Method for green extraction of waste beer yeast cell wall polysaccharide by using eutectic solvent

Abstract

The invention provides a method for green extraction of waste beer yeast cell wall polysaccharide by using a deep eutectic solvent, relates to the field of biomass resource utilization, and aims to solve the problems of environmental pollution, high cost and insufficient resource utilization in the prior art. The method comprises the following steps: pretreating a yeast raw material and a deep eutectic solvent to selectively dissolve mannan; performing solid-liquid separation for the first time to obtain insoluble beta-glucan and a first supernatant containing mannan; an anti-solvent is added into the supernate for precipitation, and mannan is obtained through secondary solid-liquid separation; and finally, treating the separated liquid to recover the eutectic solvent. The two polysaccharides are cooperatively extracted, the solvent is recycled, comprehensive utilization of resources is achieved, and the process is environmentally friendly, low in cost and high in efficiency.

Description

Technical Field

[0001] This invention relates to the field of biomass resource utilization, and in particular to a method for green extraction of polysaccharides from waste brewer's yeast cell walls using a eutectic solvent. Background Technology

[0002] Yeast β-glucan is a functional polysaccharide with important biological activities and broad application prospects in food, medicine, and chemical industries. The brewing industry generates a large amount of waste brewer's yeast as a byproduct annually. Its cell wall is mainly composed of an inner β-glucan layer and an outer mannan layer. Currently, the main method of utilizing waste brewer's yeast is as low-value-added feed, resulting in a significant waste of resources. Existing methods for extracting yeast β-glucan, such as alkaline and enzymatic methods, generally suffer from complex processes, low yields and purity, and high production costs. Furthermore, the use of strong acids and alkalis in the production process can easily cause environmental pollution.

[0003] In recent years, eutectic solvents, as a novel type of green solvent, have begun to be applied in the field of biomass treatment due to their advantages such as biodegradability, low toxicity, and simple preparation. Existing technologies have attempted to use eutectic solvents to treat yeast to extract insoluble β-glucan, which to some extent addresses the environmental issues of traditional methods.

[0004] However, this existing technology only focuses on the extraction of insoluble β-glucan, discarding the resulting supernatant directly. In fact, this supernatant contains another important polysaccharide component—mannan—which is not being effectively utilized. Furthermore, the eutectic solvent itself has a certain cost; if it cannot be effectively recovered and recycled in the process, it will significantly limit the economic feasibility of this technology. Therefore, the existing technology still suffers from insufficient resource utilization and high overall costs. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide a method for green extraction of polysaccharides from waste brewer's yeast cell walls using a eutectic solvent, in order to solve the problems of environmental pollution, high cost, and failure to comprehensively utilize the main polysaccharide components in yeast cell walls, namely β-glucan and mannan, in existing methods for extracting polysaccharides from waste brewer's yeast.

[0006] To achieve the above objectives, the present invention provides a method for green extraction of polysaccharides from the cell walls of waste brewer's yeast using a eutectic solvent, comprising the following steps: Step 1: Pre-treat the yeast raw material by mixing it with a eutectic solvent to selectively dissolve the mannan in the yeast raw material, thereby obtaining a mixture containing insoluble yeast β-glucan; Step 2: Perform a first solid-liquid separation on the mixture to obtain crude yeast β-glucan as the first insoluble substance, and a first supernatant containing dissolved mannan and the eutectic solvent; Step 3: Add an antisolvent to the first supernatant to precipitate the mannan, and perform a second solid-liquid separation to obtain a second precipitate containing the mannan and a second supernatant; Step 4: Process the second supernatant to recover the eutectic solvent.

[0007] Optionally, the eutectic solvent consists of a hydrogen bond acceptor and a hydrogen bond donor, wherein the hydrogen bond acceptor may be choline chloride, and the hydrogen bond donor may be lactic acid, formic acid, oxalic acid, or urea.

[0008] Optionally, the pretreatment conditions include: mixing the yeast raw material with the eutectic solvent at a mass ratio of 1:10 and reacting at a temperature of 70°C to 80°C for 60 minutes.

[0009] Optionally, after the first solid-liquid separation, the method further includes an enzymatic purification step of the crude yeast β-glucan.

