Method for purifying isoleucine fermentation liquor

Abstract

The invention provides a method for purifying isoleucine fermentation liquor, which adopts a mode of adding a crystallization assistant to improve the evaporative crystallization yield, namely adding ammonium magnesium phosphate into a crystallization base material or adding ammonium magnesium phosphate in the crystallization process, thereby being beneficial to improving the crystallization effect and improving the primary yield of centrifugal separation after crude product crystallization and fine product crystallization. The stacking density of the product is greatly improved, and the proportion of the obtained food-grade isoleucine product is remarkably improved.

Description

Technical Field

[0001] The present invention relates to the technical field of amino acid fermentation and fermentation broth purification, and more specifically, to a method for purifying isoleucine fermentation broth. Background Art

[0002] L-isoleucine is a neutral amino acid and one of the eight essential amino acids in the human body. Together with L-valine and L-leucine, they are collectively called branched-chain amino acids. Due to its special structure and function, it plays a particularly important role in human life metabolism. L-isoleucine is an essential amino acid that cannot be synthesized by mammalian tissues. The main function of this amino acid is to integrate into proteins and enzymes, which helps to maintain the tertiary structure of these macromolecules. It has the effect of promoting insulin secretion. Lack of L-isoleucine will cause loss of appetite, physical decline, anemia, skeletal muscle disorders and other dysfunctions. L-isoleucine has been widely used in the pharmaceutical and food fields and has been increasingly used in the sports food industry in recent years.

[0003] At present, fermentation is mainly used in industrial production to produce L-isoleucine. The known methods for separating and extracting L-isoleucine mainly include ion exchange method, chromatography method, membrane method, etc. Among them, both the ion exchange process and the chromatography process require a large amount of water, resulting in a large amount of sewage. When producing 1 ton of finished L-isoleucine, about 200 cubic meters of sewage is discharged, with a relatively high cost and a large waste of water resources. For example, CN200510123082.X discloses a clean production process for extracting L-isoleucine from fermentation broth by ion exchange method. Although it discloses the further recycling of ion exchange wastewater, it cannot change the basic problem of large water consumption in ion exchange. CN201010557488.X discloses a method for extracting L-isoleucine, which also uses the process of ion exchange and fractional purification. Although the quality has been improved, the problem of large water consumption in the ion exchange process has still not been solved.

[0004] The equipment investment cost of the chromatography process and the membrane method is high. For example, CN200910025832.8 discloses a method for extracting and separating L-isoleucine by combining membrane separation and industrial chromatography separation, with a recovery rate of about 93%. However, continuous chromatography equipment and organic membrane separation equipment with huge equipment investment are used, and there is no significant reduction in water consumption. CN201610655330.3 discloses a method for extracting L-isoleucine by using an all-membrane process. This process mainly uses flocculants and organic membranes to remove impurities. Compared with the ion exchange process, it can save a large amount of acid and alkali and reduce partial sewage discharge. However, the use of self-made flocculants will introduce other impurities. At the same time, the membrane core life of the ultrafiltration membrane is relatively short, and it needs to be cleaned with a cleaning agent after each batch, resulting in high equipment investment and operating costs. Summary of the Invention

[0005] The object of the present invention is to provide an efficient method for purifying isoleucine fermentation broth.

[0006] To achieve the object of the present invention, in a first aspect, the present invention provides the use of magnesium ammonium phosphate in the refining process of amino acid fermentation production, and the use includes: after the amino acid fermentation broth is successively subjected to membrane filtration and decolorization treatment, magnesium ammonium phosphate is added to the obtained crystallization bottom material, or magnesium ammonium phosphate is added during the crystallization process.

[0007] The amino acids include isoleucine and valine, preferably L-isoleucine.

[0008] In a second aspect, the present invention provides a method for purifying isoleucine fermentation broth. First, the pH value of the isoleucine fermentation broth is adjusted to 4-6, most of the bacteria, miscellaneous proteins and colloids are removed by ceramic membrane filtration, and the first activated carbon decolorization is carried out at pH 4-6. Then, the pH is adjusted to 5-7 for the first evaporation crystallization. Seeds are added to the crystallization evaporator, and magnesium ammonium phosphate is added to the crystallization bottom material, or magnesium ammonium phosphate is added during the crystallization process. When the isoleucine content in the crystallization evaporator is 250-260 g / L, the pot is discharged. The crystallization liquid is centrifuged to obtain crude isoleucine and crude mother liquor respectively; after the crude isoleucine is redissolved with water, the second activated carbon decolorization is carried out, and then the pH is adjusted to 5-7 for the second evaporation crystallization. Seeds are added to the crystallization evaporator, and magnesium ammonium phosphate is added to the crystallization bottom material. When the isoleucine content in the crystallization evaporator is 250-260 g / L, the pot is discharged. The crystallization liquid is centrifuged to obtain refined isoleucine, which is dried to obtain the isoleucine finished product.

