Production of pulse protein product with reduced astringency

Abstract

Pulse proteins of reduced astringency are obtained by fractionating pulse protein products which are completely soluble and heat stable in aqueous media at acid pH value of less than about 4.4 into lower molecular weight, less astringent proteins and higher molecular weight, more astringent proteins.

Claims

1. A method of preparing pulse protein product with reduced astringency when tasted in aqueous solution at a pH below about 5, which comprises: (a) extracting a pulse protein source with an aqueous calcium salt solution to cause solubilization of pulse protein from the protein source and to form an aqueous pulse protein solution, (b) separating the aqueous pulse protein solution from residual pulse protein source, (c) optionally diluting the aqueous pulse protein solution, (d) adjusting the pH of the aqueous pulse protein solution to a pH of about 1.5 to about 4.4 to produce an acidified pulse protein solution, (e) optionally clarifying the acidified pulse protein solution if it is not already clear, (f) alternatively from steps (b) to (e), optionally, diluting and then adjusting the pH of the combined aqueous pulse protein solution and residual pulse protein source to a pH of about 1.5 to about 4.4 and then separating the acidified pulse protein solution from residual pulse protein source, and (g) fractionating the proteins in the acidified pulse protein solution to separate lower molecular weight, less astringent proteins from higher molecular weight, more astringent proteins, wherein said fractionation step is effected by: (i) membrane processing the acidified aqueous pulse protein solution to fractionate the protein components of the acidified aqueous pulse protein solution into a higher molecular weight fraction in a first retentate and a lower molecular weight fraction in a first permeate, (ii) membrane processing the first permeate to retain the lower molecular weight fraction protein components in a second retentate and to permit contaminants to pass the membrane in a second permeate, (iii) drying the second retentate to provide a pulse protein product of reduced astringency.

2. The method of claim 1, wherein the membrane processing step (i) is effected by microfiltration using membranes having a pore size of about 0.05 to about 0.1 ?m or ultrafiltration using membrane with a molecular weight cut-off of about 10,000 to about 1,000,000 daltons to concentrate the acidified aqueous pulse protein solution to a protein concentration of about 50 to about 300 g/L to provide a concentrated retentate.

3. The method of claim 2, wherein the concentrated retentate is subjected to a diafiltration step using about 1 to about 40 volumes of diafiltration solution.

4. The method of claim 1, wherein the membrane processing of the first permeate in step (ii) is effected by ultrafiltration to concentrate the first permeate to a concentration of about 10 to about 300 g/L followed by optional diafiltration, or to a partial concentration of less than about 10 g/L, using membranes having a molecular weight cut-off of about 1,000 to about 100,000 daltons.

5. The method of claim 1, wherein the first retentate from step (i) is further processed by a step selected from the group consisting of: (i) drying the first retentate, and (ii) adjusting the pH of the first retentate to a pH of about 6 to about 8, and drying the pH-adjusted retentate.

6. The method of claim 1 wherein the membrane processing step (i) is effected by microfiltration using membranes having a pore size of about 0.08 to about 0.1 ?m or ultrafiltration using membrane processing of a molecular weight cut-off of about 100,000 to about 1,000,000 daltons to concentrate the acidified aqueous pulse protein solution to a protein concentrate of about 100 to about 200 g/L to provide a concentrated retentate.

7. The method of claim 2 wherein the concentrated retentate is subjected to a diafiltration step using about 2 to about 25 volumes of diafiltration solution.

8. The method of claim 4, wherein the membrane processing of a first permeate in step (ii) is effected by ultrafiltration to concentrate the first permeate to a concentration of about 100 to about 200 g/L, followed by optional diafiltration, or to a partial concentration of less than about 10 g/L, using membrane having a molecular weight cut-off of about 1,000 to about 10,000 daltons.

Description

EXAMPLES

Example 1

(1) This Example illustrates production of the reduced astringency pulse protein product of the invention utilizing methods where the acidified pulse protein solution is partially concentrated or concentrated and diafiltered prior to the precipitation of the more astringent protein by pH adjustment.

(2) a kg of b was combined with c L of reverse osmosis purified (RO) water and the mixture stirred for d minutes at ambient temperature. Insoluble material was removed and the sample partially clarified by centrifugation, yielding a protein solution having a protein concentration of e wt %. To this protein solution was added f kg of calcium chloride stock solution, prepared by dissolving 1 kg calcium chloride pellets (95.5%) per 9 L water g. Insoluble material was removed and the sample clarified by centrifugation, yielding h L of protein extract solution having a protein concentration of i wt %. j L of protein extract solution was combined with k L of RO water and the pH of the sample lowered to l with HCl solution (concentrated HCl diluted with an equal volume of water). m L of acidified protein solution was clarified by running it on a microfiltration system equipped with a 0.80 ?m pore size Membralox ceramic membrane operated at n ? C. until o L of permeate (clarified, acidified protein solution) was collected. p L of q, having a protein content of r wt % was s concentrated to t L using a PES ultrafiltration membrane having a pore size of 1,000 daltons operated at a temperature of about u ? C. v L of w concentrated protein solution was then diafiltered with x L of RO water at about y ? C. to provide z of diafiltered, aa concentrated protein solution having a protein content of ab wt %. The diafiltered, ac concentrated protein solution was diluted with ad L RO water and the pH adjusted to ae with NaOH solution, which caused the formation of a precipitate. af kg of wet precipitate was removed by centrifugation to provide ag L of protein solution with a protein content of ah wt %. The pH of the protein solution was lowered to ai and then aj L of re-acidified protein solution was polished by running the solution through a Membralox ceramic microfiltration membrane having a pore size of 0.80 ?m and operated at ak ? C. until al L of permeate was collected. am L of an was then reduced in volume to ao L by concentration on a PES ultrafiltration membrane having a pore size of 1,000 daltons operated at a temperature of about ap ? C. The resulting aq concentrated protein solution, having a protein content of ar wt % was then diafiltered with as L of RO water at about at ? C. au to provide av kg of concentrated, diafiltered protein solution having a protein content of aw wt %. This represented a yield of ax % of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. ay kg of concentrated, diafiltered protein solution was spray dried to yield a protein product, having a protein content of az % (N?6.25) d.b., termed ba bb.

