Biological yeast, method for obtaining same and uses thereof

Abstract

The present invention relates to a method for producing yeast. It relates in particular to a method for producing biological yeast, comprising the use of substrates of biological origin, in particular a biological substrate which makes it possible to supplement the nutritional requirements in the yeast in terms of phosphorus. The method of the present invention makes it possible to obtain biological yeast and biological yeast extracts in accordance with European Union Regulation (EC) 834/2007. According to the invention, the phosphorus-rich biological composition is obtained by hydrolysis and solubilization of at least one plant substrate of biological origin comprising from 2 to 18 g of phosphorus per kg of product, 60% to 80% of which is in the form of phytic acid. The preferred substrate according to the invention is wheat bran.

Claims

1. A method for preparing a biological yeast culture medium, said culture medium comprising carbon, nitrogen and phosphorus sources of biological origins, the phosphorus source being a phosphorus-containing purified solution obtained by hydrolysis and solubilization of at least one phytic acid-rich biological substrate comprising from 2 to 18 g of phosphorus per kg of substrate, 60% to 80% of which is in the form of phytic acid, said method comprising the following steps: a) milling said phytic acid-rich biological substrate, b) suspending the product obtained in a), c) heating the suspension, d) enzymatic deactivation of the suspension to obtain the phosphorus source, e) conducting protein hydrolysis on a substrate containing protein to obtain the nitrogen source, and f) adding a substrate containing molasses to obtain the carbon source, wherein said steps produce a biological yeast culture medium, and wherein said method makes it possible to obtain a biological yeast or biological yeast extract in accordance with European Union Regulation (EC) 834/2007, and wherein said at least one phytic acid-rich biological substrate comprises a phytic acid-rich biological substrate having endogenous phytase, and exogenous phytase is not added in the method.

2. The method as claimed in claim 1, wherein said phytic acid-rich substrate is chosen from the group comprising: corn gluten, fatty rice bran, rapeseed, soy cake, sunflower cake, half-white common wheat middlings, wheat bran, rye, and common wheat flour.

3. The method as claimed in claim 1, wherein the milled substrate is suspended in water in a proportion of from 100 to 250 g of milled product per kg of suspension.

4. The method as claimed in claim 1, wherein the suspension is heated at a temperature of between 40 and 50° C. for 5 to 20 hours.

5. The method as claimed in claim 1, wherein the suspension comprises a phytase.

6. The method as claimed in claim 1, wherein the phytic acid-rich substrate is wheat bran.

7. The method as claimed in claim 1, wherein the solubilization and hydrolysis method further comprises protein hydrolysis and starch saccharification of the phytic acid-rich biological substrate.

8. The method as claimed in claim 1, wherein the substrate containing protein is gluten.

9. The method as claimed in claim 8, wherein the gluten hydrolysis is carried out with an enzymatic cocktail comprising a mixture of endoproteases and exopeptidases.

10. The method as claimed in claim 1, wherein the substrate containing molasses is cane and/or beet molasses.

11. The method as claimed in claim 10, wherein the substrate containing molasses consists of a mixture of cane molasses and beet molasses in a ratio of between 50/50 and 80/20.

12. The method as claimed in claim 1, further comprising the addition of sodium carbonate, lactic or citric acids and vegetable oils to the culture medium.

13. A method for preparing a biological yeast culture medium, said culture medium comprising carbon, nitrogen and phosphorus sources of biological origins, the phosphorus source being a phosphorus-containing purified solution obtained by hydrolysis and solubilization of at least one phytic acid-rich biological substrate comprising from 2 to 18 g of phosphorus per kg of substrate, 60% to 80% of which is in the form of phytic acid, said method consisting of the following steps: a) milling said phytic acid-rich biological substrate, b) suspending the product obtained in a), c) heating the suspension, d) enzymatic deactivation of the suspension to obtain the phosphorus source, e) conducting protein hydrolysis on a substrate containing protein to obtain the nitrogen source, and f) adding a substrate containing molasses to obtain the carbon source, and optionally adding an exogenous phytase to the suspension, optionally at least one of decanting, clarifying, and filtering the suspension after said enzymatic deactivation, optionally washing at least one of decanting sludges, clarifying sludges, and filtration cakes resulting from said at least one of decanting, clarifying, and filtering to provide washing waters, optionally combining the washing waters and supernatant resulting from said at least one of decanting, clarifying, and filtering, optionally concentrating the washing waters and supernatant to provide concentrated washing waters and supernatant, and optionally harvesting the concentrated washing waters and supernatant to produce a biological yeast culture medium, wherein said steps produce a biological yeast culture medium, and wherein said method makes it possible to obtain a biological yeast or biological yeast extract in accordance with European Union Regulation (EC) 834/2007.

