METHOD FOR ISSUING PROTEINS AND SACCHARIDS FROM YEAST

Description

Technical area

[0001] The invention relates to a process for digesting yeast, in particular brewer's yeast, for isolating proteins and saccharides. State of the art

[0002] Yeasts are single-celled fungi that reproduce by budding or division and have always been of great technical importance. Yeasts are used in the production of beer, wine, spirits, food, and therapeutic substances. There are basically several types of yeast, with baker's or brewer's yeast (Saccharomyces cerevisiae) being one of the most commonly used. Accordingly, considerable quantities of yeast-containing, and especially brewer's yeast-containing, byproducts are produced.

[0003] Since yeast cell walls, in particular, contain a number of technically useful biomolecules, such as proteins, glucans, mannans, chitins, and lipids, yeast-containing waste is processed to extract these substances. The yeast cell walls are partially or completely digested to release the substances of interest.

[0004] EP 1 990 419 A1 (Tex-a-tec AG), for example, describes a process for isolating glucan, protein, mannan and lipids from yeast, wherein the yeast is treated with ultrasound in successive steps at increasing temperatures.

[0005] EP 2 272 876 A1 (Angel Yeast Co., Ltd.) discloses a process for extracting glucan and mannan from the cell wall of microorganisms, such as yeasts. The process comprises the steps of: a) treating the cells of the microorganism with an alkaline protease and a mannanase; b) separating the mixture from step a) into a heavy phase and a light phase; c) drying the heavy phase obtained from step b), thereby obtaining a glucan preparation; and d) drying the light phase obtained from step b), thereby obtaining the mannan preparation. The glucan and mannan preparations are intended for therapeutic applications.

[0006] WO 2010 / 070207 A1 (Glykos Finland Oy) describes a process for producing an immunostimulatory composition comprising a saccharide fraction, the process comprising the steps of hydrolyzing yeast cells and obtaining a soluble fraction. The resulting saccharide fraction can be incorporated as a food or beverage component or used as a pharmaceutical.

[0007] However, the methods known to date for isolating technically viable biomolecules from yeast are not entirely convincing, especially for large-scale applications. The yield-to-effort ratio, particularly in terms of energy and time requirements, is unsatisfactory in many processes, which greatly hampers commercial production.

[0008] Other processes focus on the selective extraction of individual components (e.g., glucan or mannan), while ignoring the remaining components (e.g., proteins), such as the process described in EP0950716 (Farmint Group Holding SA) or WO2006 / 121803 (Sensient Flavors Inc.). This is disadvantageous in terms of the most complete utilization of yeast-containing by-products.

[0009] There is therefore still a need for improved solutions that at least partially or completely overcome the aforementioned disadvantages. Description of the invention

[0010] The object of the invention is to provide an improved process for the extraction of biomolecules from yeast cells, in particular brewer's yeast cells. In particular, several different biomolecules should be isolated simultaneously with the highest possible yields and purities, particularly preferably proteins, glucans, and mannans. The process should also allow the most efficient extraction of the biomolecules in large-scale applications. It is desirable that the process be carried out with the lowest possible energy and time expenditure.

[0011] The solution to the problem is defined by the features of claim 1. The core of the invention is therefore a method for digesting yeast, in particular brewer's yeast, for isolating proteins and saccharides, comprising the steps: a) Providing a yeast suspension, in particular a brewer's yeast suspension; b) Physically disrupting the yeast in the yeast suspension; c) Microfiltration of the disrupted yeast suspension from step b); d) Ultrafiltration of the permeate from the microfiltration from step c), wherein a phase with proteins as the main solid component is separated as the retentate from the ultrafiltration; e) Treating the retentate from the microfiltration from step c) with a protease under basic conditions and then separating a phase with mannan as the main solid component, in particular by filtration and / or centrifugation; f) Treating the phase retained in step e) successively first under basic and then under acidic conditions and then separating a phase with glucan as the main solid component, in particular by filtration and / or centrifugation.

[0012] It has surprisingly been shown that the inventive combination of physical, chemical, and biochemical treatment steps enables good yields with relatively low energy and time requirements. This applies to all products, i.e., proteins, glucans, and mannans, with yields of up to approximately 40% each being achievable for these substances with high purity.

[0013] Further The process is suitable for large-scale applications. Tens of thousands of tons of yeast suspension per year or several thousand kilograms of yeast suspension per hour can be easily processed.

[0014] Glucans are oligo- or polysaccharides composed of D-glucose molecules linked together by glycosidic bonds. Glucans, such as those found in yeast cells, are typically beta-glucans, which have a β-glycosidic bond, specifically β-1,3-glucan.

