far less
soluble than the S701. The S300 was slightly less soluble than the
S701 after pH correction.
Example 18
[0168] This Example contains an evaluation of the clarity in a soft
drink and sports drink of the soy protein isolate produced by the
method of Example 12 (S701), soy protein isolate produced by the
PMM method of Example 13 (S300) and commercial soy protein isolates
Pro Fam 825 and Pro Fam 873.
[0169] The clarity of the 2% protein w/v dispersions prepared in
soft drink (Sprite) and sports drink (Orange Gatorade) in Example
17 was assessed using the methods described in Example 15 but with
the appropriate beverage used to blank the spectrophotometer for
the absorbance measurements at 600 nm.
[0170] The results obtained are set forth in the following Tables
36 and 37:
TABLE-US-00037 TABLE 36 Clarity (A600) of products in Sprite and
Orange Gatorade no pH correction pH correction A600 in A600 in
Orange A600 in A600 in Orange Batch Product Sprite Gatorade Sprite
Gatorade S005-K18-08A S701 0.017 0.000 0.016 0.000 S005-K24-08A
S701 0.017 0.000 0.007 0.000 S005-L08-08A S701 0.030 0.000 0.035
0.010 S005-K19-08A S300 >3.0 >3.0 1.339 1.028 Pro Fam 825
>3.0 2.972 >3.0 >3.0 Pro Fam 873 >3.0 2.961 >3.0
>3.0
TABLE-US-00038 TABLE 37 HunterLab haze readings for products in
Sprite and Orange Gatorade no pH correction pH correction haze (%)
in haze (%) in haze (%) in haze (%) in Batch Product Sprite Orange
Gatorade Sprite Orange Gatorade no protein 0.0 44.0 0.0 44.0
S005-K18-08A S701 0.0 38.5 5.4 47.6 S005-K24-08A S701 0.0 39.7 0.0
41.4 S005-L08-08A S701 0.0 40.8 8.4 48.6 S005-K19-08A S300 93.6
93.5 94.9 86.3 Pro Fam 825 93.3 93.7 90.8 91.4 Pro Fam 873 93.4
94.2 90.9 91.9
[0171] As can be seen from the results of Tables 36 and 37, the
S701 product had minimal effect on the clarity of the Sprite and
Orange Gatorade. Addition of the commercial isolates and the S300
made these beverages very cloudy, even after pH correction.
Example 19
[0172] This Example contains an evaluation of the solubility in
alcoholic beverages of the soy protein isolate produced by the
method of Example 12 (S701), soy protein isolate produced by the
PMM method of Example 13 (S300) and commercial soy protein isolates
Pro Fam 825 and Pro Fam 873. The solubility was determined with the
protein added to the beverages with no pH correction and again with
the pH of the protein fortified beverages adjusted to the level of
the original beverages.
[0173] When the solubility was assessed with no pH correction, a
sufficient amount of protein powder to supply 1 g of protein was
weighed into a beaker and a small amount of beverage was added and
stirred until a smooth paste formed. Additional beverage was added
to bring the volume to 50 ml, and then the solutions were stirred
slowly on a magnetic stirrer for 60 minutes to yield a 2% protein
w/v dispersion. The protein content of the samples was analyzed
using a LECO FP528 Nitrogen Determinator then an aliquot of the
protein containing beverages was centrifuged at 7,800 g for 10
minutes and the protein content of the supernatant measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0174] When the solubility was assessed with pH correction, the pH
of the Miller Genuine Draft beer (4.05), Bacardi Breezer Strawberry
Daiquiri (3.60) and Pomtini Vodka and Pomegranate Cooler (3.36)
without protein was measured. A sufficient amount of protein powder
to supply 1 g of protein was weighed into a beaker and a small
amount of beverage was added and stirred until a smooth paste
formed. Additional beverage was added to bring the volume to 50 ml,
and then the solutions were stirred slowly on a magnetic stirrer
for 60 minutes to yield a 2% protein w/v dispersion. The pH of the
protein containing beverages was measured and then adjusted to the
original no-protein pH with HCl or NaOH as necessary. The protein
content of the samples was analyzed using a LECO FP528 Nitrogen
Determinator then an aliquot of the protein containing beverages
was centrifuged at 7,800 g for 10 minutes and the protein content
of the supernatant measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0175] The results obtained are set forth in the following Tables
38 and 39:
TABLE-US-00039 TABLE 38 Solubility of products in alcoholic
beverages with no pH correction Solubility Solubility Solubility
(%) Miller (%) in (%) in Genuine Bacardi Pomtimi Batch Product
Draft Beer Breezer Cooler S005-K18-08A S701 98.6 100 98.9
S005-K24-08A S701 100 100 99.0 S005-L08-08A S701 100 100 100
S005-K19-08A S300 18.9 25.8 32.2 Pro Fam 825 30.1 14.8 22.3 Pro Fam
873 35.0 23.3 26.4
TABLE-US-00040 TABLE 39 Solubility of products in alcoholic
beverages with pH correction Solubility Solubility Solubility (%)
Miller (%) in (%) in Genuine Bacardi Pomtimi Batch Product Draft
Beer Breezer Cooler S005-K18-08A S701 97.2 98.9 95.3 S005-K24-08A
S701 100 98.3 97.9 S005-L08-08A S701 99.4 98.3 100 S005-K19-08A
S300 33.3 63.3 73.7 Pro Fam 825 22.4 26.1 16.0 Pro Fam 873 23.3
34.2 22.0
[0176] As can be seen from the results of Tables 38 and 39, the
S701 product was extremely soluble in the alcoholic beverages. As
the S701 is an acidified product, its addition did not alter the pH
of the beverages as much as the neutral S300 and commercial
isolates. The solubility of the S300 was somewhat improved by
correcting the pH of the beverages, but was still notably poorer
than the solubility of the S701. The commercial isolates were
poorly soluble regardless of whether the pH of the beverages was
corrected.
Example 20
[0177] This Example contains an evaluation of the clarity and heat
stability in alcoholic beverages of the soy protein isolate
produced by the method of Example 12 (S701), soy protein isolate
produced by the PMM method of Example 13 (S300) and commercial soy
protein isolates Pro Fam 825 and Pro Fam 873.
[0178] The clarity of the protein dispersions prepared in alcoholic
beverages in Example 19 was assessed using the methods described in
Example 15 but with the appropriate beverage used to blank the
spectrophotometer for the absorbance measurements at 600 nm. Heat
stability was assessed by heating an aliquot of the protein
containing alcoholic beverages to 95.degree. C. and holding the
samples at this temperature for 30 seconds. The samples were then
immediately cooled to room temperature in an ice bath and the
clarity measured again. The appropriate unheated, no-protein
beverage was used to blank the spectrophotometer for the absorbance
measurements at 600 nm.