(3) The af kg of wet precipitate collected, having a protein content of bc, represented a yield of bd % of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. be kg of this precipitate was diluted with bf kg water then the pH adjusted to bg and the mixture pasteurized at about bh for bi minutes. The bj sample was then spray dried to provide a dried protein product having a protein content of bk% (N?625) d.b. that was termed ba bl.

(4) The parameters a to bl are set forth in the following Table 1.

(5) TABLE-US-00001 TABLE 1 Parameters for the production of protein products by the precipitation fractionation method ba YP20-D23-13A YP20-D24-13A YP20-E02-13A LE03-D02-14A a 30 30 60 36 b yellow pea yellow pea yellow pea whole green protein protein protein lentil flour concentrate concentrate concentrate c 500 500 1000 600 d 30 30 30 10 e 2.69 2.68 2.67 1.27 f 63.14 65 137.34 80 g and the and the and the N/A mixture mixture mixture stirred stirred stirred 15 minutes 15 minutes 15 minutes h 459 484 978 586 i 1.60 1.41 1.55 0.68 j 459 484 978 586 k 371 317 640 368 l 2.91 3.12 3.00 3.02 m 830 790 N/A N/A n 59 59 N/A N/A o NR NR N/A N/A p 780 700 1585 975 q clarified clarified acidified acidified acidified acidified protein protein protein protein solution solution solution solution r 0.81 0.74 0.81 0.40 s partially N/A N/A partially t 120 72 215 50 u 57 57 58 58 v 120 72 215 50 w partially N/A N/A partially x 240 144 430 100 y 60 61 59 60 z 120 L 72 L 220 L 48.56 kg aa partially N/A N/A N/A ab 4.04 5.57 5.62 5.13 ac partially N/A N/A N/A ad 120 78 344 NR ae 5.63 5.73 about 5.5 6.10 af 33.50 31.12 105.36 16.14 ag 230.1 128.5 444 80 ah 0.40 0.51 0.36 0.65 ai 3.08 2.79 3.11 2.95 aj N/A N/A N/A 80 ak N/A N/A N/A 46 al N/A N/A N/A 64 am 230 150 444 64 an re-acidified re-acidified re-acidified clarified, protein protein protein re-acidified solution solution solution protein solution ao 78 25 32.5 22 ap 58 52 54 58 aq N/A N/A N/A partially ar 1.16 1.91 4.62 0.62 as 78 25 32.5 22 at 60 59 60 59 au and then N/A and then N/A further further concentrated concentrated av 34.56 29.14 24.86 21.00 aw 2.87 2.38 6.25 1.51 ax 13.5 10.1 10.2 8.0 ay 35.54 29.14 24.86 21.00 az 100.17 99.36 101.84 92.26 bb YP705 YP705 YP705 LE705 bc 12.33 11.65 10.38 11.18 bd 56.3 53.2 72.2 45.2 be 8.5 8.94 24 16.14 bf 8.5 8.94 0 8.00 bg 7.07 6.82 N/A N/A bh N/A N/A N/A 66 bi N/A N/A N/A 15 bj N/A N/A N/A pasteurized bk 102.58 102.49 101.44 102.08 bl YP705P YP705P YP705P LE705P NA = not applicable NR = not recorded

Example 2

(6) This Example illustrates production of the reduced astringency pulse protein product of the invention according to the procedure where the acidified pulse protein solution is pH adjusted to precipitate the more astringent protein.

(7) 18 kg of yellow pea protein concentrate was combined with 300 L of reverse osmosis purified (RO) water and the mixture stirred for 30 minutes at ambient temperature. Insoluble material was removed and the sample partially clarified by centrifugation, yielding a protein solution having a protein concentration of 2.47 wt %. To this protein solution was added 51.1 kg of calcium chloride stock solution, prepared by dissolving 8.0 kg calcium chloride pellets (95.5%) in 72 L water. Insoluble material was removed and the sample clarified by centrifugation, yielding 295 L of protein extract solution having a protein concentration of about 1.32 wt %. The 295 L of protein extract solution was combined with 206 L of RO water and the pH of the sample lowered to 2.75 with HCl solution (concentrated HC diluted with an equal volume of water). 495 L of acidified protein solution having a protein content of 0.66 wt %, was then adjusted to pH 5.5 using 2M NaOH solution, resulting in the formation of a precipitate. 24.92 kg of precipitate was collected by centrifugation yielding 480 L of pulse protein solution having a protein concentration of 0.20 wt %. The pH of the sample was then adjusted to about 3 with diluted HCl solution and then 480 L of re-acidified pulse protein solution was concentrated to 28 L using a PES ultrafiltration membrane having a pore size of 1,000 daltons operated at a temperature of about 58? C. 28 L of concentrated protein solution was then diafiltered with 28 L of RO water at about 63? C. and further concentrated to provide 19.94 kg of concentrated, diafiltered protein solution having a protein content of 6.52 wt %. This represented a yield of 33.4% of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. 19.94 kg of concentrated, diafiltered protein solution was spray dried to yield a protein product, having a protein content of 96.07% (N?6.25) d.b., termed YP20-E13-13A YP705.

(8) The 24.92 kg of wet precipitate collected, having a protein content of 7.83 wt % represented a yield of 50.1% of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. A14.76 kg aliquot of the precipitate was washed with an equal weight of RO water and then re-captured by centrifugation. This washed precipitate was suspended in fresh water and then spray dried. The dried protein product had a protein content of 95.02% (N?6.25) d.b. and was termed YP20-E13-13A YP705P-01. A second aliquot (10 kg) of the precipitate was suspended in water and spray dried without a wash step. The dried protein product had a protein content of 87.52 (N?6.25) d.b. and was termed YP20-E13-13A YP705P-02.