[0015] Mannans are polysaccharides that are polymers of the sugar mannose. The mannans found in yeast typically have an α(1-6)-linked backbone and α(1-2)- and α(1-3)-linked side chains that average about two sugar units in length. "Microfiltration" refers to a process for filtration through a filter medium, especially a membrane, with a pore size ≥ 0.1 µm. In contrast, the pore size in ultrafiltration is below 0.1 µm.

[0016] Unless otherwise stated, the "solids content" refers to the dry mass of the respective component. The term "major solids content" refers to the component of the solids with the largest weight percentage.

[0017] The yeast suspension provided in step a) is, in particular, a yeast suspension stored at a temperature of 0-15°C, preferably 2-12°C, especially 3-7°C or 5°C, and / or a pasteurized yeast suspension. This ensures that the suspension contains as few undesirable byproducts as possible.

[0018] In principle, the yeast suspension provided in step a) may also be a yeast suspension stored at a different temperature, in particular room temperature, and not pasteurised.

[0019] The yeast suspension is, in particular, an aqueous yeast suspension. Water is present as a continuous liquid phase within the yeast suspension.

[0020] The yeast suspension is, in particular, a brewer's yeast suspension, preferably containing used brewer's yeast. Such brewer's yeast suspensions arise, for example, as byproducts in beer production and can be used directly in the process according to the invention.

[0021] The yeast suspension preferably has a solids content of 5-30 wt.%, in particular 7-20 wt.%, based on the total weight of the yeast suspension. This has proven particularly suitable for the process according to the invention. If necessary, the solids content of the yeast suspension can be adjusted to the desired range with liquid, in particular water.

[0022] In step a), the yeast suspension has a temperature of 0-15°C, preferably 2-12°C, especially 3-7°C or 5°C. This ensures that the composition of the suspension does not change further, e.g., due to active yeast cells, and reduces the formation of undesirable byproducts.

[0023] Further preferably, step b), and optionally also steps c) and d), are carried out at a temperature of 0-15°C, preferably 2-12°C, in particular 3-7°C or 5°C. This further reduces the formation of undesirable by-products.

[0024] In principle, the yeast suspension in steps a), b), c) and / or d) can also have a different temperature, for example room temperature.

[0025] According to an advantageous embodiment, the provided yeast suspension is subjected to a washing process before step b), in which at least a portion of the liquid phase of the yeast suspension, in particular at least 50% by weight, especially at least 60% by weight, of the liquid phase of the yeast suspension, is replaced by another liquid or a replacement liquid, in particular water. The washing process preferably takes place in a centrifuge or a drum filter.

[0026] The washing process can be used to reduce the proportion of undesirable components dissolved in the liquid, if necessary, thus improving the purity and quality of the substances to be isolated. However, a washing process is optional and can be omitted.

[0027] The physical disruption in step b) is carried out in particular by means of electric fields, in a homogenizer or a grinding media mill, in particular in a ball mill.

[0028] The homogenizer is in particular a high-pressure homogenizer, which is operated in particular with a pressure of more than 1,000 bar, especially more than 1,200 bar.

[0029] In a grinding media mill, the material to be ground and freely moving grinding media, particularly balls, are circulated within a process chamber. This creates collisions between the grinding media, the walls of the process chamber, and the material to be ground. The material is primarily crushed.

[0030] Disintegration with a grinding media mill, particularly a ball mill, has proven particularly advantageous in this case, as it allows the proteins to be isolated to be released gently yet effectively. This is especially true for brewer's yeast suspensions. However, other physical disintegration methods can also be used for specific applications.

[0031] The physical disruption by electric fields in step b) can be carried out, in particular, by pulsed electric fields (also known as PEFs). This disruption technique is known per se to the person skilled in the art in other contexts.

[0032] In pulsed electric field treatment, the yeast suspension is placed between two electrodes and treated with pulsed electric fields. Pulsed electric fields are characterized by high field strengths and short durations. This results in reversible or irreversible pore formation in the cell membranes, allowing intracellular components to be released and / or extracellular substances to penetrate into the cells.

[0033] In this case, the following parameters have proven particularly suitable for controlling the disintegration of the yeast in the yeast suspension: the electric field strength [kV / cm], the inlet temperature of the yeast suspension [°C], and the specific energy input [kJ / L]. Energy input is defined as the energy required for the process in kilojoules per liter of yeast suspension. Furthermore, the energy input can be controlled via the pulse frequency [Hz], which in turn depends on the flow rate and is automatically controlled in existing systems.

[0034] The electric field strength is preferably 1 - 30 kV / cm, in particular 5 - 24 kV / cm.

