CN108715586B - Method for recycling L-cystine mother liquor through enzymatic reaction - Google Patents
Method for recycling L-cystine mother liquor through enzymatic reaction Download PDFInfo
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- CN108715586B CN108715586B CN201810676343.8A CN201810676343A CN108715586B CN 108715586 B CN108715586 B CN 108715586B CN 201810676343 A CN201810676343 A CN 201810676343A CN 108715586 B CN108715586 B CN 108715586B
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- C07D277/12—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having one double bond between ring members or between a ring member and a non-ring member with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
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Abstract
The application discloses a method for recycling L-cystine mother liquor in enzymatic reaction, which specifically comprises the following steps: evaporating the L-cystine mother liquor until the concentrations of DL-ATC and L-cystine are 5-10 g/L and 3-8 g/L respectively, adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 2-4, selectively adsorbing and eluting through cation exchange resin, collecting eluent in a certain time period, and directly entering a process cycle to realize the recycling of the L-cystine mother liquor. The method has the following benefits: (1) the waste of materials is reduced, waste materials are changed into valuable materials, and the yield is indirectly improved; (2) can obviously reduce the treatment difficulty of the waste water and save the production cost. (3) The materials do not need to be extracted independently, but directly enter the process circulation, and the operation is simple and convenient.
Description
Technical Field
The application relates to the field of post-treatment of amino acid in enzymatic reaction, in particular to a recycling method of L-cystine mother liquor in enzymatic reaction.
Background
L-cystine is one of amino acids essential to human body, and has wide application in aspects of amino acid medicines, food additives, chemical raw materials and the like. Currently, the production process of L-cystine mainly comprises a protein hydrolysis method, a chemical synthesis method, a fermentation method and an enzyme method. The enzyme method has the advantages of mild reaction conditions, good stereoselectivity, good product quality, environmental friendliness and the like, so that the enzyme method has increasingly paid attention to and becomes a hotspot of the current research.
DL-ATC, the full name DL-2-amino-delta 2-thiazoline-4-carboxylic acid, is one of the most commonly used biological enzyme methods to catalyze the conversion of L-cysteine to ultimately produce L-cystine precursor. At present, the technology for producing L-cystine by the process is successfully industrialized industrially.
The L-cystine mother liquor is waste liquor obtained by enzymatic reaction, oxidation filtration or centrifugal extraction of a substrate with DL-ATC as a main component. The waste liquid has complex components, is based on the conversion efficiency of enzyme and the limitation of an L-cystine extraction process, contains about 0.5g/L of DL-ATC, 0.8g/L of L-cystine and other useful materials, and if the waste liquid is directly used, the conversion efficiency of enzymatic reaction is seriously influenced because cell endotoxin with certain concentration and other harmful substances are accumulated; if the waste water is directly discharged, the COD (chemical oxygen demand) and ammonia nitrogen value of the waste water are improved due to the useful materials contained in the waste water, the difficulty of waste water treatment is increased, so that the production cost is increased, meanwhile, the waste of the materials is caused, and how to reasonably recycle the waste water is a technical problem which is continuously concerned in the field for a long time.
Disclosure of Invention
The invention aims to solve the problems and provides a method for recycling L-cystine mother liquor through enzymatic reaction so as to achieve the aim of fully recycling DL-ATC and L-cystine in the L-cystine mother liquor.
Therefore, the invention discloses a method for recycling an enzymatic reaction L-cystine mother liquor, which comprises the following steps:
(1) evaporating the L-cystine mother liquor until the concentrations of DL-ATC and L-cystine are 5-10 g/L and 3-8 g/L respectively;
(2) adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 2-4;
(3) filling cation exchange resin into a chromatographic column, loading the concentrated solution of the L-cystine mother liquor obtained in the step (2) onto the chromatographic column of the cation exchange resin, and stopping loading when the cation exchange resin is saturated;
(4) and sequentially carrying out gradient elution on the chromatographic columns by using hydrochloric acid solutions with different concentrations, and collecting the eluent.
According to the method, the L-cystine mother liquor is waste liquor obtained after L-cystine is extracted by performing enzymatic reaction, oxidation filtration or centrifugation on a substrate taking DL-ATC as a main component;
preferably, the content of DL-ATC in the L-cystine mother liquor is 0.2-0.8 g/L, and the content of L-cystine is 0.2-1.5 g/L, and the mother liquor is particularly suitable for the recycling method provided by the invention and has particularly outstanding treatment efficiency and quality.
