U.S. patent application number 10/517277 was filed with the patent office on 2006-09-14 for protein extraction from canola oil seed meal.
Invention is credited to Radka Milanova, E. Donald Murray, Paul S. Westdal.
Application Number | 20060205929 10/517277 |
Document ID | / |
Family ID | 30003139 |
Filed Date | 2006-09-14 |
United States Patent
Application |
20060205929 |
Kind Code |
A1 |
Milanova; Radka ; et
al. |
September 14, 2006 |
Protein extraction from canola oil seed meal
Abstract
The recovery of protein from canola oil seed meal and other oil
seed meals in the preparation of canola or other oil seed protein
isolate is improved in comparison to conventional toasted meal by
the use of a meal which has been air-desolventized at a temperature
below about 50.degree. C.
Inventors: |
Milanova; Radka; (Vancouver,
CA) ; Murray; E. Donald; (Eden Mills, CA) ;
Westdal; Paul S.; (Winnipeg, CA) |
Correspondence
Address: |
Michael I Stewart;Sim & McBurney
6th Floor
330 University Avenue
Toronto
M5G 1R7
CA
|
Family ID: |
30003139 |
Appl. No.: |
10/517277 |
Filed: |
June 19, 2003 |
PCT Filed: |
June 19, 2003 |
PCT NO: |
PCT/CA03/00923 |
371 Date: |
April 5, 2006 |
Current U.S.
Class: |
530/422 ;
530/370 |
Current CPC
Class: |
A23J 3/14 20130101; A23J
1/144 20130101; A23J 1/142 20130101 |
Class at
Publication: |
530/422 ;
530/370 |
International
Class: |
C07K 14/415 20060101
C07K014/415 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 21, 2002 |
US |
60390126 |
Aug 8, 2002 |
US |
60401782 |
Claims
1. A process of preparing a protein isolate, which comprises: (a)
crushing oil seeds to form oil and oil seed meal therefrom, (b)
solvent extracting the oil seed meal to recover residual oil
therefrom, (c) removing solvent from the extracted oil seed meal at
a temperature of below about 50.degree. C. to provide a
desolventized oil seed meal, (d) extracting the desolventized oil
seed meal to cause solubilization of protein in said desolventized
oil seed meal and to form an aqueous protein solution having a pH
of about 5 to about 6.8, (e) separating the aqueous protein
solution from residual oil seed meal, (f) increasing the protein
concentration of said aqueous protein solution while maintaining
the ionic strength substantially constant by using a selective
membrane technique to provide a concentrated protein solution, (g)
diluting said concentrated protein solution into chilled water
having a temperature of below about 15.degree. C. to cause the
formation of discrete protein particles in the aqueous phase at
least partially in the form of micelles, (h) settling the protein
micelles to form an amorphous, sticky, gelatinous, gluten-like
protein micellar mass, and (i) recovering the protein micellar mass
from supernatant, the protein micellar mass having a protein
content of at least about 90 wt % (N.times.6.25) on a dry weight
basis.
2. The process of claim 1 wherein said steps (d) to (i) are
effected in a batch mode of operation.
3. The process of claim 1 wherein said steps (d) to (i) are
effected in a semi-continuous mode of operation.
4. The process of claim 1 wherein said steps (d) to (i) are
effected in a continuous mode of operation.
5. The process of claim 2 wherein said extracting of said oil seed
meal is effected using an aqueous salt solution having an ionic
strength of at least about 0.10 and a pH of about 5 to about 6.8
and said aqueous protein solution has a protein content of about 5
to about 40 g/L.
6. The process of claim 5 wherein said salt solution has an ionic
strength of about 0.15 to about 0.6.
7. The process of claim 5 wherein said salt solution has a pH of
about 5.3 to about 6.2.
8. The process of claim 5 wherein said extracting of said oil seed
meal is effected with agitation of said aqueous salt solution for
about 10 to about 30 minutes.
9. The process of claim 8 wherein the concentration of oil seed
meal in said aqueous salt solution during said extracting step is
about 5 to about 15% w/v.
10. The process of claim 5 wherein said aqueous protein solution
resulting from the extraction step has a concentration of about 10
to about 30 g/L.
11. The process of claim 3 wherein said extraction step is effected
by: (i) continuously mixing an oil seed meal with an aqueous salt
solution having an ionic strength of at least about 0.10 and a pH
of about 5 to about 6.8 at a temperature of about 5.degree. to
about 65.degree. C., and (ii) continuously conveying said mixture
through a pipe while extracting protein from the oil seed meal to
form an aqueous protein solution having a protein content of about
5 to about 40 g/L for a period of time up to about 10 minutes.
12. The process of claim 11 wherein said salt solution has an ionic
strength of about 0.15 to about 0.8.
13. The process of claim 11 wherein the salt solution has a pH of
about 5.3 to about 6.2.
14. The process of claim 11 wherein the concentration of oil seed
meal in said aqueous salt solution in said mixing step is about 5
to about 15% w/v.
15. The process of claim 11 wherein said temperature is at least
about 35.degree. C.
16. The process of claim 11 wherein said aqueous protein solution
has a protein content of about 10 to about 30 g/L.
17. The process of claim 1 wherein said extracting of said oil seed
meal is effected using an aqueous salt solution having an ionic
strength of at least about 0.10 and a pH of about 3 to about 5 or
about 6.8 to about 9.9 and, following said separation of the
aqueous protein solution from residual oil seed meal, the pH of the
aqueous protein solution is adjusted to a pH of about 5 to about
6.8.
18. The process of claim 17 wherein said salt solution has a ionic
strength of about 0.15 to about 0.6.
19. The process of claim 17 wherein the pH of the aqueous protein
solution is adjusted to a pH of 5.3 to about 6.2.
20. The process of claim 1 wherein said oil seed meal is canola oil
seed meal and, following said separating of the aqueous protein
solution from the residual canola seed meal, the aqueous protein
solution is subjected to a pigment removal step.
21. The process of claim 20 wherein said pigment removal step is
effected by diafiltration of the aqueous protein solution.