[0035] The specific energy input is preferably 1 - 180 kJ / L, in particular 3 - 120 kJ / L.

[0036] The inlet temperature of the yeast suspension is preferably 0 - 15°C, more preferably 2 - 12°C, in particular 3 - 7°C or 5°C.

[0037] With such parameters, the yeast in the yeast suspension can be effectively digested.

[0038] Systems for performing physical cell disruption using pulsed electric fields are commercially available from various suppliers, such as the PEF Pilot Dual system (Elea-Technology; Germany).

[0039] Treatment with pulsed electric fields is preferably followed by a mechanical post-treatment. This is therefore preferably carried out between steps b) and c).

[0040] The mechanical post-treatment can be carried out using a mixer, preferably with high shear forces. A rotor-stator mixer, especially an inline rotor-stator mixer, is particularly suitable. In an inline rotor-stator mixer, the rotor and stator are typically contained in a housing with an inlet at one end and an outlet at the other. The yeast suspension to be treated can be drawn through the mixer in a continuous stream.

[0041] The shear rate during mechanical post-treatment with a mixer is selected to further disrupt the yeast cells. This allows the release of any intracellular components (proteins) that may not yet be fully released after application of the pulsed electric fields.

[0042] Due to the high shear forces, the already opened yeast cells are further "squeezed out" and intracellular components, especially proteins, can escape from the cells.

[0043] Microfiltration in step c) is preferably carried out using a filter medium with a pore size of 0.1 - 0.5 µm, preferably 0.1 - 0.2 µm, especially 0.1 µm. This has proven particularly advantageous in order to achieve, after physical disruption, a relative enrichment of the proteins to be isolated in the permeate, based on the solids content, and simultaneously an enrichment of biomass residues, e.g., cell fragments and fats, in the retentate.

[0044] In particular, the microfiltration in step c) is carried out as diafiltration, wherein a replacement liquid, in particular water, is preferably continuously added to the reservoir. In other words, during diafiltration, the liquid phase of the digested yeast suspension, which is discharged as permeate, is preferably continuously replaced, at least partially, in particular completely, by a replacement liquid, in particular water, on the reservoir side.

[0045] The digested yeast suspension is circulated primarily from a reservoir. By regulating the flow downstream of the filter medium, a transmembrane pressure can be established, which acts as the driving force in diafiltration.

[0046] Diafiltration is preferably operated in a closed circuit so that the removal of the permeate leads to the flow of the exchange solvent, in particular so that the circulating volume remains constant during diafiltration.

[0047] Specifically, the microfiltration in step c) is preferably carried out such that a proportion of proteins in the permeate of the microfiltration from step c), based on the total weight of the solids in the permeate, is preferably at least 50 wt.%, more preferably 50 - 80 wt.%, in particular 60 - 70 wt.%.

[0048] A total solids content in the permeate of the microfiltration in step c) is preferably 1 - 10 wt.%, in particular 3 - 7 wt.%, based on the total weight of the permeate.

[0049] Further preferably, the microfiltration in step c) is carried out such that a proportion of proteins in the retentate of the microfiltration from step c) based on the total weight of the solids in the retentate is less than 50 wt.%, preferably less than 40 wt.%, in particular less than 35 wt.%.

[0050] A total solids content in the retentate of the microfiltration in step c) is preferably 20 - 50 wt.%, in particular 25 - 30 wt.%, based on the total weight of the retentate.

[0051] According to another possible embodiment, the digested yeast suspension is treated with an oxidizing agent, in particular ozone (O 3 ) and / or hydrogen peroxide (H 2 O 2 ), before microfiltration step c). This can prevent, in particular, chemical reactions that lead to undesirable byproducts and / or flavor changes. However, treatment with an oxidizing agent is optional.

[0052] Furthermore, the digested yeast suspension can be treated enzymatically before microfiltration step c). This can increase the efficiency and yield of the process if necessary. However, enzymatic treatment before microfiltration step c) is optional.

[0053] The ultrafiltration in step d) is preferably carried out using a filter medium with a pore size of less than 90 nm and / or an exclusion limit (nominal molecular weight cut-off) of 1-100 kDa, especially 5-100 kDa, preferably 10-50 kDa, especially 15-25 kDa, especially 20 kDa. The exclusion limit is defined as the minimum molecular mass of molecules that are 90% retained by the filter medium, especially the membrane.

[0054] In particular, the ultrafiltration in step d) is carried out as diafiltration, with a replacement liquid, in particular water, preferably being continuously added to the reservoir. In other words, during the diafiltration in step d), the liquid phase of the permeate from the microfiltration from step c), which is removed as permeate in the diafiltration, is preferably continuously replaced, at least partially, in particular completely, on the reservoir side by a replacement liquid, in particular water.