Example 3

(9) This Example illustrates production of the reduced astringency pulse protein product of the invention according to the procedure where membrane processing is utilized to separate the less astringent proteins from the more astringent proteins. a kg of b was combined with c L of reverse osmosis purified (RO) water and the mixture stirred for 10 minutes at ambient temperature. Insoluble material was removed and the sample partially clarified by centrifugation, yielding a protein solution having a protein concentration of d wt %. To this protein solution was added e g antifoam and f kg of calcium chloride stock solution, prepared by dissolving g kg calcium chloride pellets (95.5%) in h L water. Insoluble material was removed and the sample clarified by centrifugation, yielding i L of protein extract solution having a protein concentration of j wt %. k L of protein extract solution was combined with l L of RO water and the pH of the sample lowered to about m with HCl solution (concentrated HCl diluted with an equal volume of water). n L of acidified pulse protein solution, having a protein concentration of o wt %, was concentrated to p using a polyvinylidene fluoride (PVDF) microfiltration membrane having a pore size of 0.08 ?m operated at a temperature of about q ? C. The microfiltration retentate was then diafiltered with r L of RO water at about s ? C. and then the diafiltered retentate further reduced to t kg at about u ? C. v L of microfiltration/diafiltration permeate, having a protein concentration of w wt %, was concentrated to x L using a PES ultrafiltration membrane having a pore size of 1,000 daltons operated at a temperature of about y C. The concentrated protein solution was then diafiltered with z L of RO water at about aa ? C. ab to provide ac kg of concentrated, diafiltered protein solution having a protein content of ad wt %. This represented a yield of ac % of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. af kg of concentrated, diafiltered protein solution was spray dried to yield a protein product, having a protein content of ag % (N?6.25) d.b., termed ah ai.

(10) The aj kg of ak microfiltration retentate collected, having a protein content of al wt % represented a yield of am % of the protein in the protein extract solution resulting from the clarification step after calcium chloride addition. an kg of concentrated and diafiltered microfiltration retentate was adjusted to pH ao and then spray dried to form a protein product having a protein content of ap % (N?6.25) d.b., termed ah aq

(11) The parameters a to ao are set forth in the following Table 2.

(12) TABLE-US-00002 TABLE 2 Parameters for the production of protein products by the membrane fractionation method ah YP23-H12-13A YP23-H14-13A YP23-J02-13A LE03-D01-14A a 24 24 60 36 b yellow pea yellow pea yellow pea whole green protein protein protein lentil flour concentrate concentrate concentrate c 400 400 1008 600 d 3.11 2.92 3.16 1.25 e N/A N/A 19 N/A f 54.6 56.0 135 79.36 g 6 6 20 10 h 54 54 180 90 i 398 398.8 934 604 j 1.66 1.60 about 1.90 0.61 k 398 398.8 934 604 l 269 278.2 666 398 m 3.17 3.16 2.99 3.01 n 670 490 1440 1025 o 0.86 0.91 0.83 0.30 p 65 L 28.04 kg 180 L 35 L q 59 55 55 56 r N/A N/A 180 80 s N/A N/A 55 55 t N/A N/A 140 N/A u N/A N/A 55 N/A v 600 458 about 1470 1052 w 0.18 0.29 0.31 0.27 x 28 30 40 48 y 56 54 56 54 z 140 150 200 96 aa 59 59 58 61 ab and further N/A and further and further concentrated concentrated concentrated ac 21.36 32.35 33.6 32.08 ad 3.43 2.44 5.02 2.09 ae 11.0 12.4 9.5 18.2 af 21.36 32.35 33.6 32.08 ag 101.64 98.24 99.78 93.52 ai YP706 YP706 YP706 LE706 aj 65 L 28.04 kg 140 L 32.12 ak concentrated concentrated concentrated concentrated and and diafiltered diafiltered al 7.02 9.45 6.63 4.87 am 69.0 41.5 52.3 42.4 an N/A N/A 135 32.12 ao N/A N/A about 7 7.29 ap N/A N/A 91.60 94.64 aq N/A N/A YP706B LE706B N/A = not applicable

Example 4

(13) This Example contains an evaluation of the dry colour and colour in solution of the reduced astringency pulse protein products produced by the methods of Examples 1-3.

(14) The colour of the dry powders was assessed using a HunterLab ColorQuest XE instrument in reflectance mode. The colour values are set forth in the following Table 3:

(15) TABLE-US-00003 TABLE 3 HunterLab scores for dry reduced astringency pulse protein products Sample L* a* b* YP20-D23-13A YP705 89.33 0.02 5.75 YP20-D24-13A YP705 88.55 ?0.14 5.73 YP20-E02-13A YP705 89.14 0.26 6.68 YP20-E13-13A YP705 86.90 0.90 8.55 LE03-D02-14A LE705 88.09 1.07 5.54 YP23-H12-13A YP706 88.23 ?0.09 6.35 YP23-H14-13A YP706 88.53 0.22 6.78 YP23-J02-13A YP706 87.25 0.75 7.45 LE03-D01-14A LE706 85.94 0.84 7.92

(16) As may be seen from Table 3, the reduced astringency pulse protein products were light in colour.

(17) Solutions of the reduced astringency pulse protein products were prepared by dissolving sufficient protein powder to supply 0.48 g of protein in 15 ml of RO water. The pH of the solutions was measured with a pH meter and the colour and clarity assessed using a HunterLab Color Quest XE instrument operated in transmission mode. The results are shown in the following Table 4.

(18) TABLE-US-00004 TABLE 4 pH and HunterLab scores for solutions of reduced astringency pulse protein products sample pH L* a* b* haze YP20-D23-13A YP705 3.35 97.2 ?0.10 6.42 22.9 YP20-D24-13A YP705 2.93 97.91 ?0.40 5.81 8.2 YP20-E02-13A YP705 3.39 97.76 ?0.33 5.52 9.9 YP20-E13-13A YP705 3.26 95.33 0.05 9.69 29.8 LE03-D02-14A LE705 3.21 96.33 0.66 7.18 4.5 YP23-H12-13A YP706 3.72 94.65 0.01 9.20 14.9 YP23-H14-13A YP706 3.57 96.07 ?0.25 8.99 7.7 YP23-J02-13A YP706 3.51 96.55 0.09 9.7 17.2 LE03-D01-14A LE706 3.42 93.86 0.60 12.8 21.5

(19) As may be seen from the results in Table 4, the solutions of the reduced astringency pulse protein products were light in colour and generally low in haze.

Example 5

(20) This Example contains an evaluation of the solubility in water of the reduced astringency pulse protein products produced by the methods of Examples 1 and 3. Solubility was tested based on protein solubility (termed protein method, a modified version of the procedure of Morr et al., J. Food Sci. 50:1715-1718) and total product solubility (termed pellet method).

