AU2007251849A1 - Process for the production of yeast - Google Patents

Description

NSPERFECT Affidavit of Accuracy This is to certify the following document WO 2007/131669 has been translated from German into English by staff members of TransPerfect Translations familiar with both the English and German languages and is to the best of our knowledge, ability and belief, a true and accurate translation. Sara J. Green For TransPerfect Translations Sworn to before me this Tuesday, November 04, 2008 Signature, Notary Public SUSAN C. CHRISTIAN . Notary Puojie Lummonwealth ot Massacrwsetts cunfnissiun hinreaS Aug4 14, 2W9 4T0 BOYLSrON STREET. 6TH FLOOR, BOSTON, MA 02116 T 617.52, 6936 F 617.523.0595 WWW TRANSPERFECTCOM WO 2007/131669 PCT/EP2007/004070 DESCRIPTION PROCESS FOR THE PRODUCTION OF YEAST BACKGROUND OF THE INVENTION Yeasts are the most important group of microorganisms of commercial value worldwide. They have been used by humans for thousands of years. The total quantity of yeast production when brewing and fermenting and in food production amounts to millions of tons per year. Yeast production in the European Community has a turnover of 800 million Euros and secures more than 8,000 jobs, supplier industries not even counting. The most important yeast from a commercial point of view is saccharomyces cervisiae. However, there are numerous less common yeasts with a potential for technological applications. Kluyveromyces lactis, for example, is an important strain of yeast These microbes are used in the large-scale production of chymosin (rennin) in fermenters; this rennin that replaces the conventional form obtained from slaughtered animals is widely used today in cheese production. Saccharomyces yeasts are generally rated as safe ("GRAS"). They produce two important metabolites-ethanol and carbon dioxide. Ethanol is used for the production of alcoholic beverages, as a fuel and solvent. Carbon dioxide is used as a leavening agent for bread dough, for producing carbonated beverages, as an inert gas when preserving foods and as a solvent for extractions. Yeast itself is used as a flavor enhancer in foods and as a nucleotide source in the production of substitutes for breast milk. In addition, yeast is an important source of vitamin B in human and animal nutrition. Sterile yeast extract is used in the cultivation of industrial mold fungus cultures for enzyme production or for producing starter CONFIRMATION COPY WO 2007/131669 PCT/EP2007/004070 2 cultures. Feed yeast is used in cattle and horse feed for stabilizing an anaerobic environment in the rumen. Yeast modified by genetic engineering is used for producing proteins such as insulins that are applied in human medicine. Yeast is always produced from a pure yeast culture that has been isolated by selection, breeding, or genetic modification with regard to its power in the intended application. When producing pure yeast cultures, the dual principle of using a bacteriologically flawless sample for initial inoculation and maintaining this purity over the entire yeast production cycle. Baker's yeast (saccharomyces cervisiae) has its origin in top-fermentation brewer's yeast. A pure yeast culture is obtained by selecting certain yeast strains and their properties and by subsequent cell multiplication on a nutrient substrate of molasses and various additives. Baker's yeast, for example, is characterized by high dough raising power and a low content of gluten-degrading enzymes. While a particular yeast strain is, of course, a trade secret proprietary to the yeast producer, the technological process of yeast multiplication is generally known. Multi-stage cultivation is common in bioengineering to produce a large scale of pure culture biomass. In a one-stage cultivation process in which a large substrate volume is inoculated with a small quantity of the respective microorganisms, sufficient biomass multiplication would take a long time. This would have several disadvantages: A longer dwell time in which the yeast creates an optimum environment for yeast growth (reduced redox potential, concentration of growth stimulators), a high risk of contamination with undesirable organisms, and economic inefficiency.