[0179] The results obtained are set forth in the following Tables
40 to 43:
TABLE-US-00041 TABLE 40 Clarity (A600) of products in alcoholic
beverages before and after heat treatment (no pH correction) Miller
Genuine Draft Bacardi Breezer Pomtini Cooler A600 before A600 after
A600 before A600 after A600 before A600 after Batch Product heating
heating heating heating heating heating S005-K18-08A S701 0.032
0.002 0.117 0.096 0.163 0.089 S005-K24-08A S701 0.031 0.010 0.065
0.091 0.187 0.092 S005-L08-08A S701 0.056 0.021 0.095 0.100 0.203
0.093 S005-K19-08A S300 >3.0 >3.0 >3.0 >3.0 >3.0
>3.0 Pro Fam 825 >3.0 >3.0 >3.0 >3.0 >3.0 >3.0
Pro Fam 873 >3.0 >3.0 >3.0 >3.0 >3.0 >3.0
TABLE-US-00042 TABLE 41 Clarity (A600) of products in alcoholic
beverages before and after heat treatment (with pH correction)
Miller Genuine Draft Bacardi Breezer Pomtini Cooler A600 before
A600 after A600 before A600 after A600 before A600 after Batch
Product heating heating heating heating heating heating
S005-K18-08A S701 0.082 0.071 0.076 0.036 0.208 0.160 S005-K24-08A
S701 0.035 0.034 0.059 0.045 0.213 0.150 S005-L08-08A S701 0.369
0.302 0.098 0.056 0.251 0.178 S005-K19-08A S300 >3.0 >3.0
2.444 1.830 2.498 0.707 Pro Fam 825 >3.0 >3.0 >3.0 >3.0
>3.0 >3.0 Pro Fam 873 >3.0 >3.0 >3.0 >3.0 >3.0
>3.0
TABLE-US-00043 TABLE 42 Hunter Lab haze readings for products in
alcoholic beverages before and after heat treatment (no pH
correction) Miller Genuine Draft Bacardi Breezer Pomtini Cooler
haze (%) haze (%) haze (%) haze (%) haze (%) haze (%) before after
before after before after Batch Product heating heating heating
heating heating heating no protein 0.0 N/A 29.1 N/A 17.2 N/A
S005-K18-08A S701 1.9 0.0 33.4 25.5 19.8 10.4 S005-K24-08A S701 5.4
0.7 30.3 24.9 23.5 12.4 S005-L08-08A S701 6.1 1.4 33.8 26.5 23.6
12.7 S005-K19-08A S300 93.3 93.2 94.1 94.0 95.3 93.7 Pro Fam 825
93.0 92.7 94.0 94.9 97.6 96.3 Pro Fam 873 93.2 93.1 94.4 94.3 95.1
94.8 N/A = not available
TABLE-US-00044 TABLE 43 Hunter Lab haze readings for products in
alcoholic beverages before and after heat treatment (with pH
correction) Miller Genuine Draft Bacardi Breezer Pomtini Cooler
haze (%) haze (%) haze (%) haze (%) haze (%) before haze (%) before
after before after Batch Product heating after heating heating
heating heating heating no protein 0.3 N/A 25.9 N/A N/A N/A
S005-K18-08A S701 20.0 18.1 33.5 25.6 N/A N/A S005-K24-08A S701 7.3
7.2 31.9 29.7 N/A N/A S005-L08-08A S701 20.6 14.0 35.2 31.2 N/A N/A
S005-K19-08A S300 97.0 96.3 96.7 95.6 N/A 81.9 Pro Fam 825 96.9
96.9 96.9 97.1 N/A 98.7 Pro Fam 873 97.0 97.1 97.2 96.9 N/A 99.6
N/A = not available
[0180] As can be seen from the results of Tables 40 to 43, addition
of S701 had little effect on the clarity of the alcoholic
beverages. However, beverages containing the commercial soy protein
isolates and the S300 were very cloudy. Heat treatment did not
reduce the clarity of the alcoholic beverages containing S701, and
in many cases slightly improved it. The beverages containing the
commercial isolates and the S300 stayed cloudy after heat
treatment.
Example 21
[0181] This Example contains an evaluation of the content of
specific elements in the soy protein isolate produced by the method
of Example 12 (S701), soy protein isolate produced by the PMM
method of Example 13 (S300) and commercial soy protein isolate Pro
Fam 825.
[0182] The detection of the elements calcium, phosphorus,
magnesium, potassium, sodium, iron, copper, zinc and manganese was
performed by plasma emission spectroscopy.
[0183] The results obtained are set forth in the following Table
44:
TABLE-US-00045 TABLE 44 Content of specific elements in protein
products % dry weight basis ppm Batch Product Ca P Mg K Na Fe Cu Zn
Mn S005-K18-08A S701 0.03 0.03 0.01 0.04 0.04 0.003 16 8.14 1
S005-K24-08A S701 0.08 0.04 0.01 0.02 0.04 0.004 23 5.42 3
S005-L08-08A S701 0.12 0.06 0.01 0.01 0.02 0.007 22 4.76 2
S005-K19-08A S300 0.16 0.27 0.07 0.16 1.28 0.01 31 37.77 35 Pro Fam
825 0.08 0.90 0.04 0.93 0.96 0.01 13 47.87 10
[0184] As may be seen from the results of Table 44, the content of
the elements of interest was generally lower in the S701 products
than in the S300 or the commercial isolate. The S701 products were
particularly low in phosphorus, potassium, sodium, zinc and
manganese compared to the other isolates.
Example 22
[0185] This Example contains an evaluation of the phytic acid
content of the soy protein isolate produced by the method of
Example 12 (S701), soy protein isolate produced by the PMM method
of Example 13 (S300) and commercial soy protein isolates Pro Fam
825 and 873.
[0186] Phytic acid content was determined using the method of Latta
and Eskin (J. Agric. Food Chem., 28: 1313-1315).
[0187] The results obtained are set forth in the following Table
45:
TABLE-US-00046 TABLE 45 Phytic acid content of protein products
Batch Product % Phytic acid d.b. S005-K18-08A S701 0.00
S005-K24-08A S701 0.02 S005-L08-08A S701 0.00 S005-K19-08A S300
0.62 Pro Fam 825 2.00 Pro Fam 873 1.53
[0188] As may be seen from the results in Table 45, the S701
samples were extremely low in phytic acid, while the S300 and the
commercial isolates contained significant levels of phytic
acid.
Example 23
[0189] This Example contains an evaluation of the solubility in a
reconstituted sports drink (Orange Gatorade powder) of the soy
protein isolate produced by the method of Example 12 (S701), soy
protein isolate produced by the PMM method of Example 13 (S300) and
commercial soy protein isolates Pro Fam 825 and Pro Fam 873. The
solubility was determined with the protein and beverage powder dry
blended and then dissolved in water with no pH correction and again
with the pH of the reconstituted protein/beverage mix adjusted to
the level of the powdered beverage reconstituted without
protein.
[0190] From the preparation instructions on the container of Orange
Gatorade powder, it was determined that 6.68 g of powder was
required to make 100 ml of the beverage. A sufficient amount of
protein powder to provide 2 g of protein was weighed into a 250 ml
beaker then Orange Gatorade powder (6.68 g) was added and the
mixture dry blended by swirling the beaker. Reverse osmosis (RO)
purified water (100 ml) was added to the protein-Gatorade mixture
and the sample was stirred slowly on a stir plate for 60 minutes.
When the samples were evaluated with pH correction, the pH of the
reconstituted Gatorade/protein beverage was adjusted to 3.17 (the
pH of reconstituted Orange Gatorade powder without protein) with
HCl or NaOH as necessary. The protein content of the samples was
analyzed using a LECO FP528 Nitrogen Determinator then an aliquot
of the beverages was centrifuged at 7,800 g for 10 minutes and the
protein content of the supernatant measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0191] The results obtained are set forth in the following Table
46:
TABLE-US-00047 TABLE 46 Solubility of protein products
reconstituted with Orange Gatorade powder no pH correction with pH
correction Batch Product Solubility (%) Solubility (%)
BW-S005-K18-08A S701 94.7 100 BW-S005-K24-08A S701 97.4 100
BW-S005-L08-08A S701 97.9 100 BW-S005-K19-08A S300 49.5 94.4
Pro-Fam 825 12.9 13.1 Pro-Fam 873 14.0 12.7
[0192] As can be seen in the results of Table 46, the S701 products
were extremely soluble with the Gatorade powder. The S701 was an
acidified product and so had little effect on the pH of the
reconstituted beverage. The S300 was not an acidified product and
was very soluble only when the pH was corrected. The commercial soy
isolates were also not acidified products but were poorly soluble
with the Gatorade powder whether or not the pH was corrected.