Description

DETAILED DESCRIPTION OF THE INVENTION

(1) The first subject of the present invention is a method for producing biological yeast, comprising the use of substrates of biological origins capable of providing all the nutrients required for the growth of the yeast.

(2) These substrates are preferentially molasses as a source of sugars, a source of hydrolyzed biological proteins as a source of nitrogen and at least one phosphorus-rich purified solution as a source of phosphorus.

(3) The method of the invention also comprises the use of other substances required for the growth of the yeast, chosen from those authorized by the European Regulation, such as sodium carbonate, lactic or citric acids and vegetable oils.

(4) The method of the invention uses molasses of biological origin as a source of sugar. According to one form of the invention, the molasses used is chosen from cane molasses and beet molasses. In order to take advantage of their different compositions in terms of minerals and vitamins, it is preferable according to the invention to jointly use cane and beet molasses in a ratio of between 50/50 and 80/20. According to one preferred form of the invention, the cane molasses/beet molasses ratio is between 65/35 and 75/25, and more preferentially close to 70/30.

(5) According to the invention, the biological source of nitrogen is a source of hydrolyzed biological proteins, chosen from rice, pea, potato, wheat, soy, alfalfa, spirulina and gluten proteins.

(6) According to one preferential form of the invention, the biological source of nitrogen is hydrolyzed gluten.

(7) The hydrolysis of the biological source of nitrogen can be carried out using an enzymatic cocktail which makes it possible to have a degree of solubilization of the dry matter close to 80% and a nitrogen yield of 80% to 85%. This enzymatic cocktail comprises a mixture of endoproteases and exopeptidases, preferentially all or part of the mixture NEUTRASE® a neutral, zinc metallo endo-protease from Bacillus amyloliquefaciens), ALCALASE® (a serine endopeptidase that consists primarily of subtilisin A), CRISTALASE® (a liquid preparation of purified papain, standardized with sorbitol) and FLAVOURZYME® (a peptidase preparation from Aspergillus oryzae). These enzymes, which are non-GMO, are authorized for the manufacture of protein hydrolysates.

(8) According to the invention, the provision of phosphorus is covered by the use of a solution, of biological origin, which is purified and rich in phosphorus. This solution is obtained by solubilization and hydrolysis of a phytic acid-rich plant substrate of biological origin, the phosphorus being released in the form of inorganic phosphate after solubilization and hydrolysis of the phytic acid.

(9) According to the invention, the “phytic acid-rich biological substrate” is intended to mean a plant substrate of biological origin comprising from 2 to 18 g of phosphorus per kg of substrate, 60% to 80% of which is in the form of phytic acid.

(10) The phytic acid-rich substrate according to the invention is chosen from the group of plants listed in table I below:

(11) TABLE-US-00001 TABLE I Total phosphorus contents, phytic phosphorus to total phosphorus ratio and phytase activity of various raw materials (according to Sauvant 2002). P (g/kg raw) mean Phytic Phytase Group (standard P/total activity of Name deviation) P (%) (U/kg) interest Corn gluten 8.9 (1.5) 65 0 1 Fatty rice bran 16.1 (2.1) 85 120 1 Rapeseed 6.6 (0.9) 70 0 1 Rapeseed cake 11.4 (0.9) 60 10 1 Soy cake 6.2 (0.5) 60 20 1 Sunflower cake 10.1 (1.4) 85 0 1 Half-white common wheat 8.7 (1.4) 80 2590 2 middlings Wheat bran 9.9 (1.1) 80 1770 2 Rye 3 (0.3) 65 5350 3 Low-grade common 3.6 80 3080 3 wheat flour

(12) As indicated in the table above, certain phytic acid-rich substrates exhibit a more or less marked phytase activity. The phytic acid-rich substrate of biological origin according to the invention will be chosen according to either: its high phosphorus content (group of interest 1); its high phosphorus content and its richness in phytase activity (group of interest 2); or its richness in phytase activity (group of interest 3).