22. The process of claim 20 wherein said pigment removal step is
effected by mixing a pigment adsorbing agent with the aqueous
protein solution and subsequently removing the pigment adsorbing
agent from the aqueous protein solution.
23. The process of claim 22 wherein the pigment adsorbing agent is
powdered activated carbon.
24. The process of claim 1 wherein said oil seed meal is extracted
with water and subsequent thereto salt is added to the resulting
aqueous protein solution to provide an aqueous protein solution
having an ionic strength of at least about 0.10.
25. The process of claim 1 wherein said concentration step is
effected by ultrafiltration to produce a concentrated protein
solution having a protein content of at least about 200 g/L.
26. The process of claim 25 wherein said concentration step is
effected to produce a concentrated protein solution having a
protein content of at least about 250 g/L.
27. The process of claim 25 wherein said concentrated protein
solution is warmed to a temperature of at least about 20.degree. C.
to decrease the viscosity of the concentrated protein solution but
not beyond a temperature above which the temperature of the
concentrated protein solution does not permit micelle
formation.
28. The process of claim 27 wherein said concentrated protein
solution is warmed to a temperature of about 25.degree. C. to about
40.degree. C.
29. The process of claim 2 wherein said concentrated protein
solution is diluted by about 15 fold or less by adding the
concentrated protein solution into a body of water having the
volume required to achieve the desired degree of dilution.
30. The process of claim 29 wherein said body of water has a
temperature of less than about 10.degree. C.
31. The process of claim 30 wherein said concentrated protein
solution is diluted by about 10 fold or less.
32. The process of claim 3 wherein said concentrated protein
solution is continuously mixed with said chilled water to provide a
dilution of the concentrated protein solution by about 15 fold or
less.
33. The process of claim 32 wherein said chilled water has a
temperature of less than about 10.degree. C.
34. The process of claim 33 wherein said dilution is by about 10
fold or less.
35. The process of claim 1 wherein the recovered protein micellar
mass is dried to a proteinaceous powder.
36. The process of claim 1 wherein said recovered protein micellar
mass has a protein content of at least about 100 wt %
(N.times.6.25).
37. The process of claim 1 wherein said oil seed meal is canola
seed meal and, following recovering of the protein micellar mass
therefrom, the supernatant is processed, on a batch,
semi-continuous or continuous basis, to recover additional
quantities of protein isolate therefrom.
38. The process of claim 37 wherein said additional quantities of
protein isolate are recovered from the supernatant by concentrating
the supernatant to a protein concentration of about 100 to about
400 g/L, preferably about 200 to about 300 g/L, and drying the
concentrated supernatant.
39. The process of claim 37 wherein said additional quantities of
protein isolate are recovered from the supernatant by concentrating
the supernatant to a protein concentration of about 100 to about
400 g/L, preferably about 200 to about 300 g/L, mixing the
concentrated supernatant with the recovered protein micellar mass,
and drying the mixture.
40. The process of claim 37 wherein said additional quantities of
protein isolate are recovered from the supernatant by concentrating
the supernatant to a protein concentration of about 100 to about
400 g/L, preferably about 200 to about 300 g/L, mixing a portion of
said concentrated supernatant with at least a portion of the
recovered protein micellar mass, and drying the resulting
mixture.
41. The process of claim 40 wherein the remainder of the
concentrated supernatant is dried and any remainder of the
recovered protein micellar mass is dried.
42. The process of claim 1 wherein, as an alternative to said
diluting, settling and recovering steps, the concentrated protein
solution is dialyzed to reduce the salt content thereof and to
cause the formation of protein micelles, and recovering a protein
isolate from the dialyzed concentrated protein solution having a
protein content of at least about 100 wt % (N.times.6.25) on a dry
weight basis.
43. The process of claim 42 wherein said protein isolate recovery
is effected by drying the dialyzed concentrated protein
solution.
44. The process of claim 1 wherein said oil seed meal is canola oil
seed meal.
45. The process of claim 44 wherein the canola oil seed meal is
cold pressed canola oil seed meal.
46. The process of claim 44 wherein the canola oil seed meal is
derived from a non-genetically modified canola oil seed.
47. The process of claim 1 wherein the oil seed meal is rapeseed
meal.
48. The process of claim 1 wherein said oil seed meal is mustard
seed meal.
49. The process of claim 1 wherein said solvent removal step is
effected by air-desolventizing at a temperature of about 15.degree.
to about 25.degree. C.
50. The process of claim 1 wherein said solvent removal step is
effected under vacuum.
Description
REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority under 35 USC 119(e) from
U.S. Provisional Patent Applications Nos. 60/390,126 filed Jun. 21,
2002 and 60/401,782 filed Aug. 8, 2002.
FIELD OF INVENTION
[0002] The present invention is concerned with the recovery of
protein from oil seed proteins, particularly canola oil seed
protein.
BACKGROUND OF THE INVENTION
[0003] Canola oil seed is extensively processed for the recovery of
canola oil therefrom. The canola oil seed is crushed to remove most
of the oil and the residual meal is hot solvent extracted,
generally using hexane, to recover the remainder of the oil. The
residual meal from the solvent extraction contains residual hexane
and is commonly known as "white flake" or less commonly as "marc"
meal. The solvent is recovered from the meal for reuse before the
oil seed meal is disposed of by the crusher. In the solvent
recovery process, the oil seed meal often is heated to a higher
temperature of about 120.degree. to 140.degree. C. in a procedure
termed "toasting". The resulting meal is referred to as "toasted
meal" or "high temperature produced meal".
[0004] The residual oil seed meal disposed of by the crusher
contains significant quantities of protein and often is employed as
animal feed. There have been prior procedures to recover the canola
protein from the residual canola oil seed meal in the form of a
canola protein isolate.
