Extraction process of L-aspartic acid

Abstract

The invention belongs to the technical field of organic chemistry, and particularly relates to an extraction process of L-aspartic acid. The extraction process of the L-aspartic acid comprises five steps of hydrolysis extraction, decoloration and purification, acidification separation, purification and extraction, and refining and impurity removal. According to the method, the utilization efficiency of resources is improved, the added value is increased, the production cost is reduced, meanwhile, the purity and the yield of the product L-aspartic acid are improved by improving the ion exchange extraction and separation method, and the safety and the market value of the product are improved.

Description

Technical Field

[0001] The invention belongs to the technical field of organic chemistry, and in particular relates to an extraction process of L-aspartic acid. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance the understanding of the overall background of the invention, and should not necessarily be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to a person skilled in the art.

[0003] L-Aspartic acid is an amino acid and one of the 20 standard amino acids. It plays an important physiological role in the body, especially in protein synthesis and metabolism. The main functions of aspartic acid include: (1) Protein synthesis: Aspartic acid is a component of protein and participates in the protein synthesis process; (2) Metabolic effect: In the body, it participates in amino acid metabolism as an important metabolic intermediate, especially in the urea cycle; (3) Neurotransmitter: Aspartic acid can act as a neurotransmitter in the nervous system, affecting the activity and signal transmission of nerve cells; (4) Synthesis of other molecules: It is also involved in the synthesis of other important amino acids and biological molecules, such as adenosine deoxyphosphate (ATP) and adenosine diphosphate (ADP).

[0004] Aspartic acid has many uses, including: (1) as an electrolyte supplement for amino acid infusion, potassium, calcium and other inorganic ion supplements, fatigue recovery agents, etc. Potassium magnesium aspartate injection or oral solution is used for arrhythmia and premature beats, tachycardia, hypokalemia, hypomagnesemia, heart failure, myocardial infarction, angina pectoris, hepatitis, cirrhosis and other diseases caused by cardiac glycoside poisoning. It is low-toxic and should not be injected without dilution. It should be used with caution in patients with renal insufficiency and atrioventricular block. (2) It can be used as an ammonia detoxifier, liver function promoter, fatigue recovery agent and other pharmaceutical products. It can be used to make sodium aspartate food additives and additives for various refreshing drinks. It can also be used as a biochemical reagent, culture medium and organic synthesis intermediate. (3) It is used in the synthesis of sweeteners. It is used in medicine to treat heart disease, as a liver function promoter, ammonia detoxifier, fatigue elimination agent and amino acid infusion component, etc. (4) Nutritional supplement, flavor enhancer. Added to various refreshing drinks. It is used in medicine as an ammonia detoxifier, liver function promoter, and fatigue recovery agent.

[0005] Generally speaking, aspartic acid can be prepared by two methods: synthesis and fermentation. The synthesis method mainly uses maleic acid or fumaric acid or their esters as raw materials, treats them with ammonia under pressure, and then hydrolyzes them. The fermentation method adds fumaric acid to ammonia under the action of enzymes to obtain the product in high yield. Many beans, grains, and vegetables are also rich in aspartic acid, including asparagus, ginger, Xanthoceras sorbifolia, green asparagus, spinach, beans, bean sprouts, pears, peaches, and meat. There are also reports on methods for directly separating and extracting aspartic acid.

[0006] The Wuhan Institute of Botany, Chinese Academy of Sciences, reported a method for separating L-aspartic acid from cottonseed meal hydrolysate. Cottonseed meal was hydrolyzed with hydrochloric acid. The resulting mixture of amino acids and hydrochloric acid was treated with cation exchange resin to adsorb the amino acids on the exchange column. Chloride ions and other impurities on the column were washed away with distilled water. Ammonia water was then used as the eluent to replace the acidic, neutral and some alkaline amino acids in groups. The eluate containing acidic amino acids was further separated with anion exchange resin to obtain L-aspartic acid. (Separation of L-arginine, L-glutamic acid and L-aspartic acid from cottonseed meal hydrolysate [J]. Pharmaceutical Industry, 1979, (10): 43-47.) Chinese patent publication number CN105294470A discloses a method for extracting rose aspartic acid from waste liquid after essential oil extraction from roses, the method comprising the following steps: a first step, pretreatment; a second step, acidification; a third step, separation and purification by cation exchange resin; a fourth step, decolorization and precipitation by activated carbon; and a fifth step, recrystallization.

