阅读说明：本技术 脱色的油菜籽蛋白质分离物 (Decolorized canola protein isolate ) 是由 尼恩克·尼娜·海尔克马 格拉尔杜斯·约翰尼斯·弗拉其苏斯·思莫德尔斯 于 2018-12-04 设计创作，主要内容包括：本发明公开了一种具有改善颜色的可溶性天然油菜籽蛋白质分离物,一种制备具有改善颜色的可溶性天然油菜籽蛋白质分离物的方法,以及所述油菜籽蛋白质分离物在食物产品中的用途。(The present invention discloses a soluble native rapeseed protein isolate having improved color, a process for preparing a soluble native rapeseed protein isolate having improved color, and the use of the rapeseed protein isolate in food products.)

1. A method of obtaining a natural rapeseed protein isolate, said method comprising the steps of:

i) mixing cold pressed rapeseed oil meal with an aqueous liquid at a temperature of 45 ℃ to 65 ℃;

ii) separating the aqueous liquid from the mixture obtained in step i);

iii) degreasing the aqueous liquid obtained in step ii);

iv) adjusting the pH of the defatted aqueous liquor obtained in step iii) to neutral by adding an acid or base and mixing with a precipitating agent to obtain a precipitate, wherein the precipitating agent comprises a salt of magnesium, zinc, iron or calcium;

v) removing the precipitate obtained in step iv) to obtain an aqueous liquid;

vi) concentrating and washing the aqueous liquid obtained in step v);

vii) separating the natural rapeseed protein isolate from the concentrated and washed aqueous liquor obtained in step vi) by means of drying,

characterised in that ascorbic acid or a derivative thereof and a sulphite are added before, during or after any of steps i) or ii) or iii) or iv) or v) or vi).

2. The method of claim 1, wherein the ascorbic acid or derivative thereof is L-ascorbic acid or L-calcium ascorbate or L-potassium ascorbate or L-sodium ascorbate, and wherein the sulfite is an ammonium or metal salt of sulfurous acid, an ammonium or metal salt of bisulfite, or an ammonium or metal salt of metabisulfite.

3. The method of claim 1, wherein the sulfite is sodium metabisulfite or potassium metabisulfite.

4. The method of any one of the preceding claims, wherein the L-ascorbic acid is in an amount of 0.05 to 5g/kg of the mixture of cold pressed rapeseed oil meal and aqueous liquid, and the sulfite is in an amount of 0.01 to 0.5g/kg of the mixture of cold pressed rapeseed oil meal and aqueous liquid.

5. The method of claim 4, wherein the amount of L-ascorbic acid is from 0.25 to 1g/kg of the mixture of cold pressed rapeseed oil meal and aqueous liquid, and the amount of sulfite is from 0.05 to 0.1g/kg of the mixture of cold pressed rapeseed oil meal and aqueous liquid.

6. A natural rapeseed protein isolate comprising 40 to 65 wt% cruciferous protein and 35 to 60 wt% commodity protein and having a solubility of at least 88% at a pH in the range of 3 to 10 and a temperature of 23 ± 2 ℃, the natural rapeseed protein isolate having an L value of 88 to 98 in a1 wt% solution in 0.2M phosphate buffer at pH 6 and 20 ± 2 ℃.

7. The natural rapeseed protein isolate of claim 6, comprising 5% to 65% of 12S rapeseed protein on a dry matter basis, wherein the presence of 12S is verified by blue native-PAGE.

8. The canola protein isolate of any one of claims 6-7, having an electrical conductivity of less than 9,000 μ S/cm in a 2 weight% aqueous solution at a pH in the range of 2 to 12.

9. The natural rapeseed protein isolate of any of claims 6 to 8, comprising less than 20% 7S rapeseed protein on a dry matter basis.

10. The natural rapeseed protein isolate of any of claims 6 to 9, having a C/N ratio in the range of 0.9 to 1.3.

11. The canola protein isolate of any one of claims 6-10, having a phytic acid level of less than 0.4% by weight.

12. The canola protein isolate of any one of claims 6-11, wherein the solubility of the canola protein isolate is at least 94% when measured at a pH in the range of 3 to 10 and a temperature of 23 ± 2 ℃.

13. The natural rapeseed protein isolate of any of claims 6 to 12, having a sulfite level of from 1ppm to 9 ppm.

14. Use of a natural rapeseed protein isolate according to any one of claims 6 to 13 in a food product.

15. A food product or pet food product comprising a canola protein isolate according to any one of claims 6 to 13.

Technical Field

The present invention relates to a soluble native rapeseed protein isolate having improved color, a process for preparing a soluble native rapeseed protein isolate having improved color, and the use of the rapeseed protein isolate in food products.