[0005] In U.S. Pat. Nos. 5,844,086 and 6,005,076 ("Murray II"),
assigned to the assignee hereof and the disclosures of which are
incorporated herein by reference, there is described a process for
the isolation of protein isolates from oil seed meal having a
significant fat content, including canola oil seed meal having such
content. The steps involved in this process include solubilizing
proteinaceous material from oil seed meal, which also solubilizes
fat in the meal and removing fat from the resulting aqueous protein
solution. The aqueous protein solution may be separated from the
residual oil seed meal before or after the fat removal step. The
defatted protein solution then is concentrated to increase the
protein concentration while maintaining the ionic strength
substantially constant, after which the concentrated protein
solution may be subjected to a further fat removal step. The
concentrated protein solution then is diluted to cause the
formation of a cloud-like mass of highly aggregated protein
molecules as discrete protein droplets in micellar form. The
protein micelles are allowed to settle to form an aggregated,
coalesced, dense, amorphous, sticky gluten-like protein isolate
mass, termed "protein micellar mass" or PMM, which is separated
from the residual aqueous phase and dried.
[0006] The protein isolate has a protein content (as determined by
Kjeldahl or equivalent method N.times.6.25) of at least about 90 wt
%, is substantially undenatured (as determined by differential
scanning calorimetry) and has a low residual fat content. The term
"protein content" as used herein refers to the quantity of protein
in the protein isolate expressed on a dry weight basis. The yield
of protein isolate obtained using this procedure, in terms of the
proportion of protein extracted from the oil seed meal which is
recovered as dried protein isolate was generally less than 40 wt %,
typically around 20 wt %.
[0007] The procedure described in the aforementioned patents was
developed as a modification to and improvement on the procedure for
forming a protein isolate from a variety of protein source
materials, including oil seeds, as described in U.S. Pat. No.
4,208,323 (Murray IB), the disclosure of which is incorporated
herein by reference. The oil seed meals available in 1980, when
U.S. Pat. No. 4,208,323 issued, did not have the fat contamination
levels of canola oil seed meals at the time of Murray II patents,
and, as a consequence, the procedure of U.S. Pat. No. 4,208,323
cannot produce from such oil seed meals processed according to the
Murray II process, proteinaceous materials which have more than 90
wt % protein content. There is no description of any specific
experiments in U.S. Pat. No. 4,208,323 carried out using rapeseed
(canola) meal as the starting material.
[0008] U.S. Pat. No. 4,208,323 itself was designed to be an
improvement on the process described in U.S. Pat. Nos. 4,169,090
and 4,285,862 (Murray IA), incorporated herein by reference, by the
introduction of the concentration step prior to dilution to form
the PMM. The latter step served to improve the yield of protein
isolate from around 20% for the Murray IA process.
[0009] In copending U.S. Patent Applications Nos. 60/288,415 filed
May 4, 2001, 60/326,987 filed Oct. 5, 2001, 60/331,066 filed Nov.
7, 2001, 60/333,494 filed Nov. 26, 2001, 60/374,801 filed Apr. 24,
2002 and U.S. patent application Ser. No. 10/137,391 filed May 3,
2002 (WO 02/089597), all assigned to the assignee hereof and the
disclosures of which are incorporated herein by reference, there is
described a process for producing a protein isolate of high purity,
containing at least about 100 wt % protein (N.times.6.25). In the
aforementioned US Patent Applications, the protein isolate is made
by a process in which oil seed meal is extracted with a food grade
salt solution, the resulting protein solution, after an initial
treatment with a colourant adsorbent, if desired, is concentrated
to a protein content of at least about 200 g/L, and the
concentrated protein solution is diluted in chilled water to form
protein micelles, which are allowed to settle to form an
aggregated, coalesced, dense amorphous, sticky gluten-like protein
isolate mass, termed "protein micellar mass" or PMM, which is
separated from residual aqueous phase and may be used as such or
dried.
[0010] In one embodiment of the process described above and as
specifically described in U.S. Patent Applications Nos. 60/326,987,
60/331,066, 60/333,494, 60/374,801 and Ser. No. 10/137,391, the
supernatant from the PMM settling step is processed to recover a
protein isolate comprising dried protein from wet PMM and
supernatant. This procedure may be effected by initially
concentrating the supernatant using ultrafiltration membranes,
mixing the concentrated supernatant with the wet PMM and drying the
mixture. The resulting canola protein isolate has a high purity of
at least about 90 wt %, preferably at least about 100 wt %, protein
(N.times.6.25).
[0011] In another embodiment of the process described above and as
significantly specifically described in Applications Nos.
60/331,066, 60/333,494, 60/374,801 and Ser. No. 10/137,391, the
supernatant from the PMM settling step is processed to recover a
protein from the supernatant. This procedure may be effected by
initially concentrating the supernatant using ultrafiltration
membranes and drying the concentrate. The resulting canola protein
isolate has a high purity of at least about 90 wt %, preferably at
least about 100 wt %, protein (N.times.6.25).
[0012] The procedures described in the aforementioned US Patent
Applications are essentially batch procedures. In copending U.S.
Patent Applications Nos. 60/331,646 filed Nov. 20, 2001, 60/383,809
filed May 30, 2002 and 10/298,678 filed Nov. 19, 2002, assigned to
the assignee hereof and the disclosures of which are incorporated
herein by reference, there is described a continuous process for
making canola protein isolates. In accordance therewith, canola oil
seed meal is continuously mixed with a salt solution, the mixture
is conveyed through a pipe while extracting protein from the canola
oil seed meal to form an aqueous protein solution, the aqueous
protein solution is continuously separated from residual canola oil
seed meal, the aqueous protein solution is continuously conveyed
through a selective membrane operation to increase the protein
content of the aqueous protein solution to at least about 200 g/L
while maintaining the ionic strength substantially constant, the
resulting concentrated protein solution is continuously mixed with
chilled water to cause the formation of protein micelles, and the
protein micelles are continuously permitted to settle while the
supernatant is continuously overflowed until the desired amount of
PMM has accumulated in the settling vessel. The PMM is removed from
the settling vessel and may be dried. The PMM has a protein content
of at least about 90 wt % (N.times.6.25), preferably at least about
100 wt %.
[0013] The experimentation described in such prior U.S. patent
applications is carried out on commercially-available oil seed meal
which has been desolventized in a conventional
desolventizer-toasting operation. Using such materials as the oil
seed meal for production of oil seed protein isolate, results in
extraction of less than about 30 wt % of the protein present in the
oil seed, possibly due to denaturation of protein by the high
temperature desolventizing operation.
