WO2008134936A1

WO2008134936A1 - Method for extracting threonine from threonine fermentation liquor

Info

Publication number: WO2008134936A1
Application number: PCT/CN2008/000874
Authority: WO
Inventors: Dehui Wang; Dongshu Jia
Original assignee: Changchun Dacheng Industrial Group Company Limited
Priority date: 2007-04-29
Filing date: 2008-04-29
Publication date: 2008-11-13
Also published as: CN101293847A; CN101293847B

Abstract

Disclosed is a method for extracting threonine from threonine fermentation liquor, comprising: filtrating the sterilized threonine fermentation liquor through ceramic film, decoloring the resulting threonine fermentation liquor by adding of active carbon, then pouring the threonine fermentation liquor into four-effect evaporator to be concentrated, introducing the concentrated liquor into gradient temperature drop crystalling tank to be crystallized when the Baume concentration of the liquor reaches 25~26, centrifugation separating and drying the resulting threonine crystals, and thus obtaining the threonine crystals with the purity of more than 98.5%. The primary mother liquor obtained from the crystalling step can be used in circle, and threonine crystalline product can be obtained by the processes of decoloration, concentration, temperature drop crystalling, centrifugation, re-resolution, centrifugation separation and dry of said mother liquor. The method of the invention provides a purifying yield of threomine of more than 95%, and solves the environmental problems caused by discharge of the mother liquor in large amount.

Description

Method for extracting threonine from threonine fermentation broth

The present invention relates to a method for fermenting threonine by threonine fermentation, whereby a threonine crystal having a high content of threonine can be obtained, and the threonine crystal can be used as an animal feed additive. Background technique

Threonine is one of the essential amino acids in humans and animals, but threonine cannot be synthesized in the body and must be taken from food. Therefore, in recent years, threonine has been widely used as an additive in the fields of food, health care, and feed industries. In the animal term, threonine is the second limiting amino acid. In recent years, as the word additive, as the amount of lysine has increased year by year, the amount of threonine has increased year by year. China's threonine feed additive mainly relies on imports. At present, medical threonine is mostly chemically synthesized. The production of a large amount of threonine used as a food or feed additive is mainly produced by fermentation. Compared with the chemical synthesis method, the fermentation method has a large production scale, high yield and low cost, which can meet the increasing demand. The process for producing threonine by fermentation is not mature enough, and various production steps such as filtration, concentration and crystallization are to be improved and optimized. Patent application of Tianjin University of Science and Technology

CN200610014324.4 discloses a method for separating and extracting L-threonine from a fermentation liquid, wherein the fermentation liquid is filtered by a metal membrane to improve the filtration process and improve the production efficiency, and the purity of the threonine crystal product can reach 99. %the above. The inventors have made a lot of research and experiments on the process of extracting the threonine crystal product from the fermentation liquid, and have successfully completed the industrial scale production experiment, the product quality is good, the yield is high, and there is no Environmental pollution. Summary of the invention

It is an object of the present invention to provide a method for preparing threonine crystals from a threonine fermentation broth comprising the steps of:

(1) After the threonine fermentation broth is sterilized by high temperature, impurities such as bacteria and proteins are removed by filtration through an inorganic ceramic membrane to obtain a clarified threonine fermentation liquid;

(2) decolorizing the threonine fermentation broth with activated carbon; (3) concentrating the decolorized fermentation broth under reduced pressure to a concentration of 25 Bo or more;

(4) allowing the concentrated concentrated liquid to enter the crystallization tank for cooling and crystallization;

(5) centrifugally separating the crystal slurry after cooling and crystallization to obtain crystals and a mother liquor;