WO 2007/131669 PCT/EP2007/004070 3 Yeast utilizes a wide range of carbohydrates and sugars. But there is as yet no type of yeast known that is capable of utilizing all the sugars available. The top-fermentation strain saccharomyces cerevisiae typically utilizes glucose, fructose, mannose, galactose, saccharose, maltose, maltotriose, and raffinose. Saccharomyces cerevisiae and related strains, however, cannot utilize C5 sugars such as ribose, xylose, arabinose, or cellobiose. The most common substrate for the cultivation of yeast is an aqueous solution of 8 - 10% of molasses. Molasses contains about 50% sugar. The molasses is clarified to separate any sludge and sterilized using high-pressure vapor. It is then diluted with water and kept in buffer vessels until it is needed for the fermentation process. The solution is then set to a pH value of about 4.5 to 5.0 using inorganic acids and enriched with minerals and vitamins of the B group that are needed for yeast production. Required nutrients and minerals include nitrogen, potassium, phosphate, magnesium, and calcium with traces of iron, zinc, copper, manganese, and molybdenum. The substrate is typically supplied with nitrogen by adding ammonium salts, ammonia liquor, or anhydrous ammonia. Phosphates and magnesium are added as phosphoric acid or phosphate and magnesium salts. Vitamins (biotin, inosite, pantothenic acid, and thiamine) are also required for yeast growth. The yeast cultures are aerated to achieve as great a biomass yield as possible. Yeasts are highly developed monocellular fungi. They are facultatively anaerobic: In the presence of air, they produce carbon dioxide and water from sugar and oxygen. This metabolic process is called respiration and produces a great amount of energy (ATP). Glucose is completely oxidized by respiration, and all the biochemical energy it contains is released. Glucose + Oxygen -+ Carbon Dioxide + Water + Energy (38 ATP) WO 2007/131669 PCT/EP2007/004070 4 This energy is used by yeast for keeping up vital functions and for synthesizing biomass, i.e. for yeast growth. A few other substances besides oxygen and sugar are required for producing biomass, in particular, nitrogen. In the absence of oxygen, yeast can still produce energy from sugar to keep up its vital functions. The anaerobic metabolism of yeast has been defined as fermentation by Louis Pasteur. During fermentation, carbon dioxide and ethanol are produced from sugar. Glucose oxidation is incomplete. Ethanol contains a quantity of energy so that only a small portion of the biochemical energy contained in the glucose is released: Glucose -> Carbon dioxide + Ethanol + Energy (2 ATP) Accordingly, the multiplication of yeast cells is very limited in anaerobic conditions. Industrial yeast production is based on the aerobic process. Air is blown through the solution in which the yeast is grown to create aerobic conditions. The problem the yeast producer is facing is not as simple as just supplying air during the fermentation process. If the sugar concentration in the growth medium exceeds a minor amount, the yeast will produce some alcohol even if the air supply is sufficient or abundant (Crabtree effect). This problem can be solved by adding the sugar solution to the yeast slowly over the entire fermentation process. The rate of adding the sugar solution must be selected so that the yeast uses up the sugar fast and the sugar concentration virtually is zero at any given point in time. This type of fermentation is called fed-batch fermentation. Yeast is cultivated in several stages when producing baker's yeast. For example, yeast could be transferred from a test-tube culture in substrate with 50 ml, WO 2007/131669 PCT/EP2007/004070 5 then I liter, 10 liters, 40 liters, 400 liters, 4 m 3 , 10 M 3 , and finally 200 M 3 . The initial stage of yeast growth takes place in the lab. A portion of the pure yeast culture is mixed with molasses wort in a sterilized flask and left to grow in for 2 to 4 days. The entire contents of the flask is then used to inoculate the first fermenter in the purified culture stage. Purified culture fermentations are discontinuous fermentations in which the yeast multiplies in 13 to 24 hours. One to two fermenters are used in this stage of the process. But for the fact that there are no precautions for sterile aeration and aseptic transfer to the next stage, purified culture fermentation is basically a continuation of the flask fermentation step. All growth media and nutrients are introduced to the vessel prior to inoculation. The next fermentation stage is a stock culture fermentation. The contents of the intermediate fermenter is pumped into the stock fermenter that is designed for continuous aeration. This stage is called inoculating or budding yeast fermentation since the yeast is separated by centrifuging from the major portion of the fermenter liquid after fermentation is complete, which yields a stock culture for inoculating the next stage. Aeration is powerful, and molasses and other nutrients are added in steps. The liquid from this fermenter is typically divided into several portions for pitching the concluding shipping yeast cultivation process. Alternatively, the yeast can be separated by centrifuging and stored for several days before use in the concluding shipping yeast cultivations. The introduction of the fed-batch process into yeast production marks the beginning of modern yeast industry. Fed-batch production was introduced in Germany in 1915, and today's yeast production is still relying on the fed-batch process. In a fed-batch fermentation WO 2007/131669 PCT/EP2007/004070 6 process, molasses, phosphoric acid, ammonia and minerals of yeast are added at a controlled rate. The rate is selected so that the yeast is fed just that quantity of nutrients and sugar needed to maximize multiplication and prevent the production of alcohol. At the start of each batch, a relatively considerable quantity of process water is transferred to the fermenter together with the budding yeast. The optimum quantity of sugar to be added per gram of yeast and per hour is predefined. Molasses is added to the fermenter at a rate that corresponds to the predetermined quantity of sugar to be added. When making baker's yeast, a basic supply of 16 to 18 g/l is common. Nitrogen and phosphorus can either be fed at a specific rate or intermixed with the process water prior to adding molasses. The quantities are determined by the weight ratio of sugar, nitrogen, and phosphorus. The yeast culture has ripened after 8 to 16 hours of cultivation. 2.5 g of nitrogen and 5.0 g of phosphorus or 0.3 g of nitrogen and 0.5 g of phosphorus per 100 g of sugar are required for yeast production. The cultivation temperature is typically between 25*C and 35*C, the pH value is determined by the quantities of nitrogen added. Budding yeast is produced at a pH value between 4.5 and 5.0; shipping yeast is produced at a pH value between 5.0 and 7.0. The concluding shipping yeast production has the highest degree of aeration, and molasses and other nutrients are added in steps. Considerable quantities of air must be supplied to the concluding shipping yeast production process so that these vessels are often started in a staggered pattern to reduce the size of the air compressors. The final fermentation stages take around 11 to 15 hours. After adding all the molasses required to the fermenter, the liquid is aerated for another 0.5 to 1.5 hours to allow further ripening of the yeast, which makes the yeast WO 2007/131669 PCT/EP2007/004070 7 more stable for refrigerated transport. This method also ensures that small quantities of ethanol are removed by respiration. At the end of the fermentation process, the fermenter broth is centrifuged off, washed with water and re-centrifuged, yielding a yeast milk with a solids concentration of about 18%. The yeast milk is cooled down to about 8'C and stored in a separate, refrigerated yeast milk container made of stainless steel. The yeast milk can directly be loaded onto tankers and carried to the customers that have a respective yeast milk handling system. Before 1825, yeast was only available as yeast milk. Tebbenhof introduced pressed yeast in 1825. Yeast milk production became popular again when wholesale bakeries for large-scale output developed. Alternatively, yeast can be centrifuged, and the solid yeast can be further concentrated using a filter press or rotary vacuum filter. A filter press will yield a filter cake containing 27 to 32 percent of solids. A rotary vacuum filter will yield a filter cake containing about 33 percent of solids. The filter cake is than intermixed in mixers with small quantities of water, emulgators, and flux oils to become the final product. Emulgators are added in the production of pressed yeast to give the yeast a white, creamy appearance and to inhibit the formation of water stains on the yeast cake. A small quantity of oil, typically soy bean or cottonseed oil, is added to enhance extrusion of the yeast through nozzles into endless strips of yeast cake. These strips are cut, and the yeast cakes are packaged and cooled down to a temperature of less than 8'C whereupon they are ready for transport on a refrigerated truck. 100 kg to 250 kg of budding yeast and one ton of molasses are required to produce one ton of pressed yeast with a dry-matter content of 27% (Y27).