(21) Sufficient protein powder to supply 0.5 g of protein was weighed into a beaker and wetted by mixing with about 20-25 ml of reverse osmosis (RO) purified water. 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 periodically 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% w/v protein dispersion. The protein content of the dispersions was determined by combustion analysis using a Leco Nitrogen Determinator. Aliquots (20 ml) of the dispersions were then transferred to pre-weighed centrifuge tubes that had been dried overnight in a 100? 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 supernatant. The protein content of the supernatant was measured by combustion analysis and then the supernatant and the tube lids were discarded and the pellet material dried overnight in an oven set at 100? 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:
Solubility (protein method) (%)=(% protein in supernatant/% protein in initial dispersion)?1001)
Solubility (pellet method) (%)=(1?(weight dry insoluble pellet material/((weight of 20 ml of dispersion/weight of 50 ml of dispersion)?initial weight dry protein powder)))?1002)
Values calculated as greater than 100% were reported as 100%.

(22) The natural pH values of the 1% w/v protein solutions of the protein products produced in Examples 1 and 3 are shown in Table 5:

(23) TABLE-US-00005 TABLE 5 Natural pH of reduced astringency pulse solutions prepared in water at 1% protein Batch Product Natural pH YP20-D23-13A YP705 3.36 YP20-D24-13A YP705 3.15 YP20-E02-13A YP705 3.22 LE03-D02-14A LE705 3.19 YP23-H12-13A YP706 3.74 YP23-H14-13A YP706 3.53 LE03-D01-14A LE706 3.40

(24) The solubility results obtained are set forth in the following Tables 6 and 7:

(25) TABLE-US-00006 TABLE 6 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 YP20- YP705 100 100 95.4 94.4 90.1 96.1 98.1 D23-13A YP20- YP705 98.0 100 100 100 93.7 98.1 100 D24-13A YP20- YP705 96.9 100 100 99.0 98.9 93.1 100 E02-13A LE03- LE705 98.0 100 99.1 95.9 100 99.0 96.1 D02-14A YP23- YP706 99.0 100 100 80.2 78.4 92.9 95.2 H12-13A YP23- YP706 100 100 99.0 73.2 77.8 82.7 100 H14-13A LE03- LE706 93.3 100 100 64.6 59.8 64.6 100 D01-14A

(26) TABLE-US-00007 TABLE 7 Solubility of products at different pH values based on pellet method Solubility (pellet method) (%) Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 Nat. pH YP20- YP705 97.4 98.8 98.4 94.8 92.9 93.7 98.6 D23-13A YP20- YP705 99.8 100 99.3 98.4 97.4 98.4 99.4 D24-13A YP20- YP705 99.8 99.8 100 96.4 96.9 97.9 99.1 E02-13A LE03- LE705 99.9 100 99.4 94.4 96.3 95.7 99.6 D02-14A YP23- YP706 99.8 99.9 99.1 82.0 79.7 87.8 100 H12-13A YP23- YP706 97.8 97.7 98.5 67.9 81.7 75.0 98.9 H14-13A LE03- LE706 96.8 97.2 96.3 67.1 54.7 68.6 97.4 D01-14A

(27) As can be seen from the results presented in Tables 6 and 7, the reduced astringency pulse protein products were extremely soluble in the pH range 2-4 and also quite soluble in the pH range of 5-7.

Example 6

(28) This Example contains an evaluation of the clarity in water of the reduced astringency pulse protein products produced by the methods of Examples 1 and 3.

(29) The clarity of the 1% w/v protein solutions prepared as described in Example 5 was assessed by measuring the absorbance at 600 nm (water blank), with a lower absorbance score indicating greater clarity. Analysis of the samples on a HunterLab ColorQuest XE instrument in transmission mode also provided a percentage haze reading, another measure of clarity.

(30) The clarity results are set forth in the following Tables 8 and 9:

(31) TABLE-US-00008 TABLE 8 Clarity of protein solutions at different pH values as assessed by A600 A600 Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 Nat. pH YP20- YP705 0.008 0.016 0.029 0.337 0.807 0.596 0.022 D23-13A YP20- YP705 0.013 0.012 0.021 0.076 0.309 0.213 0.012 D24-13A YP20- YP705 0.007 0.011 0.014 0.063 0.506 0.369 0.012 E02-13A LE03- LE705 0.010 0.012 0.073 0.062 0.027 0.026 0.014 D02-14A YP23- YP706 0.008 0.016 0.034 1.923 1.889 0.791 0.033 H12-13A YP23- YP706 0.011 0.015 0.024 1.931 1.690 1.577 0.018 H14-13A LE03- LE706 0.019 0.025 0.050 2.424 2.412 2.426 0.024 D01-14A

(32) TABLE-US-00009 TABLE 9 Clarity of protein solutions at different pH values as assessed by HunterLab haze analysis HunterLab haze reading (%) Batch Product pH 2 pH 3 pH 4 pH 5 pH 6 pH 7 Nat. pH YP20- YP705 0.5 5.6 13.0 73.6 90.8 85.3 9.7 D23-13A YP20- YP705 0.0 1.7 6.4 23.1 65.7 50.3 2.2 D24-13A YP20- YP705 0.0 0.8 3.2 16.0 79.5 68.5 1.0 E02-13A LE03- LE705 0.3 1.2 19.8 16.7 3.6 1.8 1.8 D02-14A YP23- YP706 0.0 1.0 4.4 96.0 95.8 87.9 4.7 H12-13A YP23- YP706 0.0 0.5 2.3 95.9 95.7 95.5 1.1 H14-13A LE03- LE706 3.3 4.9 12.6 100.3 101.3 101.3 4.3 D01-14A

(33) As can be seen from the results of Tables 8 and 9, the reduced astringency pulse protein products generally provided transparent solutions at pH 2-4.

Example 7

(34) This Example contains an evaluation of the solubility in a soft drink (Sprite) and sports drink (Orange Gatorade) of the reduced astringency pulse protein products produced by the methods of Examples 1 and 3. 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.

(35) 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 wetted by mixing with about 20-25 ml of beverage. Additional beverage was then 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 determined by combustion analysis using a Leco 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)?100.

(36) Values calculated as greater than 100% were reported as 100%.

(37) When the solubility was assessed with pH correction, the pH of the soft drink (Sprite) and sports drink (Orange Gatorade) without protein was measured. A sufficient amount of protein powder to supply 1 g of protein was weighed into a beaker and wetted by mixing with about 20-25 ml of beverage. 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 determined immediately after dispersing the protein and was adjusted to the original no-protein pH with HCl or NaOH as necessary. The pH was measured and corrected periodically during the 60 minutes stirring. After the 60 minutes of stirring, the total volume of each solution was brought to 50 ml with additional beverage, yielding a 2% protein w/v dispersion. The protein content of the samples was determined by combustion analysis using a Leco 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)?100
Values calculated as greater than 100% were reported as 100%.