[0055] The permeate from the microfiltration process from step c), which forms the feedstock for step d), is circulated from a feed tank. By regulating the flow downstream of the filter medium, a transmembrane pressure can be adjusted, which acts as the driving force for diafiltration in step d).

[0056] The diafiltration in step d) is preferably operated in a closed circuit so that the removal of the permeate leads to the flow of the exchange liquid, in particular so that the circulating volume remains constant during the diafiltration.

[0057] By ultrafiltration and diafiltration in step d), the relative solid content of the protein in the retentate can be specifically increased.

[0058] Preferably, the ultrafiltration in step d) is carried out such that a proportion of the proteins in the phase separated in step d) with proteins as the main solid component, based on the weight of the solids, is at least 75 wt.%, in particular at least 85 wt.%, preferably at least 90 wt.%.

[0059] A total solids content in the phase separated in step d) is preferably 0.5 - 7 wt.%, in particular 1 - 3 wt.%, based on the total weight of the phase separated in step d).

[0060] In particular, both the microfiltration in step c) and the ultrafiltration in step d) are carried out as diafiltration.

[0061] Particularly preferably, the phase separated in step d) containing proteins as the main solid component is dried after step d), in particular spray-dried. This makes the protein available as a powdered product with high purity. Appropriate apparatus and methods for spray drying are known per se to those skilled in the art.

[0062] Particularly preferably, the solids content, in particular the protein content, is concentrated prior to spray drying, in particular to a solids content of at least 50 wt.%, in particular at least 60 wt.%, preferably at least 70 wt. The concentration is carried out in particular in an evaporator, which partially evaporates the liquid phase. Appropriate apparatus and methods for concentration are known per se to those skilled in the art.

[0063] Optionally, the phase separated in step d) containing proteins as the main solid component can be treated with an oxidizing agent, in particular ozone and / or hydrogen peroxide, and / or contacted with an adsorbent material, e.g., activated carbon. This can, in particular, improve the product quality of the proteins. For example, undesirable discolorations and / or flavors can be chemically decomposed and / or removed.

[0064] If spray drying is carried out, the treatment with the oxidizing agent and / or contacting with the adsorbent material is preferably carried out before spray drying and, if such is carried out, before concentration.

[0065] Treatment with an oxidizing agent is particularly preferably carried out using a membrane contactor. In a membrane contactor, the fluids undergoing mass transfer are separated by a porous membrane and passed past each other. In this case, the fluids are the phase separated in step d), which contains proteins as the main solid component, and the oxidizing agent. In the membrane contactor, the oxidizing agent passes through the membrane into the phase separated in step d), which contains proteins as the main solid component. The use of a membrane contactor has proven particularly advantageous and efficient compared to other contactors. This is particularly true because the pressure drop and energy consumption in the membrane contactor are relatively low, yet the oxidizing agent can still exert its effect very effectively.

[0066] In step e), a protease is used. Proteases are enzymes that can hydrolytically cleave proteins. Proteases are also called proteolytic enzymes.

[0067] Preferably, in step e), the protease is used in a proportion of 0.0001-10 wt.%, in particular 0.001-5 wt.%, especially 0.01-1 wt.% or 0.05-0.5 wt.%, based on the solids content of the retentate of the microfiltration from step c).

[0068] The protease is preferably an alkaline protease, in particular a serine protease, most preferably a subtilisin.

[0069] Serine proteases are a subfamily of proteases that have the amino acid serine in their active center.

[0070] Subtilisin-type proteases (subtilases, subtilopeptidases, EC 3.4.21.62) are classified as serine proteases due to their catalytically active amino acids. They are naturally produced and secreted by microorganisms, particularly Bacillus species. They act as nonspecific endopeptidases, meaning they hydrolyze any acid amide bonds found within peptides or proteins. Their pH optimum is usually in the significantly alkaline range.

[0071] Proteases, and in particular the specific proteases mentioned, have proven extremely advantageous in the present process. In particular, the protease can selectively release mannan, which can be separated as a liquid phase with mannan as the main solid component.

[0072] Specifically, the process is carried out without the addition of mannanase or in the absence of it. Mannanase is an enzyme that degrades mannans.

[0073] The treatment in step e) is preferably carried out at a pH of 7.5-13, preferably 8-11, in particular 8.5-10.5, and / or at a temperature of 40-80°C, preferably 50-70°C, in particular 55-65°C. The pH is preferably adjusted to the protease used.