Background

Proteins are a major feature of human nutrition. This may be from animals (e.g. meat, fish, eggs, dairy products) or vegetables. It is generally desirable to reduce the amount of animal-based protein. The use of egg proteins is often undesirable.

The use of plant-based proteins in human nutrition is known, for example WO2008/094434 discloses the use of wheat protein isolate as an alternative to the use of egg yolk protein. However, for persons with gluten allergy, the use of wheat protein isolates may be undesirable. The use of soy-based proteins instead of whey proteins is also described, for example, in WO 2014/018922. Soy protein is widely used, but in view of some intolerance to soy products, other sources of vegetable protein are required.

Suitable alternatives include pea protein and rapeseed protein. Rapeseed is rich in oil and contains a large amount of protein, which represents 17% to 25% of the dry weight of the seed. Processing rapeseed to obtain oil for human consumption produces rapeseed meal (60%) as a by-product, containing about 30% to 40% protein. The rape used for this purpose is usually the Brassica napus (Brassica napus) and Brassica juncea (Brassica juncea) varieties. These varieties contain lower levels of erucic acid and glucosinolates and are known as Canola. Canola is an acronym for Canada and ola, meaning "low acid oil," but is now a general term defined as rapeseed oil containing < 2% erucic acid and < 30mmol/g glucosinolate. The resulting rapeseed meal is currently used as a high protein animal feed.

The protein may be obtained as a hydrolysate, concentrate and isolate. A hydrolysate is a protein that is partially broken down by exposing the protein to heat, acid, or enzymes that break the bonds connecting the amino acids. This makes the hydrolysate taste more bitter, but also makes it absorbed faster than the native (non-hydrolysed) protein during digestion. The isolate is purer than the concentrate, which means that other non-protein components have been partially removed to "isolate" the protein. Many concentrates have about 80% protein, which means that 80% of the total weight is protein on a dry basis. The isolate is typically about 90% protein (on a dry basis). This was calculated using Kjeldahl method.

The major storage proteins found in rapeseed are cruciferin (cruciferin) and nain (napin). Cruciferous proteins are globulins and are the major storage proteins in seeds. Cruciferae proteins are composed of 6 subunits and have a total molecular weight of about 300 kDa. The resistant protein is albumin and is a low molecular weight storage protein with a molecular weight of about 14 kDa. The resistant protein is more soluble and a method for isolating a more soluble resistant protein fraction, preferably to at least 85 wt.%, is disclosed in e.g. EP 1715752B 1. Resistant proteins were originally proposed for use in applications where solubility was critical. DE 102014005466 a1 also describes a process for obtaining purified cruciferous protein fractions and resistant protein fractions. During the process, a protein mixture of the two is also obtained, which has 55-60% of resistant protein and 40-45% of cruciferous protein. The solubility of the protein mixture was about 75%.

Rapeseed proteins can also be divided into different fractions according to the corresponding sedimentation coefficient in Svedberg (S), which represents the sedimentation velocity of macromolecules in centrifugal fields for rapeseed proteins, the main reported fractions are 12S, 7S and 2S. cruciferous proteins and commodity-resistant proteins are the two main families of storage proteins found in canola/rapeseed.the commodity-resistant protein is 2S albumin and the cruciferous protein is 12S globulin.furthermore, Schwenke and L inow (Nahrung (1982)26K5-K6) indicated that the reversible dissociation of 12S globulin from rapeseed (brassica napus) depends on the ionic strength. The cruciferin complex exists as a 300kDa 12S hexamer when exposed to higher ionic strength (. mu.gtoreq.0.5 mS/cm) and reversibly dissociates into 150kDa 7S trimer molecules when exposed to low ionic strength conditions.

The production of rapeseed protein isolate from rapeseed meal, as described for example in EP 1389921B1, discloses extraction of rapeseed oil seed meal with an aqueous food grade salt solution to form an aqueous protein solution, followed by separation of the two protein fractions by micelles. WO 2013/000066 discloses a rapeseed protein product having a protein content of at least about 60 wt.% and having a low phytic acid content, with preference for having equal parts of 2S and 7S and a minor content of 12S. EP 1720415 discloses a process for preparing rapeseed protein isolate comprising 25 to 55 wt% 2S rapeseed protein, 47 to 75 wt% 7S rapeseed protein and 0 to 15 wt% 12S rapeseed protein. This method requires the use of high levels of salt, which is not problematic in aquaculture, but is not applicable to human nutrition. In WO2016/042001 a process is described in which the protein component need not be isolated, but solubility can still be maintained over a broader pH range.