[0009] Further, when carrying out the first evaporation crystallization, the addition amount of the seeds is 1-2% (preferably 2%) of the mass of the material in the crystallization evaporator.

[0010] Further, when carrying out the first evaporation crystallization, the addition amount of magnesium ammonium phosphate is 0.1%-0.2% of the mass of the material in the crystallization evaporator.

[0011] Further, when carrying out the second evaporation crystallization, the addition amount of the seeds is 1-2% (preferably 1%) of the mass of the material in the crystallization evaporator.

[0012] Further, when carrying out the second evaporation crystallization, the addition amount of magnesium ammonium phosphate is 0.1%-0.2% of the mass of the material in the crystallization evaporator.

[0013] Specifically, the method includes the following steps:

[0014] (1) Separation of denatured bacteria in the isoleucine fermentation broth to obtain a supernatant: Take 31 L of isoleucine fermentation broth with an isoleucine content of 55.2 g / L, adjust the pH to 4.5 (using concentrated sulfuric acid) to denature and precipitate the protein, and then separate the bacteria through a ceramic membrane to obtain a retentate containing bacteria and a supernatant without bacteria;

[0015] (2) Primary decolorization of the clear liquid: Perform activated carbon decolorization on the clear liquid obtained in step (1), adding 0.6 kg of activated carbon to every 100 L of the clear liquid; stir and decolorize at 65 - 70 °C for 30 - 60 min (preferably stir and decolorize at 65 °C for 30 min), and the light transmittance of the decolorized liquid is 81.2%;

[0016] (3) Primary evaporation crystallization after decolorization of the clear liquid: Adjust the pH value of the decolorized clear liquid obtained in step (2) to 6.0 with sodium hydroxide, add it to a crystallization evaporator, and add 0.1% of magnesium ammonium phosphate based on the mass of the material as a crystallization aid, then perform evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the content of isoleucine evaporates to 60 - 65 g / L, add food-grade isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 2% of the mass of the material in the crystallization evaporator; control the vacuum degree of the evaporator at 0.090 - 0.095 MPa, the stirring speed before adding the crystal seeds is 20 - 50 r / min (preferably 50 r / min), and the stirring speed after adding the crystal seeds is 80 - 100 r / min (preferably 100 r / min); when the content of isoleucine in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water with a volume of 5% of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0017] (4) Centrifugally separate the crystallization liquid to obtain crude isoleucine: After discharging the crystallization liquid from the tank, separate it directly without cooling, wash the mother liquor on the surface of the crystals with water, continue centrifuging after adding water, and separate to obtain crude isoleucine and crude mother liquor;

[0018] (5) Redissolve the crude isoleucine: Dissolve the crude isoleucine separated in step (4) with water, control the water temperature at 65 - 66 °C, and prepare a redissolved solution with a content of 50 g / L;

[0019] (6) Secondary decolorization of the crude product dissolved solution: Adjust the pH of the redissolved solution obtained in step (5) to 5.0 with concentrated sulfuric acid, perform secondary decolorization, adding 0.3 kg of activated carbon to every 100 L of the dissolved solution; stir and decolorize at 65 - 70 °C for 30 - 60 min (preferably stir and decolorize at 65 °C for 30 min), and the light transmittance of the decolorized liquid is 93.2%;

[0020] (7) Secondary evaporation crystallization of the decolorized clear liquid: Adjust the pH of the decolorized clear liquid obtained in step (6) to 6.0 with sodium hydroxide, add it to the crystallization evaporator, and add ammonium magnesium phosphate accounting for 0.1% of the material mass as a crystallization aid, and then carry out evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the isoleucine content evaporates to 60 - 65 g / L, add food-grade isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 1% of the material mass in the crystallization evaporator; the vacuum degree of the evaporator is controlled at 0.090 - 0.095 MPa, the stirring speed before adding crystal seeds is 20 - 50 revolutions (preferably 50 revolutions / min) / min, and the stirring speed after adding crystal seeds is 80 - 100 revolutions / min (preferably 100 revolutions / min); when the isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water accounting for 5% of the volume of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0021] (8) Centrifugally separate the crystallization liquid to obtain high-quality isoleucine: After discharging the crystallization liquid in step (7) into the tank, directly separate it without cooling, wash the mother liquor on the crystal surface with water, centrifuge and spin dry, then wash the crystals with 30% alcohol, continue to centrifuge after washing with alcohol, and separate to obtain high-quality isoleucine, and obtain the finished product of food-grade isoleucine after drying.

[0022] Furthermore, step (4) also includes: Mix the separated crude mother liquor with the retentate containing bacteria obtained in step (1), concentrate it to 50 - 55% (preferably 50%) dry matter, and spray granulate to obtain feed-grade isoleucine.

[0023] In the present invention, the strain used for isoleucine fermentation can be Corynebacterium glutamicum.