[0074] In particular, in step e), the pH is adjusted by adding a base, preferably a base with a pK B value of less than 4.75, especially less than 1. The pK B value represents the negative decimal logarithm of the base constant KB . It is particularly preferably an inorganic base, especially NaOH. Such bases have proven particularly suitable.

[0075] The base is preferably used in diluted form, e.g. with a proportion of 30 - 60 wt.% in water.

[0076] The treatment in step e) typically lasts from 10 minutes to 18 hours, especially 3 to 12 hours. This allows for optimized yield.

[0077] In another embodiment, a pasty fraction with a small amount of liquid can be processed in step e). This allows the enzymatic step to be optimized in terms of yield, depending on the composition of the yeast suspension and the reaction procedure.

[0078] In step f), the retained phase is preferably treated in a first time interval, preferably for 1-5 hours, at a pH of 7.5-13, preferably 8-11, in particular 8.5-10.5, and / or at a temperature of 40-95°C, preferably 65-85°C, in particular 75-85°C. Subsequently, the retained phase is preferably treated in a second time interval, preferably for 0.5-2 hours, at a pH of 2-6.5, preferably 3-5, in particular 3.5-4.5, and / or at a temperature of 50-95°C, preferably 80-95°C, in particular 80-90°C.

[0079] The "phase retained in step e)" referred to in step f) refers specifically to the phase remaining after separation of the phase containing mannan as the main solid component in step e). In particular, the "phase retained in step e)" thus corresponds to the retentate from step c), from which the phase containing mannan as the main solid component was separated.

[0080] If the phase containing mannan as the main solid component is separated in step e), for example, by filtration, the "phase retained in step e)" referred to in step f) is, in particular, the retentate from step e). In this case, the phase containing mannan as the main solid component in step e) forms the permeate that is separated.

[0081] Particularly preferably, in step f), the pH value in the first time interval is selected such that it corresponds to the pH value present during the treatment in step e).

[0082] Furthermore, it is preferred if in step f) the temperature in the first time interval is higher than the temperature present during the treatment in step e).

[0083] Preferably, in step f) the temperature in the first time interval is lower than in the second time interval and / or the first time interval is longer than the second time interval.

[0084] In step f), if necessary, the pH is adjusted by adding a base, preferably NaOH, and / or by adding an acid, preferably H 2 SO 4 .

[0085] The base is preferably a base as described above in step e). Particularly preferably, the base in step f) is the same base as in step e).

[0086] The acid is in particular an acid with a pK a value of less than 4.75, especially less than 1. The pK a value represents the negative decimal logarithm of the acid constant KS. It is particularly preferably a mineral acid, in particular H 2 SO 4 .

[0087] The acid is preferably used in diluted form, e.g. with a proportion of 30 - 60 wt.% in water.

[0088] These aforementioned measures, especially in combination, have proven particularly advantageous in terms of the efficiency of the process.

[0089] The separation of the phase containing mannan as the main solid component in step e) is preferably carried out by filtration, in particular a combined microfiltration and ultrafiltration. Likewise, the separation of the phase containing glucan as the main solid component in step f) is preferably carried out by filtration, in particular a combined microfiltration and ultrafiltration.

[0090] The phase remaining after separation of the phase containing mannan in step e) is further treated in step f) in particular as the "retained phase".

[0091] Microfiltration is preferably carried out using a filter medium with a pore size of 0.1-0.5 µm, in particular 0.1-0.2 µm, preferably 0.1 µm. Ultrafiltration is preferably carried out using a filter medium with a pore size of less than 90 nm and / or a nominal molecular weight cut-off of 1-100 kDa, in particular 5-100 kDa, preferably 10-50 kDa, in particular 15-25 kDa, especially 20 kDa.

[0092] The microfiltration in step e) and / or the ultrafiltration in step f) are preferably carried out as diafiltration, in particular in the same manner as described for steps c) and d).

[0093] In step e), in particular the phase with mannan as the main solid component in the form of the permeate is removed from the microfiltration, fed to the ultrafiltration and the relative proportion of mannan in the retentate of the ultrafiltration is increased.

[0094] In step f), in particular, the phase with glucan as the main solid component in the form of the permeate is removed from the microfiltration, fed to the ultrafiltration and the relative proportion of glucan in the retentate of the ultrafiltration is increased.

[0095] During diafiltration in steps e) and / or f), the liquids are circulated in particular from the reaction vessel in which steps e) and / or f) take place, with exchange liquid preferably being continuously supplied on the receiver side.

[0096] According to a further advantageous embodiment, the separation in steps e) and / or f) is carried out by a drum filter and / or a centrifuge, particularly in step f).