[0007] The Chinese patent with publication number CN116354839A discloses a method for extracting aspartic acid from Robinia pseudoacacia. The extraction method is: using fresh Robinia pseudoacacia roots as raw materials, peeling the root bark, crushing, using 50% ethanol reflux extraction, decompressing and concentrating the extract, cooling to room temperature, and standing to obtain crude aspartic acid crystals; dissolving the crude aspartic acid crystals in boiling water, filtering, cooling to room temperature, and standing to obtain the finished aspartic acid.

[0008] Many studies have shown that spinach is rich in aspartic acid, but there is currently no report on the method of extracting aspartic acid from spinach. Summary of the invention

[0009] The invention overcomes the shortcomings of the prior art and provides a process for extracting L-aspartic acid from spinach, which improves the utilization efficiency of resources, increases added value, reduces production costs, and also improves the purity and yield of the product L-aspartic acid by improving the extraction method.

[0010] Specifically, the method for extracting L-aspartic acid from spinach is as follows: (1) Hydrolysis extraction: Wash and cut 1000 parts by weight of spinach into sections, add 2-4 times boiling water and boil for 2-5 minutes, filter and take the filtrate; (2) Decolorization and purification: The filtrate is concentrated to 1 / 5~1 / 2 of the original volume, cooled, and 0.5~1 times the mass fraction of 85%~95% ethanol is added, shaken and evenly mixed, filtered, and the solvent is removed by heating under reduced pressure rotary evaporation. The water bath temperature is controlled at 30~35℃ for 5~15min; 35~45℃ for 10~20min; 45~55℃ for 10~20min; 25~30℃ for 10~20min; 40~50℃ for 10~20min; 55~70℃ for 5~15min; 70~80℃ for 5~15min to obtain solution A; 0.005~0.012 parts by weight of calcium chloride is added to solution A, stirred evenly, and filtered to obtain solution B; (3) Acidification separation: add acid to solution B to adjust the pH to 2.0-3.0, cool, adsorb using H-type 732 cation exchange resin, rinse with deionized water for 2-4 times, elute with ammonia-glycerol-nonadienol as eluent for 3-5 times, and collect the eluate; (4) Purification and extraction: Add 0.01-0.03 parts by weight of activated carbon to the eluent for decolorization, filter, concentrate the filtrate to half of its original volume, add 0.2-0.8 times the volume of ether, extract, and take the lower layer.

[0011] (5) Refining and removing impurities: Place the lower layer of liquid in a water bath and evaporate under reduced pressure. Dry the crystals, grind them, wash them with ethanol 2 to 3 times, and dry them.

[0012] Furthermore, the acid in step (3) is one of hydrochloric acid and sulfuric acid.

[0013] Furthermore, the mass fractions of ammonia water, glycerol and nonadienol in the eluent are 4.5% to 6.5%, 0.5% to 1.5% and 0.1% to 0.5%. In a preferred embodiment, the eluent is 5.5% ammonia water-1.0% glycerol-0.4% nonadienol.

[0014] The effects of glycerol include: (1) polarity adjustment: glycerol has strong polarity and can interact with the polar groups of aspartic acid, thereby affecting the elution behavior of aspartic acid and cation exchange resin; (2) fluidity adjustment: glycerol has a certain viscosity, which can change the fluidity of the eluent, affecting the contact time with the resin and the elution efficiency. The effects of ammonia water include: (1) pH adjustment: ammonia water can adjust the pH value of the solution, change the charge state of the ion exchange resin, and affect the interaction between aspartic acid and the resin; (2) ionic strength: the ammonia ions in ammonia water can compete with the ions on the resin, thereby helping to elute aspartic acid. A suitable mixture of ammonia water and glycerol can play a synergistic role as an eluent to achieve more efficient elution. Glycerol can adjust fluidity and polarity, while ammonia water can change pH and ionic strength, thereby better eluting aspartic acid. It can also improve selectivity: by adjusting the ratio of glycerol and ammonia water, the properties of the eluent can be finely adjusted, thereby achieving more accurate separation of the target compound and improving specific selectivity and purity. In addition, it can reduce nonspecific interactions, reduce background interference, and avoid competition between some other compounds and resins, thereby increasing the yield and purity of aspartic acid. Finally, stability can also be improved: the mixture of glycerol and ammonia water can make the eluent more stable, reduce fluctuations that may occur during the elution process, and improve the consistency and repeatability of the separation process. However, nonadienol has good solubility, especially for some non-polar and weakly polar compounds. In addition, nonadienol can also reduce the surface tension of the liquid, which helps to improve the elution efficiency. Through continuous attempts, the inventor unexpectedly discovered that nonadienol can effectively synergize with ammonia water to improve the desorption ability, thereby improving the separation efficiency. Ammonia water provides an alkaline environment, which can enhance the selective elution of certain compounds. The combination with nonadienol can be optimized for specific target molecules.

