Preparation and purification method of carboxymethyl chitin

Abstract

The invention discloses a preparation and purification method of carboxymethyl chitin, which comprises the following steps: crushing a proper amount of blocky chitin raw material, adding the crushed blocky chitin raw material into an alcohol solution, fully stirring to fully swell the chitin, adding a pre-prepared alkali solution into the solution, carrying out alkalization reaction, adding pre-prepared carboxylation reaction liquid in batches, and carrying out solid-liquid separation to obtain the carboxymethyl chitin. The preparation method comprises the following steps: fully carrying out carboxymethylation reaction, standing, removing supernate, filtering to obtain a crude carboxymethyl chitin product, carrying out alkali-assisted alcohol precipitation, adding the precipitate into a low-concentration alcohol solution, adjusting the pH value to be neutral by using acid in a stirring state, adding a high-concentration alcohol solution, uniformly stirring, filtering, removing supernate and the like. According to the method, a traditional heterogeneous reaction system preparation process is optimized, the alkali-assisted alcohol precipitation and step-by-step elution technology is utilized for purification treatment, the dosage of ethanol is saved, the problems that in the prior art, carboxymethyl chitin is high in purification cost, high in salt content and the like are solved, and the process is simple, efficient, low in cost, good in purification effect and suitable for industrial production.

Description

Technical Field

[0001] The invention belongs to the technical field of biomaterials and relates to a method for preparing and purifying carboxymethyl chitosan. Background Art

[0002] Chitin (CT), also known as chitin and chitin, is widely found in the shells of shrimps, crabs and insects, as well as in the cell walls of fungi and plants. It is a natural organic compound second only to cellulose in number on Earth, with good biocompatibility, biodegradability and antibacterial properties. However, there are strong hydrogen bonds within and between chitin molecules, and it is only soluble in concentrated acids, concentrated alkalis and some organic solvents, which greatly limits the development and application of chitin. Therefore, it is necessary to develop its water-soluble derivatives to expand its application range. Carboxymethyl chitin (CMC), as an important chitin derivative, introduces carboxymethyl groups into chitin molecules to optimize the hydrophilic properties of chitin, greatly improving the moisture retention and film-forming properties of chitin. At the same time, this water-soluble chitin derivative inherits many important functional properties of chitin, such as good antibacterial properties, strong anti-infection ability, lipid-lowering and fresh-keeping, etc. It is widely used in beauty, daily chemicals, medicine, textiles, environmental protection, health products and other fields.

[0003] The preparation of carboxymethyl chitosan is generally divided into homogeneous system and heterogeneous system. In the homogeneous system, the raw materials are completely dissolved in the reaction liquid for preparation. This method can easily lead to a serious decrease in molecular weight, affect the biological properties of the material, and the process is complicated. Therefore, industrial preparation usually adopts a solid-liquid heterogeneous system for reaction. This reaction method can easily lead to incomplete reaction and requires purification by elution process.

[0004] At present, carboxymethyl chitosan is often purified and impurized by conventional alcohol precipitation, but this purification method consumes a large amount of ethanol and is difficult to meet industrial production. For example, the patent number "CN 114106217 B" is named "a method for preparing 6-O-carboxymethyl chitosan with high degree of substitution", the reaction is completed and the product is filtered, washed with anhydrous ethanol, then dissolved in water and an acidic solution is added to adjust the pH value to neutral, stirred and dissolved, and then washed and extracted repeatedly with anhydrous ethanol, centrifuged, and freeze-dried to obtain 6-O-carboxymethyl chitosan. The patent adopts conventional alcohol precipitation to purify, but elution and alcohol precipitation will consume a large amount of ethanol. As described in Examples 1 to 4, 1g of chitosan raw material corresponds to 200mL of water for dissolution and pH adjustment and alcohol precipitation. Only this step requires at least 600mL of anhydrous ethanol to achieve raw material alcohol precipitation, which is too costly and cannot be adapted to industrial preparation.

[0005] Patent No. "CN 110734505 B" is named "A method for preparing carboxymethyl chitosan". Carboxymethyl chitosan is prepared by a secondary reaction method. The crude carboxymethyl chitosan is dissolved in a mixture of physiological saline and water in a volume ratio of (4-8): (11-15), filtered after dissolution, and anhydrous ethanol is dripped into the mixture. When a white precipitate begins to appear in the solution, the dripping is stopped, and then the solution is dripped into the remaining anhydrous ethanol, so that the final volume ratio of the solution to anhydrous ethanol is (0.5-1): (3-6); the precipitate is collected, washed, and dried to obtain the finished carboxymethyl chitosan. Among them, 5g of the secondary reaction crude product consumes 250mL of 80% ethanol and 250mL of 90% ethanol for washing, and 1500mL of anhydrous ethanol is consumed for alcohol precipitation. The process is complicated, the time cost and the raw material cost are too high, and it cannot meet the requirements of industrial preparation.