(13) In addition to the phosphorus, essentially present in the form of phytic acid, these compounds also contain proteins or even starch, compounds that are very useful for yeast growth.

(14) For example, the average composition of wheat bran and of soy cake is:

(15) for wheat bran: dry matter content: 87% on product as is (TQ) phosphorus: approximately 1% (TQ), i.e. approximately 10 g of phosphorus per kg of product, in the form of phytic acid at 80%. Phytase activity 1770 U/kg proteins: 15% to 18% (TQ) starch: 20% (TQ)

(16) for soy cake: dry matter content: 88% to 93% (TQ) phosphorus: 0.6% (TQ), (i.e. approximately 6 g of phosphorus per kg) in the form of phytic acid (60%) and of phospholipids. Phytase activity 20 U/kg proteins: 41% (TQ).

(17) It is necessary to hydrolyze these compounds in order to release the substances that can be assimilated by the yeast, since neither the phytic acid nor the proteins and the starch can be assimilated as they are by the yeast.

(18) Phytic acid, of chemical formula C.sub.6H.sub.18O.sub.24P.sub.6, consists of an inositol ring and 6 phosphate groups (InsP6). Under the action of a phytase, the phytic acid is hydrolyzed in the form of inorganic monophosphate and of myoinositol phosphates having a lower degree of phosphorylation (InsP5 to InsP1) and free myoinositol in certain cases as described in EP 1 910 531 B1.

(19) The phytase used for this hydrolysis may be endogenous to one of the rich substrates used in the mixture to be hydrolyzed, or exogenous in the case where the mixture of plants to be hydrolyzed is deficient in endogenous phytase activity.

(20) It should be noted that wheat bran does not require the use of exogenous phytase. The solubilization of the phosphorus of wheat bran without recourse to exogenous phytase activity is particularly advantageous since the provision of phytase not derived from GMOs (biological regulation obligation) is difficult.

(21) The mixture of plants, such as those described in table I, can allow the provision of phytase activity even though the plant used might not at first glance be described as phosphorus-rich (substrates of group of interest 3 of table I).

(22) The solubilization and hydrolysis of phytic acid are carried out according to the mode comprising at least the following steps: milling the phytic acid-rich biological substrate, suspending in water and heating the suspension, and enzymatic deactivation.

(23) By way of indication, the milling is carried out using a hammer mill equipped with an 800 μm grill. However, any type of mill can be used.

(24) The suspending in water is carried out in a proportion of from 100 to 250 g and preferably from 160 to 180 g of milled product per kg of final suspension. The suspension is heated at a temperature of between 40 and 50° C. for 5 to 20 hours.

(25) For the enzymatic deactivation, the suspension is brought to a temperature of 90° C. for a period ranging from 15 to 30 minutes, also enabling pasteurization of the substrate.

(26) It is sometimes necessary to add an exogenous phytase to the suspension so as to reinforce or supplement the endogenous phytase activity. In this case, the exogenous phytase is used in a proportion of 15 to 25 g per kg of milled product.

(27) At the end of these treatments, the suspensions are decanted and/or clarified and/or filtered.

(28) In order to maximize the recovery of the solubilized matter, the decanting sludges, the centrifugal clarification sludges or the filtration cakes can be washed, and the washing waters are then combined with the initial supernatant. All of the solutes can then be concentrated.

(29) The analysis of the harvested concentrated supernatants and washing waters shows that 80% to 90% of the phosphorus contained in the initial substrate is solubilized.

(30) According to one preferred mode of the invention, the phytic acid-rich substrate is wheat bran.

(31) In certain cases, the solubilization of the phosphorus can be added to using more conventional treatments of protein hydrolysis (mixture of endo and exopeptidases) and of starch saccharification (use of a mixture of amylase and amyloglucosidase).

(32) Another subject of the invention is the use of a phosphorus-rich purified solution, as described above, in a method for producing yeast, in particular in a method for producing biological yeasts, comprising the use of substrates of biological origin.

(33) The method for producing the yeast of the invention is carried out under the culture conditions normally used for producing conventional yeast, likewise with regard to the operating conditions for recovering, drying and packaging the yeasts produced.

(34) The present invention makes it possible in particular to uncouple the provisions from the main ingredients required for the yeast growth, namely the sugar, nitrogen source, phosphorus source and air. This uncoupling is desired in order to control the final composition of the yeast manufactured.