Example 24
[0193] This Example contains an evaluation of the clarity in a
reconstituted sports drink (Orange Gatorade powder) of the soy
protein isolate produced by the method of Example 12 (S701), soy
protein isolate produced by the PMM method of Example 13 (S300) and
commercial soy protein isolates Pro Fam 825 and Pro Fam 873. The
clarity was determined with the protein and beverage powder dry
blended and then dissolved in water with no pH correction and again
with the pH of the reconstituted protein/beverage mix adjusted to
the level of the powdered beverage reconstituted without
protein.
[0194] The clarity of the protein dispersions prepared in
reconstituted sports drink (Orange Gatorade powder) in Example 23
was assessed using the methods described in Example 15 but with the
appropriate beverage used to blank the spectrophotometer for the
absorbance measurements at 600 nm.
[0195] The results obtained are set forth in the following Table
47:
TABLE-US-00048 TABLE 47 Clarity of protein products reconstituted
with Orange Gatorade powder no pH correction with pH correction
Batch Product A600 haze (%) A600 haze (%) no protein 0.000 31.7
0.000 31.7 BW-S005-K18-08A S701 0.040 32.6 0.001 32.0
BW-S005-K24-08A S701 0.043 33.3 0.006 33.8 BW-S005-L08-08A S701
0.077 37.4 0.038 37.1 BW-S005-K19-08A S300 >3.0 94.0 1.23 83.9
Pro-Fam 825 >3.0 94.3 >3.0 97.1 Pro-Fam 873 >3.0 94.4
>3.0 97.3
[0196] As may be seen by the results of Table 47, the S701 products
had little effect on the haze level in the reconstituted Orange
Gatorade powder. The reconstituted Orange Gatorade powder with S300
and the commercial isolates was very hazy regardless of whether the
pH was corrected.
Example 25
[0197] This Example contains an evaluation of the solubility in a
reconstituted soft drink (Raspberry Ice Crystal Light powder) of
the soy protein isolate produced by the method of Example 12
(S701), soy protein isolate produced by the PMM method of Example
13 (S300) and commercial soy protein isolates Pro Fam 825 and Pro
Fam 873. The solubility was determined with the protein and
beverage powder dry blended and then dissolved in water with no pH
correction and again with the pH of the reconstituted
protein/beverage mix adjusted to the level of the powdered beverage
reconstituted without protein.
[0198] From the preparation instructions on the package of
Raspberry Ice Crystal Light powder, it was determined that 0.53 g
of powder was required to make 100 ml of the beverage. A sufficient
amount of protein powder to provide 2 g of protein was weighed into
a 250 ml beaker then Raspberry Ice Crystal Light powder (0.53 g)
was added and the mixture dry blended by swirling the beaker. RO
water (100 ml) was added to the protein-Crystal Light mixture and
the sample was stirred slowly on a stir plate for 60 minutes. When
the samples were evaluated with pH correction, the pH of the
reconstituted Crystal Light/protein beverage was adjusted to 3.27
(the pH of reconstituted Raspberry Ice Crystal Light powder without
protein) with HCl or NaOH as necessary. The protein content of the
samples was analyzed using a LECO FP528 Nitrogen Determinator then
an aliquot of the beverages was centrifuged at 7,800 g for 10
minutes and the protein content of the supernatant measured.
Solubility (%)=(% protein in supernatant/% protein in initial
dispersion).times.100
[0199] The results obtained are set out in the following Table
48:
TABLE-US-00049 TABLE 48 Solubility of protein products
reconstituted with Raspberry Ice Crystal Light powder no pH
correction with pH correction Batch Product Solubility (%)
Solubility (%) BW-S005-K18-08A S701 100 97.2 BW-S005-K24-08A S701
99.0 96.0 BW-S005-L08-08A S701 97.9 100 BW-S005-K19-08A S300 8.5
92.3 Pro-Fam 825 11.9 23.3 Pro-Fam 873 14.8 19.2
[0200] As can be seen from the results in Table 48, the S701
products were extremely soluble when reconstituted with the Crystal
Light powder. The S701 was an acidified product and so had little
effect on the pH of the reconstituted beverage. The S300 was not an
acidified product and was very soluble only when the pH was
corrected. The commercial soy isolates were not acidified products
but were poorly soluble when reconstituted with the Crystal Light
powder whether or not the pH was corrected.
Example 26
[0201] This Example contains an evaluation of the clarity in a
reconstituted soft drink (Raspberry Ice Crystal Light powder) of
the soy protein isolate produced by the method of Example 12
(S701), soy protein isolate produced by the PMM method of Example
13 (S300) and commercial soy protein isolates Pro Fam 825 and Pro
Fam 873. The clarity was determined with the protein and beverage
powder dry blended and then dissolved in water with no pH
correction and again with the pH of the reconstituted
protein/beverage mix adjusted to the level of the powdered beverage
reconstituted without protein.
[0202] The clarity of the protein dispersions prepared in
reconstituted soft drink (Raspberry Ice Crystal Light powder) in
Example 25 was assessed using the methods described in Example 15
but with the appropriate beverage used to blank the
spectrophotometer for the absorbance measurements at 600 nm.
[0203] The results obtained are set forth in the following Table
49:
TABLE-US-00050 TABLE 49 Clarity of protein products reconstituted
with Raspberry Ice Crystal Light powder no pH correction with pH
correction Batch Product A600 haze (%) A600 haze (%) no protein
0.000 0.3 0.000 0.3 BW-S005-K18-08A S701 0.003 0.5 0.000 2.8
BW-S005-K24-08A S701 0.000 0.7 0.000 2.8 BW-S005-L08-08A S701 0.026
4.6 0.000 4.7 BW-S005-K19-08A S300 >3.0 100.1 1.296 81.7 Pro-Fam
825 >3.0 95.8 >3.0 97.4 Pro-Fam 873 >3.0 96.9 >3.0
98.7
[0204] As may be seen by the results of Table 49, the S701 products
had little effect on the clarity of the reconstituted Raspberry Ice
Crystal Light. The reconstituted Raspberry Ice Crystal Light with
S300 and the commercial isolates were very hazy regardless of
whether the pH was corrected.
Example 27
[0205] This Example describes the production of novel soy protein
products having a protein content of less than 90% by weight
(N.times.6.25) d.b. in accordance with one embodiment of the
invention.
[0206] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of `c` M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy flour was removed and the resulting protein
solution was partially clarified by centrifugation to produce `d` L
of partially clarified protein solution having a protein content of
`e` % by weight. The partially clarified protein solution was
diluted with 1 volume of water and adjusted to pH 3 with HCl.
[0207] The diluted and acidified protein solution was further
clarified by filtration, providing `f` L of solution having a
protein content of `g` by weight.
[0208] The `h` L of protein solution was reduced in volume to `i` L
by concentration on a polyvinylidene fluoride (PVDF) membrane
having a molecular weight cutoff of `j` Daltons. At volume
reduction factors (VRF) 5, 7, and 10, 200 ml samples of the
retentate were taken and freeze dried.
[0209] The parameters `a` to `j` are set forth in the following
Table 50:
TABLE-US-00051 TABLE 50 a 20 b 200 c 0.15 d 138 e 2.57 f 304 g 1.20
h 304 i 28 j 5,000
[0210] Dried samples were analyzed for protein content using a Leco
FP 528 Nitrogen Determinator and moisture content using an oven
drying method. Samples of the soy protein products, sufficient to
supply 0.48 g of protein, were then dissolved in water (15 ml) and
the pH adjusted to 3 as necessary. The colour and clarity of the
solutions was measured using a HunterLab ColorQuest XE instrument.
The results obtained are set forth in Table 51 below.
TABLE-US-00052 TABLE 51 Protein content of dry products and colour
and haze readings for solutions of soy protein product in water %
protein Colour Analysis (N .times. 6.25) d.b. L* a* b* Haze (%) VRF
5 73.0% 94.88 -1.54 13.31 6.6% VRF 7 78.1% 94.43 -1.55 14.31 5.1%
VRF 10 82.4% 94.29 -1.50 14.50 7.0%
[0211] As can be seen from the results in Table 51, the partially
purified soy protein products gave solutions of acceptable colour
and very good clarity when solubilized in water at pH 3.