[0024] The isoleucine fermentation method refers to Comparative Experimental Example 1 of Chinese Invention Patent CN202110539756.3.

[0025] By means of the above technical solutions, the present invention has at least the following advantages and beneficial effects:

[0026] The invention removes protein by adjusting the pH value of fermentation liquid to 4-6, performs activated carbon decolorization when the pH value is 4-6, improves the decolorization effect, then adjusts the pH value to 5-7 for evaporation and crystallization, adds crystal seeds into a crystallization evaporator, adds magnesium ammonium phosphate into a crystallization base material, or adds magnesium ammonium phosphate into a crystallization process, forms an environment suitable for crystal growth by reasonably setting the adding time and adding amount of crystal seeds and crystallization aids, makes the L-isoleucine crystal form uniformly granular, greatly improves the separation effect of mother liquor and crystal after crystallization, realizes that the yield of food-grade L-isoleucine reaches more than 82% without adopting ion exchange process and chromatography process, and improves the yield of high value-added products; the refined mother liquor is recycled for primary decolorization, the primary mother liquor is mixed with bacterial protein to prepare low-content feed, and the total extraction yield reaches more than 96%.

[0027] In the extraction process of the present invention, no ion exchange process and chromatography process are adopted, and the main effluent water is evaporated condensed water, which enters into fermentation ingredients and ceramic membrane dialysis water, and no sewage is discharged; at the same time, the amount of primary water taken is greatly reduced; and the pH is adjusted to between 4 and 7, the amount of acid and alkali is small, and it is environmentally friendly; there is no other complex and high-cost equipment investment, the operating cost is low, and it is conducive to large-scale industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is the existing L-isoleucine production technology route.

[0029] Figure 2 This is the L-isoleucine production technology route of the present invention. DETAILED DESCRIPTION

[0030] The present invention aims to provide a simple and high-yield L-isoleucine extraction method. Different from the prior art, the present invention adopts a method of adding a crystallization aid to increase the primary yield of evaporation crystallization, thereby increasing the yield of L-isoleucine.

[0031] The present invention also provides a crystallization aid: ammonium magnesium phosphate. Adding ammonium magnesium phosphate to the crystallization base material or adding ammonium magnesium phosphate during the crystallization process helps to improve the crystallization effect, improve the primary yield of centrifugal separation after crude product crystallization and fine product crystallization, greatly improve the product bulk density, and significantly increase the proportion of the obtained food-grade L-isoleucine product.

[0032] The present invention adopts the following technical solution:

[0033] (1) The cells in the L-isoleucine fermentation broth are denatured and separated to obtain a clear liquid; (2) The clear liquid is decolorized once; (3) The decolorized clear liquid is evaporated and crystallized once; (4) The crystallization liquid is centrifuged to separate the crude L-isoleucine; (5) The crude L-isoleucine is redissolved; (6) The crude product solution is decolorized twice; (7) The decolorized clear liquid is evaporated and crystallized twice; (8) The crystallization liquid is centrifuged to separate the fine L-isoleucine; (9) The fine product is dried to obtain a food-grade L-isoleucine product.

[0034] The following examples are used to illustrate the present invention, but are not used to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0035] The strain used for fermenting and preparing isoleucine in the following examples is Corynebacterium glutamicum, see CN202010189011.4. The strain is provided by Langfang Meihua Biotechnology Development Co., Ltd.

[0036] Ammonium magnesium phosphate is purchased from Aiko Reagent.

[0037] Extraction method of L-isoleucine in Comparative Example 1

[0038] As Figure 1 shown, the extraction method of L-isoleucine includes the following steps: (1) The cells in the fermentation broth are denatured and separated to obtain a clear liquid; (2) The clear liquid is decolorized once; (3) The decolorized clear liquid is evaporated and crystallized once; (4) The crystallization liquid is centrifuged to separate the crude L-isoleucine; (5) The crude L-isoleucine is redissolved; (6) The crude product solution is decolorized twice; (7) The decolorized clear liquid is evaporated and crystallized twice; (8) The crystallization liquid is centrifuged to separate the fine L-isoleucine; (9) The fine product is dried to obtain a food-grade L-isoleucine product. Specifically as follows:

[0039] (1) The cells in the fermentation broth are denatured and separated to obtain a clear liquid: Take 30 L of L-isoleucine fermentation broth with an L-isoleucine content of 53.0 g / L, adjust the pH to 4.5 with concentrated sulfuric acid to denature and precipitate the protein, and then pass through a ceramic membrane to separate the cells, obtaining a retentate containing cells and a clear liquid without cells;

[0040] (2) The clear liquid is decolorized once: The clear liquid obtained in step (1) is decolorized with activated carbon. 0.6 kg of activated carbon is added to every 100 L of clear liquid; Stir and decolorize at 65 °C for 30 min, and the light transmittance of the decolorized liquid is 80.5%;