WO 2007/131669 PCT/EP2007/004070 8 When producing active dry yeast, pressed yeast is molded into cylinders that are dried in a fluidized-bed drier. Active dry yeast can be stored at room temperature. The molasses substrate is replaced with starch substrates when producing organic yeast. Wheat, corn, or potato starch are dextrinized into a sugar solution using technical enzymes. Organic baker's yeast produces less carbon dioxide since the final product contains about 30% of starch. Fed-batch fermentations are not completely sterile. It is inefficient to use pressure vessels for ensuring the sterility of the huge volumes of air needed in these fermenters or for creating sterile conditions for all transfer processes through numerous lines, pumps, and centrifuges. The equipment is thoroughly cleaned, pipes and containers are charged with steam, and the air is filtered to ensure conditions are as aseptic as possible. However, it is almost impossible to avoid a certain form of microbial contamination, and the presence of bacteria such as leuconostoc, pediococcus, aerococcus, bacillus, lactococcus and e.coli have been reported. The fed-batch stages are cultivated for a limited period of time (10 to 20 hours). The relatively short cultivation period and a large yeast inoculum are to prevent foreign. organisms from growing into a visible infection. Presently, controlling infection seems to be limited to physical methods such as the use of wash separators for removing as many bacteria as possible with the yeast wort due to the difference in density when harvesting yeast cells. Another preventive measure occasionally proposed is to reduce the pH value but optimum yeast growth can only be achieved at a pH value around 6. Sometimes sodium chloride is used to support the preservation of pressed yeast over a longer period of time.