[0010] Optionally, the enzymatic purification uses papain and nuclease to remove protein and nucleic acid impurities.

[0011] Optionally, the antisolvent is deionized water.

[0012] Optionally, the volume ratio of the antisolvent to the first supernatant is 3:2.

[0013] Optionally, at least one of the first solid-liquid separation and the second solid-liquid separation is carried out by centrifugation, filtration or pressure filtration.

[0014] Optionally, the step of recovering the eutectic solvent includes removing the water content of the antisolvent from the second supernatant by means of heating evaporation or vacuum distillation.

[0015] Optionally, when vacuum distillation is used, the steps include: The antisolvent is first recovered by distillation at a first temperature. The eutectic solvent is then recovered by distillation at a second temperature, which is higher than the first temperature.

[0016] Compared with the prior art, the present invention has the following beneficial effects: (1) High resource utilization rate. Through a continuous process, this invention not only extracts high-value yeast β-glucan, but also recovers mannan dissolved in eutectic solvent, and achieves synergistic extraction of the two major polysaccharide components of waste yeast cell wall, which significantly improves the comprehensive utilization value of raw materials.

[0017] (2) Green and environmentally friendly, low cost. This invention uses biodegradable and non-toxic eutectic solvents to replace strong alkalis and high-concentration organic solvents in traditional processes, reducing environmental pollution at the source. More importantly, by effectively recovering and recycling the eutectic solvent, the cost of solvents is greatly reduced and the economic feasibility of the method is improved.

[0018] (3) The process is highly efficient. Compared with traditional methods, this invention achieves higher crude extract content while ensuring β-glucan yield, and also obtains mannan product, resulting in higher overall process efficiency and output value. Attached Figure Description

[0019] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0020] Figure 1 This is a flowchart of a method for green extraction of polysaccharides from waste brewer's yeast cell walls using eutectic solvents; Figure 2 These are ion chromatograms for the determination of monosaccharide composition of crude extracts by alkaline method, ultrasonic method, and DES method (Note: Figure A shows the ion chromatogram of the mixed standard sample; Figure B shows the ion chromatogram of the crude extract by alkaline method; Figure C shows the ion chromatogram of the crude extract by ultrasonic-enzymatic method; Figure D shows the ion chromatogram of the crude extract by DES method). Detailed Implementation

[0021] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be described in detail below. Obviously, the described embodiments are merely some embodiments of this invention, and not all embodiments. Based on the embodiments of this invention, all other implementation methods obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0022] Based on the deficiencies described in the background art, the method for green extraction of polysaccharides from waste brewer's yeast cell walls using a eutectic solvent according to the present invention will be described in detail below.

[0023] This invention provides a method for the green extraction of polysaccharides from the cell walls of waste brewer's yeast using a eutectic solvent. (Refer to...) Figure 1 This diagram macroscopically illustrates the complete process path employed in this invention in the form of a flowchart. It aims to elaborate on the basic implementation scheme of this method, the core of which lies in achieving efficient separation and extraction of two main polysaccharide components from the cell walls of waste brewer's yeast through a continuous and integrated process, while recovering key solvents, thereby achieving the goals of comprehensive resource utilization, environmental protection, and cost control.

[0024] The initial stage of this method involves preparing a key medium for selective dissolution—a eutectic solvent. In this invention, choline chloride is selected as the hydrogen bond acceptor, and lactic acid as the hydrogen bond donor.

[0025] It is understandable that choline chloride is an inexpensive, non-toxic, and biodegradable quaternary ammonium salt, while lactic acid is an organic acid that naturally exists in organisms. The eutectic solvent formed by the combination of the two has excellent biocompatibility and environmental friendliness.

[0026] Specifically, the preparation process is as follows: Accurately weigh chemically pure choline chloride and lactic acid, ensuring a molar ratio of 1:2. Place the weighed solid powders in a clean, dry beaker and stir initially with a glass rod. Then, place the beaker in an 80°C constant-temperature water bath and turn on the magnetic stirring function, continuously stirring at a medium speed. Under the combined effect of this temperature and mechanical stirring, the original hydrogen bonds between choline chloride and lactic acid molecules are weakened, subsequently forming a new, more stable hydrogen bond network, and the mixture gradually changes from a solid to a liquid state. Continue stirring for about 30 to 40 minutes until the mixture is completely melted, forming a homogeneous, clear, transparent viscous liquid without any solid particles. At this point, the eutectic solvent is ready. Store at room temperature for later use.