SUMMARY OF THE INVENTION
[0014] It has now surprisingly been found that the amount of
protein which can be extracted from canola oil seed protein meal
can be significantly increased if the extraction is effected on
ambient temperature desolventized meal. The ability to extract more
protein from the meal improves the overall economics of the
process. In addition a product of improved quality is obtained.
[0015] In accordance with one aspect of the present invention,
there is provided a process of preparing a protein isolate, which
comprises (a) crushing oil seeds to form oil and oil seed meal
therefrom, (b) solvent extracting the oil seed meal to recover
residual oil therefrom, (c) removing solvent from the extracted oil
seed meal at a temperature of below about 50.degree. C. to provide
a desolventized oil seed meal, (d) extracting the desolventized oil
seed meal to cause solubilization of protein in the desolventized
oil seed meal and to form an aqueous protein solution having a pH
of about 5 to about 6.8, (e) separating the aqueous protein
solution from residual oil seed meal, (f) increasing the protein
concentration of the aqueous protein solution while maintaining the
ionic strength substantially constant by using a selective membrane
technique to provide a concentrated protein solution, (g) diluting
the concentrated protein solution into chilled water having a
temperature of below about 15.degree. C. to cause the formation of
discrete protein particles in the aqueous phase at least partially
in the form of micelles, (h) settling the protein micelles to form
an amorphous, sticky, gelatinous, gluten-like protein micellar
mass, and (i) recovering the protein micellar mass from
supernatant, the protein micellar mass having a protein content of
at least about 90 wt % (N.times.6.25) on a dry weight basis.
[0016] The present invention uses white flake or marc meal which
has been desolventized at moderate temperatures below about
50.degree. C., preferably at about 15.degree. to about 30.degree.
C. Desolventizing may be effected by air drying the meal or by
vacuum stripping.
[0017] The protein may be extracted and recovered from the ambient
temperature desolventized meal by either a batch process, a
semi-batch process or a continuous process as generally described
in the aforementioned U.S. patent applications.
[0018] The protein isolate produced according to the process herein
may be used in conventional applications of protein isolates, such
as, protein fortification of processed foods, emulsification of
oils, body formers in baked goods and foaming agents in products
which entrap gases. In addition, the protein isolate may be formed
into protein fibers, useful in meat analogs, may be used as an egg
white substitute or extender in food products where egg white is
used as a binder. The canola protein isolate may be used as
nutritional supplements. Other uses of the canola protein isolate
are in pet foods, animal feed and in industrial and cosmetic
applications and in personal care products.
BRIEF DESCRIPTION OF DRAWINGS
[0019] FIGS. 1 to 3 are HPLC chromatograms of extractions of canola
oil seed meal which has been air-desolventized meal at room
temperature using 0.05 M NaCl (FIG. 1) and 0.10 M NaCl (FIG. 2) and
at 60.degree. C. in the absence of salt (FIG. 3).
GENERAL DESCRIPTION OF INVENTION
[0020] The process of the invention commences with oil seed,
particularly canola oil seed, although the process may be applied
to other oil seeds, such as soybean, traditional rapeseed,
traditional flax, linola, sunflower and mustard oil seed meals. The
invention is more particularly described herein with respect to
canola oil seed meal.
[0021] The oil seed is washed to recover oil therefrom. Following
separation of the oil, the residual meal is solvent extracted,
usually using hexane, to recover residual amounts of oil from the
meal. The resulting meal then is desolventized in accordance with
the present invention at a temperature below about 50.degree. C.,
preferably at about 15.degree. to about 30.degree. C. By effecting
desolventizing in this manner, it has been found that the amount of
protein which can be extracted from the meal is significantly
increased.
[0022] The oil seed meal which is processed in this manner may be
processed as described in the Murray I or II patents to recover
protein isolate from the oil seed meal, details of which are
described therein. Preferably, the procedure described in the
aforementioned copending U.S. Patent Applications Nos. 60/288,415,
60/326,987, 60/331,066, 60/333,494, 60/372,165, 60/374,801 and Ser.
No. 10/137,391 (WO 02/089567) is employed since there are obtained
thereby improved yields of dried protein isolate, in terms of the
proportion of the protein extracted from the oil seed meal which is
recovered as protein isolate and a protein isolate of high protein
content is obtained, usually at least about 100 wt % as determined
by the Kjeldahl method as percent nitrogen (N) and multiplied by a
factor of 6.25. Alternatively, the continuous process described in
the aforementioned U.S. Applications Nos. 60/331,646, 60/383,809
and Ser. No. 10/298,678 may be employed. Details of these preferred
procedures as applied to canola protein isolate are described
below.
[0023] It will be understood that the processing of the oil seed to
recover oil therefrom may be effected in a different facility from
that at which the protein isolate is recovered from the oil seed
meal. Alternatively, the operations may be combined at a single
facility.
[0024] The initial step of the process of separating the canola
protein isolate involves solubilizing proteinaceous material from
canola oil seed meal. The proteinaceous material recovered from
canola seed meal may be the protein naturally occurring in canola
seed or other oil seed or the proteinaceous material may be a
protein modified by genetic manipulation but possessing
characteristic hydrophobic and polar properties of the natural
protein. Canola oil seed is also known as rapeseed or oil seed
rape.
[0025] Protein solubilization is effected most efficiently by using
a food grade salt solution since the presence of the salt enhances
the removal of soluble protein from the oil seed meal. Where the
canola protein isolate is intended for non-food uses, non-food
grade chemicals may be employed. The food grade salt usually is
sodium chloride, although other salts, such as, potassium chloride,
may be used. The food grade salt solution has an ionic strength of
at least about 0.10, preferably at least about 0.15, to enable
solubilization of significant quantities of protein to be effected.
As the ionic strength of the salt solution increases, the degree of
solubilization of protein in the oil seed meal initially increases
until a maximum value is achieved. Any subsequent increase in ionic
strength does not increase the total protein solubilized. The ionic
strength of the food grade salt solution which causes maximum
protein solubilization varies depending on the salt concerned and
the oil seed meal chosen.