[0041] (3) Primary evaporation crystallization after defecation of the clear liquid: Adjust the pH value of the defecated clear liquid obtained in step (2) to 6.0 with sodium hydroxide and add it to a crystallization evaporator for evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the L-isoleucine content evaporates to 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 2.0% (weight 27.4 g) of the mass of the material in the crystallization evaporator; the evaporator controls the vacuum degree to be 0.090 - 0.095 Mpa, the stirring speed before adding crystal seeds is 50 rpm, and the stirring speed after adding crystal seeds is 100 rpm; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water with a volume of 5% of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0042] (4) Centrifugal separation of the crystallization liquid to obtain crude L-isoleucine: After discharging the crystallization liquid from the tank, directly separate it without cooling, add washing water with a volume ratio of 10%, wash the mother liquor on the crystal surface, continue centrifuging for 30 min after adding water, and separate to obtain crude L-isoleucine with a moisture content of 11.5%;

[0043] Mix the separated crude mother liquor with the bacterial body retention liquid obtained in step (1), concentrate to 50% dry matter, spray granulate to obtain 396.8 g of feed-grade L-isoleucine, and the L-isoleucine content reaches 60.5%;

[0044] (5) Redissolution of the crude L-isoleucine: Dissolve the crude L-isoleucine separated in step (4) with water, control the water temperature at 65 - 66 °C, and prepare a redissolved solution with a content of 50 g / L;

[0045] (6) Secondary defecation of the crude product dissolution liquid: Adjust the pH of the redissolved solution obtained in step (5) to 5.0 with concentrated sulfuric acid for secondary defecation, add 0.3 kg of activated carbon to every 100 L of the dissolution liquid; stir and defecate at 65 °C for 30 min, and the light transmittance of the defecated liquid is 92.5%;

[0046] (7) Secondary evaporation crystallization of the defecated clear liquid: Adjust the pH of the defecated clear liquid obtained in step (6) to 6.0 with sodium hydroxide and add it to a crystallization evaporator for evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the L-isoleucine content evaporates to 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 1.0% (weight 13.0 g) of the mass of the material in the crystallization evaporator; the evaporator controls the vacuum degree to be 0.090 - 0.095 MPa, the stirring speed before adding crystal seeds is 30 rpm, and the stirring speed after adding crystal seeds is 100 rpm; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water with a volume of 5% of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0047] (8) The crystal liquid is centrifuged to obtain high-quality L-isoleucine: After the crystal liquid in step (7) is discharged from the tank, it is separated directly without cooling, 5% by volume of washing water is added to wash the mother liquor on the crystal surface, and then it is centrifuged and dried. Then, 2% by volume of 30% alcohol is added to wash the crystals. After alcohol washing, it is centrifuged for another 30 min to obtain high-quality L-isoleucine, with a water content of 5.2%.

[0048] The refined mother liquor obtained by separation is recycled to step (2) of the next round for continuous treatment, which can improve the overall yield of L-isoleucine.

[0049] (9) The high-quality product is dried to obtain food-grade L-isoleucine product: The high-quality L-isoleucine obtained in step (8) is dried at 105 °C to obtain qualified food-grade L-isoleucine finished product.

[0050] The mass of the food-grade L-isoleucine finished product after drying is 1085.0 g, the bulk density of the finished product is 0.525 g / cm 3 , and after adding the recycled refined mother liquor, the total is 1274.8 g, the liquid phase content is 98.1%, meeting the USP standard. The food-grade yield is 80.2%, the feed-grade yield is 15.1%, and the total yield is 95.3%.

[0051] Purification method of L-isoleucine fermentation broth with crystal growth aid added to the bottom material in Example 1

[0052] As Figure 2 shown, the purification method of L-isoleucine fermentation broth includes the following steps: (1) The bacteria in the fermentation broth are denatured and separated to obtain a clear liquid; (2) The clear liquid is decolorized for the first time; (3) The clear liquid is evaporated and crystallized for the first time after decolorization; (4) The crystal liquid is centrifuged to obtain crude L-isoleucine; (5) The crude L-isoleucine is redissolved; (6) The dissolved solution of the crude product is decolorized for the second time; (7) The decolorized clear liquid is evaporated and crystallized for the second time; (8) The crystal liquid is centrifuged to obtain high-quality L-isoleucine; (9) The high-quality product is dried to obtain food-grade L-isoleucine product. Specifically as follows:

[0053] (1) The bacteria in the fermentation broth are denatured and separated to obtain a clear liquid: Take 31 L of L-isoleucine fermentation broth with an L-isoleucine content of 55.2 g / L, adjust the pH to 4.5 with concentrated sulfuric acid to denature and precipitate the protein, and then separate the bacteria through a ceramic membrane to obtain a retentate containing bacteria and a clear liquid without bacteria;