(6) Dry the crystal to obtain a finished product of threonine crystal. In the method of the present invention, the step (1) further comprises: after subjecting the threonine fermentation broth to high temperature sterilization, adjusting the pH to 4.0 to 5.0, preferably about 4.5, with sulfuric acid, and then removing the cells by filtration through an inorganic ceramic membrane. With respect to impurities such as proteins, the membrane can retain a protein having a molecular weight of 300 kD, and a threonine fermentation liquid having a light transmittance of 87% or more is obtained. The bacterial protein removed by membrane filtration can be made into a fertilizer after being filtered by a three-stage membrane. The inorganic ceramic membrane is obtained by sintering an oxide such as alumina, titania or zirconia at a high temperature, and is a porous ceramic filter material having a porous structure in which the porous support layer, the filter layer and the microporous membrane layer are asymmetrically distributed. Filtration accuracy can range from microfiltration to ultrafiltration to sodium filtration. Ceramic membrane filtration is a fluid separation process in the form of "cross-flow filtration" in which a raw material liquid flows at a high speed in a membrane tube, and a clarified permeate containing a small molecular component is driven through the membrane in a direction perpendicular thereto. The turbid concentrate containing the macromolecular component is trapped by the membrane, thereby allowing the fluid to be separated, concentrated and purified. Compared with traditional filter materials, inorganic ceramic membranes have many advantages, such as excellent chemical stability, high temperature resistance, high mechanical strength, narrow pore size distribution, high separation precision, easy cleaning, etc., compared with other filter materials, inorganic ceramics. When the membrane is separated, the obtained separation product is high in purity and easier to concentrate, so it is widely used in pharmaceutical, bioengineering, electronics and other industries. When the inorganic ceramic membrane is used for the filtration operation, the operating temperature and pressure can be carried out according to the instructions of the product specification or the conventional operating conditions. Preferably, for example, the membrane filtration temperature is about 75 ° C, and the pressure inlet pressure is about 0.7 mpa/m. ² , the filming pressure is about 0.4mpa/m ² . In the method of the present invention, the activated carbon used in the decolorization step of the activated carbon in the step (2) may be a conventionally used activated carbon, preferably two activated carbons having different particle sizes are used together, and more preferably the particle size is 200~ 300 mesh and 18 mesh activated carbon are used together. Among them, 200~300 mesh activated carbon can be used to remove about 80% of the color. With 18 mesh activated carbon, about 15% of the color can be removed. Decolorization with activated carbon can greatly improve the decolorization efficiency of activated carbon. The light transmittance can reach more than 99%, which is beneficial to Large crystal grains are formed. Those skilled in the art can easily determine the ratio of the two activated carbon particles according to the condition of the material. Generally, the ratio of the activated carbon having a particle size of 200 to 300 mesh and 18 mesh is 4:3 (volume ratio). In the method of the present invention, the concentration under reduced pressure according to the step (3) can be carried out by a conventional concentration method, preferably by using a multi-effect falling film evaporation apparatus and method, for example, a four-effect falling film evaporator for concentration, and a four-effect falling film. The temperatures of the evaporation system are approximately: one-effect evaporation temperature of 90 ° C, two-effect evaporation temperature of 60 Ό, three-effect evaporation temperature of 80 ° C, and four-effect evaporation temperature of 70 ° C. When the concentration of the obtained concentrated liquid reaches 25 Bo or more, preferably 25 to 26 Bo, the concentration step can be completed.

In the method of the present invention, the temperature-lowering crystallization described in the step (4) is carried out by using a gradient cooling crystallization method in the crystallization tank, and the gradient cooling crystallization condition is an initial temperature of 70 ° C and a cooling rate of 2.5-2.7 ° C / h. , the minimum temperature is 5-10 ° C, maintained for 15-30 hours, while stirring, the small grains are further grown to facilitate separation.

The preferred conditions for the gradient cooling crystallization are: an initial temperature of 70 ° C, a cooling rate of 2.57 ° C / h, a minimum temperature of 9 ° C, a temperature of 20 h, and a stirring speed of about 30 rpm. In order to ensure the obtained crystal of threonine of a suitable size, it is preferred to carry out the crystallization operation in the above temperature and pH range, and the temperature and pH stability should be maintained as much as possible, and it is preferred that the maximum fluctuation of the temperature does not exceed the soil C, The maximum fluctuation of pH does not exceed ± 0.1. According to the method for extracting threonine crystals from the threonine fermentation broth according to the present invention, the purity of the resulting threonine crystal can reach 98.5% or more, and the threonine extraction rate can reach 95% or more. Preferably, the method for extracting threonine crystals from the threonine fermentation broth of the present invention comprises the following steps:

(1) After the threonine fermentation broth is sterilized by high temperature, the pH is adjusted to 4.5 with sulfuric acid, and then the inorganic protein membrane is used to filter and remove impurities such as bacterial proteins, and the membrane can retain a protein having a molecular weight of 300 kD to obtain a light transmittance. More than 87% of threonine fermentation broth;

(2) Decolorizing the above-mentioned threonine fermentation liquid with two kinds of activated carbons having different particle sizes, preferably using 200-300 mesh and 18-mesh activated carbon for decolorization, and the light transmittance of the fermentation liquid after decolorization reaches 99% or more;