[0027] It should be noted that the solvent is formed by strong hydrogen bonds between the hydrogen bond acceptor (chloride ions of choline chloride) and the hydrogen bond donor (hydroxyl and carboxyl groups of lactic acid), which makes the melting point of the mixture much lower than that of any single component.

[0028] Next, we will proceed with the formal extraction process of the method.

[0029] Reference Figure 1 In step S101, the yeast raw material is first provided.

[0030] The yeast raw material used in this invention is waste brewer's yeast sludge, a byproduct of brewery production. To facilitate subsequent processing and accurate measurement, the yeast sludge can be pre-treated, for example, by drying it in a 60°C oven with forced air or by freeze-drying technology, to remove most of the moisture, thereby obtaining dry yeast powder.

[0031] Subsequently, in step S102, the dry yeast powder is mixed with a eutectic solvent for pretreatment.

[0032] The prepared dry yeast powder was mixed with a choline chloride-lactic acid eutectic solvent kept at 80°C, with a mass ratio of 1:10 (1 gram of dry yeast powder to 10 grams of eutectic solvent) to ensure adequate solvation. The mixture was placed in a constant-temperature reactor at 80°C and mechanically stirred continuously at approximately 200 rpm for 60 minutes.

[0033] This step is crucial for achieving selective separation. The underlying principle is that the yeast cell wall is a complex network structure composed of β-glucan, mannan, proteins, and a small amount of chitin. At 80°C, the choline chloride-lactic acid eutectic solvent, with its moderate viscosity and strong hydrogen bonding ability, can effectively penetrate and disrupt the integrity of the yeast cell wall.

[0034] More importantly, this solvent exhibits significant differences in solubility for different polysaccharides: it can form effective hydrogen bonds with mannan molecules, which have a relatively loose structure and more branched chains, thus selectively dissolving them from the cell wall skeleton into the solvent; however, its solubility for β-glucan long chains, which have a more regular structure, higher crystallinity, and stronger intermolecular hydrogen bonds, is very limited. Therefore, after 60 minutes of thorough stirring, most of the mannan in the mixture is transferred to the liquid phase, while β-glucan remains essentially insoluble, dispersed in the solvent in solid form.

[0035] After the pretreatment is completed, the first solid-liquid separation is performed in step S103.

[0036] The pretreated viscous mixture is removed from the reactor and immediately separated to prevent changes in the product properties. In this invention, high-speed centrifugation is used.

[0037] Specifically, the mixture was dispensed into centrifuge tubes and centrifuged for 10 minutes at 4°C and 5000 rpm using a high-speed refrigerated centrifuge. After centrifugation, the mixture was clearly observed to be divided into two parts: the lower layer was a compacted solid precipitate, i.e., the first insoluble matter; the upper layer was a clear, slightly brownish viscous liquid, i.e., the first supernatant.

[0038] The two phases obtained from this separation will be processed through different pathways. The solid fraction, the first insoluble substance, mainly consists of crudely extracted yeast β-glucan, corresponding to... Figure 1 Step S104 in the process.

[0039] As an optional implementation, in order to obtain a product with higher purity, the present invention also includes a purification step.

[0040] First, the collected first insoluble matter is washed multiple times with deionized water to remove the eutectic solvent adhering to and entrained on the surface.

[0041] Subsequently, it was resuspended in a certain volume of distilled water to form a suspension with a solid-liquid ratio of 1:5 (mass / volume), and the pH of the suspension was adjusted to 6.0 using dilute acid or dilute alkali to provide the optimal environment for subsequent enzyme reactions.

[0042] Next, enzymatic purification is performed in step S105. A complex enzyme preparation consisting of papain and nuclease is added to the suspension to achieve a final mass concentration of 0.50% (w / v) of the enzyme preparation in the suspension.

[0043] Papain, a broad-spectrum proteolytic enzyme, effectively degrades residual protein impurities in the crude extract, while nucleases degrade residual nucleic acids. The mixture was placed in a constant-temperature shaker at 50°C for 3 hours for hydrolysis, with slow shaking to promote the reaction.