[0054] (2) The clear liquid is decolorized for the first time: The clear liquid obtained in step (1) is decolorized with activated carbon. 0.6 kg of activated carbon is added to every 100 L of clear liquid; Stir and decolorize at 65 °C for 30 min, and the light transmittance of the decolorized liquid is 81.2%;

[0055] (3) Primary evaporation crystallization after supernatant decolorization: Adjust the pH value of the decolorized supernatant obtained in step (2) to 6.0 with sodium hydroxide, add it to a crystallization evaporator, and add ammonium magnesium phosphate accounting for 0.1% of the material mass (weight: 1.5 g) as a crystallization aid, and then carry out evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the L-isoleucine content evaporates to 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 2% of the material mass in the crystallization evaporator (weight: 29.6 g); the vacuum degree of the evaporator is controlled at 0.090 - 0.095 MPa, the stirring speed before adding crystal seeds is 50 rpm, and the stirring speed after adding crystal seeds is 100 rpm; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water accounting for 5% of the volume of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0056] (4) Centrifugal separation of the crystallization liquid to obtain crude L-isoleucine: After the crystallization liquid is discharged from the tank, it is separated directly without cooling. Add washing water with a volume ratio of 10%, wash the mother liquor on the crystal surface, continue centrifuging for 30 min after adding water, and separate to obtain crude L-isoleucine with a water content of 11.5%;

[0057] Mix the separated crude mother liquor with the thallus retention liquid obtained in step (1), concentrate to 50% dry matter, spray granulate to obtain 427.1 g of feed-grade L-isoleucine, and the L-isoleucine content reaches 60.1%;

[0058] (5) Redissolution of crude L-isoleucine: Dissolve the crude L-isoleucine separated in step (4) in water, control the water temperature at 65 - 66 °C, and prepare a redissolved solution with a content of 50 g / L;

[0059] (6) Secondary decolorization of the crude product dissolution solution: Adjust the pH of the redissolved solution obtained in step (5) to 5.0 with concentrated sulfuric acid for secondary decolorization, and add 0.3 kg of activated carbon to every 100 L of the dissolution solution; stir and decolorize at 65 °C for 30 min, and the light transmittance of the decolorized solution is 93.2%;

[0060] (7) Secondary evaporation crystallization of the decolorized clear liquid: Adjust the pH of the decolorized clear liquid obtained in step (6) to 6.0 with sodium hydroxide, add it to the crystallization evaporator, and add ammonium magnesium phosphate accounting for 0.1% of the material mass (weighing 1.3 g) as a crystallization aid, and then carry out evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the L-isoleucine content evaporates to 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of the crystal seeds is 1% of the material mass in the crystallization evaporator (weighing 13.2 g); the vacuum degree of the evaporator is controlled at 0.090 - 0.095 Mpa, the stirring speed before adding crystal seeds is 50 r / min, and the stirring speed after adding crystal seeds is 100 r / min; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water accounting for 5% of the volume of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank;

[0061] (8) Centrifugal separation of the crystallization liquid to obtain high-quality L-isoleucine: After discharging the crystallization liquid in step (7) into the tank, separate it directly without cooling, add washing water with a volume ratio of 5%, wash the mother liquor on the crystal surface, centrifuge and spin dry, then add 30% alcohol with a volume ratio of 2% to wash the crystals, continue to centrifuge for 30 min after alcohol washing, and separate to obtain high-quality L-isoleucine, with the water content being 4.9%;

[0062] The separated high-quality mother liquor is recycled to step (2) of the next round for continuous treatment, which can improve the overall yield of L-isoleucine.

[0063] (10) Drying the high-quality product to obtain food-grade L-isoleucine product: Dry the high-quality L-isoleucine obtained in step (8) at 105 °C to obtain qualified food-grade L-isoleucine finished product.

[0064] Experimental results: The mass of the food-grade L-isoleucine finished product after drying is 1199.1 g, the bulk density of the finished product is 0.545 g / cm 3 , and after adding the recycled high-quality mother liquor, the total is 1389.0 g, the liquid phase content is 98.7%, meeting the USP standard. The food-grade yield is 81.2%, the feed-grade yield is 15.0%, and the total yield is 96.2%.

[0065] Purification method of L-isoleucine fermentation broth with crystallization aid added to the bottom material in Example 2

[0066] Refer to Example 1, in step (3), adjust the amount of ammonium magnesium phosphate added as the crystallization aid to 0.2% of the material mass (weighing 3.0 g), and in step (7), adjust the amount of ammonium magnesium phosphate added as the crystallization aid to 0.2% of the material mass (weighing 2.6 g), and other operation methods are the same as those in Example 1.