[0026] In view of the greater degree of dilution required for
protein precipitation with increasing ionic strengths, it is
usually preferred to utilize an ionic strength value less than
about 0.8, and more preferably a value of about 0.15 to about
0.6.
[0027] In a batch process, the salt solubilization of the protein
is effected at a temperature of at least about 5.degree. and
preferably up to about 35.degree. C., preferably accompanied by
agitation to decrease the solubilization time, which is usually
about 10 to about 60 minutes. It is preferred to effect the
solubilization to extract substantially the maximum amount of
protein from the oil seed meal, so as to provide an overall high
product yield.
[0028] The lower temperature limit of about 5.degree. C. is chosen
since solubilization is impractically slow below this temperature
while the upper preferred temperature limit of about 35.degree. C.
is chosen since the process becomes uneconomic at higher
temperature levels in a batch mode.
[0029] In a continuous process, the extraction of the protein from
the canola oil seed meal is carried out in any manner consistent
with effecting a continuous extraction of protein from the canola
oil seed meal. In one embodiment, the canola oil seed meal is
continuously mixed with a 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 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 the maximum amount of protein from the
canola oil seed meal. The solubilization in the continuous
procedure preferably is effected at elevated temperatures,
preferably above about 35.degree. C., generally up to about
65.degree. C. or more.
[0030] The aqueous food grade salt solution and the canola oil seed
meal have a natural pH of about 5 to about 6.8 to enable a protein
isolate to be formed by the micellar route, as described in more
detail below.
[0031] At and close to the limits of the pH range, protein isolate
formation occurs only partly through the micelle route and in lower
yields than attainable elsewhere in the pH range. For these
reasons, pH values of about 5.3 to about 6.2 are preferred.
[0032] The pH of the food grade salt solution may be adjusted to
any desired value within the range of about 5 to about 6.8 for use
in the extraction step by the use of any convenient food grade
acid, usually hydrochloric acid, or food grade alkali, usually
sodium hydroxide, as required. Where the canola protein isolate is
intended for non-food uses, then non-food grade chemicals may be
used.
[0033] The concentration of oil seed meal in the food grade salt
solution during the solubilization step may vary widely. Typical
concentration values are about 5 to about 15% w/v.
[0034] The protein extraction step with the aqueous salt solution
has the additional effect of solubilizing fats which may be present
in the canola meal, which then results in the fats being present in
the aqueous phase.
[0035] The protein solution resulting from the extraction step
generally has a protein concentration of about 5 to about 40 g/L,
preferably about 10 to about 30 g/L.
[0036] The aqueous phase resulting from the extraction step then
may be separated from the residual canola meal, in any convenient
manner, such as by employing vacuum filtration, followed by
centrifugation and/or filtration to remove residual meal. The
separated residual meal may be dried for disposal.
[0037] The colour of the final canola protein isolate can be
improved to obtain a lighter and less intense yellow colour by the
mixing of powdered activated carbon or other pigment adsorbing
agent with the separated aqueous protein solution and subsequently
removing the adsorbent, conveniently by filtration, to provide a
protein solution. Diafiltration of the separated aqueous protein
solution, before or after concentration, as described below, also
may be used for pigment removal.
[0038] Such pigment removal step may be carried out under any
convenient conditions, generally at the ambient temperature of the
separated aqueous protein solution, employing any suitable pigment
adsorbing agent. 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.
[0039] Where the canola seed meal contains significant quantities
of fat, as described in the aforementioned U.S. Pat. Nos. 5,844,086
and 6,005,076, then the defatting steps described therein may be
effected on the separated aqueous protein solution and on the
concentrated aqueous protein solution discussed below. When the
colour improvement step is carried out, such step may be effected
after the first defatting step.
[0040] As an alternative to extracting the oil seed meal with an
aqueous food grade salt solution, such extraction may be made using
water alone, although the utilization of water alone tends to
extract less protein from the oil seed meal than the aqueous food
grade salt solution. Where such alternative is employed, then the
food grade salt, in the concentrations discussed above, may be
added to the protein solution after separation from the residual
oil seed meal in order to maintain the protein in solution during
the concentration step described below. When a colour removal step
and/or a first fat removal step is carried out, the food grade salt
generally is added after completion of such operations.
[0041] Another alternative procedure is to extract the oil seed
meal with the food grade salt solution at a relatively high pH
value above about 6.8, generally up to about 9.8. The pH of the
food grade salt solution, may be adjusted in pH to the alkaline
value by the use of any convenient food-grade alkali, such as
aqueous sodium hydroxide solution. Where such alternative is
employed, the aqueous phase resulting from the oil seed meal
extraction step then is separated from the residual canola meal, in
any convenient manner, such as by employing vacuum filtration,
followed by centrifugation and/or filtration to remove residual
meal. The separated residual meal may be dried for disposal.
[0042] The aqueous protein solution resulting from the high pH
extraction step then is pH adjusted to the range of about 5 to
about 6.8, preferably about 5.3 to about 6.2, as discussed above,
prior to further processing as discussed below. Such pH adjustment
may be effected using any convenient food grade acid, such as
hydrochloric acid.
[0043] The aqueous protein solution then is concentrated to
increase the protein concentration thereof while maintaining the
ionic strength thereof substantially constant. Such concentration
generally is effected to provide a concentrated protein solution
having a protein concentration of at least about 200 g/L,
preferably at least about 250 g/L.
[0044] 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 cut-off, such as about 3000 to about 50,000
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.
[0045] The concentration step may be effected at any convenient
temperature, generally about 20.degree. to about 60.degree. C., and
for the period of time to effect the desired degree of
concentration. The temperature and other conditions used to some
degree depend upon the membrane equipment used to effect the
concentration and the desired protein concentration of the
solution.
[0046] The concentrating of the protein solution to a concentration
above about 200 g/L in this step not only increases the process
yield to levels above about 40% in terms of the proportion of
extracted protein which is recovered as dried protein isolate,
preferably above about 80%, but also decreases the salt
concentration of the final protein isolate after drying. The
ability to control the salt concentration of the isolate is
important in applications of the isolate where variations in salt
concentrations affect the functional and sensory properties in a
specific food application
[0047] 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 and anti-nutritional factors, as well as
any low molecular weight forms of the protein. 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.