(3) The decolorized fermentation broth enters the four-effect falling film evaporator, so that the concentration of the fermentation liquid reaches 25~26Bo°; (4) The above fermentation broth is passed into a gradient cooling crystallization tank for temperature crystallization, and the gradient crystallization conditions are: starting temperature 70 ° C, cooling rate is 2.5-2.7 ° C / h, minimum temperature 5 - 10 ° C, Maintain for 15-30 hours while stirring;

(5) centrifugally separating the crystal slurry after cooling and crystallizing to obtain crystals and a mother liquor;

(6) Drying the crystal to obtain a finished product of threonine, and the purity of the obtained threonine crystal can reach 98.5% or more. Further, in the method for extracting threonine according to the present invention, the primary mother liquor recovered in the step (5) can be recovered and reused, and the reuse method comprises the following steps: First, fermenting the sterilized and filtered threonine The clear liquid is combined and injected into the program of the method of the present invention for recycling; the second is to directly decolorize, concentrate, gradient crystallize, and centrifuge to obtain threonine crystal and secondary mother liquid, and the obtained crystal is obtained. The finished threonine crystals are obtained by redissolving, crystallizing and separating and drying the crystals with water. The method described in the latter is basically the same as the method for treating the fermentation liquid concentrate described above, that is, another independent process system is provided, and the operating conditions of each step are substantially the same as those described above. - Specifically, the method for obtaining threonine by recrystallization from a mother liquor in the latter method is: decolorizing, concentrating, gradient cooling, and centrifuging the primary mother liquor to obtain threonine crystals and a second mother liquor. The threonine content of the obtained threonine crystal is about 60%; the crystal is redissolved with water, that is, the crystal is dissolved by steam at 70-80 ° C with high-temperature hot water, for example, the obtained crystal is 2 tons. The ratio of lm ³ water is dissolved in high temperature hot water at 70-80 ° C with steam, and then cooled again by gradient cooling, separation and drying to obtain the finished threonine crystal. Further, the primary mother liquor obtained in the step (5) is subjected to decolorization, concentration, gradient cooling crystallization and centrifugation to obtain a threonine product, and a secondary mother liquor is obtained in the above centrifugation process, and the secondary mother liquor is also obtained. Recyclable, for example, can be used to make fertilizer. Further, the secondary condensate of the concentration step in the method of the present invention can be fully utilized, and can be used as condensed water or as washing water in a post-treatment operation of threonine crystals. The method of the invention has high yield and the threonine extraction rate can reach more than 95%; the yield, that is, the ratio of the total amount of the finished crystal product to the total amount of threonine in the clear liquid can reach above 91%; the product quality is good, the obtained product The purity of threonine crystals is above 98.5 %; the crystal product particles are moderate, which is advantageous for separation, and is also advantageous for mixing with other ingredients in feed processing; The comprehensive utilization of various by-products greatly reduces the production cost. For example, the primary mother liquor, the secondary mother liquor, and the bacterial protein dosage form in the production process can be comprehensively utilized as needed, and the threonine is recovered or made into a secondary product. Grade products, such as fertilizers; the secondary condensate obtained in the concentration step can be used as condensed water or washing water in the post-treatment of the threonine crystallization process. The method of the invention is simple, easy to popularize, and can meet the increasing demand, and the production method of the invention is a typical environment-friendly production process, which is beneficial to environmental protection. - BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is a flow chart showing the preparation of threonine crystals from a threonine fermentation broth according to the present invention. BEST MODE FOR CARRYING OUT THE INVENTION

The following examples are given to further illustrate the process of the invention. However, these examples do not limit the scope of the invention in any way. Example 1 Preparation of threonine crystals from threonine fermentation broth

The specific operation steps are carried out according to the flow shown in FIG.

1. Preparation of threonine fermentation broth according to a conventional method: using corn sugar as a main raw material, adding ammonium sulfate, ammonia water and supplemented with corn syrup as a nitrogen source, and adding a bacterium of the genus Brevibacterium to ferment a threonine fermentation liquid, the fermentation broth The basic parameters are:

pH 7~8

Threonine content: 7~12g/100ml

Dry matter: 12~14g/100ml)

2, the filtered broth: take threonine broth 300m ^3, wherein the threonine content of 9g / 100ml, pH value of 7.00, the 100 ° C high temperature sterilization, adjusted with food grade concentrated sulfuric acid to pH 4.5, followed by The pump is pumped into a 300KD inorganic ceramic membrane (France, Novartis Orelis, model: K99BW) for filtration to remove impurities such as bacterial proteins. The pumping pressure is 7kg, the pumping pressure is 4kg, and the temperature is 70°C. If the pressure is too high, the membrane will be broken and the filtration will be lost.