Example 28
[0212] This Example describes the production of a novel soy protein
product having a protein content of less than 90% by weight
(N.times.6.25) d.b. in accordance with one embodiment of the
invention
[0213] `a` g of defatted, minimally heat processed soy flour was
added to `b` ml of `c` M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy flour was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`d` ml of clarified protein solution having a protein content of e'
% by weight.
[0214] An `f` ml aliquot of filtrate was diluted with 1 volume of
water and adjusted to pH 3 with HCl. The resulting `g` ml solution
having a protein content of `h` % by weight was then concentrated
on a PES membrane having a molecular weight cutoff of `i` Daltons.
At volume reduction factors of 1.25, 2 and at the final
concentration point, samples of the retentate were taken and freeze
dried. Another sample of retentate was also taken after 2 volumes
of diafiltration (DF) with water and freeze dried.
[0215] The parameters `a` to `i` for the run are set forth in the
following Table 52:
TABLE-US-00053 TABLE 52 a 100 b 1,000 c 0.15 d 800 e 2.91 f 500 g
1,000 h 1.20 i 10,000
[0216] Dried samples were analyzed for protein content using a Leco
FP 528 Nitrogen Determinator and moisture content using an oven
drying method. Samples of the soy protein products, sufficient to
supply 0.48 g of protein, were then dissolved in water (15 ml) and
the pH adjusted to 3 as necessary. The colour and clarity of the
solutions was measured using a HunterLab ColorQuest XE instrument.
The results obtained are set forth in Table 53 below:
TABLE-US-00054 TABLE 53 Protein content of dry products and colour
and haze readings for solutions of soy protein product in water %
protein Colour Analysis (N .times. 6.25) d.b. L* a* b* Haze (%) VRF
1.25 50.6% 98.58 -1.66 6.90 0.0 VRF 2 57.7% 98.46 -1.66 7.19 0.0
end retentate 81.6% 98.28 -1.59 7.65 0.0 before DF end retentate
after 90.4% 98.42 -1.45 6.96 0.0 DF (2 vol.)
[0217] As can be seen from the results in Table 53, the partially
purified soy protein products gave solutions of very good colour
and clarity when solubilized in water at pH 3.
Example 29
[0218] This Example illustrates the effect of membrane type and
pore size on the trypsin inhibitor activity of the novel, acid
soluble soy protein isolate (S701) provided herein.
[0219] Protein content determinations were performed using a Leco
FP528 Nitrogen Determinator. Trypsin inhibitor activity was
determined using the method of Kakade et al. Cereal Chem.,
51:376-381 (1974), modified to initially solubilize the sample in
water.
[0220] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0221] The filtered protein solution was then added to e' volume(s)
of reverse osmosis purified water and the pH of the sample lowered
to 3.01 with diluted HCl. The diluted and acidified filtrate was
not heat treated.
[0222] The diluted and acidified protein extract solution was
reduced in volume from `f` L to `g` L by concentration on a `h`
membrane, having a molecular weight cutoff of `i` Daltons, operated
at a temperature of approximately `j`.degree. C. The acidified
protein solution, with a protein content of `k` wt %, was
diafiltered with `l` L of reverse osmosis purified water, with the
diafiltration operation conducted at approximately `m`.degree. C.
The resulting diafiltered protein solution was then further
concentrated to provide a solution with a protein content of `n` %
by weight which represented a yield of `o` wt % of the initial
filtered protein solution. The acidified, diafiltered, concentrated
protein solution was then dried to yield a product found to have a
protein content of `p` wt % (N.times.6.25) d.b. and a phytic acid
content of `q` wt % d.b. The dry product was found to have a
trypsin inhibitor activity of `r` trypsin inhibitor units/mg
protein (N.times.6.25). The product was given designation `s`
S701.
[0223] The parameters `a` to `s` for two runs are set forth in the
following Table 54:
TABLE-US-00055 TABLE 54 Parameters for the runs to produce S701 in
this Example s S008-D01-09A S008-D02-09A a 60 60 b 600 600 c 420
450 d 2.41 2.87 e 1 1 f 840 900 g 136 180 h PES PES i 100,000
10,000 j 30 30 k 7.55 5.93 l 400 600 m 31 30 n 15.28 15.13 o 82.3
73.5 p 100.74 100.17 q 0.00 0.00 r 82 120
[0224] As can be seen from the data in Table 54, the isolate
produced using the membrane with the larger pore size had a lower
trypsin inhibitor activity.
[0225] When the S701 isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 54A:
TABLE-US-00056 TABLE 54A pH, colour and haze readings for solutions
of S701 in water Sample pH L* a* b* haze (%) S008-D01-09A S701 3.23
95.44 -0.37 8.75 9.4 S008-D02-08A S701 3.16 95.38 -0.36 8.79
10.8
Example 30
[0226] This Example illustrates the effectiveness of concentration
and diafiltration steps in lowering the trypsin inhibitor
activity.
[0227] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0228] The filtered protein solution was then added to `e`
volume(s) of reverse osmosis purified water and the pH of the
sample lowered to `f` with diluted HCl. The diluted and acidified
solution was then heat treated at 90.degree. C. for 1 minute.
[0229] The diluted, acidified and heat treated protein extract
solution was reduced in volume from `g` L to `h` L by concentration
on a `i` membrane, having a molecular weight cutoff of `j` Daltons,
operated at a temperature of approximately `k`.degree. C. At this
point the acidified protein solution, with a protein content `l` wt
%, was diafiltered with `m` L of reverse osmosis (RO) purified
water, with the diafiltration operation conducted at approximately
`n`.degree. C. The diafiltered solution was then concentrated to a
volume of `o` L and diafiltered with an additional `p` L of RO
water, with the diafiltration operation conducted at approximately
`q`.degree. C. After this second diafiltration, the protein
solution was concentrated from a protein content of `r` % by weight
to a protein content of approximately `s` % by weight then diluted
to a protein content of `t` % by weight with water to facilitate
spray drying. The protein solution before spray drying was
recovered in a yield of `u` wt % of the initial filtered protein
solution. The acidified, diafiltered, concentrated protein solution
was then dried to yield a product found to have a protein content
of `v` wt % (N.times.6.25) d.b. and a phytic acid content of `w` wt
% d.b. The dry product was found to have a trypsin inhibitor
activity of `x` trypsin inhibitor units (TIU)/mg protein
(N.times.6.25). The product was given designation `y` S701H.
[0230] The parameters `a` to `y` for three runs are set forth in
the following Table 55:
TABLE-US-00057 TABLE 55 Parameters for the runs to produce S701H y
S010-G20-09A S010-G22-09A S010-G29-09A a 22.68 22.68 22.68 b 300
300 300 c 280 290 289 d 1.54 1.64 1.47 e 1 1 1 f 3.07 3.07 2.92 g
560 580 580 h 99 92 93 i PES PVDF PES j 100,000 100,000 100,000 k
30 30 31 l 4.20 4.42 4.44 m 100 100 100 n 31 30 30 o 49 39 46 p 300
300 300 q 31 30 30 r 6.92 7.54 8.75 s 16 16 18 t 6.90 6.92 8.39 u
83.3 76.1 84.5 v 100.80 99.41 101.56 w 0.05 0.05 0.06 x 18.2 18.9
17.4
[0231] Table 56 shows the reduction in trypsin inhibitor activity
of the protein solution at various points in the process.