[0048] Depending on the temperature employed in the concentration
step, the concentrated protein solution may be warmed to a
temperature of at least about 200, and up to about 60.degree. C.,
preferably about 25.degree. to about 40.degree. C., to decrease the
viscosity of the concentrated protein solution to facilitate
performance of the subsequent dilution step and micelle formation.
The concentrated protein solution should not be heated beyond a
temperature above which the temperature of the concentrated protein
solution does not permit micelle formation on dilution by chilled
water. The concentrated protein solution may be subject to a
farther defatting operation, if required, as described in the
aforementioned U.S. Pat. Nos. 5,844,086 and 6,005,076.
[0049] The concentrated protein solution resulting from the
concentration step and optional defatting step then is diluted to
effect micelle formation by mixing the concentrated protein
solution with chilled water having the volume required to achieve
the degree of dilution desired. Depending on the proportion of
canola protein desired to be obtained by the micelle route and the
proportion from the supernatant, the degree of dilution of the
concentrated protein solution may be varied. With higher dilution
levels, in general, a greater proportion of the canola protein
remains in the aqueous phase.
[0050] When it is desired to provide the greatest proportion of the
protein by the micelle route, the concentrated protein solution is
diluted by about 15 fold or less, preferably about 10 fold or
less.
[0051] The chilled water with which the concentrated protein
solution is mixed has a temperature of less than about 15.degree.
C., generally about 3.degree. to about 15.degree. C., preferably
less than about 10.degree. C., since improved yields of protein
isolate in the form of protein micellar mass are attained with
these colder temperatures at the dilution factors used.
[0052] In a batch operation, the batch of concentrated protein
solution is added to a static body of chilled water having the
desired volume, as discussed above. The dilution of the
concentrated protein solution and consequential decrease in ionic
strength causes the formation of a cloud-like mass of highly
associated protein molecules in the form of discrete protein
droplets in micellar form. In the batch procedure, the protein
micelles are allowed to settle in the body of chilled water to form
an aggregated, coalesced, dense, amorphous sticky gluten-like
protein micellar mass PMM. The settling may be assisted, such as by
centrifugation. Such induced settling decreases the liquid content
of the protein micellar mass, thereby decreasing the moisture
content generally from about 70% by weight to about 95% by weight
to a value of generally about 50% by weight to about 80% by weight
of the total micellar mass. Decreasing the moisture content of the
micellar mass in this way also decreases the occluded salt content
of the micellar mass, and hence the salt content of dried
isolate.
[0053] Alternatively, the dilution operation may be carried out
continuously by continuously passing the concentrated protein
solution to one inlet of a T-shaped pipe, while the diluting water
is fed to the other inlet of the T-shaped pipe, permitting mixing
in the pipe. The diluting water is fed into the T-shaped pipe at a
rate sufficient to achieve the desired degree of dilution.
[0054] The mixing of the concentrated protein solution and the
diluting water in the pipe initiates the formation of protein
micelles and the mixture is continuously fed from the outlet from
the T-shaped pipe into a settling vessel, from which, when full,
supernatant is permitted to overflow. The mixture preferably is fed
into the body of liquid in the settling vessel in a manner which
minimizes turbulence within the body of liquid.
[0055] In the continuous procedure, the protein micelles are
allowed to settle in the settling vessel to form an aggregated,
coalesced, dense, amorphous, sticky, gluten-like protein micellar
mass (PMM) and the procedure is continued until a desired quantity
of the PMM has accumulated in the bottom of the settling vessel,
whereupon the accumulated PMM is removed from the settling
vessel.
[0056] The combination of process parameters of concentrating of
the protein solution to a protein content of at least about 200 g/L
and the use of a dilution factor less than about 15, result in
higher yields, often significantly higher yields, in terms of
recovery of protein in the form of protein micellar mass from the
original meal extract, and much purer isolates in terms of protein
content than achieved using any of the known prior art protein
isolate forming procedures discussed in the aforementioned U.S.
patent applications.
[0057] The settled isolate is separated from the residual aqueous
phase or supernatant, by such methods as decantation of the
residual aqueous phase from the settled mass or centrifugation. The
PMM may be used in the wet form or may be dried, by any convenient
technique, such as spray drying, freeze drying or vacuum drum
drying, to a dry form. The dry PMM has a high protein content, in
excess of about 90 wt % protein, preferably at least about 100 wt %
protein (calculated as Kjeldahl N.times.6.25), and is substantially
undenatured (as determined by differential scanning calorimetry).
The dry PMM isolated from fatty oil seed meal also has a low
residual fat content, when the procedures of the aforementioned
U.S. Pat. Nos. 5,844,086 and 6,005,076 are employed, which may be
below about 1 wt %.
[0058] The supernatant from the PMM formation and settling step
contains significant amounts of canola protein, not precipitated in
the dilution step, and is processed to recover canola protein
isolate therefrom. The supernatant from the dilution step,
following removal of the PMM, is concentrated to increase the
protein concentration thereof. Such concentration is effected using
any convenient selective membrane technique, such as
ultrafiltration, using membranes with a suitable molecular weight
cut-off permitting low molecular weight species, including the food
grade salt and other non-proteinaceous low molecular weight
materials extracted from the protein source material, to pass
through the membrane, while retaining canola protein in the
solution. Ultrafiltration membranes having a molecular weight
cut-off of about 3000 to 10,000 daltons, having regard to differing
membrane materials and configuration, may be used. Concentration of
the supernatant in this way also reduces the volume of liquid
required to be dried to recover the protein. The supernatant
generally is concentrated to a protein concentration of about 100
to about 400 g/L, preferably about 200 to about 300 g/L, prior to
drying. Such concentration operation may be carried out in a batch
mode or in a continuous operation, as described above for the
protein solution concentration step.
[0059] The concentrated supernatant may be dried by any convenient
technique, such as spray drying, freeze drying or vacuum
drum-drying, to a dry form to provide a further canola protein
isolate. Such further canola protein isolate has a high protein
content, in excess of about 90 wt %, preferably at least about 100
wt % protein (calculated as N.times.6.25) and is substantially
undenatured (as determined by differential scanning
calorimetry).