[0097] In a particular embodiment, the separation in step e) is carried out by a combined microfiltration and ultrafiltration, as described above, and the separation in step f) is carried out by a drum filter and / or a centrifuge, in particular a centrifuge.

[0098] Preferably, the phase separated in step e) containing mannan as the main solid component is dried after step e), in particular spray-dried, and / or the phase separated in step f) containing glucan as the main solid component is dried after step f), in particular spray-dried. This makes both mannan and glucan available as powdered products with high purity.

[0099] Preferably, the solids content, in particular the content of mannan and / or glucan, is concentrated before spray drying, in particular using an evaporator.

[0100] Optionally, the phase separated in step e) with mannan as the main solid component can be treated after step e) with an oxidizing agent, in particular ozone and / or hydrogen peroxide, and / or contacted with an adsorbent material, e.g. activated carbon.

[0101] Likewise, the phase separated in step f) with glucan as the main solid component can optionally be treated after step f) with an oxidizing agent, in particular ozone and / or hydrogen peroxide, and / or contacted with an adsorbent material, e.g. activated carbon.

[0102] As with protein separation, this can improve the product quality of mannan and / or glucan.

[0103] If spray drying is carried out after steps e) and / or f), the treatment with the oxidizing agent and / or the contacting with the adsorbent material preferably takes place before spray drying and, if such takes place, before concentration.

[0104] Treatment with an oxidizing agent is particularly preferably carried out using a membrane contactor. The advantages of this method have been described above in connection with the protein stream.

[0105] Further advantageous embodiments and combinations of features of the invention emerge from the following detailed description and the entirety of the patent claims. Short description of the drawings

[0106] The figures used to explain the example Fig. 1 and 2 show the process flow diagram of a process according to the invention for digesting brewer's yeast for the purpose of isolating proteins (P), mannan (M) and glucan (G).

[0107] In principle, identical parts in the figures are provided with identical reference symbols. Ways to implement the invention

[0108] The Fig. 1 and 2show the process flow diagram of a process according to the invention for digesting brewer's yeast for the purpose of isolating proteins (P), mannan (M) and glucan (G).

[0109] In a first step 100, a cooled aqueous brewer's yeast suspension BS (temperature = 5°C; solids concentration = 15 wt%) is delivered by a tank truck 1 to be temporarily stored in a storage tank 2 at 5°C in the following step 101.

[0110] The brewer's yeast suspension BS is then subjected to a washing process 102 in a centrifuge 3. The liquid phase of the brewer's yeast suspension BS is replaced by water W and fed to a storage tank 4.

[0111] From the storage tank 4, the washed brewer's yeast suspension is then fed into a ball mill 5, where the brewer's yeast cells in the suspension are physically disrupted in the next process step 103. The disrupted brewer's yeast suspension is then subjected to a microfiltration step 104. The microfiltration is carried out as diafiltration using a filter medium with a pore size of 0.1 µm. The disrupted brewer's yeast suspension is circulated from a storage tank 6 by a pump 7 through the microfilter 8, with water W continuously being added to the storage tank as an exchange fluid. The transmembrane pressure, which can be regulated by the permeate outflow, is approximately 1.5 bar. The proportion of proteins in the permeate p1 of the microfiltration, based on the total weight of the solids in the permeate p1, is, for example, 60-70 wt.%.

[0112] The permeate p1 from the microfiltration step 104 is then subjected to an ultrafiltration step 105. The ultrafiltration is carried out as diafiltration using a filter medium with a nominal molecular weight cut-off of 20 kDa. The permeate p1 is circulated from an additional reservoir 9 via a pump 10 through the ultrafilter 11, with water W being continuously added to the reservoir as an exchange fluid. The transmembrane pressure, which can be regulated by the permeate outflow, is approximately 2 bar. The ultrafiltration step 105 increases the relative solids content of the protein in the retentate r2, so that the solids content of the proteins, based on the total solids content in the remaining retentate r2, is approximately 90 wt.%.

[0113] The permeate p2 of the ultrafiltration step 105 is discharged as wastewater (this can be recycled if necessary).

[0114] The retentate r2 from the ultrafiltration step 105, containing proteins as the main solid component, is subjected to a drying process 106. After passing through an evaporator 12, the retentate r2 is fed to a spray-drying device 13. This results in the protein as a powdered product P.

[0115] The protein yield over the entire process is approximately 38 wt% based on the content of available proteins in the brewer's yeast suspension BS.