TABLE-US-00058 TABLE 56 Trypsin inhibitor activity at various
points in the process (TIU/mg protein (N .times. 6.25)) sample
S010- S010- S010- G20-09A G22-09A G29-09A after heat treatment 72.5
79.8 101.9 concentrated solution before 1.sup.st DF 61.0 73.8 47.3
concentrated solution after 1.sup.st DF 48.5 50.3 42.0 solution
after 2.sup.nd DF 30.5 35.2 24.9 after dilution for spray drying
24.8 30.9 22.7
[0232] As may be seen from the results in Table 56, reductions in
trypsin inhibitor activity were achieved at all points in the
concentration and diafiltration process.
[0233] When the S701H isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 56A:
TABLE-US-00059 TABLE 56A pH, colour and haze readings for solutions
of S701H in water Sample pH L* a* b* haze (%) S010-G20-09A S701H
3.33 95.80 -0.35 7.58 6.9 S010-G22-09A S701H 3.37 96.82 -0.45 6.84
2.4 S010-G29-09A S701H 3.45 96.55 -0.52 7.21 4.2
Example 31
[0234] This Example illustrates a reduction in the trypsin
inhibitor activity level resulting from the inclusion of the
optional dilution step. Use of the dilution step results in greater
membrane processing and is in essence resulting in extra
diafiltration.
[0235] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation to produce `c` L of
clarified protein solution having a protein content of `d` % by
weight.
[0236] The clarified protein solution was then added to `e`
volume(s) of reverse osmosis purified water and the pH of the
sample T. The sample was then polished by filtration, providing `g`
L of filtered protein solution with a protein content of `h` % by
weight. The pH of the sample T. The filtrate was not heat
treated.
[0237] The filtered protein extract solution was reduced in volume
from `j` L to `k` L by concentration on a `l` membrane, having a
molecular weight cutoff of `m` Daltons, operated at a temperature
of approximately `n`.degree. C. The concentrated, acidified protein
solution was diafiltered with `o` L of reverse osmosis purified
water, with the diafiltration operation conducted at approximately
`p`.degree. C. The resulting acidified, diafiltered, concentrated
protein solution had a protein content of `q` % by weight and
represented a yield of `r` wt % of the initial clarified protein
solution. The acidified, diafiltered, concentrated protein solution
was then dried to yield a product found to have a protein content
of `s` wt % (N.times.6.25) d.b. and a phytic acid content of `t` wt
% d.b. The dry product was found to have a trypsin inhibitor
activity of trypsin inhibitor units/mg protein (N.times.6.25). The
product was given designation `v` S701.
[0238] The parameters `a` to `v` for three runs are set forth in
the following Table 57:
TABLE-US-00060 TABLE 57 Parameters for the runs to produce S701 v
S005-A08-09A S005-A15-09A S005-A27-09A a 20 20 20 b 200 200 200 c
138 147 159 d 2.57 2.54 2.61 e 1 1 0 f lowered to 3.00 lowered to
2.91 was not changed with diluted HCl with diluted H.sub.3PO.sub.4
g 311 325 205 h 1.2 0.88 2.09 i was not changed was not changed was
lowered to 3.07 with diluted HCl j 304 325 205 k 28 28 25 l PVDF
PVDF PVDF m 5,000 5,000 5,000 n 30 28 30 o 140 140 125 p 30 29 30 q
10.03 12.87 12.04 r 79.4 59.0 76.4 s 100.87 102.60 102.26 t 0.34 ND
0.11 u 66 57 90
[0239] As can be seen from the results presented in Table 57, the
runs with a dilution step generated product with a lower trypsin
inhibitor activity than the run where no dilution step was
employed.
[0240] When the S701 isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 57A:
TABLE-US-00061 TABLE 57A pH, colour and haze readings for solutions
of S701 in water Sample pH L* a* b* haze (%) S005-A08-09A S701 3.15
95.44 -0.68 10.47 4.4 S005-A15-09A S701 3.32 95.45 -0.74 10.54 2.5
S005-A27-09A S701 3.07 96.19 -1.35 11.60 3.8
Example 32
[0241] This Example illustrates the effect of the temperature of
membrane processing on the trypsin inhibitor activity of the novel,
acid soluble soy protein isolate (S701H) provided herein.
[0242] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0243] The filtered protein solution was then added to `e`
volume(s) of reverse osmosis purified water and the pH of the
sample lowered to `f` with diluted HCl. The diluted and acidified
solution was then heat treated at 90.degree. C. for 1 minute.
[0244] The diluted, acidified and heat treated protein extract
solution was reduced in volume from `g` L to `h` L by concentration
on a `i` membrane, having a molecular weight cutoff of `j` Daltons,
operated at a temperature of approximately `k`.degree. C. At this
point the acidified protein solution, with a protein content of `l`
% wt, was diafiltered with `m` L of reverse osmosis (RO) purified
water, with the diafiltration operation conducted at approximately
`n`.degree. C. The diafiltered solution was then further
concentrated to a volume of `o` L and diafiltered with an
additional `p` L of RO water, with the diafiltration operation
conducted at approximately `q`.degree. C. After this second
diafiltration, the protein solution was concentrated from a protein
content of `r` % by weight to a protein content of approximately
`s` % by weight then diluted to a protein content of `t` % by
weight with water to facilitate spray drying. The protein solution
before spray drying was recovered in a yield of wt % of the initial
filtered protein solution. The acidified, diafiltered, concentrated
protein solution was then dried to yield a product found to have a
protein content of `v` wt % (N.times.6.25) d.b. and a phytic acid
content of `w` wt % d.b. The dry product was found to have a
trypsin inhibitor activity of `x` trypsin inhibitor units (TIU)/mg
protein (N.times.6.25). The product was given designation `y`
S701H.
[0245] The parameters `a` to `y` for two runs are set forth in the
following Table 58:
TABLE-US-00062 TABLE 58 Parameters for the runs to produce S701H y
S010-G06-09A S010-G14-09A a 22.5 22.68 b 300 300 c 290 280 d 1.45
1.71 e 1 1 f 2.90 3.00 g 580 560 h 85 85 i PES PES j 100,000
100,000 k 49 30 l 4.39 4.71 m 100 100 n 50 30 o 42 42 p 300 300 q
51 30 r 8.29 7.61 s N/A 17 t 6.94 7.48 u 78.1 70.8 v 102.61 100.01
w 0.00 0.02 x 15.3 35.2 N/A = not available
[0246] As may be seen from the results in Table 58, the run
conducted with membrane processing at about 50.degree. C. had a
lower trypsin inhibitor activity than the run with membrane
processing conducted at about 30.degree. C.
[0247] When the S701H isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 58A:
TABLE-US-00063 TABLE 58A pH, colour and haze readings for solutions
of S701H in water Sample pH L* a* b* haze (%) S010-G06-09A S701H
3.28 96.11 -0.22 6.19 8.9 S010-G14-09A S701H 3.42 96.50 -0.54 7.27
4.5
Example 33
[0248] This Example illustrates the effect of a heat treatment on
the trypsin inhibitor activity of the novel, acid soluble soy
protein isolate (S701 and S701H).
[0249] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation to produce `c` L of
clarified protein solution having a protein content of `d` % by
weight.
[0250] The clarified protein solution was then added to `e`
volume(s) of reverse osmosis purified water and the pH of the
sample lowered to `f` with diluted HCl. The sample was heat treated
at `g`.degree. C. for `h` minutes and then polished by filtration,
providing L of filtered protein solution with a protein content of
`j` % by weight.
[0251] The filtered protein extract solution was reduced in volume
from `k` L to `l` L by concentration on a `m` membrane, having a
molecular weight cutoff of `n` Daltons, operated at a temperature
of approximately `o`.degree. C. The concentrated, acidified protein
solution was diafiltered with `p` L of reverse osmosis purified
water, with the diafiltration operation conducted at approximately
`q`.degree. C. The resulting acidified, diafiltered, concentrated
protein solution had a protein content of `r` % by weight and
represented a yield of `s` wt % of the initial clarified protein
solution. The acidified, diafiltered, concentrated protein solution
was then dried to yield a product found to have a protein content
of `t` wt % (N.times.6.25) d.b. and a phytic acid content of `u` wt
% d.b. The dry product was found to have a trypsin inhibitor
activity of `v` trypsin inhibitor units/mg protein (N.times.6.25).