(38) The results obtained are set forth in the following Table 10:

(39) TABLE-US-00010 TABLE 10 Solubility of reduced astringency pulse protein products in Sprite and Orange Gatorade no pH correction pH correction Solubility Solubility Solubility (%) in Solubility (%) in (%) in Orange (%) in Orange Batch Product Sprite Gatorade Sprite Gatorade YP20-D23-13A YP705 100 98.0 97.0 100 YP20-D24-13A YP705 100 97.5 99.5 99.0 YP20-E02-13A YP705 100 100 100 100 LE03-D02-14A LE705 100 100 98.5 100 YP23-H12-13A YP706 100 99.0 97.0 96.0 YP23-H14-13A YP706 98.5 99.5 98.0 92.1 LE03-D01-14A LE706 92.6 98.9 93.3 100

(40) As can be seen from the results of Table 10, the reduced astringency pulse protein products were highly soluble in the Sprite and the Orange Gatorade.

Example 8

(41) This Example contains an evaluation of the clarity in a soft drink and sports drink of the reduced astringency pulse protein products produced by the methods of Examples 1 and 3.

(42) The clarity of the 2% w/v protein dispersions prepared in soft drink (Sprite) and sports drink (Orange Gatorade) in Example 7 were assessed using the HunterLab haze method described in Example 6.

(43) The results obtained are set forth in the following Table 11:

(44) TABLE-US-00011 TABLE 11 HunterLab haze readings for reduced astringency pulse protein products in Sprite and Orange Gatorade no pH correction pH correction Haze Haze Haze (%) in Haze (%) in (%) in Orange (%) in Orange Batch Product Sprite Gatorade Sprite Gatorade no protein 0.0 82.6 0.0 82.6 YP20-D23-13A YP705 17.8 70.6 21.8 72.2 YP20-D24-13A YP705 9.4 79.7 12.5 76.3 YP20-E02-13A YP705 8.5 86.2 20.2 86.5 LE03-D02-14A LE705 1.4 85.4 1.7 85.0 YP23-H12-13A YP706 10.2 84.7 6.4 79.9 YP23-H14-13A YP706 4.5 80.6 7.3 78.7 LE03-D01-14A LE706 11.5 77.5 12.1 78.9

(45) As can be seen from the results of Table 11, the addition of the reduced astringency pulse protein products to the soft drink and sports drink added little or no haziness.

Example 9

(46) This Example contains an evaluation of the heat stability in water of the reduced astringency pulse protein products produced by the methods of Examples 1 and 3.

(47) 2% w/v protein solutions of the protein products were prepared in RO water. The pH of the solutions was determined with a pH meter and then adjusted to about 3.0 with HCl solution. The clarity of the solutions was assessed by haze measurement with the HunterLab Color Quest XE instrument operated in transmission mode. The solutions were then heated to 95? 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.

(48) The clarity of the protein solutions before and after heating is set forth in the following Table 12:

(49) TABLE-US-00012 TABLE 12 Effect of heat treatment on clarity of 2% w/v protein solutions of reduced astringency pulse protein products haze before heat haze after heat Batch Product treatment (%) treatment (%) YP20-D23-13A YP705 13.0 0.0 YP20-D24-13A YP705 4.2 0.0 YP20-E02-13A YP705 5.5 1.4 LE03-D02-14A LE705 1.0 0.0 YP23-H12-13A YP706 5.0 2.0 YP23-H14-13A YP706 3.3 2.2 LE03-D01-14A LE706 6.3 1.6

(50) As can be seen from the results in Table 13, the solutions of reduced astringency pulse protein product were substantially clear before heat treatment and the level of haze was actually reduced by the heat treatment.

Example 10

(51) This Example illustrates the production of pulse protein products by the method described in U.S. patent application Ser. No. 13/556,357.

(52) a kg of b was combined with c L of d at e and agitated for f minutes. g kg of calcium chloride pellets (95.5%) dissolved in h L of RO water was then added and the mixture stirred for an additional i minutes. The residual solids were removed by centrifugation to produce a centrate having a protein content of j % by weight. k L of centrate was added to l L of RO water at m and the pH of the sample lowered to n with diluted HCl. The diluted and acidified centrate was further clarified by filtration to provide a clear protein solution with a protein content of o % by weight.

(53) The filtered protein solution was reduced in volume from p L to q L by concentration on a polyethersulfone membrane, having a molecular weight cutoff of r daltons, operated at a temperature of about s ? C. At this point the protein solution, with a protein content of t wt %, was diafiltered with u L of RO water, with the diafiltration operation conducted at about v ? C. The diafiltered protein solution was then further concentrated to w kg, having a protein content of x wt %, then diluted with RO water to a protein content of y wt % to facilitate spray drying. The protein solution before spray drying, having a weight of z kg was recovered in a yield of aa % of the initial centrate that was further processed. The concentrated and diafiltered protein solution was then dried to yield a product found to have a protein content of ab wt % (N?6.25) d.b. The product was given designation ac.

(54) The parameters a to ac are set forth in the following Table 13.

(55) TABLE-US-00013 TABLE 13 Parameters for the runs to produce pulse 701 products ac YP01-E19-11A YP05-A18-12A LE01-J24-13A YP701 YP701 LE701 a 20 70 20 b Yellow split Yellow split whole green pea flour pea flour lentil flour c 200 300 200 d 0.15M CaCl.sub.2 RO water 0.13M CaCl.sub.2 e 60? C. 30? C. Ambient temperature f 30 60 30 g 0 4.52 0 h 0 10 0 i 0 30 0 j 1.32 2.92 1.65 k 186.5 223.3 146.2 l 225.8 223.0 147.7 m 60? C. Ambient Ambient temperature temperature n 3.34 3.04 2.65 o 0.58 1.25 0.62 p 400 550 295 q 35 101 25 r 100,000 10,000 100,000 s 58 53 30 t 4.94 4.05 4.23 u 350 202 250 v 60 53 32 w 21.52 34.78 21.60 x 7.54 10.02 4.69 y N/A 5.00 N/A z 21.52 57.90 21.60 aa 65.9 44.5 41.9 ab 103.19 101.99 103.11 N/A = not applicable

Example 11

(56) This Example illustrates a comparison of the astringency level of the YP20-D24-13A YP705 prepared as described in Example 1 with that of the YP01-E19-11A YP701 prepared as described in Example 10.