[0116] The retentate r1 of the microfiltration process 104 is further treated step by step enzymatically and chemically, which results in Fig. 2 is shown. Fig. 2 ends with the position marked with an asterisk (*) at the position marked with an asterisk in Fig. 1 to.

[0117] In a proteolytic digestion step 107, the retentate r1 is treated in a reactor 14, for example, for 8 hours with 0.2 wt.% (based on the solids content of the retentate r1) of a protease E in the form of subtilisin. The treatment takes place at a pH of approximately 9.5, with the pH being adjusted by adding base B in the form of a 45% aqueous solution of NaOH and kept constant during the protease treatment.

[0118] Proteolytic digestion selectively releases mannan, which is then separated by microfiltration and subsequent ultrafiltration in the form of an aqueous phase with mannan as the main solid component. Both microfiltration and ultrafiltration are performed as diafiltration steps. During microfiltration, the liquid to be filtered is circulated from reactor 14 via microfiltration filter 16 (pore size: 0.1 µm) by a pump 15, with exchange fluid continuously being added to the reservoir. The permeate from the microfiltration stage is then fed to the reservoir tank 17 of the ultrafiltration stage, from which the liquid is circulated via ultrafilter 19 (cutoff point: 20 kDa) by a pump 18.Ultrafiltration increases the relative solids content of mannan in the retentate from the ultrafiltration stage, resulting in a solids content of mannan of approximately 65 wt.% relative to the total solids content in the remaining retentate. The permeate from the ultrafiltration stage is discharged as wastewater (this can be recycled if necessary).

[0119] The retentate from the ultrafiltration stage, with mannan as the main solid component, is subjected to a drying process 110. After passing through an evaporator 20, the retentate is fed to a spray-drying device 21. This results in the mannan as a powdered product M.

[0120] The yield of mannan over the entire process is approximately 39 wt.% based on the content of available mannan in the brewer's yeast suspension BS.

[0121] The phase retained in reactor 14 is then heated to a temperature of approximately 80°C and subjected to a basic extraction 108 at a pH of 9.5 for a first time interval of approximately 3 hours. The pH is kept constant by controlled addition of dilute NaOH.

[0122] Subsequently, the temperature is increased again to approximately 85°C and the pH is lowered to 4 by adding 50% H 2 SO 4 in water. The phase retained in the reactor is then subjected to acid extraction 109 over a period of 1 hour.

[0123] After basic and acidic extraction, a liquid phase containing glucan as the main component is separated by diafiltration through the two filter stages (microfilter 16 and ultrafilter 18), similar to mannan. This results in a solids content of approximately 72 wt.% of the total solids content of the retentate from the diafiltration stage.

[0124] The retentate from the ultrafiltration stage, with glucan as the main solid component, is also subjected to a drying process 110, wherein the retentate is fed to the spray drying device 21 after passing through the evaporator 20. This results in the glucan as powdered product G.

[0125] The yield of glucan over the entire process is approximately 42 wt.% based on the content of available glucan in the brewer's yeast suspension BS.

[0126] However, the invention is not limited to the embodiment shown. It can therefore be modified as desired within the scope of the invention.

[0127] For example, other yeast suspensions can be used instead of a brewer's yeast suspension. Non-essential process steps, such as the washing process 102, can also be omitted. Furthermore, cell disruption can be performed using pulsed electric fields instead of a ball mill 5. It is also possible, among other things, to separate the mannan and / or glucan using a drum filter or a centrifuge instead of the described filter stages (microfilter 16 and ultrafilter 18).

[0128] Furthermore, a membrane contactor MK or MK' can optionally be provided upstream of the evaporator 12 and / or the evaporator 20. This allows the corresponding phases to be treated with an oxidizing agent, for example, ozone and / or hydrogen peroxide, before spray drying.

[0129] In summary, a novel and particularly advantageous method for digesting yeast, in particular brewer's yeast, for the isolation of proteins and saccharides has been provided, which is particularly suitable for large-scale approaches.

Claims

1. Method for digesting yeast, in particular brewer's yeast, for the isolation of proteins and saccharides comprising the steps: a) Providing a yeast suspension, in particular a brewer's yeast suspension; b) Physical digestion of the yeast in the yeast suspension; c) Microfiltration of the digested yeast suspension from step b); d) Ultrafiltration of the permeate of the microfiltration from step c) wherein a phase with proteins as the main solid component is separated as the retentate of the ultrafiltration; e) Treating the retentate of the microfiltration from step c) with a protease under basic conditions and subsequently separating a phase with mannan as the main solid component, in particular by filtration and / or centrifugation; f) treating the phase retained in step e) successively first at basic and then at acidic conditions and subsequently separating a phase with glucan as the main solid component, in particular by filtration and / or centrifugation.