WO 2007/131669 PCT/EP2007/004070 9 Controlling infection in yeast production is a must for ensuring the purity of the yeast culture. Bacterial contamination is particularly interfering when yeast is used in fermentation processes such as in beer or ethanol production. In this case, the yeast could be the source of infection for the fermentation process. Yeast producers are fully aware that the number of bacterial contaminations per gram of yeast milk, pressed yeast, or active dry yeast is a decisive factor determining the quality of the yeast. This invention provides a method of controlling bacterial contamination during the production and storage of yeast. U.S. patent publication 6,326,185 BI describes a method of decontaminating brewer's yeast used in fermentation so that this yeast can be used in subsequent fermentations. The yeast is brought into contact with tetrahydroiso-a acids up to a final concentration of 40 ppm while the pH value of the mixture is set to about 2.0 to 2.6. The mixture is then kept at a specific temperature over a specific period of time. U.S. patent publication 1,477,132 relates to a yeast composition, in particular a yeast composition for rice fermentation, and a production method therefor. Cooked rice is intermixed with hop juice. The rice is treated with yeast so that the yeast can grow. Hop bitter acid is added to prevent oxidation, degradation and other toxic metabolic processes caused by bacteria. WO 2004/072291 A2 relates to the use of hop acids in bioethanol production for the inhibition of microorganisms. This application only mentions in general that the method for controlling microorganisms using hop bitter acid can also be applied when multiplying yeast.

WO 2007/131669 P'T/LV2UU7/UU4U7U 10 It is the problem of this invention to provide a method for the commercial production of yeast in which the activity of microorganisms can be prevented or at least decreased in a relatively cost-efficient manner. It is another problem of this invention to provide a method for the commercial production of yeast in which physical methods of decreasing the activity of unwanted microorganisms in yeast can be avoided or at least be simplified. It is another problem of this invention to provide a method for the commercial production of yeast in which remedial building work, such as pressure vessels at the stage of fed-batch production of the yeast, can be avoided or at least be simplified. It is another problem of this invention to provide a method for the commercial production of yeast in which the required intensity of cleaning the equipment can be reduced. It is another problem of this invention to provide a method for the commercial production of yeast, in particular shipping yeast, in which the use of chemical antifoaming agents can be reduced or avoided. BRIEF SUMMARY OF THE INVENTION According to the invention, yeast is brought into contact with a sufficient quantity of an auxiliary processing agent that is either a natural substance or derived from a natural substance for inhibiting or at least decreasing the activity of microorganisms during the commercial production of budding and/or shipping yeast. This measure can effectively replace other expensive measures aimed at inhibiting or at least decreasing the activity of unwanted microorganisms.