[0015] Furthermore, for sufficient elution, the elution flow rate is 110-160 ml / min. In a preferred embodiment, the elution flow rate is 130 ml / min.

[0016] In a preferred extraction scheme, the filtrate after cooling in step (1) is concentrated to 1 / 4 to 1 / 3 of the original volume.

[0017] Compared with the prior art, the technical effects of the present invention are: It is well known that spinach contains L-aspartic acid, which is soluble in boiling water. The present invention utilizes waste water from blanching spinach to extract L-aspartic acid, which can reduce the waste of raw materials, improve the utilization efficiency of resources, and be used to develop new products or applications, thereby increasing added value. By recycling the effective ingredients in the waste water, the demand for new raw materials can be reduced, thereby reducing production costs.

[0018] The present invention combines the prior art method for extracting L-aspartic acid from natural products, explores and improves an ion exchange extraction method suitable for spinach wastewater, improves the extraction efficiency and purity of the product, improves the safety of the product, and increases the production capacity and market value. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 : Infrared characterization of the product of Example 1.

[0020] Figure 2 : Mass spectrometry characterization of the product of Example 1.

[0021] Figure 3 : The yield and content of L-aspartic acid obtained from groups A to H.

[0022] Figure 4 : Product purity of Examples 1~3 and Comparative Examples 1~6. DETAILED DESCRIPTION

[0023] In order to make the purpose and technical solution of the present invention clearer, the present invention is further described below in conjunction with embodiments, but the protection scope of the present invention is not limited to these embodiments, and the embodiments are only used to explain the present invention. It should be understood by those skilled in the art that any changes or equivalent substitutions that do not deviate from the concept of the present invention are included in the protection scope of the present invention.

[0024] Study on single factor experiment of eluent In order to improve the purity and yield of L-aspartic acid, the eluent in the ion exchange method was optimized and screened. The other methods were the same as in Example 1, and the variable factor was the eluent, which was expressed in mass fraction.

[0025] A: 4.5% ammonia-0.5% glycerol-0.0% nonadienol solution B: 5.0% ammonia water-0.0% glycerol 0.4% nonadienol solution C: 5.5% ammonia-0.2% glycerol-0.8% nonadienol solution D: 6.0% ammonia-0.5% glycerol-0.1% nonadienol solution E: 6.5% ammonia-1.0% glycerol-0.3% nonadienol solution F: 5.0% ammonia-1.5% glycerol-0.5% nonadienol solution G: 5.0% ammonia-2.5% glycerol-1.0% nonadienol solution H: 5.0% ammonia-3.5% glycerol-1.5% nonadienol solution Figure 3The yield and content (purity) of L-aspartic acid obtained from Group A to Group H. The purity of L-aspartic acid extracted with 4.5% ammonia water as the eluent was 98.2% and the yield was 0.201g. In order to improve the purity and yield of L-aspartic acid, the eluent was improved, and glycerol and nonadienol were added in appropriate amounts. The suitable concentration of glycerol was screened to be 0.5%~1.5% by mass fraction, and the suitable mass fraction of nonadienol was 0.1%~0.5%. On the one hand, nonadienol can reduce the surface tension of the liquid, thereby helping to improve the elution efficiency. On the other hand, the polarity of glycerol can interact with the polarity of aspartic acid, affecting its elution behavior on the resin, and can also help reduce the nonspecific interaction between the target molecule and the ion exchange resin, thereby making the elution more effective. In addition, ammonia water-glycerol-nonadienol can play a synergistic role in enhancing elution. However, high concentration of glycerol will also affect the purity of L-aspartic acid and the yield of the product. Experimental screening showed that the most suitable eluents were ammonia water, glycerol, and nonadienol with mass fractions of 4.5%~6.5%, 0.5%~1.5%, and 0.1%~0.5%, respectively.