2. Method according to claim 1, wherein the yeast suspension provided in step a) has a solids content of 5-30 % by weight, in particular 7-20 % by weight, based on the total weight of the yeast suspension.

3. Method according to any one of the preceding claims, wherein the yeast suspension provided in step a) has a temperature of 0 - 15°C, preferably 2 - 12°C, more particularly 3 - 7°C or 5°C.

4. Method according to one of the preceding claims, wherein the yeast suspension is subjected to a washing process before step b), in which at least part of the liquid phase of the yeast suspension, in particular at least 50% by weight, in particular at least 60% by weight, of the liquid phase of the yeast suspension, is replaced by another liquid, in particular water.

5. Method according to any one of the preceding claims, wherein the physical digestion in step b) is carried out in a homogenizer or a grinding media mill, in particular a ball mill.

6. Method according to any one of the preceding claims, wherein the physical digestion in step b) is performed by pulsed electric fields.

7. Method according to claim 6, wherein the electric field strength is 1 - 30 kV / cm, in particular 5 - 24 kV / cm, and / or the specific energy input is 1 - 180 kJ / L, in particular 3 - 120 kJ / L.

8. Method according to one of the preceding claims, wherein the microfiltration in step c) is carried out with a filter medium having a pore size of 0.1 - 0.5 µm, preferably 0.1 - 0.2 µm, in particular 0.1 µm, and wherein the ultrafiltration in step d) is carried out with a filter medium having a pore size of less than 90 nm and / or an exclusion limit of 1 - 100 kDa, preferably 10 - 50 kDa, in particular 15 - 25 kDa, in particular 20 kDa.

9. Method according to one of the preceding claims, wherein the microfiltration in step c) and / or the ultrafiltration in step d) is carried out as diafiltration, wherein water is preferably continuously supplied on the feed side.

10. Method according to any one of the preceding claims, wherein the phase separated in step d) with proteins as the main solid component is dried, in particular spray-dried, after step d).

11. Method according to one of the preceding claims, wherein in step e) protease is used in a proportion of 0.0001-10 % by weight, in particular 0.001-5 % by weight, more particularly 0.01-1 % by weight or 0.05-0.5 % by weight, based on the solids content of the retentate of the microfiltration from step c), wherein the protease is preferably a subtilisin and wherein the treatment in step e) is carried out at a pH of 7.5 - 13, preferably 8 - 11, in particular 8.5 - 10.5, and at a temperature of 40 - 80°C, preferably 50 - 70°C, in particular 55 - 65°C.

12. Method according to any one of the preceding claims, wherein the treatment with protease in step e) lasts 10 min to 18 hours, in particular 3 - 12 hours.

13. Method according to any one of the preceding claims, wherein in step f) the retained phase is treated in a first time interval, preferably for 1 - 5 hours, at a pH of 7.5 - 13, preferably 8 - 11, in particular 8.5 - 10. 5, and a temperature of 40 - 95°C, preferably 65 - 85°C, in particular 75 - 85°C, and subsequently treated in a second time interval, preferably for 0.5 - 2 hours, at a pH of 2 - 6.5, preferably 3 - 5, in particular 3.5 - 4.5, and a temperature of 50 - 95°C, preferably 80 - 95°C, in particular 80 - 90°C.

14. Method according to claim 13, wherein in step f) the pH in the first time interval is equal to the pH present during the treatment in step e), and wherein in step f) the temperature in the first time interval is lower than in the second time interval and / or the first time interval is longer than the second time interval.

15. Method according to one of the preceding claims, wherein the separation of the phase with mannan as the main solid component in step e) and / or the separation of a phase with glucan as the main solid component in step f) is carried out by filtration, in particular a combined microfiltration and ultrafiltration.

16. Method according to claim 15, wherein the microfiltration is carried out with a filter medium having a pore size of 0.1 - 0.5 µm, in particular 0.1 - 0.2 µm, preferably 0.1 µm, and wherein the ultrafiltration is carried out with a filter medium having a pore size of less than 90 nm and / or an exclusion limit of 1 - 100 kDa, preferably 10 - 50 kDa, in particular 15 - 25 kDa, in particular 20 kDa.

17. Method according to any one of claims 15 - 16, wherein the microfiltration in step e) and / or the ultrafiltration in step f) is carried out as diafiltration.

18. Method according to any one of the preceding claims, wherein the phase separated in step e) with mannan as the main solids component after step e) is dried, in particular spray-dried, and / or wherein the phase separated after step f) with glucan as the main solids component after step f) is dried, in particular spray-dried.