[0060] If desired, at least a portion of the wet PMM may be
combined with at least a portion of the concentrated supernatant
prior to drying the combined protein streams by any convenient
technique to provide a combined canola protein isolate composition
according to one invention. The relative proportions of the
proteinaceous materials mixed together may be chosen to provide a
canola protein isolate composition having a desired profile of
2S/7S/12S proteins. Alternatively, the dried protein isolates may
be combined in any desired proportions-to provide any desired
specific 2S/7S/12S protein profile in the mixture. The combined
canola protein isolate composition has a high protein content, in
excess of about 90 wt %, preferably at least about 100 wt %,
(calculated as N.times.6.25) and is substantially undenatured (as
determined by differential scanning calorimetry).
[0061] In another alternative procedure, where a portion only of
the concentrated supernatant is mixed with a part only of the PMM
and the resulting mixture dried, the remainder of the concentrated
supernatant may be dried as may any of the remainder of the PMM.
Further, dried PMM and dried supernatant also may be dry mixed in
any desired relative proportions, as discussed above.
[0062] By operating in this manner, a number of canola protein
isolates may be recovered, in the form of dried PMM, dried
supernatant and dried mixtures of various proportions by weight of
PMM and supernatant, generally from about 5:95 to about 95:5 by
weight, which may be desirable for attaining differing functional
and nutritional properties.
[0063] As an alternative to dilution of the concentrated protein
solution into chilled water and processing of the resulting
precipitate and supernatant as described above, protein may be
recovered from the concentrated protein solution by dialyzing the
concentrated protein solution to reduce the salt content thereof.
The reduction of the salt content of the concentrated protein
solution results in the formation of protein micelles in the
dialysis tubing. Following dialysis, the protein micelles may be
permitted to settle, collected and dried, as discussed above. The
supernatant from the protein micelle settling step may be
processed, as discussed above, to recover further protein
therefrom. Alternatively, the contents of the dialysis tubing may
be directly dried. The latter alternative procedure is useful where
small laboratory scale quantities of protein are desired.
EXAMPLES
Example 1
[0064] This Example illustrates the process of the invention.
[0065] 75 g samples of canola oil seed meal which had been
air-desolventized at ambient temperature (20.degree. C.) were added
to 500 ml samples of 0.15 M NaCl solution at ambient or room
temperature (RT), 55.degree. C., 60.degree. C. and 65.degree. C.,
agitated for 30 minutes while maintaining the temperature of the
solution substantially constant to provide aqueous protein
solutions. Samples of aqueous protein solution were taken at 5, 10,
15, 20 and 30 minutes for analysis. The spent meal was separated by
centrifugation at 10,000.times.g for 5 minutes and
freeze-dried.
[0066] The protein concentrations of the various aqueous protein
solutions obtained in these experiments were determined and the
results appear in the following Table I: TABLE-US-00001 TABLE I
Protein Concentration in Extracts (wt %) Extraction Time (min) RT*
55.degree. C. 60.degree. C. 65.degree. C. 5 2.97 3.33 3.33 3.37 10
3.21 3.39 3.52 3.40 15 3.22 3.47 3.59 3.41 20 3.21 3.51 3.53 3.39
30 3.17 3.46 3.63 3.14 *Room Temperature (20.degree. C.)
[0067] As may be seen from this data, extraction at elevated
temperature proceeded faster than at room temperature. Extraction
in terms of maximum protein concentration reached equilibrium
within 5 minutes at elevated temperatures, while extraction at room
temperature usually took 10 minutes. As the extraction temperature
rose from room temperature to 60.degree. C., the protein
concentration of the extract increased by over 10% while a further
rise in temperature resulted in a slightly decreased
extractability.
[0068] Based on the protein concentration data set forth in Table
I, protein extractabilities were calculated and the results appear
in the following Table II: TABLE-US-00002 TABLE II Protein
Extractability at Different Temperatures* Temperature (.degree. C.)
Extractability (wt %) RT 50.1 55 54.0 60 55.9 65 53.9 *Defined as
percentage of the amount of protein extracted as of the total
amount of protein in the meal
[0069] As may be seen from this data, the extractability of the
protein in the canola oil seed meal exceeded 50 wt % at all
temperatures tested, a considerable improvement over the maximum 30
wt % achieved with commercial toasted canola oil seed meal.
Example 2
[0070] This Example shows the effects of certain parameters on
protein extractability.
[0071] In a first set of experiments, 50 g samples of (a) canola
oil seed meal which had been air-desolventized at ambient
temperature (20.degree. C.) or (b) commercial canola oil seed meal
which had been desolventized by conventional toasting (toasted
commercial meal) were added to 500 mL samples of 0.05 M or 0.10 M
NaCl solution at room temperature (20.degree. C.) and stirred for
15 minutes. The slurry was centrifuged at 5000.times.g for 10
minutes to remove the spent meal.
[0072] In a second set of experiments, 500 mL of water with no salt
added was first heated to 60.degree. C. on a hot plate stirrer and
then (a) 50 g of canola oil seed meal which had been
air-desolventized at ambient temperature (20.degree. C.) Marc meal)
or (b) commercial canola oil seed meal which had been desolventized
by conventional toasting (commercial meal) was added and stirred
for 15 minutes while the temperature was maintained. The extract
was separated from the spent meal by centrifugation at 5000.times.g
for 10 minutes.
[0073] The protein concentration of the various aqueous protein
solutions obtained in these experiments were determined and appear
in the following Table V: TABLE-US-00003 TABLE V Protein
Concentrations in Extracts (wt %) 0.05 M saline 0.10 M saline
60.degree. C. water Ambient temperature 2.09 2.04 1.38
desolventized meal Toasted commercial meal 0.75 0.85 0.60
[0074] The protein extractability from the meals was determined
from the protein concentration data of Table V and this data is
presented in Table VI: TABLE-US-00004 TABLE VI Protein
Extractability (wt %)* 0.05 M saline 0.10 M saline 60.degree. C.
water Ambient temperature 49.6 48.4 32.7 desolventized meal Toasted
commercial meal 17.0 20.0 14.0 *Defined as percentage of the amount
of protein extracted as of the total amount of protein in the
meal.