(57) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP705 solution was 3.09 and it was adjusted to about 3.50 with food grade sodium hydroxide solution. The initial pH of the YP701 solution was 3.92 and it was adjusted to about 3.50 with food grade hydrochloric acid. An informal panel of seven panellists was asked to blindly taste the samples and indicate which was less astringent.

(58) Five out of seven panellists indicated that the YP20-D24-13A YP705 was less astringent.

Example 12

(59) This Example illustrates a comparison of the astringency level of the YP20-E02-13A YP705 prepared as described in Example 1 with that of the YP01-E19-11A YP701 prepared as described in Example 10.

(60) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP705 solution was 3.38 and it was adjusted to about 3.50 with food grade sodium hydroxide solution. The initial pH of the YP701 solution was 3.94 and it was adjusted to about 3.50 with food grade hydrochloric acid. An informal panel of seven panellists was asked to blindly taste the samples and indicate which was less astringent.

(61) Five out of seven panellists indicated that the YP20-E02-13A YP705 was less astringent.

Example 13

(62) This Example illustrates a comparison of the astringency level of the YP20-E13-13A YP705 prepared as described in Example 2 with that of the YP05-A18-12A YP701 prepared as described in Example 10.

(63) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The two samples had pH values within 0.1 units of each other so no pH adjustment was done. An informal panel of eight panellists was asked to blindly taste the samples and indicate which was less astringent. The experiment was conducted a second time with a panel often members. The cumulative results are presented below.

(64) Eleven out of eighteen panellists indicated that the YP20-E13-13A YP705 was less astringent.

Example 14

(65) This Example illustrates a comparison of the astringency level of the YP20-H12-13A YP706 prepared as described in Example 1 with that of the YP05-A18-12A YP701 prepared as described in Example 10.

(66) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP706 solution was 3.72 and it was adjusted to about 3.50 with food grade hydrochloric acid. The initial pH of the YP701 solution was 3.17 and it was adjusted to about 3.50 with food grade sodium hydroxide solution. An informal panel of seven panellists was asked to blindly taste the samples and indicate which was less astringent.

(67) Four out of seven panellists indicated that the YP20-H12-13A YP706 was less astringent.

Example 15

(68) This Example illustrates a comparison of the astringency level of the YP20-H14-13A YP706 prepared as described in Example 1 with that of the YP05-A18-12A YP701 prepared as described in Example 10.

(69) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP701 solution was 3.12 and it was adjusted to 3.48 with food grade sodium hydroxide solution. The pH of the YP706 solution was 3.46. An informal panel of seven panellists was asked to blindly taste the samples and indicate which was less astringent.

(70) Five out of seven panellists indicated that the YP20-H14-13A YP706 was less astringent.

Example 16

(71) This Example illustrates a comparison of the astringency level of the LE03-D02-14A LE705 prepared as described in Example 1 with that of the LE01-J24-13A YP701 prepared as described in Example 10.

(72) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the LE705 solution was 3.17 and it was adjusted to 3.47 with food grade sodium hydroxide solution. The initial pH of the LE701 solution was 3.81 and it was adjusted to 3.52 with food grade hydrochloric acid. An informal panel of eight panellists was asked to blindly taste the samples and indicate which was less astringent.

(73) Six out of eight panellists indicated that the LE03-D02-14A LE705 was less astringent.

Example 17

(74) This Example illustrates a comparison of the astringency level of the LE03-D01-14A LE706 prepared as described in Example 3 with that of the LE01-J24-13A LE701 prepared as described in Example 10.

(75) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the LE706 solution was 3.37 and it was adjusted to about 3.5 with food grade sodium hydroxide solution. The pH of the LE701 solution was 3.84 and it was adjusted to about 3.5 with food grade hydrochloric acid solution. An informal panel of eight panellists was asked to blindly taste the samples and indicate which was less astringent.

(76) Five out of eight panellists indicated that the LE03-D01-14A LE706 was less astringent.

Example 18

(77) This Example contains an evaluation of the dry colour and colour in solution of the co-products of the production of reduced astringency pulse protein products, prepared according to the methods of Examples 1-3.

(78) The colour of the dry powders was assessed using a HunterLab ColorQuest XE instrument in reflectance mode. The colour values are set forth in the following Table 14:

(79) TABLE-US-00014 TABLE 14 HunterLab scores for dry protein products Sample L* a* b* YP20-D23-13A YP705P 84.78 1.30 9.87 YP20-D24-13A YP705P 88.97 0.21 6.08 YP20-E02-13A YP705P 89.06 0.22 6.37 YP20-E13-13A YP705P-01 82.64 1.99 12.53 YP20-E13-13A YP705P-02 83.61 1.80 11.06 LE03-D02-14A LE705P 74.27 1.53 8.32 YP23-J02-13A YP706B 81.57 1.32 10.45 LE03-D01-14A LE706B 78.19 1.96 8.35

(80) As may be seen from the results in Table 14, the co-products generally were darker, redder and more yellow than the reduced astringency pulse protein products.

(81) Solutions of the co-products from the preparation of reduced astringency pulse protein products were prepared by dissolving sufficient protein powder to supply 0.48 g of protein in 15 ml of RO water. The pH of the solutions was measured with a pH meter and the colour and clarity assessed using a HunterLab Color Quest XE instrument operated in transmission mode. The results are shown in the following Table 15.

(82) TABLE-US-00015 TABLE 15 pH and HunterLab scores for solutions of pulse protein products sample pH L* a* b* haze YP20-D23-13A YP705P 5.81 43.87 5.5 28.43 97.1 YP20-D24-13A YP705P 6.13 40.94 6.82 30.44 97.3 YP20-E02-13A YP705P 4.95 39.68 6.79 31.08 99.2 YP20-E13-13A YP705P-01 5.29 39.32 8.4 33.01 96.5 YP20-E13-13A YP705P-02 5.03 32.10 10.7 34.12 96.4 LE03-D02-14A LE705P 6.40 11.69 11.81 17.59 97.9 YP23-J02-13A YP706B 7.39 41.26 7.88 31.65 95.7 LE03-D01-14A LE706B 7.09 38.09 7.75 25.18 97.3

(83) As may be seen from the results in Table 15, the solutions of the co-products were all very high in haze. The solutions were also darker, redder and more yellow than the solutions of the reduced astringency pulse products.