The method of the present invention, the evaporation in step (1) may be carried out using a variety of known equipment, preferably MVR equipment.
Preferably, the concentration is 5-20 times, the specific selection depends on the concentration of DL-ATC and L-cystine in the initial mother liquor, and after the concentration in the step (1), the invention further finds that the concentration is obviously beneficial to improving the treatment efficiency of the cation exchange resin and reducing the amount of wastewater.
According to the method, the pH value of the L-cystine mother liquor concentrated solution is adjusted to 2-4, preferably 3 in the step (2), the complete dissolution of the L-cystine and the subsequent adsorption separation of the L-cystine by using the cation exchange resin are facilitated in the pH value range, and the maximum adsorption capacity of the cation exchange resin is realized.
In the method of the present invention, preferably, the cation exchange resin is a D001 type cation exchange resin or a 732 sodium type cation exchange resin;
the exchange resins are all known commercial products, such as those available from the national drug group, the arotinoid chemical industry and the like.
The method also comprises the step of pretreating the resin, and specifically comprises the following steps:
soaking the cation exchange resin in saturated NaCl solution 3-5 times the volume of the resin for 18-20 hr, rinsing with clear water, soaking in 2-4% NaoH solution for 2-4 hr, rinsing with clear water to neutrality, soaking in 4-6% HCl solution for 4-8 hr, and rinsing with clear water to neutrality. According to the method, the eluent used in the step (4) is 0.1-0.3 mol/L hydrochloric acid solution and 0.8-1.5 mol/L hydrochloric acid solution;
wherein, hydrochloric acid solution with hydrochloric acid gradient concentration of 0.1mol/L-0.3mol/L is used for eluting L-cystine, and hydrochloric acid solution with concentration of 0.8mol/L-1.5mol/L is used for eluting DL-ATC;
preferably, the eluent used in the step (4) is 0.1-0.2 mol/L hydrochloric acid solution and 1.0-1.2 mol/L hydrochloric acid solution, so as to separate L-cystine and DL-ATC more effectively and ensure the separation effect.
The present invention preferably provides a method of gradient elution as follows:
washing the chromatographic column with 0.1-0.3 mol/L hydrochloric acid, collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent, changing to 0.8-1.5 mol/L hydrochloric acid when the concentration of DL-ATC reaches 0.2g/L, and separately collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent until the majority of DL-ATC is washed out;
by adopting the elution method, the invention can fully recover the residual DL-ATC and L-cystine in the enzymatic reaction L-cystine mother liquor.
The volume of each eluent and the concentration of the L-cystine and the DL-ATC in the eluent are respectively detected, and the recovery rate of the DL-ATC is more than 80 percent, the recovery rate of the L-cystine is more than 90 percent, and the COD (chemical oxygen demand) and ammonia nitrogen values before and after the concentration of the L-cystine mother liquor are reduced to 75 percent at most by detection and calculation.
The method also comprises the step of recycling the eluent.
Specifically, the DL-ATC eluent obtained by the method can be used for preparing a substrate of L-cystine by using a substrate with DL-ATC as a main component through an enzymatic reaction; the L-cystine eluent can be further refined and extracted.
In the method, the concentration monitoring of DL-ATC and L-cystine in each step is detected by HPLC.
The implementation of the invention has the following beneficial effects: (1) after the adsorption of the ion exchange resin, the COD and ammonia nitrogen of the L-cystine mother liquor are effectively reduced, the wastewater treatment difficulty is reduced, the production cost is saved, and the method is simple and efficient; (2) can fully recover DL-ATC and L-cystine in the L-cystine mother liquor; (3) the DL-ATC and L-cystine eluent does not need to be extracted independently, and directly enters the process circulation, so the operation is simple and convenient.
Drawings
FIG. 1 is a schematic flow diagram of the recycling of the recovered DL-ATC and L-cystine eluate in the process.
Detailed Description
The present invention is further illustrated by the following examples, in which it is noted that all reagents/materials used in the process of the present invention are commercially available.
Example 1:
in this example, a D001 type cation exchange resin is selected as a working resin, and a pretreatment method of the resin is provided, specifically as follows:
taking D001 type cation exchange resin as a working resin, soaking the resin in a saturated NaCl solution with the volume 3 times that of the resin for 18 hours, then rinsing the resin with clear water, then soaking the resin in a 3% NaoH solution for 3 hours, still rinsing the resin to be neutral with clear water, finally soaking the resin in a 5% HCl solution for 6 hours, and rinsing the resin to be neutral with clear water for later use.