WO 2007/131669 PCT/EP2007/004070 11 The auxiliary processing agent that is either a natural substance or derived from a natural substance is selected from at least one member of the group including hop bitter acid, colophonium, and myristic acid or derivatives thereof. All these acids have a bactericidal effect. It is preferred that the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the carbohydrate substrate for cultivation prior to adding the yeast culture. It is preferred that the auxiliary processing agent that is either a natural substance or derived from a natural substance is added prior to any heat treatment of the substrate. It is preferred that the auxiliary processing agent that is either a natural substance or derived from a natural substance is added continuously. In addition, or alternatively, the auxiliary processing agent that is either a natural substance or derived from a natural substance is added before the step of producing budding yeast. Since the budding yeast is washed before it is transferred to the process step of producing shipping yeast, hop bitter acid and, in particular, tetrahydroiso-a acid wit its bitter taste, can surprisingly be used without any negative effect on the taste of the finished yeast product. In addition, or alternatively, the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the step of producing shipping yeast. In addition, or alternatively, the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the step of producing yeast milk. Depending on the pH value of the respective production step, specific acids or derivatives thereof are provided for the best effect. It is preferred that the auxiliary processing agent that is either a natural substance or derived from a natural substance is added discontinuously, in other words, as a shock dose, preferably at the beginning of the production step. The concentration WO 2007/131669 PCT/EP2007/004070 12 of the auxiliary processing agent that is either a natural substance or derived from a natural substance is higher at the beginning of the production step and decreases over time. If microorganisms are introduced to a production step by a contaminated nutrient or contaminated equipment and would thus interfere with the yeast production process, such activity by microorganisms is prevented from the start, and the yeast production process will neither be impaired by the initial contamination nor by contamination introduced later. Since the auxiliary processing agent that is either a natural substance or derived from a natural substance is added as a shock dose at the beginning of the production step, its concentration is diluted in the course of the yeast production process so that the bitter taste of the hop bitter acid is decreased or completely avoided. This is particularly true if the production process of budding yeast is a fed-batch method. The concentration of the auxiliary processing agent that is either a natural substance or derived from a natural substance decreases in the course of the fed-batch process. The auxiliary processing agent that is either a natural substance or derived from a natural substance is preferably selected from at least one member of the group of hop bitter acids that includes alpha acids, beta acids, rho-iso-ct acids, iso-a acids, hexahydroiso-a acids, tetrahydroiso-a acids, and hexahydro-beta acids. It is preferred that tetrahydroiso-a acids or beta acids are added alone or in combination during the process step of producing budding yeast as the auxiliary processing agent that is either a natural substance or derived from a natural substance. Tetrahydroiso-a acids are the most effective hop compound at the pH value of 4 to 5 that is typically found when producing budding yeast. This is why it is most useful to add tetrahydroiso-a acids in this process step.