The product was given designation `w`.
[0252] The parameters `a` to `w` for two runs are set forth in the
following Table 59:
TABLE-US-00064 TABLE 59 Parameters for the runs to produce S701 and
S701H w S005-L09-08A S701H S005-A08-09A S701 a 20 20 b 200 200 c
142.1 138 d 3.07 2.57 e 1 1 f 3.14 3.00 g 75 -- h 10 -- i 280 311 j
1.23 1.20 k 280 304 l 25 28 m PVDF PVDF n 5,000 5,000 o 30 30 p 125
140 q 28 30 r 11.74 10.03 s 84.0 79.4 t 102.66 100.87 u 0.17 0.34 v
40.5 66
[0253] As may be seen from the results in Table 59, the isolate
prepared by the process that included a heat treatment step had a
lower trypsin inhibitor activity.
[0254] When the S701 and S701H isolate samples were dissolved in
water, the resulting low pH solutions were transparent and very
light in colour as may be seen from the results set forth in the
following Table 59A:
TABLE-US-00065 TABLE 59A pH, colour and haze readings for solutions
of S701 and S701H in water Sample pH L* a* b* haze (%) S005-A08-09A
S701 3.15 95.44 -0.68 10.47 4.4 S005-L09-08A S701H 3.03 95.49 -0.70
12.10 3.0
Example 34
[0255] This Example also illustrates the effect of a heat treatment
on the trypsin inhibitor activity of the novel, acid soluble soy
protein isolate (S701 and S701H) provided herein.
[0256] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0257] The filtered protein solution was then added to `e`
volume(s) of reverse osmosis purified water and the pH of the
sample lowered to `f` with diluted HCl. The diluted and acidified
solution was then heat treated at `g`.degree. C. for `h`
minute(s).
[0258] The protein extract solution was reduced in volume from `i`
L to `j` L by concentration on a `k` membrane, having a molecular
weight cutoff of `l` Daltons, operated at a temperature of
approximately `m`.degree. C. At this point the acidified protein
solution, with a protein content of `n` wt %, was diafiltered with
`o` L of reverse osmosis (RO) purified water, with the
diafiltration operation conducted at approximately `p`.degree. C.
The diafiltered solution was then further concentrated to a volume
of `q` L and diafiltered with an additional `r` L of RO water, with
the diafiltration operation conducted at approximately `s` .degree.
C. After this second diafiltration, the protein solution was
concentrated from a protein content of `t` % by weight to a protein
content of approximately % by weight then diluted to a protein
content of `v` % by weight with water to facilitate spray drying.
The protein solution before spray drying was recovered in a yield
of `w` wt % of the initial filtered protein solution. The
acidified, diafiltered, concentrated protein solution was then
dried to yield a product found to have a protein content of `x` wt
% (N.times.6.25) d.b. and a phytic acid content of `y` wt % d.b.
The dry product was found to have a trypsin inhibitor activity of
`z` trypsin inhibitor units (TIU)/mg protein (N.times.6.25). The
product was given designation `aa`.
[0259] The parameters `a` to `aa` for two runs are set forth in the
following Table 60:
TABLE-US-00066 TABLE 60 Parameters for the runs to produce S701 and
S701H aa S010-G06-09A S701H S010-G13-09A S701 a 22.5 22.68 b 300
300 c 290 275 d 1.45 1.63 e 1 1 f 2.90 2.89 g 90 -- h 1 -- i 580
560 j 85 85 k PES PES l 100,000 100,000 m 49 50 n 4.39 4.37 o 100
100 p 50 50 q 42 42 r 300 300 s 51 51 t 8.29 7.61 u N/A 14 v 6.94
6.90 w 78.1 69.2 x 102.61 100.54 y 0.00 0.04 z 15.3 26.5 N/A = not
available
[0260] As may be seen from the results in Table 60, the isolate
prepared by the process that included a heat treatment step had a
lower trypsin inhibitor activity.
[0261] When the S701 and S701H isolate samples were dissolved in
water, the resulting low pH solutions were transparent and very
light in colour as may be seen from the results set forth in the
following Table 60A.
TABLE-US-00067 TABLE 60A pH, colour and haze readings for solutions
of S701 and S701H in water Sample pH L* a* b* haze (%) S010-G06-09A
S701H 3.28 96.11 -0.22 6.19 8.9 S010-G13-09A S701 3.42 95.43 -0.02
6.74 9.3
Example 35
[0262] This Example illustrates the effect of using soy protein
sources exposed to different levels of heat treatment on the
trypsin inhibitor activity of the novel, acid soluble soy protein
isolate (S701) provided herein.
[0263] `a` kg of defatted, minimally heat processed soy flour
(S005), defatted, moderately heat treated soy flour (S007) or
defatted, fully heat treated soy flour (S006) was added to `b` L of
0.15 M CaCl.sub.2 solution at ambient temperature and agitated for
30 minutes to provide an aqueous protein solution. The residual soy
meal was removed and the resulting protein solution was clarified
by centrifugation to produce `c` L of clarified protein solution
having a protein content of `d` % by weight.
[0264] The clarified protein solution was then added to e'
volume(s) of reverse osmosis purified water and the pH of the
sample lowered to `f` with diluted `g`. The sample was then
polished by filtration, providing `h` L of filtered protein
solution with a protein content of `i` % by weight. The filtrate
was not heat treated.
[0265] The filtered protein extract solution was reduced in volume
from `j` L to `k` L by concentration on a `l` membrane, having a
molecular weight cutoff of `m` Daltons, operated at a temperature
of approximately `n`.degree. C. The concentrated, acidified protein
solution was diafiltered with `o` L of reverse osmosis purified
water, with the diafiltration operation conducted at approximately
`p`.degree. C. The resulting acidified, diafiltered, concentrated
protein solution had a protein content of `q` % by weight and
represented a yield of `r` wt % of the initial clarified protein
solution. The acidified, diafiltered, concentrated protein solution
was then dried to yield a product found to have a protein content
of `s` wt % (N.times.6.25) d.b. and a phytic acid content of `t` wt
% d.b. The dry product was found to have a trypsin inhibitor
activity of `u` trypsin inhibitor units/mg protein (N.times.6.25).
The product was given designation `v` S701.
[0266] The parameters `a` to `v` for three runs are set forth in
the following Table 61:
TABLE-US-00068 TABLE 61 Parameters for the runs to produce S701 v
S005-A15-09A S007-A26-09A S006-A21-09A a 20 20 20 b 200 200 200 c
147 169 175 d 2.54 1.50 1.00 e 1 1 1 f 2.91 3.00 2.86 g
H.sub.3PO.sub.4 HCl HCl h 325 390 414 i 0.88 0.66 0.33 j 325 390
414 k 28 25 27 l PVDF PVDF PVDF m 5,000 5,000 5,000 n 28 30 30 o
140 125 135 p 29 28 29 q 12.87 9.54 4.59 r 59.0 73.2 48.0 s 102.60
101.07 97.25 t N.D. 0.20 0.00 u 57 40.5 12 N.D. = not
determined
[0267] As can be seen from the data presented in Table 61, the
greater the heat treatment of the soy protein source, the lower the
trypsin inhibitor activity in the final isolate.
[0268] When the S701 isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 61A.
TABLE-US-00069 TABLE 61A pH, colour and haze readings for solutions
of S701 in water Sample pH L* a* b* haze (%) S005-A15-09A S701 3.32
95.45 -0.74 10.54 2.5 S007-A26-09A S701 3.14 95.90 -0.29 7.27 9.1
S006-A21-09A S701 3.31 93.82 0.32 10.82 8.9
Example 36
[0269] This Example illustrates the effect of sodium sulfite
addition with the soy protein source on the trypsin inhibitor
activity of the novel, acid soluble soy protein isolate (S701H)
provided herein.