[0044] After the enzymatic hydrolysis reaction is complete, the solids are collected again by centrifugation or filtration, and washed with plenty of distilled water to thoroughly remove degraded small molecule impurities and enzyme preparations. Finally, the washed solids are freeze-dried to obtain a high-purity white powdery yeast β-glucan product in step S106.

[0045] Accordingly, the other part of the product obtained after the first solid-liquid separation (step S103) – the first supernatant – is processed to extract mannan. This supernatant is a mixed solution containing dissolved mannan, a eutectic solvent, and small amounts of water and other soluble impurities of yeast.

[0046] In step S107, an antisolvent is added to the supernatant. In this invention, deionized water is selected as the antisolvent.

[0047] It is understandable that an antisolvent is a solvent that is miscible with the original solvent, but in which the target solute has very low solubility. The first supernatant and deionized water are mixed at a volume ratio of 2:3, that is, 3 volumes of deionized water are slowly added to every 2 volumes of the first supernatant, and the mixture is stirred continuously during the addition process to ensure uniform mixing.

[0048] The principle is that the addition of deionized water greatly changes the polarity and solvation ability of the entire solvent system, destroys the original "eutectic solvent-mannan" solvation structure, and causes the solubility of mannan to drop sharply, thus precipitating out of the solution in the form of a solid, usually white flocculent precipitate.

[0049] After complete precipitation, a second solid-liquid separation is performed in step S108. Centrifugation is also used, with the mixture containing the mannan precipitate centrifuged at 5000 rpm for 10 minutes. After centrifugation, crude mannan as the second precipitate (corresponding to step S109) and a liquid as the second supernatant are obtained. The collected mannan precipitate is washed and dried to obtain the mannan product.

[0050] As a closed loop of this process, step S110 involves processing the second supernatant obtained from the second solid-liquid separation to achieve the recovery of the eutectic solvent.

[0051] The second supernatant is a mixture of a eutectic solvent and a small amount of water. Since the boiling point of water (100°C) is much lower than the decomposition temperature of the eutectic solvent, it can be separated by evaporation.

[0052] In this invention, the second supernatant is placed in an open, heat-resistant container and heated in an oven at 105°C for evaporation. At this temperature, water evaporates and escapes, while the thermally stable eutectic solvent remains. Drying continues until the liquid volume in the container no longer changes; the resulting viscous liquid is the recovered eutectic solvent. Testing shows that the recovered solvent's physicochemical properties are essentially consistent with those of the freshly prepared solvent, allowing it to be directly used for the next batch of yeast pretreatment. This significantly reduces solvent costs and improves the overall economic efficiency and environmental friendliness of the process.

[0053] Experimental Example 1: DES Method (1) DES preparation: Mix choline chloride and lactic acid in a 1:2 molar ratio, pour into a round-bottom flask, stir in an 80°C constant temperature water bath until a homogeneous, stable and transparent solution is formed in the round-bottom flask, and store at room temperature.

[0054] (2) Yeast β-glucan extraction: Yeast cells and DES were mixed at a ratio of 1:10 (w:w), stirred at 80℃ for 1 h, and centrifuged (5000 rpm, 10 min) to obtain insoluble matter P1 and supernatant S1. Precipitate P1 was added to distilled water at a ratio of 1:5, and the pH was adjusted to 6.0. Papain and nuclease were added at a mass ratio of 0.50% (w / v) for hydrolysis for 3 h. After centrifugation (5000 rpm, 10 min), distilled water was added at a ratio of 1:10 (v / v) for resuspending. After two centrifugations (5000 rpm, 10 min), the mixture was freeze-dried to obtain crude yeast β-glucan extracted by the DES method (DES-extracted yeast β-glucan, DEB), and its weight was recorded as G. D .

[0055] (3) DES recovery: The antisolvent and supernatant S1 were mixed evenly at a ratio of 2:3 (v:v), and centrifuged (5000 rpm, 10 min) to obtain supernatant S2 and precipitate P2 (mannan). The separated supernatant S2 was dried overnight in an oven at 105°C until the volume remained constant, which produced the recovered eutectic solvent S3.