WO 2007/131669 PCT/EP2007/004070 13 The initial concentration of tetrahydroiso-a acids is in the range from I - 1000 ppm, preferably 10 - 100 ppm, preferably 20 - 60 ppm, and most preferably 15 - 30 ppm. This concentration provides the bactericidal effect. On the other hand, the hop bitter acid is sufficiently diluted throughout the production process that it does not impair the taste of the yeast. The final concentration of tetrahydroiso-a acids in the treated container is in the range from 5 - 20 ppm, preferably 3 - 15 ppm, and most preferably 2 - 10 ppm, which is sufficient for inhibiting bacteria growth. Beta acids are preferably added as an auxiliary processing agent that is either a natural substance or derived from a natural substance to the step of producing shipping yeast. It has been found that beta acids contribute to the prevention of yeast cell flocculation caused by electrochemical interaction with bacteria. In addition, beta acids are best qualified for use in the shipping yeast production step because they are most effective at a pH value of 5 to 7, the typical pH value at the step of producing shipping yeast. It has also been determined that beta acids have an antifoaming effect in the production step of shipping yeast. The use of beta acids that are added as an auxiliary processing agent that is either a natural substance or derived from a natural substance in the step of producing shipping yeast therefore helps to reduce or avoid the use of chemical antifoaming agents. The initial concentration of beta acids is in the range from I - 1000 ppm, preferably 10 - 100 ppm, and most preferably 40 - 60 ppm. The final concentration of beta acids is in the range from 8 - 300 ppm, preferably 3 - 30 ppm, and most preferably I - 12 ppm. Alternatively or in addition, colophonium and/or myristic acid may be added as an auxiliary processing agent that is either a natural substance or derived from a natural substance during WO 2007/131669 PCT/EP2007/004070 14 the production of shipping yeast. These acids reach their best performance at a pH of 5.0 to 7.0 as well. For best results, the initial concentration of colophonium and/or myristic acid is in the range from I - 1000 ppm, preferably 5 - 500 ppm, and most preferably 10 - 100 ppm. It has been found that colophonium and/or myristic acid can also be added to the carbohydrate substrate at the beginning of the yeast production process. It is preferred that the pH value during this step of producing shipping yeast does not drop below 5.0, more preferably not below 6.0. DESCRIPTION Embodiments of the method according to the invention are described below with reference to the figures. Wherein: Fig. 1 shows an example of the production of baker's yeast on molasses substrate; Fig. 2 represents a diagram showing the inhibition of bacteria growth in MRS broth at pH 5 using tetrahydroiso-a acids and beta acids. Fig. I shows a diagrammatic view (flowchart) of the production process of baker's yeast. A mixture of sugar cane and beet molasses is sterilized by heat treatment with high-pressure steam, and its pH value is set by acidification with sulphuric acid. The molasses is clarified to remove any sludge. Clarified, sterile molasses is used as so-called carbohydrate substrate to produce yeast biomass.

WO 2007/131669 PCT/EP2007/004070 15 According to a first embodiment of the invention, hop bitter acids, in particular, tetrahydroiso-a acids, are added to the clarified sterile molasses (see line A in Fig. 1) to prevent any activity by gram-positive bacteria, especially in the subsequent processes. The substrate typically is an aqueous solution containing 8 - 10% molasses. The molasses contains about 50% sugar. The clarified sterile molasses is introduced to the purified culture fermenter in which the molasses is intermixed with process water and purified culture. According to another embodiment of the invention, hop bitter acids, in particular, tetrahydroiso-a acids, are added to the purified culture fermenter (see line B in Fig. 1) to prevent or at least decrease the activity of unwanted microorganisms. The mixture is then transferred from the aqueous molasses solution and the purified yeast culture to a budding yeast fermenter. In the budding yeast fermenter, the solution is set to a pH value of about 4.0 to 5.0 and enriched with minerals and vitamins of the B group that are needed for proper yeast growth. Required nutrients and minerals include nitrogen, potassium, phosphate, magnesium, and calcium with traces of iron, zinc, copper, manganese, and molybdenum. The substrate is typically supplied with nitrogen by adding ammonium salts, ammonia liquor, or anhydrous ammonia to the input material. Phosphates and magnesium are added as phosphoric acid or phosphate and magnesium salts. The yeast cultures in the budding yeast production process are also aerated to achieve a maximum biomass yield. According to another embodiment of the invention, hop bitter acids are introduced to the budding yeast fermenter (see line C in Fig. 1) to prevent or at least decrease the activity of unwanted microorganisms.