The method for recycling the enzymatic reaction L-cystine mother liquor provided by the embodiment comprises the following steps:
filling the treated ion exchange resin into an 80L chromatographic column;
the concentrations of L-cystine and DL-ATC of the starting solution of the L-cystine mother liquor are respectively 0.96g/L and 0.78g/L through HPLC detection;
evaporating and concentrating the L-cystine mother liquor by about 10 times by using MVR equipment, and respectively detecting the concentrations of the L-cystine and DL-ATC to be 8.8g/L and 6.5g/L by HPLC;
adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 3.0;
pumping L-cystine mother liquor into ion exchange resin chromatographic column at constant speed of 1.5L/min, collecting effluent liquid at the other end, detecting concentrations of L-cystine and DL-ATC, and stopping loading the column when the outflow of L-cystine and DL-ATC is detected.
Washing the chromatographic column with clear water in the volume of the chromatographic column, and replacing the residual solution of the chromatographic column;
washing the chromatographic column with 0.1mol/L hydrochloric acid, collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent, when the concentration of DL-ATC reaches 0.2g/L, eluting the chromatographic column with 1.2mol/L hydrochloric acid, separately collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent until the majority of DL-ATC is eluted;
the volume of each eluent and the concentration of L-cystine and DL-ATC in each eluent were measured, wherein the recovery rate of DL-ATC was 83.9%, and the recovery rate of L-cystine was 94.6%. The COD and ammonia nitrogen values before and after the concentration of the L-cystine mother liquor are reduced by about 74.8 percent.
The two sections of eluents are respectively used in the preparation of the substrate and the refining process of the crude L-cystine, and the overall flow scheme of the embodiment is shown in figure 1.
Example 2:
in this example, 732 sodium type cation exchange resin is selected as the working resin, and a pretreatment method of the resin is provided, specifically as follows:
soaking 732 sodium type cation exchange resin as working resin in saturated NaCl solution in 3 times the volume of the resin for 18 hr, rinsing with clear water, soaking in 3% NaoH solution for 3 hr, rinsing with clear water to neutrality, soaking in 5% HCl solution for 6 hr, and rinsing with clear water to neutrality.
The method for recycling the enzymatic reaction L-cystine mother liquor provided by the embodiment comprises the following steps:
filling the treated ion exchange resin into an 80L chromatographic column;
the concentrations of the L-cystine and DL-ATC of the L-cystine mother liquor are respectively 0.96g/L and 0.78g/L through HPLC detection;
evaporating and concentrating the L-cystine mother liquor by about 10 times by using MVR equipment, and respectively detecting the concentrations of the L-cystine and DL-ATC to be 8.8g/L and 6.5g/L by HPLC;
adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 3.0;
pumping L-cystine mother liquor into ion exchange resin chromatographic column at constant speed of 1.5L/min, collecting effluent liquid at the other end, detecting concentrations of L-cystine and DL-ATC, and stopping loading the column when the outflow of L-cystine and DL-ATC is detected.
Washing the chromatographic column with clear water in the volume of the chromatographic column, and replacing the residual solution of the chromatographic column;
washing the chromatographic column with 0.1mol/L hydrochloric acid, collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent, when the concentration of DL-ATC reaches 0.2g/L, eluting the chromatographic column with 1.2mol/L hydrochloric acid, separately collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent until the majority of DL-ATC is eluted;
the volume of each eluent and the concentration of L-cystine and DL-ATC in each eluent were respectively detected, wherein the recovery rate of DL-ATC is 82.4%, and the recovery rate of L-cystine is 91.6%. The COD and ammonia nitrogen values before and after the concentration of the L-cystine mother liquor are reduced by about 64 percent.
The two sections of eluents are respectively used in the preparation of the substrate and the refining process of the crude L-cystine, and the overall flow scheme of the embodiment is shown in figure 1.
Example 3:
in this example, 732 sodium type cation exchange resin is selected as the working resin, and a pretreatment method of the resin is provided, specifically as follows:
soaking 732 sodium type cation exchange resin as working resin in saturated NaCl solution in 3 times the volume of the resin for 18 hr, rinsing with clear water, soaking in 3% NaoH solution for 3 hr, rinsing with clear water to neutrality, soaking in 5% HCl solution for 6 hr, and rinsing with clear water to neutrality.