[0067] Experimental results: The mass of the food-grade L-isoleucine finished product after drying is 1223.2 g, the bulk density of the finished product is 0.564 g / cm3 After the refined mother liquor was recycled, the total amount was 1404.8 g, the liquid-phase content was 98.5%, meeting the USP standard. The food-grade yield was 82.1%, the feed-grade yield was 14.4%, and the total yield was 96.5%. This shows that increasing the addition amount of the crystallization aid magnesium ammonium phosphate can improve the food-grade L-isoleucine yield and increase the bulk density of the finished product.

[0068] Example 3: Influence of Changing the Addition Timing of the Crystallization Aid on the Purification Process of L-Isoleucine Fermentation Broth

[0069] Referring to Example 1, steps (3) and (7) were adjusted as follows, and other operation methods were the same as those in Example 1.

[0070] Step (3): Adjust the pH value of the decolorized clear liquid obtained in step (2) to 6.0 with sodium hydroxide, add it to the crystallization evaporator, and perform evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the evaporation reaches the L-isoleucine content of 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator. The addition amount of the crystal seeds is 2% of the mass of the material in the crystallization evaporator (weighing 29.6 g), and add 0.1% of the mass of the material (weighing 1.5 g) of magnesium ammonium phosphate as the crystallization aid; the vacuum degree of the evaporator is controlled at 0.090 - 0.095 MPa, the stirring speed before adding the crystal seeds is 50 r / min, and the stirring speed after adding the crystal seeds is 100 r / min; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water with a volume of 5% of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank.

[0071] Step (7): Adjust the pH to 6.0 with sodium hydroxide for the decolorized clear liquid obtained in step (6), add it to the crystallization evaporator, and perform evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C. When the evaporation reaches the L-isoleucine content of 60 - 65 g / L, add food-grade L-isoleucine as crystal seeds to the crystallization evaporator. The addition amount of the crystal seeds is 1% of the mass of the material in the crystallization evaporator (weighing 13.2 g), and add 0.1% of the mass of the material (weighing 1.3 g) of magnesium ammonium phosphate as the crystallization aid; the vacuum degree of the evaporator is controlled at 0.090 - 0.095 Mpa, the stirring speed before adding the crystal seeds is 50 r / min, and the stirring speed after adding the crystal seeds is 100 r / min; when the L-isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add water with a volume of 5% of the concentrated liquid in the crystallizer to dissolve the fine crystals, and then discharge the tank.

[0072] Experimental results: The mass of the dried food-grade L-isoleucine finished product is 1210.6 g, and the bulk density of the finished product is 0.554 g / cm 3, after the refined mother liquor was recycled, the total amount was 1394.8 g, the liquid phase content was 98.9%, meeting the USP standard. The food-grade yield was 81.5%, the feed-grade yield was 14.9%, and the total yield was 96.4%. This shows that changing the addition timing of the crystallization aid magnesium ammonium phosphate can also increase the food-grade L-isoleucine yield and improve the bulk density of the finished product.

[0073] Purification method of L-valine in Comparative Example 2

[0074] The purification method of L-valine includes the following steps: (1) separating the denatured bacteria in the fermentation broth to obtain a clear liquid; (2) decolorizing the clear liquid; (3) performing evaporation crystallization on the decolorized clear liquid; (4) centrifuging the crystallization liquid to obtain a wet product; (5) drying the wet product to obtain an L-valine product. Specifically as follows:

[0075] (1) Separating the denatured bacteria in the fermentation broth to obtain a clear liquid: Taking 25 L of L-valine fermentation broth with an L-valine content of 85.0 g / L, adjusting the pH to 5.0 with concentrated sulfuric acid to denature and precipitate the protein, and then separating the bacteria through a ceramic membrane to obtain a retentate containing bacteria and a clear liquid without bacteria;

[0076] (2) Decolorizing the clear liquid: Decolorizing the clear liquid obtained in step (1) with activated carbon, adding 0.3 kg of activated carbon to every 100 L of clear liquid; stirring and decolorizing at 70 °C for 60 min, and the light transmittance of the decolorized liquid was 45.9%;

[0077] (3) Performing evaporation crystallization on the decolorized clear liquid: Adjusting the pH value of the decolorized clear liquid obtained in step (2) to 6.0 with sodium hydroxide and adding it to a crystallization evaporator for evaporation crystallization; the temperature of the material in the crystallization evaporator was 65 - 70 °C, controlling the vacuum degree to be 0.090 - 0.095 MPa, and the stirring speed was 60 revolutions per minute; when the L-valine content in the crystallization evaporator reached 550 - 600 g / L, stop evaporation, add 5% by volume of water to dissolve the fine crystals, and then discharge the tank;

[0078] (4) Centrifuging the crystallization liquid to obtain a wet L-valine product: Gradually cooling the crystallization liquid to 10 °C at a cooling rate of 5 °C per hour, then centrifuging for 20 minutes at a speed of 3000 revolutions per minute, adding 10% by volume of washing water to wash the mother liquor on the crystal surface, and continuing to centrifuge for 30 min after adding water to separate the wet L-valine product with a water content of 8.3%;

[0079] (5) Drying the wet product to obtain a food-grade L-valine product: Drying the wet L-isoleucine product obtained in step (4) at 105 °C to obtain a qualified L-valine finished product.