Example 19

(84) This Example contains an evaluation of the solubility in water of the co-products of the production of the reduced astringency pulse products, prepared by the methods of Examples 1 and 3. Solubility was tested based on protein solubility (termed protein method, a modified version of the procedure of Morr et al., J. Food Sci. 50:1715-1718).

(85) Sufficient protein powder to supply 0.5 g of protein was weighed into a beaker and wetted by mixing with about 20-25 ml of reverse osmosis (RO) purified water. 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 (6, 6.5, 7, 7.5 or 8) with diluted NaOH or HCl. The pH was then measured and corrected periodically 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% w/v protein dispersion. The protein content of the dispersions was determined by combustion analysis using a Leco Nitrogen Determinator. The samples were then centrifuged at 7,800 g for 10 minutes, which sedimented insoluble material and yielded a supernatant. The protein content of the supernatant was measured by combustion analysis.

(86) Solubility of the product was then calculated:
Solubility (protein method) (%)=(% protein in supernatant/% protein in initial dispersion)?1001)
Values calculated as greater than 100% were reported as 100%.

(87) The solubility results obtained are set forth in the following Table 16:

(88) TABLE-US-00016 TABLE 16 Solubility of products at different pH values based on protein method Solubility (protein method) (%) Batch Product pH 6 pH 6.5 pH 7 pH 7.5 pH 8 YP20-D23-13A YP705P 5.7 2.9 9.9 12.0 11.8 YP20-D24-13A YP705P 13.0 9.9 15.2 11.7 15.3 LE03-D02-14A LE705P 13.6 10.9 11.0 11.7 9.6 YP23-J02-13A YP706B 16.5 15.5 20.4 17.7 19.6 LE03-D01-14A LE706B 2.0 1.8 4.7 9.3 5.1

(89) As may be seen from the results in Table 16, the co-products of the production of the reduced astringency pulse protein products were poorly soluble over the pH range of 2 to 7.

Example 20

(90) This Example contains an evaluation of the water binding capacity of the co-products of the production of the reduced astringency pulse products, prepared by the methods of Examples 1 and 3.

(91) Protein powder (1 g) was weighed into centrifuge tubes (50 ml) of known weight. To this powder was added approximately 20 ml of reverse osmosis purified (RO) water at the natural pH. The contents of the tubes were mixed using a vortex mixer at moderate speed for 1 minute. The samples were incubated at room temperature for 5 minutes then mixed with the vortex mixer for 30 seconds. This was followed by incubation at room temperature for another 5 minutes followed by another 30 seconds of vortex mixing. The samples were then centrifuged at 1,000 g for 15 minutes at 20? C. After centrifugation, the supernatant was carefully poured off, ensuring that all solid material remained in the tube. The centrifuge tube was then re-weighed and the weight of water saturated sample was determined.

(92) Water binding capacity (WBC) was calculated as:
WBC (ml/g)=(mass of water saturated sample?mass of initial sample)(mass of initial sample?total solids content of sample)

(93) The water binding capacity results obtained are set forth in the following Table 17.

(94) TABLE-US-00017 TABLE 17 Water binding capacity of various products product WBC (ml/g) YP20-D23-13A YP705P 2.60 YP20-D24-13A YP705P 2.59 LE03-D02-14A LE705P 3.90 YP23-J02-13A YP706B 2.88 LE03-D01-14A LE706B 2.74

(95) As may be seen from the results of Table 17, all of the co-products of the production of the reduced astringency pulse protein products had moderate water binding capacities.

Example 21

(96) This Example illustrates the preparation of a pulse protein isolate by conventional isoelectric precipitation.

(97) 20 kg of yellow pea protein concentrate was added to 200 L of RO water at ambient temperature and the pH adjusted to about 8.5 by the addition of sodium hydroxide solution. The sample was agitated for 30 minutes to provide an aqueous protein solution. The pH of the extraction was monitored and maintained at about 8.5 throughout the 30 minutes. The residual pea protein concentrate was removed and the resulting protein solution clarified by centrifugation and filtration to produce 240 L of filtered protein solution having a protein content of 3.52% by weight. The pH of the protein solution was adjusted to about 4.5 by the addition of HCl that had been diluted with an equal volume of water and a precipitate formed. The precipitate was collected by centrifugation then washed by re-suspending it in 2 volumes of RO water. The washed precipitate was then collected by centrifugation. A total of 30.68 kg of washed precipitate was obtained with a protein content of 22.55 wt %. This represented a yield of 81.9% of the protein in the clarified extract solution. An aliquot of 15.34 kg of the washed precipitate was combined with 15.4 kg of RO Water and then the pH of the sample adjusted to about 7 with sodium hydroxide solution. The pH adjusted sample was then spray dried to yield an isolate with a protein content of 90.22% (N?6.25) d.b. The product was designated YP12-K13-12A conventional IEP pH 7.

Example 22

(98) This Example is a sensory evaluation of the YP20-D23-13A YP705P product prepared as described in Example 1 with the conventional pea protein isolate product prepared as described in Example 21.

(99) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP12-K13-12A conventional IEP pH 7 solution was 7.08. The initial pH of the YP705P solution was 5.77 and it was adjusted to 7.08 with food grade sodium hydroxide solution. An informal panel of eight panelists was asked to blindly taste the samples and indicate which had a cleaner flavour and which sample they preferred.

(100) Seven out of eight panellists preferred the YP20-D23-13A YP705P and seven out of eight found it to have a cleaner flavour.

Example 23

(101) This Example is a sensory evaluation of the YP20-D24-13A YP705P product prepared as described in Example 1 with the conventional pea protein isolate product prepared as described in Example 21.

(102) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The initial pH of the YP12-K13-12A conventional IEP pH 7 solution was 7.06. The initial pH of the YP705P solution was 6.18 and it was adjusted to 7.10 with food grade sodium hydroxide solution. An informal panel of nine panelists was asked to blindly taste the samples and indicate which had a cleaner flavour and which sample they preferred.

(103) All nine panellists preferred the YP20-D24-13A YP705P and found it to have a cleaner flavour.

Example 24

(104) This Example is a sensory evaluation of the YP23-J02-13A YP706B product prepared as described in Example 3 with the conventional pea protein isolate product prepared as described in Example 21.