(35) Another subject of the invention is the biological yeast as produced by means of the method of the invention.

(36) Another subject of the invention is the use of the biological yeast of the invention in the bread making field and the alcohol production field, and more generally for human food and animal feed.

(37) The yeasts of the invention are particularly useful for producing biological yeast extracts.

(38) Another subject of the invention is a biological yeast extract obtained from the biological yeasts of the invention.

(39) The following examples illustrate the invention without limiting the scope thereof.

EXAMPLES

Example 1

Solubilization and Hydrolysis of Gluten

(40) It is known that hydrolysis of plant proteins by a purified protease (of papain or alcalase type) can be improved by adding yeast undergoing autolysis (EP0578572A, US201202888587).

(41) In order to demonstrate the effectiveness of the enzymatic cocktail proposed by the invention, the applicant carried out the following three tests: Recipe 1: gluten of Blattman origin, biological brewer's yeast in which autolysis has been brought about, papain Recipe 2: gluten of Celnat origin, biological baker's yeast in which autolysis has been brought about, papain (CRISTALASE®), NEUTRASE® and ALCALASE® Recipe 3: gluten of Celnat and/or Blattman origin, NEUTRASE®, ALCALASE®, FLAVOURZYME®.

(42) Firstly (recipe 1), only biological brewer's yeast in which autolysis has been brought about was used in combination with papain (CRISTALASE®). Next, secondly, and in order to compensate for the loss of proteolytic activities of the brewer's yeast cells, two supplementary enzymes were used (NEUTRASE® and ALCALASE®) in combination with biological baker's yeast in which autolysis has been brought about (recipe 2).

(43) Finally, the papain is replaced with Flavourzyme (mixture of endoprotease and exopeptidase) and the use of biological baker's cream yeast in which autolysis has previously been brought about was abandoned.

(44) The matter balances of these 3 hydrolysis recipes are correlated in table II.

(45) TABLE-US-00002 TABLE II Overall results of the various gluten hydrolysis recipes used during the biological yeast tests Recipe Recipe 1 Recipe 2 Recipe 3 Raw materials (in kg DM/T nonclarified final hydrolysate) Gluten nature Blattman Celnat Celnat Celnat Blattman DM content 93.9% 93.4% 93.6% 94.1% 92.6% concentration 23 kg DM/T 103 kg DM/T 167 kg DM/T 138 kg DM/T 152 kg DM/T DM ratio 50%   90%  100%  100%  100% Yeast nature Brewer's Baker's concentration 23 kg DM/T 10 kg DM/T DM ratio   50%   10% Enzymes (in kg DM/T DM to be hydrolyzed) Protein hydrolysis Papain 3.34 kg DM/T 21.43 kg DM/T Neutrase® 19.10 kg DM/T 3.20 kg DM/T 4.78 kg DM/T 4.85 kg DM/T ALCALASE® 17.85 kg DM/T 2.58 kg DM/T 3.86 kg DM/T 3.92 kg DM/T FLAVOURZYME® 0.11 kg DM/T 0.16 kg DM/T 0.16 kg DM/T Solubilization Dry matter content (kg DM/T 43.0 kg DM/T 99.0 kg DM/T 153.3 kg DM/T 128.2 kg DM/T 135.5 kg DM/T supernatant) yield (% of DM   81%   73%   80%   83%   79% involved) Nitrogen yield (% of N   86%   82%   80%   84%   85% involved) Amount of sludge after lab centrifugation 4000 G-10 min (in kg DM/T nonclarified hydrolysate) amount 7 kg DM/T 31 kg DM/T 38 kg DM/T 25 kg DM/T 29 kg DM/T sludge DMS   10%   28%   30%   27%   28% content

(46) In terms of solubilization of the dry matter implemented, only recipe 2 exhibited a lower effectiveness (73% compared with 80-83%).

(47) In terms of solubilization of the nitrogen, no clear difference is apparent between the three recipes tested. Use of Flavourzyme therefore appears, overall, to effectively replace the use of cream yeast (recycled biological or brewer's yeast, in which autolysis has been brought about beforehand).

(48) Recipe 3 enables a nitrogen yield close to 85%, which is largely sufficient, and a degree of protein degradation of 39% which is quite satisfactory, the degree of degradation being the ratio between the amino nitrogen content of the hydrolysate (resulting from the hydrolysis) and the total nitrogen content of the hydrolysate.