[0080] The mass of the dried L-valine finished product was 1756.0 g, and the bulk density of the finished product was 0.536 g / cm 3, the yield is 82.6%. Purification method of L-valine fermentation broth with crystal growth promoter added to the bottom material in Example 4

[0081] The purification method of L-valine comprises the following steps: (1) denaturing and separating the bacteria in the fermentation broth to obtain a supernatant; (2) decolorizing the supernatant; (3) evaporating and crystallizing the decolorized supernatant; (4) centrifuging the crystallization solution to obtain a wet product; (5) drying the wet product to obtain an L-valine product. Specifically as follows:

[0082] (1) Denaturing and separating the bacteria in the fermentation broth to obtain a supernatant: Take 25 L of L-valine fermentation broth with an L-valine content of 85.0 g / L, adjust the pH to 5.0 with concentrated sulfuric acid to denature and precipitate the protein, and then separate the bacteria through a ceramic membrane to obtain a retentate containing bacteria and a supernatant without bacteria;

[0083] (2) Decolorizing the supernatant: Decolorize the supernatant obtained in step (1) with activated carbon, adding 0.3 kg of activated carbon to every 100 L of supernatant; stirring and decolorizing at 70 °C for 60 min, and the light transmittance of the decolorized solution is 45.9%;

[0084] (3) Evaporating and crystallizing the decolorized supernatant: Adjust the pH value of the decolorized supernatant obtained in step (2) to 6.0 with sodium hydroxide, add it to a crystallization evaporator, and add 0.2% (weight 3.2 g) of magnesium ammonium phosphate based on the mass of the material as a crystal growth promoter, and carry out evaporation crystallization; the temperature of the material in the crystallization evaporator is 65 - 70 °C, control the vacuum degree to be 0.090 - 0.095 MPa, and the stirring speed is 60 rpm; when the L-valine content in the crystallization evaporator is 550 - 600 g / L, stop evaporation, add 5% by volume of water to dissolve the fine crystals, and then discharge the tank;

[0085] (4) Centrifuging the crystallization solution to obtain a wet L-valine product: Gradually cool the crystallization solution to 10 °C at a cooling rate of 5 °C per hour, then centrifuge for 20 minutes at a speed of 3000 rpm, add wash water with a volume ratio of 10%, wash the mother liquor on the crystal surface, continue to centrifuge for 30 min after adding water, and separate to obtain a wet L-valine product with a water content of 8.3%;

[0086] (5) Drying the wet product to obtain a food-grade L-valine product: Dry the wet L-isoleucine product obtained in step (4) at 105 °C to obtain a qualified L-valine finished product.

[0087] The mass of the dried L-valine finished product is 1779.2 g, and the bulk density of the finished product is 0.551 g / cm 3 , the yield is 83.7%.

[0088] Although the present invention has been described in detail above with general descriptions and specific embodiments, based on the present invention, some modifications or improvements can be made thereto, which are obvious to those skilled in the art. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

Claims

1. Application of magnesium ammonium phosphate in the refining process of amino acid fermentation production, characterized in that, The application includes: after the amino acid fermentation broth is successively subjected to membrane filtration and decolorization treatment, magnesium ammonium phosphate is added to the obtained crystallization substrate, or magnesium ammonium phosphate is added during the crystallization process.

2. The application according to claim 1, wherein The amino acids include isoleucine and valine.

3. A method for purifying isoleucine fermentation broth, characterized in that, First, adjust the pH value of the isoleucine fermentation broth to 4 - 6, remove most of the bacteria, miscellaneous proteins and colloids by ceramic membrane filtration, and perform the first activated carbon decolorization at pH 4 - 6. Then adjust the pH to 5 - 7 for the first evaporation crystallization. Add crystal seeds into the crystallization evaporator, and add magnesium ammonium phosphate to the crystallization substrate or during the crystallization process. When the isoleucine content in the crystallization evaporator is 250 - 260 g / L, discharge the tank. The crystallization liquid is centrifuged to obtain crude isoleucine and crude mother liquor respectively; after the crude isoleucine is redissolved in water, perform the second activated carbon decolorization, then adjust the pH to 5 - 7 for the second evaporation crystallization. Add crystal seeds into the crystallization evaporator, and add magnesium ammonium phosphate to the crystallization substrate. When the isoleucine content in the crystallization evaporator is 250 - 260 g / L, discharge the tank. The crystallization liquid is centrifuged to obtain refined isoleucine, which is dried to obtain the finished product of isoleucine.

4. The method according to claim 3, characterized in that, When performing the first evaporation crystallization, the addition amount of crystal seeds is 1 - 2% of the mass of the material in the crystallization evaporator.