WO 2007/131669 PCT/EP2007/004070 16 These acids are preferably added as a shock dosage at the beginning of the budding yeast fermentation process. The final concentration of the hop bitter acids, in particular, tetrahydroiso-ax acids, is between 5 and 10 ppm. It has been determined that tetrahydroiso-a acids are most effective in suppressing or at least decreasing the activity of microorganisms in the budding yeast production step. Even small quantities of tetrahydroiso-a acids are sufficient to achieve the desired result. The budding yeast production process is carried out as a fed-batch process in which molasses, phosphoric acid, ammonia, and minerals are added to the yeast at a controlled rate. Thus the initial concentration of hop bitter acids decreases in the course of the fed-batch production process. As a result, the hop bitter acid concentration in the fermenter decreases and any bitterness of the final product that might be caused by the hop bitter acid is surprisingly prevented after the washing step. The budding yeast is then washed in the budding yeast wash separators where hop bitter acids, i.e. tetrahydroiso-a acids, are partially removed, which in addition prevents any bitterness in the final product. The budding yeast is then stored and prepared for the production of shipping yeast in shipping yeast fermenters. According to another embodiment of the invention, hop bitter acids, in particular, beta acids, are added in the step of producing shipping yeast (see line D in Fig. 1). Since the pH value in the production step of shipping yeast is in the range from 5 to 7, beta acids are most effective here. The use of beta acids in the shipping yeast production process also provides an antifoaming effect. As a result, the use of chemical antifoaming agents can be decreased or even avoided. In addition, adding beta acids has the advantageous effect that flocculation of yeast cells that is often caused by electrochemical interaction with bacterial yeast cells WO 2007/131669 PCT/EP2007/004070 17 can be prevented since beta acids prevent adhesion of such unwanted microorganisms to the surface of the yeast cells. After completion of the shipping yeast fermentation process, the shipping yeast is washed and further processed into yeast milk, pressed yeast, or eventually dry yeast. According to another embodiment of the invention not illustrated in Fig. 1, auxiliary processing agents that are either a natural substance or derived from a natural substance may be added to yeast milk to fight any activity of unwanted microorganisms as may be caused by re-infection with the goal to extend the storage quality of the shipping yeast product. It goes without saying that the invention covers, according to the invention, the addition of hop bitter acids in the steps A, B, C, and D in combination (A, B, C and D), as individual measure (e.g. C only), or in partial combination (such as C and D). Alternatively or in addition to hop bitter acid, colophonium and/or myristic acid can be added as an auxiliary processing agent that is either a natural substance or derived from a natural substance in the shipping yeast production step (see line D in Fig. 1). Fig. 2 reveals the inhibiting effect of tetrahydroiso-a acids at a concentration of about 5 ppm on the activity of bacteria. It has been found that beta acids become less effective as compared to tetrahydroiso-a acids at a lower pH value, which means that about triple the amount of beta acid as compared to tetrahydroiso-a acid has to be applied to achieve a similar efficiency.

Claims (21)

1. An improved method of producing yeast, in particular, baker's yeast, wherein yeast is brought into contact with a sufficient quantity of an auxiliary processing agent that is either a natural substance or derived from a natural substance for suppressing or at least decreasing the activity of unwanted microorganisms during the commercial production of budding and/or shipping yeast.

2. The method according to claim I wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is selected from at least one member of the group including hop bitter acid, colophonium, and myristic acid or derivatives thereof.

3. The method according to claim 1 or 2 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to a carbohydrate source for yeast cultivation prior to adding the yeast culture.

4. The method according to claim 3 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added before or after the heat treatment of the carbohydrate substrate yeast multiplication.

5. The method according to claim 3 or 4 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added continuously.

6. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the budding yeast production step.

7. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the shipping yeast production step.

8. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance WO 2007/131669 PCT/EP2007/004070 19 is added to the yeast milk production step or to the finished yeast product.

9. The method according to any one of claims 6 to 8 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added discontinuously, preferably at the beginning of the respective production step.

10. The method according to claim 6 wherein the budding yeast production process is a fed-batch process and the concentration of the auxiliary processing agent that is either a natural substance or derived from a natural substance decreases in the course of the fed-batch process.

11. The method according to claim 6 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the washing medium of the budding yeast.

12. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is preferably selected from at least one member of the group of hop bitter acids that includes alpha acids, beta acids, rho-iso-a acids, iso-a acids, hexahydroiso-a acids, tetrahydroiso-a acids, and hexahydro-beta acids.

13. The method according to claim 12 wherein tetrahydroiso-a acids or beta acids are added in the budding yeast production step as an auxiliary processing agent that is either a natural substance or derived from a natural substance.