The method for recycling the enzymatic reaction L-cystine mother liquor provided by the embodiment comprises the following steps:
filling the treated ion exchange resin into an 80L chromatographic column;
the concentrations of the L-cystine and DL-ATC of the L-cystine mother liquor are respectively 0.96g/L and 0.78g/L through HPLC detection;
evaporating and concentrating the L-cystine mother liquor by about 10 times by using MVR equipment, and respectively detecting the concentrations of the L-cystine and DL-ATC to be 8.8g/L and 6.5g/L by HPLC;
adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 2.0;
pumping L-cystine mother liquor into ion exchange resin chromatographic column at constant speed of 1.5L/min, collecting effluent liquid at the other end, detecting concentrations of L-cystine and DL-ATC, and stopping loading the column when the outflow of L-cystine and DL-ATC is detected.
Washing the chromatographic column with clear water in the volume of the chromatographic column, and replacing the residual solution of the chromatographic column;
washing the chromatographic column with 0.2mol/L hydrochloric acid, collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent, when the concentration of DL-ATC reaches 0.2g/L, eluting the chromatographic column with 1.0mol/L hydrochloric acid, and separately collecting and detecting the concentrations of L-cystine and DL-ATC in the eluent until the majority of DL-ATC is eluted;
the volume of each eluent and the concentration of L-cystine and DL-ATC in each eluent were respectively detected, wherein the recovery rate of DL-ATC is 80.9%, and the recovery rate of L-cystine is 90.2%. The COD and ammonia nitrogen values before and after the concentration of the L-cystine mother liquor are reduced by about 59 percent.
The two sections of eluents are respectively used in the preparation of the substrate and the refining process of the crude L-cystine, and the overall flow scheme of the embodiment is shown in figure 1.
The disclosure of the present application is only a few specific embodiments, but the present application is not limited to these, and any variations that can be made by those skilled in the art are intended to fall within the scope of the present application.
Claims (6)
1. A method for recycling L-cystine mother liquor in enzymatic reaction is characterized by comprising the following steps:
(1) evaporating the L-cystine mother liquor until the concentrations of DL-ATC and L-cystine are 5-10 g/L and 3-8 g/L respectively;
(2) adjusting the pH value of the concentrated solution of the L-cystine mother liquor to 2-4;
(3) filling cation exchange resin into a chromatographic column, loading the L-cystine mother liquor concentrated solution obtained in the step (2) onto the chromatographic column of the ion exchange resin, and stopping loading when the ion exchange resin is saturated;
(4) sequentially carrying out gradient elution on the chromatographic columns by using hydrochloric acid solutions with different concentrations, carrying out gradient elution on the chromatographic columns by using 0.1-0.3 mol/L hydrochloric acid to obtain eluted liquid, collecting and detecting the concentrations of L-cystine and DL-ATC, changing to 0.8-1.5 mol/L hydrochloric acid to obtain eluted liquid when the concentration of DL-ATC reaches 0.2g/L, and separately collecting and detecting the concentrations of L-cystine and DL-ATC in the eluted liquid until the vast majority of DL-ATC is eluted; collecting the eluent.
2. The recycling method according to claim 1, wherein the mother liquor of L-cystine contains DL-ATC 0.2-0.8 g/L and L-cystine 0.2-1.5 g/L.
3. The recycling method according to claim 1, wherein the pH of the concentrate of L-cystine mother liquor is adjusted to 3.
4. The recycling method according to any one of claims 1 to 3, wherein the cation exchange resin is a D001 type cation exchange resin or a 732 sodium type cation exchange resin.
5. The recycling method according to claim 1, wherein the eluents used in the step (4) are 0.1mol/L to 0.2mol/L hydrochloric acid solution and 1.0mol/L to 1.2mol/L hydrochloric acid solution.
6. The recycling method according to any of claims 1 to 3, wherein the DL-ATC eluate obtained in step (4) is used for substrate preparation for preparing L-cystine by enzymatic reaction of substrate containing DL-ATC as main component; the L-cystine eluent can be further refined and extracted.
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| CN105294522A (en) * | 2014-06-18 | 2016-02-03 | 四平市精细化学品有限公司 | Preparation method for D-cysteine hydrochloride monohydrate with high optical purity |
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| CN105294522A (en) * | 2014-06-18 | 2016-02-03 | 四平市精细化学品有限公司 | Preparation method for D-cysteine hydrochloride monohydrate with high optical purity |
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| 从胱氨酸母液中提取L-精氨酸工艺概述;翁连进等;《化工科技》;20041231;第12卷(第1期);第47-49页 * |
| 胱氨酸母液中L-精氨酸提取工艺研究;甘林火等;《食品工业科技》;20051231;第26卷(第3期);第157-159页 * |
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