It has been found that high purity rapeseed protein isolate has a fundamental broad functionality in food products, which is unique among protein materials. The ability to utilize plant-derived proteins in food products enables the provision of a truly vegetarian food product in the absence of any available substitute for the protein and/or animal-derived protein.

A problem associated with rapeseed protein isolates produced by the above procedures is that they have a relatively dark yellow or tan color and often have an undesirable flavor. For example, coloration often makes food products visually unappealing, which makes the acceptance of plant-based proteins unnecessarily problematic in human nutrition.

Drawings

Figure 1 shows the colour of the solutions obtained after treatment of rapeseed protein extract with different concentrations of L-ascorbic acid or L-cysteine at different time intervals the X-axis: incubation time in hours the Y-axis: values 100-L0-notations ○ ═ control, ● ═ L-ascorbic acid (0.5g/kg), ■ ═ L-ascorbic acid (1.0g/kg), ▲ ═ L-cysteine (0.5g/kg), ◆ ═ L-cysteine (1.0 g/kg).

Fig. 2 shows the colour of the solutions obtained after incubation of rapeseed protein extract at 56 ℃, prepared with different concentrations of L-ascorbic acid and/or sodium metabisulphite at different time intervals in the extract, X axis: incubation time in hours, Y axis: 100-L values, it is noted that after measurement at t ═ 3h, the experiment was temporarily stopped by storing the samples at-20 ℃ for a longer period of time (10 weeks), after which the measurement was continued by continuing incubation at 56 ℃ at the time shown in the figure, i.e. the time period for which the samples were frozen is omitted in the figure, the notations: ○ ═ control; ◇ ═ L-ascorbic acid (0.5g/kg), □ ═ sodium metabisulphite (0.1g/kg), ● ═ L-ascorbic acid (0.25g/kg) plus sodium metabisulphite (360.05 g/kg), 24 ═ sodium metabisulphite (0.1g/kg) plus sodium metabisulphite (■ g/kg), 632 ═ ascorbic acid plus sodium metabisulphite (0.5g/kg) plus 0.5g/kg ═ ascorbic acid plus 0.5g/kg ═ sodium metabisulphite (9 kg).

Detailed Description

The problem of coloration in rapeseed protein isolate is reported to be due to phenolic compounds the amount of phenolic compounds contained in rapeseed is about ten times the amount of phenolic compounds present in soybean after oxidation, phenolic compounds can lead to development of dark colour in WO 2004/000032 the problem is solved by advocating a multifaceted approach, this includes processing rapeseed (especially inactivation of myrosinase in dehulled seeds), treating meal (especially extraction of phenols and other coloring components using a solvent), utilizing a specific form of meal (especially meal or roast meal with solvent removed with air), using specific extraction conditions (especially by addition of antioxidant or coloring component absorbent), processing the extract (especially diafiltration in the presence of antioxidant), and recovering isolate (especially by extraction of isolate with solvent or addition of concentrated solution to cooling water), among the various possible approaches WO 2004/000032 describes the use of antioxidants (such as sodium sulfite or ascorbic acid) to prevent oxidation, this effect is reported by the use of reduced absorbance at 420nm (a420, oxidized phenols) and retention of the value at 35330 nm and the actual value of the value is reported to be a white value when the value is found to be between the value of the value found by the experimental tests of the value of the antioxidant alone, especially the value found for the protein isolate of alumina, especially the value of alumina found, especially the antioxidant found to be about 100. the whiteness value of canola 330. the value of the value found after the cellulosic protein isolate, thus showing that is found to be a 2. the value of the value found to be more than the value found for the value found to be about 100. the value found to be about 21. the value of the value found to be more than the value found to be about 21. the value of the value found to be found.

In a first aspect of the present invention, there is provided a process for obtaining a natural rapeseed protein isolate, said process comprising the steps of:

i) mixing cold pressed rapeseed oil meal with an aqueous liquid at a temperature of 45 ℃ to 65 ℃;

ii) separating the aqueous liquid from the mixture obtained in step i);

iii) degreasing the aqueous liquid obtained in step ii);

iv) adjusting the pH of the defatted aqueous liquor obtained in step iii) to neutral by adding an acid or base and mixing with a precipitating agent to obtain a precipitate, wherein the precipitating agent comprises a salt of magnesium, zinc, iron or calcium;

v) removing the precipitate obtained in step iv) to obtain an aqueous liquid;

vi) concentrating and washing the aqueous liquid obtained in step v);

vii) isolating the natural rapeseed protein isolate from the concentrated and washed aqueous liquor obtained in step vi) by means of drying.