14. The method according to claim 13 wherein the final concentration of tetrahydroiso-a acids preferably is in the range from 5-20 ppm, preferably 3-15 ppm and most preferably 2-10 ppm.

15. The method according to claim 7 wherein beta acids are added to the shipping yeast production step as an auxiliary processing agent that is either a natural substance or derived from a natural substance. WO 2007/131669 PCT/EP2007/004070 20

16. The method according to claim 15 wherein the final concentration of beta acids preferably is in the range from 8 - 300 ppm, preferably 3 - 30 ppm and most preferably 1 - 12 ppm.

17. The method according to claim I wherein colophonium and/or myristic acid is/are added to the shipping yeast production step as an auxiliary processing agent that is either a natural substance or derived from a natural substance.

18. The method according to claim 17 wherein the initial concentration of colophonium and/or myristic acid is in the range from I - 1000 ppm, preferably 5 - 500 ppm, and most preferably 10 - 100 ppm.

19. The method according to claim I wherein colophonium and/or myristic acid is added to the carbohydrate substrate.

20. The method according to claim 1 wherein the pH value during the shipping yeast production process does not drop below 5.0, preferably not below 6.0. WO 2007/131669 PCT/EP2007/004070

21 Received at the International Bureau on October 8, 2007 (10/08/2007) 1. An improved method of producing yeast, in particular, baker's yeast, wherein yeast is brought into contact with a sufficient quantity of an auxiliary processing agent that is either a natural substance or derived from a natural substance for suppressing or at least decreasing the activity of unwanted microorganisms during the commercial production of budding and/or shipping yeast, the auxiliary processing agent that is either a natural substance or derived from a natural substance being selected from at least one member of the group including hop bitter acids or a derivative thereof. 2. The method according to claim 1 or 2 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to a carbohydrate source for yeast cultivation prior to adding the yeast culture. 3. The method according to claim 2 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added before or after the heat treatment of the carbohydrate substrate yeast multiplication. 4. The method according to claim 2 or 3 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added continuously. 5. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the budding yeast production step. 6. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the shipping yeast production step. 7. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the yeast milk production step or to the finished yeast product. AMENDED SHEET (ARTICLE 19) WO 2007/131669 PCT/EP2007/004070 22 8. The method according to any one of claims 5 to 7 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added discontinuously, preferably at the beginning of the respective production step. 9. The method according to claim 5 wherein the budding yeast production process is a fed batch process and the concentration of the auxiliary processing agent that is either a natural substance or derived from a natural substance decreases in the course of the fed batch process. 10. The method according to claim 5 wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is added to the washing medium of the budding yeast. 11. The method according to any one of the preceding claims wherein the auxiliary processing agent that is either a natural substance or derived from a natural substance is preferably selected from at least one member of the group of hop bitter acids that includes alpha acids, beta acids, rho-iso-a acids, iso-a acids, hexahydroiso-a acids, tetrahydroiso-a acids, and hexahydro-beta acids. 12. The method according to claim 11 wherein tetrahydroiso-a acids or beta acids are added in the budding yeast production step as an auxiliary processing agent that is either a natural substance or derived from a natural substance. 13. The method according to claim 12 wherein the final concentration of tetrahydroiso-a acids preferably is in the range from 5 - 20 ppm, preferably 3 - 15 ppm and most preferably 2 - 10 ppm. 14. The method according to claim 6 wherein beta acids are added to the shipping yeast production step as an auxiliary processing agent that is either a natural substance or derived from a natural substance. AMENDED SHEET (ARTICLE 19) WO 2007/131669 PCT/EP2007/004070 23 15. The method according to claim 14 wherein the final concentration of beta acids preferably is in the range from 8 - 300 ppm, preferably 3 - 30 ppm and most preferably 1 - 12 ppm. 16. The method according to claim I wherein the pH value during the shipping yeast production process does not drop below 5.0, preferably not below 6.0. AMENDED SHEET (ARTICLE 19)