3. Decolorization of the fermentation broth: The clarified fermentation broth is added with 200-300 mesh activated carbon at a temperature of 70 ° C, pH 4.5, clarified fermentation broth at a ratio of 1%, filtered, and then passed through two 18-mesh activated carbon. The column is decolored, and the light transmittance of the decolorized fermentation broth is above 99%, which is favorable for forming large crystal grains. The carbon column has a diameter of 3 m, a height of 5 m, and a pumping flow rate of 150 m ³ /h. 4. Concentration of fermentation broth: The decolorized fermentation broth enters the four-effect falling film evaporation system, the evaporation temperature of one effect is 90Ό, the evaporation temperature of the second effect is 60°C, and the wave of the second-effect evaporation is between about 9~10. the three-way evaporation temperature of 80 ° C, four-effect evaporation temperature of 70 ° C, to obtain a concentrate 145m ^3, Baume concentration of 25 ~ 26Bo °. At the same time, a secondary condensate of 130 m ^{3 is obtained} , which can be stored as condensed water or washing water for use.

5. Gradient cooling crystallization: The crystal slurry obtained in the previous step is pumped from the second-effect evaporation pump into the gradient cooling crystallization tank, and the cooling water tube bundle of the crystallization tank is passed through the groundwater to cool down, the cooling rate is 2.570 ° C / h, and the stirring speed is 30 rpm. However, when the temperature is lower than 15 °C, due to the limitation of groundwater temperature, the ice water is used to cool down, and the cooling rate is constant. However, when the temperature reaches 9X:, the temperature is stirred for 20 hours to make the small grains grow further, so as to facilitate Separation.

6. Post-treatment of crystallization: pump the crystal slurry into the piston to push the centrifuge to separate the crystal and the primary mother liquor, and wash it three times with the condensed water obtained in step 4 to wash away impurities such as inorganic salts and pigments attached to the crystal surface. In order to improve the purity of the crystal, the threonine crystal is obtained after drying, and the crystal is dried and weighs 23.00 tons, and the content is 98.5% or more. The centrifuge used for centrifugation of crystal is a self-unloading three-legged centrifuge with a separation factor of 0.5-0.6 and a screen mesh of 250 mesh. The screen is too small to dehydrate the crystal material, and it is easy to run the crystal material and waste. energy.

7, comprehensive utilization

(1) Comprehensive utilization of primary mother liquor: The washing water and the primary mother liquor are combined for about 30 m ³ and the threonine content is 8 g/100 ml, which is again decolorized, concentrated, cooled and crystallized and separated, and the separated crystals are lm ³ Water: The ratio of 2 tons of material is re-dissolved (ie, redissolved), the soluble inorganic salt and pigment on the surface of the crystal are dissolved in water, and then crystallized, separated and dried to obtain 2.20 tons of finished threonine crystal. The content of threonine is 98.5% or more, compared with the crystal obtained in the previous step, the crystal grains are small, and it is easy to mix with other ingredients in the subsequent feed processing. The operating conditions employed therein are the same as those described above.

(2) Comprehensive utilization of the secondary mother liquor: A secondary mother liquor is obtained when the crystal is separated, wherein the threonine content is 7.90 g/100 ml and the volume is 2.5 m ³ . The above secondary mother liquor and the bacterial cells are mixed and granulated into a fertilizer.

(3) Comprehensive utilization of bacterial proteins: The bacterial proteins removed by membrane filtration are filtered through a tertiary membrane. After that, the pump is pumped along the pipeline, wherein the clear liquid is 270m ³ , 30m ³ wet cells, the threonine content is 1.2g/100ml, the wet cells are sent to the by-product workshop for drying, and after drying, the bacterial protein is dried. , used to make fertilizer.

(4) Comprehensive utilization of secondary condensate: The 145 m ³ secondary condensate of the concentration step is used as washing water in the post-treatment of threonine crystals.