In the above method, ascorbic acid or a derivative thereof and a sulfite are added before, during or after any of steps i) or ii) or iii) or iv) or v) or vi). The use of ascorbic acid (derivative) in combination with sulphite has not been reported or suggested in the prior art and, as further outlined below, may lead to unexpected effects.

As noted above, rapeseed protein isolate is produced from cold pressed rapeseed meal, a by-product of rapeseed oil production.

The process starts with an extraction step i) in which rapeseed meal is mixed with a brine solution (e.g. 0% to 5% sodium chloride) at a temperature between 4 ℃ and 75 ℃, more preferably between 20 ℃ and 75 ℃ and most preferably between 45 ℃ and 65 ℃. Preferably, the mixing is performed in step i) such that the ratio between the cold pressed rapeseed meal and the aqueous liquid is from 1:2 to 1:30 (w/w). The ratio of meal to water is preferably in the range of 1:5 to 1:40, more preferably in the range of 1:5 to 1: 20.

After a period of time of 5min to 2 hours, the protein-rich solution is separated from the insoluble material in a separation step ii). The protein-rich solution is hereinafter referred to as extract.

Preferably, the pH of the extract is adjusted to neutral and the extract is further processed to clarify the material and remove non-proteinaceous material. In the defatting step iii), the residual fat and the formed precipitate are removed by a solid/liquid separation step (e.g. filtration or centrifugation). Preferably, the degreasing in step iii) is performed by means of centrifugation.

The extract is then concentrated and washed in an ultrafiltration/diafiltration (UF/DF) step vi). The purpose of the UF/DF step is to concentrate the proteins and remove anti-nutritional factors (e.g., polyphenols, residual phytate, glucosinolates). The concentration and washing in step vi) are preferably carried out by means of ultrafiltration and diafiltration.

Finally, in step vii), the washed concentrate may be dried in a suitable dryer, such as a spray dryer (single or multi stage), using an inlet temperature in the range of 150 ℃ to 200 ℃ and an outlet temperature in the range of 50 ℃ to 100 ℃, thereby producing rapeseed protein isolate.

In one embodiment, the ascorbic acid or derivative thereof is L-ascorbic acid or L-calcium ascorbate or L-potassium ascorbate or L-sodium ascorbate in another embodiment, the sulfite is an ammonium or metal salt of sulfurous acid (sulfite), an ammonium or metal salt of bisulphite or an ammonium or metal salt of metabisulfite.

The amount of ascorbic acid or a derivative thereof can vary within wide ranges. A suitable example is where the amount of ascorbic acid is from 0.05g/kg to 5g/kg, or from 0.25g/kg to 1g/kg, relative to the mixture of cold pressed rapeseed oil meal and aqueous liquid. The amount of sulfite is from 0.01g/kg to 0.5g/kg, or from 0.05g/kg to 0.1g/kg, relative to the mixture of cold pressed rapeseed oil meal and aqueous liquor. Alternatively, the amounts of ascorbic acid and sulfite are expressed in percentages relative to the total weight of the composition. Thus, for ascorbic acid this may be in the range of 0.005% to 0.5% (w/w), or 0.025% to 0.1% (w/w), and for sulphite this may be in the range of 0.001% to 0.05% (w/w), or 0.005% to 0.01% (w/w).

In another aspect, the application of ascorbic acid results in less initial removal of color, but the less initial removal of color is more stable over time and ultimately results in significantly lower color values.

In another embodiment, the process steps i) -vi) are carried out at 4h, preferably at 30min-3.5h, under conditions where the color of the extract (expressed as 100-L) and thus the color of the final product is much lower than the color of the untreated extract, but also lower than the color of the extract treated with ascorbic acid or sulfite alone.

An advantage of the process of the first aspect is that no significant reduction of the proteins of interest, in particular of cruciferous and commodity-resistant proteins, is observed. This is particularly surprising for resistant proteins known to be susceptible to degradation. Under the above conditions, the concentration of the drug-resistant protein is maintained to be more than 95% of the initial concentration of the drug-resistant protein of the control.

Another advantage of the method of the first aspect is that the untreated clear solution of rapeseed protein isolate obtained during the process has a tendency to develop dark precipitates over time, which does not occur with the samples obtained during the method of the present invention.

Preferably, the rapeseed protein isolate is obtained in a process without a fractionation step for separating cruciferous proteins and commodity proteins.

Preferably, the canola protein isolate is obtained by a process in which the levels of tolerance protein and cruciferin are maintained substantially constant (i.e., the levels of tolerance protein (2S) or cruciferin (12S) are neither intentionally increased nor intentionally decreased).