[0270] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 20 minutes. `c` kg of sodium sulfite was then
added and the mixture agitated for an additional 10 minutes to
provide an aqueous protein solution. The residual soy meal was
removed and the resulting protein solution was clarified by
centrifugation and filtration to produce `d` L of filtered protein
solution having a protein content of `e` % by weight.
[0271] The filtered protein solution was then added to `f`
volume(s) of reverse osmosis (RO) purified water and the pH of the
sample lowered to `g` with diluted HCl. The diluted and acidified
solution was then heat treated at 90.degree. C. for 1 minute.
[0272] The diluted, acidified and heat treated protein extract
solution was reduced in volume from `h` L to `i` L by concentration
on a `j` membrane, having a molecular weight cutoff of `k` Daltons,
operated at a temperature of approximately `l`.degree. C. At this
point the acidified protein solution, with protein content `m` wt %
was diafiltered with `n` L of RO water, with the diafiltration
operation conducted at approximately `o`.degree. C. The diafiltered
solution was then further concentrated to a volume of `p` L and
diafiltered with an additional `q` L of RO water, with the
diafiltration operation conducted at approximately `r`.degree. C.
After this second diafiltration, the protein solution was
concentrated from a protein content of `s` % by weight to a protein
content of approximately `t` % by weight then diluted to a protein
content of `u` % by weight with water to facilitate spray drying.
The protein solution before spray drying was recovered in a yield
of `v` wt % of the initial filtered protein solution. The
acidified, diafiltered, concentrated protein solution was then
dried to yield a product found to have a protein content of `w` wt
% (N.times.6.25) d.b. and a phytic acid content of `x` wt % d.b.
The dry product was found to have a trypsin inhibitor activity of
`y` trypsin inhibitor units (TIU)/mg protein (N.times.6.25). The
product was given designation `z` S701H.
[0273] The parameters `a` to `z` for two runs are set forth in the
following Table 62:
TABLE-US-00070 TABLE 62 Parameters for the runs to produce S701H z
S010-G21-09A S010-G28-09A a 22.5 22.68 b 300 300 c 0.6 0 d 280
270.4 e 1.89 1.95 f 1 1 g 2.98 2.96 h 560 535 i 92 98 j PES PES k
100,000 100,000 l 31 30 m 5.15 3.74 n 100 100 o 31 30 p 46 49 q 300
300 r 30 30 s 8.18 6.93 t 17 15 u 7.45 7.16 v 75.6 59.4 w 100.67
100.44 x 0.00 0.48 y 7.5 27.7
[0274] As can be seen from the results of Table 62, the process
employing the added sodium sulfite yielded an isolate with a lower
trypsin inhibitor activity.
[0275] When the S701H isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 62A.
TABLE-US-00071 TABLE 62A pH, colour and haze readings for solutions
of S701H in water Sample pH L* a* b* haze (%) S010-G21-09A S701H
3.36 97.28 -0.39 4.58 5.4 S010-G28-09A S701H 3.41 96.05 -0.44 7.70
5.2
Example 37
[0276] This Example illustrates the effect of sodium sulfite
addition to the diafiltered concentrated protein solution before
drying on the trypsin inhibitor activity of the novel, acid soluble
soy protein isolate (S701H) provided herein.
[0277] `a` kg of defatted, minimally heat processed soy flour was
added to `b` L of 0.15 M CaCl.sub.2 solution at ambient temperature
and agitated for 30 minutes to provide an aqueous protein solution.
The residual soy meal was removed and the resulting protein
solution was clarified by centrifugation and filtration to produce
`c` L of filtered protein solution having a protein content of `d`
% by weight.
[0278] The filtered protein solution was then added to `e`
volume(s) of reverse osmosis (RO) purified water and the pH of the
sample lowered to `f` with diluted HCl. The diluted and acidified
solution was then heat treated at 90.degree. C. for 1 minute.
[0279] The diluted, acidified and heat treated protein extract
solution was reduced in volume from `g` L to `h` L by concentration
on a `i` membrane, having a molecular weight cutoff of `j` Daltons,
operated at a temperature of approximately `k`.degree. C. At this
point the acidified protein solution, with a protein content of `l`
wt % was diafiltered with `m` L of RO water, with the diafiltration
operation conducted at approximately `n`.degree. C. The diafiltered
solution was then further concentrated to a volume of `o` L and
diafiltered with an additional `p` L of RO water, with the
diafiltration operation conducted at approximately `q`.degree. C.
After this second diafiltration, the protein solution was
concentrated from a protein content of `r` % by weight to a protein
content of approximately `s` % by weight then diluted to a protein
content of `t` % by weight with water to facilitate spray drying.
The protein solution before spray drying was recovered in a yield
of `u` wt % of the initial filtered protein solution. The
acidified, diafiltered, concentrated protein solution was then
split in two portions. The control portion (25.1 kg) was dried to
yield a product found to have a protein content of `v` wt %
(N.times.6.25) d.b., a phytic acid content of `w` wt % d.b. and a
trypsin inhibitor activity of `x` trypsin inhibitor units (TIU)/mg
protein (N.times.6.25). The product was given designation `y`
S701H-01. To the other portion of the acidified, diafiltered
concentrated protein solution (25.1 kg) was added `z` mg of sodium
sulfite. This sample was then dried to yield a product found to
have a protein content of `aa` wt % (N.times.6.25) d.b., a phytic
acid content of "ab" wt % d.b. and a trypsin inhibitor activity of
`ac` trypsin inhibitor units (TIU)/mg protein (N.times.6.25). The
product was given designation `y` S701H-02.
[0280] The parameters `a` to `ac` for one run are set forth in the
following Table 63:
TABLE-US-00072 TABLE 63 Parameters for the run to produce S701H-01
and S701H-02 y S010-G16-09A a 22.68 b 300 c 280 d 1.60 e 1 f 2.98 g
560 h 76 i PES j 100,000 k 50 l 4.51 m 100 n 50 o 38 p 300 q 50 r
7.97 s 16 t 6.94 u 77.7 v 102.17 w 0.02 x 15.1 z 17.57 aa 102.61 ab
0.00 ac 6.8
[0281] As may be seen from the results of Table 63, addition of
sodium sulfite to the acidified, diafiltered concentrated protein
solution prior to drying reduced the trypsin inhibitor activity of
the isolate.
[0282] When the S701H isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 63A.
TABLE-US-00073 TABLE 63A pH, colour and haze readings for solutions
of S701H in water Sample pH L* a* b* haze (%) S010-G16-09A S701H-01
3.38 96.13 -0.29 6.71 7.1 S010-G16-09A S701H-02 3.38 96.21 -0.29
6.68 6.2
Example 38
[0283] This Example illustrates the effect of pH of membrane
processing on the trypsin inhibitor activity of the novel, acid
soluble soy protein isolate (S701) provided herein.
[0284] `a` kg of defatted, soy white flake was added to `b` L of
0.15 M CaCl.sub.2 solution at ambient temperature and agitated for
30 minutes. The residual soy flake was removed and the resulting
protein solution was clarified by centrifugation and filtration to
produce `c` L of filtered protein solution having a protein content
of `d` % by weight.
[0285] The filtered protein solution was then added to `e`
volume(s) of reverse osmosis (RO) purified water and the pH of the
sample lowered to `f` with diluted HCl. The diluted and acidified
solution was not heat treated.
[0286] The diluted and acidified protein extract solution was
reduced in volume from `g` L to `h` L by concentration on a `i`
membrane, having a molecular weight cutoff of `j` Daltons, operated
at a temperature of approximately `k`.degree. C. At this point the
acidified protein solution, with protein content `l` wt % was
diafiltered with `m` L of `n` RO water, with the diafiltration
operation conducted at approximately `o`.degree. C. The diafiltered
solution was then further concentrated to a volume of `p` L and
diafiltered with an additional `q` L of `r` RO water, with the
diafiltration operation conducted at approximately `s`.degree. C.
