Isolation of proteins by ultrafiltration

• US 5,658,714 A
• Filed: 12/23/1992
• Issued: 08/19/1997
• Est. Priority Date: 02/28/1991
• Status: Expired due to Fees

• Current Assignee: Abbott Laboratories Incorporated
• Original Assignee: Abbott Laboratories Incorporated
• Inventors: Schimpf, Karen Joyce, Westfall, Peter Hilary, Nardelli, Christy Ann
• Primary Examiner(s): Russel, Jeffrey E.
• Application Number:
  US07/997,616
• Time in Patent Office: 1,700 Days
• Field of Search: 514/2, 530/350, 530/370, 530/372, 530/373, 530/374, 530/375, 530/376, 530/377, 530/378, 530/412, 530/414, 530/418, 530/427
• US Class Current: 530/378
• CPC Class Codes: A23J 1/14 from leguminous or other vegetable seeds; from press-cake or oil-bearing seeds
• CPC Class Codes: A23J 3/16 from soybean
• CPC Class Codes: A23L 11/30 Removing undesirable substances, e.g. bitter substances

Abstract

A high quality soy protein isolate with a significant reduction in phytate and aluminum is prepared via ultrafiltration. Defatted soy flour slurry is prepared and adjusted to a pH such that the protein becomes solubilized. The solubilized protein can pass through the ultrafiltration membrane. The ultrafiltration system rejects phytate and aluminum. Once the soluble protein passes through the ultrafiltration system the soy protein isolate is then precipitated from the clear permeate stream by adjusting the pH within the isoelectric range of soy proteins.

Claims

1. A method of isolating protein from a flour comprising the steps of:

(a) providing an aqueous slurry of a flour;

(b) adjusting the pH of the slurry to be in the range of about 7.0 to about 10.0 at a temperature in the range of about 38°

about 54°

C. such that the protein in the flour becomes solubilized;

(c) passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and aluminum;

(d) adjusting the pH of the permeate collected in step (c) to be in the range of about 3.5 to about 6.0 such that the protein precipitates; and

(e) collecting the protein precipitate of step (d), the phytate content of the precipitated protein of step (e) being less than 1.0%, by weight, as determined by the following procedure;

(i) a protein isolate sample is weighed and extracted in 2.4% by vol. hydrochloric acid for two hours;

(ii) the pH of the sample is then adjusted to be greater than 8, followed by quantitative transfer and dilution;

iii) the diluted sample is filtered through #2V filter paper and the filtrate is collected;

iv) an aliquot of the filtrate is injected into a mixed mode column and separation is achieved utilizing a 56 mM to 150 mM sodium hydroxide gradient, in the presence of 5% isopropyl alcohol by vol. at a flow rate of 1.0 ml per minute;

(v) using suppressed conductivity to detect phytate in the sample; and

vi) the phytate concentration of the sample is determined by comparing chromatographic data of the sample with standards of known concentrations of phytate.

2. A method of isolating protein from a flour according to claim 1 wherein step (e) comprises centrifuging the product of step (d) to separate the protein precipitate from the remainder of the permeate.

3. A method of isolating protein from a flour according to claim 1 further comprising the step (f) of transforming the protein precipitate collected in step (e) into a powdered form.

4. A method of isolating protein from a flour according to claim 2 further comprising the step (f) of transforming the protein precipitate collected in step (e) into a powdered form.

5. A method of isolating protein from a flour according to claim 1 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

6. A method of isolating protein from a flour according to claim 2 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

7. A method of isolating protein from a flour according to claim 3 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

8. A method of isolating protein from a flour according to claim 4 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

9. A method of isolating protein from a flour according to claim 1 wherein the flour in said slurry is milled from at least one plant selected from the group of plants consisting of maize, rice, sorghum, cowpeas, soybeans, cassava, coyam, sesame, peanuts, peas, cotton and yams.

18. A method of isolating protein from flour according to claim 1 wherein in step (c) the ultrafiltration device comprises a membrane having an initial permeability to water of about 8.0.