The method of the invention is characterized in that it is very suitable for large scale applications. Thus, in one embodiment, the process is carried out on a scale of at least 500kg, preferably from 500kg to 10,000kg or from 1,000kg to 5,000 kg.

In a second aspect of the present invention, there is provided a natural rapeseed protein isolate comprising 40 to 65% by weight cruciferous protein and 35 to 60% by weight commodity protein and having a solubility of at least 88% at a pH in the range 3 to 10 and a temperature of 23 ± 2 ℃, at a pH in the range 3 to 10 and a solubility at a temperature of 23 ± 2 ℃, which natural rapeseed protein isolate has an L value of 88 to 98 in a 1% by weight solution in 0.2M phosphate buffer at a pH of 6 and 20 ± 2 ℃, which range can be achieved according to the method of the present invention and has not been reported so far.a typically acceptable subrange of L for a 1% by weight aqueous solution is 88.5 to 95, or 89 to 93, or 91 ± 2. thus, by performing the method of the first aspect, i.e. combining ascorbic acid or a derivative thereof together with sulfite during the preparation of the rapeseed protein isolate, a product having a lower color (i.e. a higher whiteness than any rapeseed protein isolate obtainable or reported so far has a positive impact on the application of the rapeseed protein isolate in a line.

In one embodiment, the Native rapeseed protein isolate comprises 5% to 65% 12S rapeseed protein on a dry matter basis, wherein the presence of 12S is verified by Blue Native PAGE. Preferably, the native rapeseed protein isolate comprises 10% to 65%, most preferably 15% to 65%, especially 25% to 65%, and most especially 35% to 65% (on a dry matter basis) of 12S rapeseed protein, wherein the presence of 12S is verified by native PAGE. As mentioned above, a certain amount of 12S protein is not necessarily identical to cruciferous proteins, since the 300kDa 12S hexamer of a cruciferous protein can dissociate into 150kDa 7S trimer molecules. Preferably, the natural rapeseed protein isolate of the present invention comprises less than 20% 7S rapeseed protein on a dry matter basis.

In one embodiment, the canola protein isolate has a ratio of cruciferous protein to tolerance protein (C/N ratio) of 0.9 to 1.3.

In one embodiment, the solubility of the canola protein isolate is at least 88%, preferably at least 90%, more preferably at least 94%, and most preferably at least 96% when measured at a pH in the range of 3 to 10 and a temperature of 23 ± 2 ℃. This is also known as the Soluble Solids Index (SSI).

For use in human food consumption, the canola protein isolate preferably contains low levels of salt. This can be determined by measuring the conductivity. Preferably, the electrical conductivity of the canola protein isolate in a 2 wt% aqueous solution is less than 9,000 μ S/cm at a pH in the range of 2 to 12. More preferably, the electrical conductivity of the canola protein isolate in a 2 wt% aqueous solution is less than 4,000 μ S/cm at a pH in the range of 2.5 to 11.5. For comparison, the conductivity of a 5g/l aqueous sodium chloride solution was about 9,400. mu.S/cm.

In another embodiment, the canola protein isolate has a phytic acid level of less than 0.4 wt%, more preferably less than 0.3 wt%, and most preferably less than 0.15 wt%.

In yet another embodiment, the protein content of the canola protein isolate is at least 90 wt% (calculated as kjeldahl nitrogen × 6.25.25), more preferably at least 94 wt%, most preferably at least 96 wt%, especially at least 98 wt% on a dry weight basis.

Preferably, the canola protein isolate is substantially unhydrolyzed. By substantially unhydrolyzed is meant that the protein is not intentionally hydrolyzed.

The present invention allows tuning the amount of ascorbic acid and sulfite added in the process and by doing so, high whiteness can be achieved, i.e. high L values, while controlling the sulfite content of the final product to be below 10ppm, e.g. below 8ppm, or between 1ppm and 9ppm or between 2ppm and 8 ppm.

In a third aspect, the canola protein isolate may be used in any nutritional food application, whether for human, pet or veterinary applications, including as a foaming agent to replace egg white, as an emulsifier to replace egg yolk in mayonnaise, for example, and simply as a nutritional component that provides excellent amino acid characteristics. Thus, the present invention provides the use of the natural rapeseed protein isolate according to the second aspect of the invention as a foaming agent or emulsifier for a food product. Accordingly, the present invention provides a food product or pet food product comprising the canola protein isolate according to the second aspect of the invention. The rapeseed protein isolate of the present invention can be used as a protein additive in food products (e.g., bars, chocolate, ice cream, etc.).