[0056] (4) Method for determining yeast β-glucan The monosaccharide composition was determined using high-performance anion exchange chromatography (HPAEC) combined with an electrochemical detector (ED). The simplified procedure is as follows: 5 mg of sample was added to 3 M trifluoroacetic acid (TFA), and the mixture was heated at 120 °C for 3 hours. The hydrolysis product was dissolved in 5 mL of ultrapure water by vortexing, and 50 μL was diluted with 950 μL of ultrapure water. 1 mL of the solution was centrifuged at 12000 rpm for 5 minutes, and the supernatant was filtered through a 0.22 μm microporous membrane. Detection was performed using an HPAEC-ED column equipped with a Dionex Carbopac PA20 column (150 × 3.0 mm). The mobile phase consisted of A (water), B (15 mM sodium hydroxide), and C (15 mM sodium hydroxide + 100 mM sodium acetate). The flow rate was 0.3 mL / min, the injection volume was 25 μL, and the column temperature was 30 °C.

[0057] Elution gradient settings: A / B / C ratio 98.8:1.2:0 (volume ratio) at 0 minutes; A / B / C ratio 98.8:1.2:0 (volume ratio) at 18 minutes; A / B / C ratio 50:50:0 (volume ratio) at 20 minutes; A / B / C ratio 50:50:0 (volume ratio) at 30 minutes; A / B / C ratio 0:0:100 (volume ratio) at 30.1 minutes; A / B / C ratio 0:0:100 (volume ratio) at 46 minutes; A / B / C ratio 0:0:100 (volume ratio) at 46.1 minutes; A / B / C ratio 0:100:0 (volume ratio) at 50 minutes; A / B / C ratio 98.8:1.2:0 (volume ratio) at 50.1 minutes; A / B / C ratio 98.8:1.2:0 (volume ratio) at 80 minutes.

[0058] Calculation of crude yeast β-glucan content: Yeast β-glucan content (%) = weight of crude extract after freeze-drying (g) / weight of dry yeast before extraction (g) Yeast β-glucan yield (%) = Crude extract content (%) × Crude extract yeast β-glucan content (µg / mg) / 1000 (5) Statistical analysis Each method was repeated three times. SPSS software was used to test the normality of the data. The data were found to be normally distributed. Further analysis using one-way ANOVA and Duncan's post-hoc multiple comparisons was performed. Data are presented as means, and all statistical results were considered statistically significant at p < 0.05.

[0059] Comparative Example 1: Alkali Method The alkaline method is a common method for extracting polysaccharides from yeast cell walls. The yeast sludge in the experimental materials was provided by China Resources Snow Breweries (China) Co., Ltd., and the papain and nuclease mixture was purchased from Pangbo Biotechnology Co., Ltd.

[0060] Add 10 mL of 1 M NaOH to 2.5 g of dry yeast and heat to 80°C for 1 hour. Centrifuge the suspension at 5000 rpm for 10 minutes, discard the supernatant, and repeat the above steps once.

[0061] After alkali treatment, the solid was resuspended, and the pH was adjusted to 4.5 with HCl. The acidified suspension was heated to 75°C and held for 2 hours, then centrifuged again (5000 rpm, 10 minutes). The insoluble residue obtained from centrifuging the suspension (5000 rpm, 10 minutes) was washed twice with distilled water. The precipitate after centrifugation and washing was added to distilled water at a ratio of 1:5, the pH was adjusted to 6.0, and the mixture was preheated to 55.0°C in a constant temperature shaking water bath. Papain and nuclease were added at a mass ratio of 0.50% (w / v) for hydrolysis for 3 hours, followed immediately by boiling in a water bath for 10 minutes to inactivate the enzymes.

[0062] The suspension was centrifuged (5000 rpm, 10 min) to separate insoluble residues, washed twice with distilled water, and freeze-dried for 48 h to obtain alkali-extracted yeast β-glucan (AEB), weighed and recorded as G. A .

[0063] Comparative Example 2: Ultrasonic Method Weigh 2.5 g of dry yeast, add 5 mL of distilled water to prepare a suspension, and adjust the pH to 7.