[0075] Table VI shows that the protein extractability of the Marc
meal at both salt concentrations were comparable with a 15 wt %
meal and 0.15 M salt concentration at room temperature (see Table
II above). The protein extraction of the Marc meal at 0.05 M NaCl
was comparable with that at 0.10 M NaCl. In the case of no salt
added, the protein extractability was substantially lower at the
elevated temperature than that using 0.05 and 0.10 M salt at room
temperature. In all cases, however, the protein extractability and
protein concentrations were significantly higher than obtained with
toasted commercial meal.
[0076] A third set of experiments was performed at room temperature
in the same manner as the room temperature experiments described
above but a salt concentration of 0.01M, 0.02M, 0.03M, 0.04M and
0.05M. The protein extractabilities were determined for each
extract and the results appear in the following VII: TABLE-US-00005
TABLE VII Protein Extractability of Marc Meal at Low Salt
Concentration Salt Concentration (M) Protein Extractability (wt %)
0.05 49.6 0.04 43.4 0.03 38.8 0.02 40.3 0.01 38.5
[0077] As may be seen from the data presented in Table VII, a
substantial decrease in protein extractability was observed between
salt concentrations, of 0.04M and 0.05M, suggesting that a minimum
salt concentration to obtain a good yield of protein in the extract
solution is 0.05M.
[0078] A Varian high pressure liquid chromatography column (HPLC),
using a 30 cm BioSep S3000 Size Exclusion Chromatography (SEC)
column containing hydrophilic-bonded silica rigid support media,
5-micron diameter, 290-Angstrom pore size, capable of separating
globular proteins from 5,000 to 700,000 dalton size, was run with a
series of standards of protein origin to determine the residence
time (RT) of each component, as measured at A280 nm, at an elution
flow rate of 1.0 mL/min. The BioRad standard proteins cover a range
from 17,000 daltons (myoglobulin) to 670,000 daltons
(thyroglobulin) with Vitamin B12 added as a low molecular mass
marker at 1,350 daltons. Each component is measured at 280 nm at an
elution flow rate of 1.0 mL/min. Saline solution, pH adjusted and
containing sodium azide as an antibacterial agent, was used as the
column solvent and to dissolve dry samples. Eluant was discarded
after UV detection as only 25 to 50 microliters of sample are
required per run. The HPLC Prostar system automatically calculated
retention times and peak areas and printed out a summary
report.
[0079] Samples of the extracts prepared as described in this
Example were run on each column. The peak area counts were
converted to percentage for each peak. All peaks on different runs
were taken into calculation and then the three major protein
fractions, 12S, 7S and 2S, were recalculated separately. The
results obtained are shown in the graphical data of FIGS. 1 to
3.
[0080] Each chromatogram showed a distinct peak representing 7S
canola protein fraction and a small bump of 12S canola protein
fraction. The peak for the 2S canola protein fraction was present
among peaks for other components of the extract. The peaks in the
lower molecular weight end of the chromatogram were not properly
identified, but likely correspond to non-protein nitrogenous
compounds, such as short peptides and free amino acids, as well as
other meal components, such as phenolic compounds, glucosinolates
and phytates.
Example 3
[0081] This Example further illustrates the preparation of a canola
protein isolate using air-desolventized canola oil seed meal.
[0082] 160 kg of marc canola meal which had been air-desolventized
at 20.degree. C. was added to 1602 L of 0.15 M NaCl at 17.6.degree.
C. and agitated for 30 minutes to provide an aqueous protein
solution having a protein content of 21.4 g/L. 0.05 wt % of
ascorbic acid was added after 15 minutes of the extraction time.
The percentage protein in the meal which was extracted was
51.6%.
[0083] The residual canola meal was removed and washed on a vacuum
filter belt. The resulting protein solution was clarified by
centrifugation and filtration to produce 1270 L of a clarified
protein solution having a protein content of 16.2 g/L.
[0084] 1270 L of the protein extract solution was reduced in volume
to 71 L by concentration on an ultrafiltration system using 5000
dalton molecular weight cut-off membranes. The protein extract
solution then was diafiltered on a diafiltration system using 5000
dalton molecular weight cut-off membranes with 5000 L (5 retentate
volumes) of 0.15 M saline solution containing 0.05 wt % ascorbic
acid to a fmal volume of 31 L with a protein content of 226 g/L.
The retentate was pasteurized at 60.degree. C. for 10 minutes.
[0085] The concentrated and diafiltered solution was divided into
three batches of 30 L, 30 L and 8 L respectively. A first batch at
30.degree. C. was diluted 1:15 into 450 L of filtered water at
4.degree. C. A white cloud of protein micelles formed immediately
and was allowed to settle. The upper diluting water was removed.
This procedure was repeated for the second and third batches. The
precipitated, viscous, sticky mass (PMM) was removed from the
bottom of the vessel. The dried protein was found to have a protein
content of 102.4 wt % (N.times.6.25) d.b. (Percentage nitrogen
values were determined using a Leco FP 328 Nitrogen Determinator).
The product was given designation BW-AA020-C17-03A-C300.
[0086] 988 L of supernatant from the protein micelle formation were
concentrated to 38 L on a ultrafiltration system using 5000 dalton
molecular weight cut-off membranes. The concentrated supernatant
then was dilafiltered on a diafiltration system using 5000 dalton
molecular weight cut-off membranes with 130 L (4 retentate volumes)
of water to a final volume of 38 L with a protein content of 194
g/L.
[0087] The concentrated and diafiltered solution was diluted to a
pumpable consistency and was then spray dried. The dried protein
was found to have a protein content of 97.6 wt % (N.times.6.25)
d.b. The product was given designation BW-AA020-C17-03A-C200.
SUMMARY OF DISCLOSURE
[0088] In summary of this disclosure, the present invention
provides an improved process for making oil seed protein isolates
from oil seed meals by using an ambient temperature desolventized
meal to provide a greater degree of extraction of protein from the
meal leading to economic benefits. Modifications are possible
within the scope of this invention.
* * * * *