[0026] Example 1 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it well, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for three times to remove impurities. Use a mixture of 5.5% ammonia water, 1.0% glycerol, and 0.4% nonadienol as the eluent. Elution is performed four times at a flow rate of 130 ml / min, and the eluate is collected. (4) Purification and extraction: Add 20 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0027] (5) Purification and impurity removal: The lower layer liquid is placed in a water bath at 30℃~60℃ and evaporated under reduced pressure. The temperature can be increased step by step until crystals appear and no more distillates are produced. The crystals are dried, ground, washed with ethanol 2~3 times, and dried to obtain 0.235g. The product is characterized by infrared spectroscopy and mass spectrometry. Figure 1 and Figure 2 .

[0028] Example 2 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 2 times boiling water and boil for 2 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 2 of its original volume, cool it, add 0.5 times the amount of 85% ethanol by mass, shake it, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min; 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 5g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.0. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water twice to remove impurities. Use a mixture of 4.5% ammonia water-0.5% glycerol-0.1% nonadienol as the eluent at a flow rate of 110 ml / min for two elutions, and collect the eluate. (4) Purification and extraction: Add 10 g of activated carbon to the eluent for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.2 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0029] (5) Purification and impurity removal: The lower layer liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no more distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.221 g.

[0030] Example 3 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 4 times boiling water and boil for 5 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 5 of its original volume, cool it, add 1 times the amount of 95% ethanol by mass, shake it evenly, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 12g of calcium chloride to solution A, stir it evenly, filter out the precipitate, and obtain solution B; (3) Acidification separation: add sulfuric acid to solution B to adjust the pH to 3.0. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for 4 times to remove impurities. Use a mixture of 6.5% ammonia water, 1.5% glycerol and 0.5% nonadienol as the eluent. Elution is performed 5 times at a flow rate of 160 ml / min, and the eluate is collected. (4) Purification and extraction: Add 30 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.8 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0031] (5) Refining and impurity removal: The lower layer liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no more distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.240 g.

[0032] Comparative Example 1 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it evenly, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it evenly, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for three times to remove impurities. Use a mixture of 5.5% ammonia water, 1.0% glycerol, and 0.4% nonadienol as the eluent. Elution is performed four times at a flow rate of 130 ml / min, and the eluate is collected. (4) Purification and extraction: Add 0.02 times the mass of activated carbon to the eluent for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0033] (5) Purification and impurity removal: The lower layer of liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no more distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.173 g.

[0034] Comparative Example 2 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it well, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for three times to remove impurities. Use 4.0% ammonia water as eluent at a flow rate of 130 ml / min for four elutions, and collect the eluate. (4) Purification and extraction: Add 20 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0035] (5) Refining and impurity removal: The lower layer liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no more distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.181 g.

[0036] Comparative Example 3 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it well, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for three times to remove impurities. Use a mixture of 5.5% ammonia water and 1.5% potassium chloride as the eluent at a flow rate of 130 ml / min for four elutions, and collect the eluate. (4) Purification and extraction: Add 20 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0037] (5) Refining and impurity removal: The lower layer liquid is placed in a water bath and evaporated under reduced pressure at 30°C~60°C. The temperature can be increased step by step until crystals appear and no distillate is produced. The crystals are dried, ground, washed with ethanol 2~3 times, and dried to obtain 0.177g.

[0038] Comparative Example 4 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it well, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption, rinse with deionized water for 3 times, use pH 2.2, 0.1 mol / L citric acid buffer as eluent, elution flow rate is 130 ml / min, perform elution 4 times, and collect the eluate; (4) Purification and extraction: Add 20 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0039] (5) Purification and impurity removal: The lower layer liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.152 g.