10. A method of isolating protein from soy flour comprising the steps of:

(a) providing an aqueous slurry comprising 1% to 30% by weight of soy flour;

(b) adjusting the pH of the slurry to be in the range of about 7.5 to about 9.5 at a temperature in the range of about 38°

to about 54°

C. such that protein in the slurry becomes solubilized;

(c) passing the slurry through an ultrafiltration device and collecting a permeate containing protein and carbohydrates and a retentate containing phytate and aluminum;

(d) adjusting the pH of the permeate collected in step (c) to be in the range of 3.5 to 6.0 and allowing the protein to precipitate; and

(e) separating the precipitated protein from the remainder of the permeate, the phytate content of the precipitated protein of step (e) being less than 1.0%, by weight, as determined by the following procedure;

(i) a protein isolate sample is weighed and extracted in 2.4% by vol. hydrochloric acid for two hours;

(ii) the pH of the sample is then adjusted to be greater than 8, followed by quantitative transfer and dilution;

(iii) the diluted sample is filtered through #2V filter paper and the filtrate is collected;

(iv) an aliquot of the filtrate is injected into a mixed mode column and separation is achieved utilizing a 56 mM to 150 mM sodium hydroxide gradient, in the presence of 5% isopropyl alcohol by vol. at a flow rate of 1.0 ml per minute;

(v) using suppressed conductivity to detect phytate in the sample; and

(vi) the phytate concentration of the sample is determined by comparing chromatographic data of the sample with standards of known concentrations of phytate.

11. A method of isolating proteins from soy flour according to claim 10 further comprising the step (f) of transforming the precipitated protein into a powdered form.

12. A method of isolating protein from soy flour according to claim 10 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and aluminum and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

13. A method of isolating protein from soy flour according to claim 11 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and aluminum and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that permeates are collected and passing said retentate through the ultrafiltration device.

19. A method of isolating protein from soy flour according to claim 10 wherein in step (c) the ultrafiltration device comprises a membrane having an initial permeability to water of about 8.0.

20. A method of isolating protein from soy flour according to claim 11 wherein in step (c) the ultrafiltration device comprises a membrane having an initial permeability to water of about 8.0.

14. A method of isolating protein from soy flour comprising the steps of:

(a) providing an aqueous slurry comprising 7% to 15% by weight of defatted soy flour;

(b) adjusting the pH of the slurry to be in the range of 7.5 to 9.5 at a temperature in the range of about 38°

to about 54°

C. such that protein in the slurry becomes solubilized;

(c) passing the slurry through an ultrafiltration device and collecting a permeate containing protein and carbohydrates, and a retentate containing phytate and aluminum;

(d) adjusting the pH of the permeate collected in step (c) to be in the range of 4.2 to 4.7 and allowing the protein to precipitate; and

,

(e) separating the precipitated protein from the remainder of the permeate, the phytate content of the precipitated protein of step (e) being less than 1.0%, by weight, as determined by the following procedure;

(i) a protein isolate sample is weighed and extracted in 2.4% by vol. hydrochloric acid for two hours;

(ii) the pH of the sample is then adjusted to be greater than 8, followed by quantitative transfer and dilution;

(iii) the diluted sample is filtered through #2V filter paper and the filtrate is collected;

(iv) an aliquot of the filtrate is injected into a mixed mode column and separation is achieved utilizing a 56 mM to 150 mM sodium hydroxide gradient, in the presence of 5% isopropyl alcohol at a flow rate of 1.0 ml per minute;

(v) Using suppressed conductivity to detect phytate in the sample; and

(vi) the phytate concentration of the sample is determined by comparing chromatographic data of the sample with standards of known concentrations of phytate.

15. A method of isolating proteins from soy flour according to claim 14 further comprising the step (f) of transforming the precipitated protein into a powdered form.

16. A method of isolating protein from a soy flour according to claim 14 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

17. A method of isolating protein from a flour according to claim 15 wherein step (c) comprises passing the slurry through an ultrafiltration device and collecting a permeate containing protein and a retentate containing phytate and thereafter adding a diafiltration fluid to the retentate at substantially the same rate that the permeate is collected and passing said retentate through the ultrafiltration device.

21. A method of isolating protein from soy flour according to claim 14 wherein in step (c) the ultrafiltration device comprises a membrane having an initial permeability to water of about 8.0.

22. A method of isolating protein from soy flour according to claim 15 wherein in step (c) the ultrafiltration device comprises a membrane having an initial permeability to water of about 8.0.