Example 2

Solubilization and Hydrolysis of Wheat Bran and of Soy Cake

(49) The operating conditions of the various treatments applied and also the results obtained are described hereinafter: Wheat bran: milling of the bran. By way of illustration, a hammer mill equipped with an 800 μm grill was used during the pilot tests; suspending in water in a proportion of 165 g of milled bran per kg of suspension; heating at 40 to 50° C. for 5 to 20 hours; no addition of exogenous phytase, the reaction takes place by virtue of the endogenous phytase activity of the bran; enzymatic deactivation at 90° C. for 30 minutes. Soy cake: milling of the cake. By way of illustration, a hammer mill equipped with an 800 μm grill was used during the pilot tests; suspending in water in a proportion of 165 g of milled cake per kg of suspension; heating at 40 to 50° C.; addition of exogenous phytase (Sumizyme PHY) at the dose of 20 g per kg of crude cake used, the endogenous phytase activity being virtually zero; treatment duration 5 to 20 hours, temperature regulated; enzymatic deactivation at 90° C. for 30 minutes.

(50) At the end of these treatments, the bran or cake suspensions are decanted and/or clarified and/or filtered.

(51) In order to maximize the recovery of the solubilized matter, the decanting sludges, the clarifying sludges or the filtration cakes can be washed, and the washing waters are then combined with the initial supernatant. All of the solutes can then be concentrated.

(52) The analysis of the harvested concentrated supernatants and washing waters shows that 80% to 90% of the phosphorus contained in the initial substrate is solubilized.

(53) The use of exogenous phytase for the treatment of the wheat bran does not improve the solubilization/recovery yield.

(54) The solubilization of the wheat bran phosphorus without recourse to exogenous phytase activity is particularly advantageous since the provision of phytase not derived from GMOs (biological regulation obligation) is difficult.

(55) The solubilization of the phosphorus can then be added to by more conventional treatments of bran or cake protein hydrolysis (use of a mixture of endo and exopeptidases) and of wheat bran starch saccharification (use of a mixture of amylase and amyloglucosidase).

(56) By way of example, the implementation of these various treatments makes it possible to prepare clarified hydrolysates of which the compositions are summarized in table III.

(57) TABLE-US-00003  III Characteristics of the various wheat bran or soy cake hydrolysates obtained after successive treatments in order to hydrolyze the phytic acid then the proteins then the starch Soy cake decoction Wheat bran decoction phytic acid hydrolysis by phytic acid hydrolysis exogenous enzymes Characteristic of the + protein + starch + protein clarified juice hydrolysis hydrolysis hydrolysis Dry matter g/kg 51.3 81.2 81.4 83.5 116.6 Nitrogen % DM 4.9 3.8 4.0 4.2 8.3 P.sub.2O.sub.5 % DM 13.7 6.8 7.1 3.7 2.2 Sugar monomer g/kg 15.6 23.8 43.2

(58) It is particularly interesting to note that the P.sub.2O.sub.5/DM contents of the clarified hydrolysates derived from wheat bran are high.

Example 3

Production of Biological Yeast

(59) The various hydrolysates resulting from the treatment of the wheat bran or of the soy cake (example 2) were used for the production of biological yeast, according to a conventional industrial method for producing compressed yeasts.

(60) As a whole, the tests proceeded very well at all levels of the manufacturing cycle tested.

(61) The growth yields observed during the parent yeast stages are comparable to those normally noted with yeast autolysate.

(62) No P.sub.2O.sub.5 deficiency was noted within prefermentation or in first generation (G1 parent yeast) (table IV).

(63) The growth yields are very good.

(64) The fermentative activity of this series of tests is overall very satisfactory and equivalent to that of the reference tests. The preservation of the friability of the compressed yeasts produced is good.

(65) In conclusion, the phosphorus resulting from the hydrolysis of the phytic acid contained in the wheat bran, or the soy cake, can therefore indeed be assimilated by the baker's yeast and allows the manufacture of a quality yeast.

(66) TABLE-US-00004 TABLE IV Phosphorus content noted during the production of biological yeast (the phosphorus content is expressed as % P.sub.2O.sub.5/yeast dry matter) Wheat bran Soy cake hydrolysate hydrolysate Prefermentation 5.5 4.4 Parent yeast (G1) 1.8 1.8 Commercial yeast (G2) 1.7 to 2.1 1.5 to 1.6