[0040] Comparative Example 5 L-Aspartic Acid Extraction Process (1) Hydrolysis extraction: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and set aside the filtrate; (2) Decolorization and purification: Heat the filtrate and concentrate it to 1 / 4 of its original volume, cool it, add 0.8 times the amount of 90% ethanol by mass, shake it well, filter it, and remove the solvent by vacuum distillation. The water bath heating program for vacuum distillation is: 30~35℃, 5~15min; 35~45℃, 10~20min; 45~55℃, 10~20min, 25~30℃, 10~20min; 40~50℃, 10~20min; 55~70℃, 5~15min; 70~80℃, 5~15min to obtain solution A; add 8g of calcium chloride to solution A, stir it well, filter out the precipitate, and obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5. After cooling, use H-type 732 cation exchange resin for adsorption. Rinse with deionized water for three times to remove impurities. Use a mixture of 5.5% ammonia water, 1.0% sodium hyaluronate, and 0.4% nonadienol as the eluent. The elution flow rate is 130 ml / min. Perform elution four times and collect the eluate. (4) Purification and extraction: Add 20 g of activated carbon to the eluate for decolorization, filter, heat and concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, let stand, and take the lower layer of liquid.

[0041] (5) Purification and impurity removal: The lower layer liquid is placed in a water bath and subjected to reduced pressure rotary evaporation at 30°C to 60°C. The temperature can be increased step by step until crystals appear and no distillate is produced. The crystals are dried, ground, washed with ethanol 2 to 3 times, and dried to obtain 0.114 g.

[0042] Comparative Example 6 L-Aspartic Acid Extraction Process (1) Pretreatment: weigh 1000 g of spinach, wash and cut into sections, add 3 times boiling water and boil for 3 min, filter and obtain a mixed solution; (2) Acidification: Add concentrated hydrochloric acid to the primary mixed solution and adjust the pH to 1.5-2.5 to obtain a secondary mixed solution; (3) Separation and purification: Place H-type 732 cation exchange resin in a 200-400 mesh filter cloth, place the filter cloth in the secondary mixed solution, stir the secondary mixed solution until the exchange adsorption is saturated, wash the 732 cation exchange resin with deionized water until the ninhydrin reaction is negative; wet-pack the 732 cation exchange resin column, elute the 732 cation exchange resin column with 4% ammonia solution at an elution rate of 1.0 l / h, collect the tertiary mixed solution when the ninhydrin reaction is positive; concentrate the tertiary mixed solution under reduced pressure to obtain a quaternary mixed solution; (4) Decolorization and precipitation: Add 1% activated carbon to the fourth mixed solution for decolorization for 20 minutes, and filter to obtain a fifth mixed solution; add 1 / 4 volume of ethanol to the fifth mixed solution, slowly stir until cooled, place at 4-7°C for 36-48 hours, filter, wash the filter cake with anhydrous ethanol, and dry to obtain a light yellow crude amino acid; (5) Recrystallization: Add the crude amino acid and 50 times the mass of deionized water to a round-bottom flask equipped with a condensation reflux device, heat and reflux, wait for the crude amino acid to dissolve, add anhydrous ethanol to the round-bottom flask at 70-80°C, stop adding anhydrous ethanol when the solution is saturated, stop heating, seal and refrigerate at 4-8°C, add aspartic acid seed crystals, continue to refrigerate at 0-4°C for 12-24h, filter, wash the crystals with 0-4°C anhydrous ethanol, dry at 60°C for 2h to obtain light yellow crystals, and repeat this process once.

[0043] Example 1~3 Product Detection According to GB 29938 / AJI92, the products obtained from the L-aspartic acid extraction process of Examples 1 to 3 were tested and evaluated in various aspects.

[0044] Table 1 Aspartic acid detection report obtained in Examples 1 to 3 Table 1 shows that the L-aspartic acid prepared by the methods of Examples 1 to 3 of the present invention has high purity and meets the GB 9938 / AJI92 standard, indicating that the method is reliable and practical.

[0045] Purity of the extracted products of Examples 1 to 3 and Comparative Examples 1 to 4 The purity of the products extracted from Examples 1 to 3, Comparative Examples 1 to 5, and the product obtained by the method disclosed in Chinese Patent CN105294470 B (Comparative Example 6) was determined by GB 9938 / AJI92 standard. Figure 4 It is shown that 5 parallel experiments were conducted for each embodiment, comparative example and prior art patent group to verify the stability and accuracy of the method.