[0064] The yeast suspension was sonicated for 30 min at 40% power (260W) using an ultrasonic cell disruptor. The insoluble residue obtained by centrifugation (5000 rpm, 10 min) was washed twice with distilled water. The precipitate after centrifugation and washing was added to distilled water at a ratio of 1:5, the pH was adjusted to 6.0, and the mixture was preheated to 55.0°C in a constant temperature shaking water bath. Papain and nuclease were added at a mass ratio of 0.50% (w / v) for hydrolysis for 3 h, followed immediately by boiling in a water bath for 10 min to inactivate the enzymes.

[0065] The suspension was centrifuged (5000 rpm, 10 min) to separate insoluble residues, washed twice with distilled water, and freeze-dried for 48 h to obtain ultrasonic-enzyme extracted yeast β-glucan (UEEB), which was weighed and recorded as G. U .

[0066] Experimental results: 1. Comparison of crude yeast β-glucan content extracted by three pretreatment methods: Table 1 shows the crude yeast β-glucan content extracted by the three pretreatment methods. The crude yeast β-glucan content of DES pretreatment was significantly higher than that of the alkaline method and the ultrasonic enzymatic method.

[0067] Table 1. Content of crude yeast β-glucan extracted by alkaline method, ultrasonic method and DES method Note: a, b, c Significant differences exist between different letters in the same row's shoulder insignia (n = 3). 2. Comparison of crude yields of yeast β-glucan extracted by three pretreatment methods: Figure 2 The table shows the ion chromatograms for the determination of monosaccharide composition of the crude extracts by the three methods. Table 2 shows the results of monosaccharide composition of the three crude extracts. Table 3 shows the results of yeast β-glucan yield. The results show that there is no significant difference in the yield of yeast β-glucan by the alkali method, ultrasonic method, and DES method. Table 2. Monosaccharide composition results of crude extracts obtained by alkaline method, ultrasonic method, and DES method. Table 3. Yields of yeast β-glucan obtained by alkaline method, ultrasonic method, and DES method The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A method for green extraction of polysaccharides from the cell walls of waste brewer's yeast using a eutectic solvent, characterized in that, Includes the following steps: Step 1: Pre-treat the yeast raw material by mixing it with a eutectic solvent to selectively dissolve the mannan in the yeast raw material, thereby obtaining a mixture containing insoluble yeast β-glucan; Step 2: Perform a first solid-liquid separation on the mixture to obtain crude yeast β-glucan as the first insoluble substance, and a first supernatant containing dissolved mannan and the eutectic solvent; Step 3: Add an antisolvent to the first supernatant to precipitate the mannan, and perform a second solid-liquid separation to obtain a second precipitate containing the mannan and a second supernatant; Step 4: Process the second supernatant to recover the eutectic solvent.

2. The method according to claim 1, characterized in that, The eutectic solvent consists of a hydrogen bond acceptor and a hydrogen bond donor, wherein the hydrogen bond acceptor may be choline chloride, and the hydrogen bond donor may be lactic acid, formic acid, oxalic acid, or urea.

3. The method according to claim 1 or 2, characterized in that, The pretreatment conditions include: mixing the yeast raw material with the eutectic solvent at a mass ratio of 1:10 and reacting at a temperature of 70°C to 80°C for 60 minutes.

4. The method according to claim 1, characterized in that, Following the first solid-liquid separation, the process further includes an enzymatic purification step of the crude yeast β-glucan.

5. The method according to claim 4, characterized in that, The enzymatic purification process uses papain and nuclease to remove protein and nucleic acid impurities.

6. The method according to claim 1, characterized in that, The antisolvent is deionized water.

7. The method according to claim 6, characterized in that, The volume ratio of the antisolvent to the first supernatant is 3:

2.

8. The method according to claim 1, characterized in that, At least one of the first solid-liquid separation and the second solid-liquid separation is carried out by centrifugation, filtration or pressure filtration.

9. The method according to claim 1, characterized in that, The step of recovering the eutectic solvent includes removing the water content of the antisolvent from the second supernatant by means of heating evaporation or vacuum distillation.

10. The method according to claim 9, characterized in that, When vacuum distillation is used, the steps include: The antisolvent is first recovered by distillation at a first temperature. The eutectic solvent is then recovered by distillation at a second temperature, which is higher than the first temperature.

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