8, the product

Finished product of threonine: 25.2 tons, yield of finished product: 91.94%; bacterial protein (after drying)

5 tons, mixed with the second mother liquor to make fertilizer. The finished product of the threonine crystal obtained by the method of the present embodiment has a purity of 98.5% or more and has the following dry matter composition:

Free threonine > 98.5% by weight

Ammonium salt <0.04% by weight

Moisture <1% by weight

Arsenic <0.001 ppm (by weight) The threonine extraction rate of this example was 95% or more. The embodiments of the present invention have been described in detail above, and it is obvious to those skilled in the art that many modifications and changes can be made without departing from the spirit of the invention. All such variations and modifications are within the scope of the invention.

Claims

The claims are:

1. A method for extracting threonine crystals from a threonine fermentation broth, the method comprising the steps of:

(2) decolorizing the threonine fermentation broth with activated carbon;

(3) concentrating the decolorized fermentation broth under reduced pressure to a concentration of 25 Bo or more;

(6) Dry the crystal to obtain a finished product of threonine crystal.

The method according to claim 1, wherein said step (1) is carried out by first adjusting the high temperature sterilized fermentation broth to a pH of about 4.5 with sulfuric acid before filtering through the inorganic ceramic membrane.

The method according to claim 2, wherein said inorganic ceramic film of said step (1) is capable of retaining a protein having a molecular weight of 300 kD.

4. The method according to claim 2, wherein said activated carbon is decolored by using two activated carbons having different particle sizes, preferably 200 to 300 mesh and 18 mesh activated carbon, wherein said 200 to 300 mesh and 18 mesh activated carbon are used. The volume ratio is 4:3; the transmittance of the fermentation liquid after decolorization by activated carbon reaches 99% or more.

5. The method according to claim 2, wherein said reduced pressure concentration is a four-effect falling film evaporator; preferably, said four-effect falling film evaporation system has operating temperatures of: one-effect evaporation temperature of 90 ° C The second-effect evaporation temperature is 60 ° C, the three-effect evaporation temperature is 80 ° C, and the four-effect evaporation temperature is 70 ° C; after the vacuum concentration step, the concentration of the fermentation liquid reaches 25 Bo ° or more; preferably 25 to 26 Bo ^0.

6. A method according to claim 2 wherein said temperature-lowering crystallization employs batch operation and gradient temperature crystallization; wherein said gradient cooling crystallization is carried out under agitation.

7. The method according to claim 6, wherein said gradient cooling crystallization is carried out in a crystallization tank, said gradient cooling crystallization conditions are: starting temperature 70 ° C, cooling rate is 2.5-2.7 ° C / h , the lowest temperature is 5-10 ° C, and is maintained for 15-30 hours while stirring; preferably, the gradient cooling crystallization conditions are: starting temperature 70 ° C, cooling rate is 2.57 ° C / h, minimum temperature 9 °C, maintaining this temperature for 20 hours, the stirring speed is about 30 rev / min; more preferred The maximum fluctuation of the temperature in the crystallization operation does not exceed 1 ° C of the soil, and the maximum fluctuation of the pH does not exceed ± 0.1.

8. A method according to any of claims 1-7, wherein the method comprises the steps of:

(1) After the threonine fermentation broth is autoclaved, the pH is adjusted to 4.5 with sulfuric acid, and then the inorganic protein membrane is used to filter and remove impurities such as bacterial proteins, and the membrane can retain a protein having a molecular weight of 300 kD;

(2) Decolorizing the above-mentioned threonine fermentation broth with two kinds of activated carbons having different particle sizes, preferably using decolorization of activated carbon of 200-300 mesh and 18 mesh;

(3) The decolorized fermentation broth enters the four-effect falling film evaporator, so that the concentration of the fermentation liquid reaches 25~26Bo°;

(4) The above fermentation broth is passed into a gradient cooling crystallization tank for gradient cooling crystallization. The gradient crystallization conditions are: initial temperature 70 ° C, cooling rate 2.5-2.7 ° C / h, minimum temperature 5-10 ° C , maintain for 15-30 hours while stirring;

(6) Dry the crystal to obtain a finished product of threonine crystal.

9. The method according to claim 8, wherein the primary mother liquor obtained in the step (5) can be reused or combined with the sterile filtered threonine fermentation broth for recycling; preferably The primary mother liquor is decolorized, concentrated, gradient crystallized, and centrifuged to obtain threonine crystals and a second mother liquor, and the obtained crystals are redissolved, crystallized and separated and dried to obtain a finished threonine crystal; The re-dissolution step is to dissolve the crystals by adding steam at 70-80 ° C with high-temperature hot water.

10. A process according to claim 9 wherein the secondary mother liquor obtained during the separation of threonine crystallization is useful in the manufacture of fertilizers.