(105) Samples were prepared for sensory evaluation by dissolving sufficient protein powder to supply 5 g protein in 250 ml of purified drinking water. The pH of the YP12-K13-12A IEP pH 7 solution was 7.09. The pH of the solution of YP23-J02-13A YP706B was adjusted to 7.04 with food grade hydrochloric acid. An informal panel of eight panellists was asked to blindly taste the samples and indicate which had a cleaner flavour and which sample they preferred. The experiment was conducted a second time with a panel having 7 members. The cumulative results are presented below.

(106) Eleven out of fifteen panellists found the YP23-J02-13A YP706B to have the cleaner flavour. Ten out of fifteen panellists preferred the YP23-J02-13A YP706B.

Example 25

(107) This Example illustrates the molecular weight profile of the pulse protein products prepared as described in Examples 1-3 as well as the molecular weight profile of some commercial yellow pea protein products (Propulse (Nutri-Pea, Portage 1a Prairie, MB), Nutralys S85F (Roquette America, Inc. Keokuk, Iowa) and Pisane C9 (Cosucra Groupe Warcoing S.A., Belgium). These protein products were chosen as they are among the most highly purified pea protein ingredients currently commercially available.

(108) Molecular weight profiles were determined by size exclusion chromatography using a Varian ProStar HPLC system equipped with a 300?7.8 mm Phenomenex BioSep S-2000 series column. The column contained hydrophilic bonded silica rigid support media, 5 micron diameter, with 145 Angstrom pore size.

(109) Before the pulse protein samples were analyzed, a standard curve was prepared using a Biorad protein standard (Biorad product #151-1901) containing proteins with known molecular weights between 17,000 Daltons (myoglobulin) and 670,000 Daltons (thyroglobulin) with Vitamin B12 added as a low molecular weight marker at 1,350 Daltons. A 0.9% w/v solution of the protein standard was prepared in water, filtered with a 0.45 ?m pore size filter disc then a 50 ?L aliquot run on the column using a mobile phase of 0.05M phosphate/0.15M NaCl, pH 6 containing 0.02% sodium azide. The mobile phase flow rate was 1 ml/min and components were detected based on absorbance at 280 nm. Based on the retention times of these molecules of known molecular weight, a regression formula was developed relating the natural log of the molecular weight to the retention time in minutes.
Retention time (min)=?0.955?ln (molecular weight)+18.502 (r.sup.2=0.98)

(110) For the analysis of the pulse protein samples, 0.05M NaCl, pH 3.5 containing 0.02% sodium azide was used as the mobile phase and also to dissolve dry samples. Protein samples were mixed with mobile phase solution to a concentration of 1% w/v, placed on a shaker for at least 1 hour then filtered using 0.45 ?m pore size filter discs. Sample injection size was 50 ?L. The mobile phase flow rate was 1 mL/minute and components were detected based on absorbance at 280 nm.

(111) The above regression formula relating molecular weight and retention time was used to calculate retention times that corresponded to molecular weights of 100,000 Da, 15,000 Da, 5,000 Da and 1,000 Da. The HPLC ProStar system was used to calculate the peak areas lying within these retention time ranges and the percentage of protein ((range peak area/total protein peak area)?100) falling in a given molecular weight range was calculated. Note that the data was not corrected by protein response factor.

(112) The molecular weight profiles of the products prepared as described in Examples 1-3 and the commercial products are shown in Table 18.

(113) TABLE-US-00018 TABLE 18 Molecular weight profile of pulse protein products % 15,000- % 5,000- % 1,000- product % >100,000 Da 100,000 Da 15,000 Da 5,000 Da YP20-D23-13A YP705 31 33 31 5 YP20-D24-13A YP705 30 36 29 5 YP20-E02-13A YP705 31 37 28 4 YP20-E13-13A YP705 66 16 14 4 LE03-D02-14A LE705 37 38 16 9 YP23-H12-13A YP706 21 30 42 7 YP23-H14-13A YP706 28 29 36 7 YP23-J02-13A YP706 16 28 48 8 LE03-D01-14A LE706 39 34 18 9 YP20-D23-13A YP705P 22 29 34 15 YP20-D24-13A YP705P 21 30 33 17 YP20-E02-13A YP705P 24 32 30 15 YP20-E13-13A YP705P-01 27 26 19 29 YP20-E13-13A YP705P-02 38 22 17 24 LE03-D02-14A LE705P 35 37 22 6 YP23-J02-13A YP706B 38 28 14 20 LE03-D01-14A LE706B 75 16 3 5 Nutralys S85F 7 29 9 56 Pisane C9 5 31 29 36 Propulse 13 25 18 45

(114) As may be seen from the results presented in Table 18, the molecular weight profiles of the products prepared according to Examples 1-3 were different from the molecular weight profiles of the commercial yellow pea protein products.

Example 26

(115) This Example contains an evaluation of the phytic acid content of the pulse protein products produced as described in Examples 1 to 3. Phytic acid content was determined using the method of Latta and Eskin (J. Agric. Food Chem., 28: 1313-1315).

(116) The results obtained are set forth in the following Table 19.

(117) TABLE-US-00019 TABLE 19 Phytic acid content of protein products product % phytic acid d.b. YP20-D23-13A YP705 0.00 YP20-D24-13A YP705 0.00 YP20-E02-13A YP705 0.02 YP20-E13-13A YP705 0.00 LE03-D02-14A LE705 0.19 YP23-H12-13A YP706 0.00 YP23-H14-13A YP706 0.00 YP23-J02-13A YP706 0.01 LE03-D01-14A LE706 0.29 YP20-D23-13A YP705P 0.02 YP20-D24-13A YP705P 0.01 YP20-E02-13A YP705P 0.06 YP20-E13-13A YP705P-01 0.00 YP20-E13-13A YP705P-02 0.00 LE03-D02-14A LE705P 0.23 YP23-J02-13A YP706B 0.10 LE03-D01-14A LE706B 0.21

(118) As may be seen from the results in Table 19, all of the products tested were low in phytic acid content.

SUMMARY OF THE DISCLOSURE

(119) In summary of this disclosure, the present invention provides pulse protein products, preferably pulse protein isolates, which have reduced astringency when tasted in an acidic solution such as an acidic beverage. Modifications are possible within the scope of this invention.