The diafiltered protein solution had a protein content of `t` % by
weight and was recovered in a yield of wt % of the initial filtered
protein solution. The acidified, diafiltered, concentrated protein
solution was then `v` and dried to yield a product found to have a
protein content of `w` wt % (N.times.6.25) d.b. and a phytic acid
content of `x` wt % d.b. The dry product was found to have a
trypsin inhibitor activity of `y` trypsin inhibitor units (TIU)/mg
protein (N.times.6.25). The product was given designation `z`
S701.
[0287] The parameters `a` to `z` for two runs are set forth in the
following Table 64:
TABLE-US-00074 TABLE 64 Parameters for the runs to produce S701 z
S013-I15-09A S013-I24-09A a 30 30 b 300 300 c 265 280 d 1.87 1.90 e
1 1 f 2.85 2.01 g 540 560 h 84 93 i PES PES j 100,000 100,000 k 30
30 l 5.57 5.15 m 100 115 n natural pH pH 2 o 30 29 p 42 47 q 300
345 r natural pH pH 2 s 29 29 t 9.84 9.99 u 87.5 88.3 v -- adjusted
to pH 3.00 with diluted NaOH w 100.46 98.97 x 0.13 0.41 y 70.0
55.7
[0288] As may be seen from the results of Table 64, the isolate
prepared with membrane processing at approximately pH 2 had a lower
trypsin inhibitor activity than the isolate prepared with membrane
processing at approximately pH 3.
[0289] When the S701 isolate samples were dissolved in water, the
resulting low pH solutions were transparent and very light in
colour as may be seen from the results set forth in the following
Table 64A.
TABLE-US-00075 TABLE 64A pH, colour and haze readings for solutions
of S701 in water sample pH L* a* b* haze (%) S013-I15-09A S701 3.29
94.36 -0.21 10.07 11.2 S013-I24-09A S701 2.97 96.53 -0.66 7.22
7.1
Example 39
[0290] This Example compares the solution colour values determined
for products produced according to the present invention with the
solution colour values of typical commercial soy protein
products.
[0291] The solution colour values for solutions of products
produced in the Examples herein in accordance with the present
invention were determined using a HunterLab ColorQuest XE
instrument operated in transmission mode and such data is set forth
in Examples 4, 5, 6, 10, 11, 12, 27, 28, 29, 30, 31, 32, 33, 34,
35, 36, 37 and 38.
[0292] The solution colour values were also determined for a
selection of commercial soy protein products and the results
compared. As in the trials cited above, the protein solution was
prepared by dissolving sufficient protein product to supply 0.48 g
of protein in 15 ml of RO water. Since the samples provided in the
Examples produced solutions having a natural pH of about 3, the
tests were done both at the natural pH of the solution provided
from the commercial samples and at an adjusted pH of 3. The results
obtained are set forth in the following Tables 65 and 66.
TABLE-US-00076 TABLE 65 Colour values for solutions of commercial
soy protein products in water at natural pH sample pH L* a* b*
Solae Supro XT-40 7.34 35.52 5.97 31.04 Solae Supro XT-34N 7.13
39.37 6.39 34.22 Solae Supro 670 7.44 37.87 6.43 31.87 ADM Pro Fam
825 7.13 43.03 5.72 32.00 ADM Pro Fam 873 7.12 45.07 4.66 30.87 ADM
Pro Fam 781 7.09 42.48 4.72 29.38
TABLE-US-00077 TABLE 66 Colour values for 3.2% protein w/v
solutions of commercial soy protein products in water at pH 3
sample L* a* b* Solae Supro XT-40 36.65 4.62 27.24 Solae Supro
XT-34N 46.56 3.95 30.08 Solae Supro 670 36.44 6.67 31.71 ADM Pro
Fam 825 38.08 6.83 31.82 ADM Pro Fam 873 44.73 4.47 29.67 ADM Pro
Fam 781 42.90 4.33 27.94
[0293] As may be seen from the data in Tables 65 and 66 and by
comparison with the solution colour values provided in the Examples
for products of the invention, the solution colour for the products
of the invention was much lighter than for the commercial products,
exhibiting notably lower a* (less red) and b* (less yellow) values.
The aqueous solutions of the products provided herein exhibited a
much higher L* value as a result of significantly lower haze in the
solutions, when compared to solutions of the poorly soluble
commercial products.
Example 40
[0294] This Example illustrates the solubility of the novel soy
protein products at pH 7 to 8 as well as the clarity and heat
stability of solutions produced at such pH values.
[0295] The solubility of batches S005-K18-08A S701 and S010-G14-09A
S701H, prepared as described respectively in Examples 12 and 32, in
water using the "protein method" described in Example 14, was
determined at pH levels of 7, 7.5 and 8.
[0296] The solubilities of the products is shown in the following
Table 67:
TABLE-US-00078 TABLE 67 Solubility of products at pH 7 to 8 based
on protein method S005-K18-08A S701 S010-G14-09A S701H pH
solubility (%) solubility (%) 7 94.4 (see Example 14) 100 7.5 100
92.5 8 97.6 100
[0297] As can be seen from the results in Table 67, the products
were both highly soluble over the pH range of 7 to 8.
[0298] The clarity of these isolates was assessed using the method
described in Example 15. The clarity readings for 1% protein w/v
solutions are shown in the following Tables 68 and 69:
TABLE-US-00079 TABLE 68 A600 readings for solutions at pH 7 to 8 pH
S005-K18-08A S701 A600 S010-G14-09A S701H A600 7 0.225 (see Example
15) 0.198 7.5 0.035 0.082 8 0.026 0.040
TABLE-US-00080 TABLE 69 HunterLab haze readings (%) for solutions
at pH 7 to 8 pH S005-K18-08A S701 haze (%) S010-G14-09A S701H haze
(%) 7 not determined 24.9 7.5 1.3 9.0 8 0.5 3.1
[0299] As can be seen from the results shown in Tables 68 and 69,
the protein solutions were quite clear in the pH range 7.5 to
8.
[0300] The heat stability of the products was tested using the
method described in Example 16, but with the pH of the protein
solutions adjusted to 7, 7.5 or 8 prior to heat treatment. The
results obtained are set forth in the following Tables 70 and
71:
TABLE-US-00081 TABLE 70 Effect of heat treatment on clarity of
solutions of S005-K18-08A S701 at pH 7 to 8 pH % haze before heat
treatment % haze after heat treatment 7 96.2 95.5 7.5 13 10.1 8 8.2
5.1
TABLE-US-00082 TABLE 71 Effect of heat treatment on clarity of
solutions of S005-G14-09A S701H at pH 7 to 8 pH % haze before heat
treatment % haze after heat treatment 7 95.6 13.5 7.5 95.9 10 8
74.1 9.6
[0301] Both samples appeared to be heat stable in the pH range 7 to
8. The S701H gave cloudy solutions at all pH values tested that
cleared upon heating. The S701 gave a cloudy solution at pH 7 that
had approximately the same level of haze after heating. The S701
gave clear solutions at pH 7.5 and 8 that remained clear after heat
treatment.
[0302] As can be seen from the data presented in this Example, the
S701 product was soluble at pH 7 to 8, transparent at pH 7.5 to 8
and heat stable at pH 7 to 8.
SUMMARY OF THE DISCLOSURE
[0303] In summary of this disclosure, the present invention
provides a novel soy protein product, which may be in the form of
an isolate, which is completely soluble and forms transparent heat
stable solutions at acid pH and is useful in the protein
fortification of aqueous systems, including soft drinks and sports
drinks, without leading to protein precipitation. Modifications are
possible within the scope of this invention.
* * * * *