5. The method according to claim 3, wherein When performing the first evaporation crystallization, the addition amount of magnesium ammonium phosphate is 0.1% - 0.2% of the mass of the material in the crystallization evaporator.

6. The method according to claim 3, characterized in that, When performing the second evaporation crystallization, the addition amount of crystal seeds is 1 - 2% of the mass of the material in the crystallization evaporator.

7. The method according to claim 3, characterized in that, When performing the second evaporation crystallization, the addition amount of magnesium ammonium phosphate is 0.1% - 0.2% of the mass of the material in the crystallization evaporator.

8. The method according to any one of claims 1-7, characterized in that, It includes the following steps: (1) Separation of denatured bacteria in the isoleucine fermentation broth to obtain clear liquid: Take 31 L of isoleucine fermentation broth with an isoleucine content of 55.2 g / L, adjust the pH to 4.5 to denature and sediment the protein, and then separate the bacteria through a ceramic membrane to obtain the retentate containing bacteria and the clear liquid without bacteria. (2) Primary decolorization of the clear liquid: Perform activated carbon decolorization on the clear liquid obtained in step (1), add 0.6 kg of activated carbon to every 100 L of clear liquid; stir and decolorize at 65 - 70 °C for 30 - 60 min. (3) Primary evaporation crystallization after decolorization of the clear liquid: Adjust the pH value of the decolorized clear liquid obtained in step (2) to 6.0, add it to the crystallization evaporator, and add 0.1% of the mass of the material of magnesium ammonium phosphate as a crystallization aid for evaporation crystallization. The temperature of the material in the crystallization evaporator is 65 - 70 °C. When the evaporation reaches an isoleucine content of 60 - 65 g / L, add food - grade isoleucine as crystal seeds to the crystallization evaporator, and the addition amount of crystal seeds is 2% of the mass of the material in the crystallization evaporator; control the vacuum degree of the evaporator at 0.090 - 0.095 MPa, the stirring speed before adding crystal seeds is 20 - 50 r / min, and the stirring speed after adding crystal seeds is 80 - 100 r / min; when the isoleucine content in the crystallization evaporator is 250 - 260 g / L, stop evaporation, add 5% volume of water to dissolve the fine crystals, and then discharge the tank. (4) The crude isoleucine is obtained by centrifuging the crystallization solution: After the crystallization solution is discharged from the tank, it is separated directly without cooling. The mother liquor on the crystal surface is washed with water, and then centrifuged continuously after adding water to obtain the crude isoleucine and the crude mother liquor; (5) Redissolution of the crude isoleucine: The crude isoleucine obtained in step (4) is dissolved in water, and the water temperature is controlled at 65 - 66 °C to prepare a redissolved solution with a content of 50 g / L; (6) Secondary decolorization of the crude product dissolution solution: The pH of the redissolved solution obtained in step (5) is adjusted to 5.0 for secondary decolorization. 0.3 kg of activated carbon is added to every 100 L of the dissolution solution, and it is stirred and decolorized at 65 - 70 °C for 30 - 60 min; (7) Secondary evaporation crystallization of the decolorized clear liquid: The pH of the decolorized clear liquid obtained in step (6) is adjusted to 6.0, added to a crystallization evaporator, and 0.1% of magnesium ammonium phosphate based on the mass of the material is added as a crystallization aid for evaporation crystallization; The temperature of the material in the crystallization evaporator is 65 - 70 °C. When the evaporation reaches an isoleucine content of 60 - 65 g / L, food - grade isoleucine is added to the crystallization evaporator as crystal seeds, and the addition amount of the crystal seeds is 1% of the mass of the material in the crystallization evaporator. The vacuum degree of the evaporator is controlled at 0.090 - 0.095 MPa. The stirring speed before adding the crystal seeds is 20 - 50 r / min, and the stirring speed after adding the crystal seeds is 80 - 100 r / min. When the isoleucine content in the crystallization evaporator is 250 - 260 g / L, evaporation is stopped, 5% by volume of water is added to dissolve the fine crystals, and then it is discharged from the tank; (8) The refined isoleucine is obtained by centrifuging the crystallization solution: After the crystallization solution in step (7) is discharged from the tank, it is separated directly without cooling. The mother liquor on the crystal surface is washed with water, centrifuged and dried, and then the crystals are washed with 30% alcohol. After washing with alcohol, it is centrifuged continuously to obtain the refined isoleucine, which is dried to obtain the finished product of food - grade isoleucine.

9. The method according to claim 8, characterized in that, (4) also includes: After mixing the separated crude mother liquor with the retentate containing bacteria obtained in step (1), it is concentrated to 50 - 55% dry matter and spray - granulated to obtain feed - grade isoleucine.

10. The method according to any one of claims 3-9, characterized in that, The strain used for isoleucine fermentation is Corynebacterium glutamicum.

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