Claims

1. A process for extracting L-aspartic acid, characterized in that: The extraction process comprises the following steps: (1) Hydrolysis extraction: Wash and cut 1000 parts by weight of spinach into sections, add 2-4 times boiling water and boil for 2-5 minutes, filter and take the filtrate; (2) Decolorization and purification: The filtrate is concentrated to 1 / 5 to 1 / 2 of the original volume, cooled, and 0.5 to 1 times the amount of 85% to 95% ethanol is added, shaken and evenly mixed, filtered, and heated by vacuum rotary evaporation to remove the solvent to obtain solution A; 0.005 to 0.012 parts by weight of calcium chloride is added to solution A, stirred and evenly mixed, and filtered to obtain solution B; (3) Acidification separation: add acid to solution B to adjust the pH to 2.0-3.0, cool, use a cation exchange column for adsorption, rinse with deionized water for 2-4 times to remove impurities, use ammonia-glycerol-nonadienol as the eluent, perform elution 3-5 times, and collect the eluate; (4) Purification and extraction: Add 0.01-0.03 parts by weight of activated carbon to the eluent for decolorization, filter, concentrate the filtrate to half of its original volume, add 0.2-0.8 times the volume of ether, extract, and take the lower layer; (5) Refining and removing impurities: Place the lower layer of liquid in a water bath and evaporate under reduced pressure. Dry the crystals, grind them, wash them with ethanol 2 to 3 times, and dry them.

2. The extraction process of L-aspartic acid according to claim 1, characterized in that: The reduced pressure rotary evaporation heating program in step (2) is: 30-35°C, 5-15 min; 35-45°C, 10-20 min; 45-55°C, 10-20 min; 25-30°C, 10-20 min; 40-50°C, 10-20 min; 55-70°C, 5-15 min; 70-80°C, 5-15 min.

3. The extraction process of L-aspartic acid according to claim 1, characterized in that, The acid in step (3) is one of hydrochloric acid and sulfuric acid.

4. The extraction process of L-aspartic acid according to claim 1, characterized in that: The mass fractions of ammonia water, glycerol and nonadienol in the eluent are 4.5%-6.5%, 0.5%-1.5% and 0.1%-0.5%.

5. The extraction process of L-aspartic acid according to claim 4, characterized in that, The eluent is 5.5% ammonia water-1.0% glycerol-0.4% nonadienol.

6. The extraction process of L-aspartic acid according to claim 1, characterized in that: The cation exchange column is H-type 732 cation exchange resin.

7. The extraction process of L-aspartic acid according to claim 1, characterized in that: The elution flow rate is 110-160 ml / min.

8. The process for extracting L-aspartic acid according to claim 1 or 7, characterized in that: The elution flow rate is 130 ml / min.

9. The extraction process of L-aspartic acid according to claim 1, characterized in that: The filtrate after cooling in the step (1) is concentrated to 1 / 4 to 1 / 3 of the original volume.

10. The process for extracting L-aspartic acid according to claim 1, characterized in that: The extraction process comprises the following steps: (1) Hydrolysis extraction: Wash and cut 1000 parts by weight of spinach into sections, add 3 times boiling water and boil for 3 min, filter and take the filtrate; (2) Decolorization and purification: The filtrate was concentrated to 1 / 4 of the original volume, cooled, and 0.8 times the mass fraction of 90% ethanol was added, shaken and evenly mixed, filtered, and the solvent was removed by vacuum rotary evaporation. The heating program was: 30-35°C, 5-15 min; 35-45°C, 10-20 min; 45-55°C, 10-20 min; 25-30°C, 10-20 min; 40-50°C, 10-20 min; 55-70°C, 5-15 min; 70-80°C, 5-15 min to obtain solution A; 0.008 parts by weight of calcium chloride was added to solution A, stirred evenly, and filtered to obtain solution B; (3) Acidification separation: Add hydrochloric acid to solution B to adjust the pH to 2.5, cool, and use H-type 732 cation exchange resin for adsorption. Rinse with deionized water to remove impurities three times, use 5.5% ammonia water-1.0% glycerol-0.4% nonadienol as eluent, elute four times at a flow rate of 130 ml / min, and collect the eluate; (4) Purification and extraction: Add 0.02 parts by weight of activated carbon to the eluate for decolorization, filter, concentrate the filtrate to half of its original volume, add 0.6 times the volume of ether, extract, and take the lower layer; (5) Refining and removing impurities: Place the lower layer of liquid in a water bath at 30℃~60℃ and evaporate under reduced pressure. Dry the crystals, grind them, wash them with ethanol 2~3 times, and dry them.

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