[0006] The invention optimizes the preparation process of the traditional heterogeneous reaction system, and uses alkali-assisted alcohol precipitation and step-by-step elution technology for purification, which effectively saves the amount of ethanol used and solves the problems of high purification cost and high salt content of carboxymethyl chitosan in the prior art. The process is simple, efficient, low-cost, and has good purification effect, and is suitable for industrial production. Summary of the invention

[0007] In view of the problems existing in the prior art, the present invention provides a method for preparing and purifying carboxymethyl chitosan. The method effectively connects the preparation process and the purification process, alkali-assisted alcohol precipitation, and combines step-by-step elution to effectively save the amount of ethanol, thereby overcoming the problems of high purification cost and high salt content of carboxymethyl chitosan in the prior art.

[0008] The specific technical solutions of the present invention are as follows:

[0009] The invention discloses a method for preparing and purifying carboxymethyl chitosan, which comprises the following steps:

[0010] 1) Pretreatment: Take the blocky chitosan raw material, crush it, add it into the alcohol solution and stir it thoroughly to make the chitosan powder fully swell;

[0011] 2) Alkalization: adding a pre-prepared alkaline solution to the above solution to carry out an alkalization reaction;

[0012] 3) Carboxymethylation: After the alkalization reaction is completed, the pre-prepared carboxylation reaction solution is added in batches to fully carry out the carboxymethylation reaction. After standing, the supernatant is removed and filtered to obtain a crude carboxymethyl chitosan product;

[0013] 4) Alkali-assisted alcohol precipitation: add an appropriate amount of purified water to the crude carboxymethyl chitosan under stirring, and after it is completely dissolved, purify and filter 1 to 3 times, remove the filter residue, slowly add alcohol solution under stirring, and filter and collect the precipitate after the alcohol precipitation is complete;

[0014] 5) Stepwise elution: add the above precipitate to a low concentration alcohol solution, adjust the pH to 6-8 with acid under stirring, then add a high concentration alcohol solution, stir evenly and filter, remove the supernatant; add a high concentration alcohol solution again, stir evenly and filter, remove the supernatant;

[0015] 6) Post-treatment: After elution, add anhydrous ethanol for dehydration, filter, and dry to obtain the pure product.

[0016] As a further improvement, in step 1) of the present invention, the chitosan is crushed to a mesh size of 20 to 60 mesh; the mass ratio of chitosan to alcohol solution is 1:(6 to 12); the pretreatment stirring time is 0.5 h to 2.5 h, and the reaction temperature is 20° C. to 65° C.

[0017] As a further improvement, in step 1) of the present invention, the alcohol solution is one or both of isopropanol and ethanol, and the alcohol concentration is 85% to 100%.

[0018] As a further improvement, in step 2) of the present invention, the mass ratio of chitosan to alkaline solution is 1:(2.5-5); the alkalization reaction time is 1h-2.5h, and the reaction temperature is 20°C-65°C.

[0019] As a further improvement, in step 2) of the present invention, the alkaline solution is composed of an alkali and water, and the alkali is one or both of sodium hydroxide and potassium hydroxide, and the concentration is 40% to 60%.

[0020] As a further improvement, in step 3) of the present invention, the mass ratio of chitosan to carboxylation reaction liquid is 1:(4.5-8); when adding the carboxylation reaction liquid, the temperature needs to be appropriately lowered to control the reaction system temperature not higher than 65°C. After the carboxylation reaction liquid is added, the reaction is stirred for 0.5h-2h, and the reaction temperature is 20°C-65°C.

[0021] As a further improvement, in step 3) of the present invention, in the carboxylation reaction solution, the solute is one or both of chloroacetic acid and sodium chloroacetate, with a concentration of 15% to 35%; and the solvent is one or both of isopropanol and anhydrous ethanol.

[0022] As a further improvement, in step 4) of the present invention, the mass ratio of chitosan, purified water and alcohol solution is 1:(40-100):(30-80); the alcohol solution is one or both of isopropanol and ethanol, and the alcohol concentration is 85%-100%.

[0023] As a further improvement, in step 4) of the present invention, the pore size of the filter element or filter disc used for purification filtration is 25 μm to 75 μm.

[0024] As a further improvement, in step 5) of the present invention, the alcohol solution is one or both of isopropanol and ethanol, which can be divided into low concentration 50% to 80% and high concentration 85% to 100% according to the alcohol concentration. The mass ratio of chitosan to the low concentration alcohol solution and the high concentration alcohol solution is 1:(6 to 10):(6 to 10); in step 6), the mass ratio of chitosan to anhydrous ethanol is 1:(6 to 10).

[0025] Compared with the prior art, the present invention has the following beneficial effects:

[0026] 1. The present invention preferably optimizes the reaction process on a heterogeneous basis, disperses the raw materials by pretreatment and fully swells them, sets the stirring time to 0.5h to 2.5h, and the temperature to 20°C to 65°C, so as to avoid the strong permeability of the high-concentration alkaline solution causing local damage to the molecular structure of chitin, make the alkalinization reaction more uniform, improve the efficiency of subsequent reactions, and shorten the preparation time. At the same time, the degradation of the raw materials due to long-term alkaline treatment is avoided, and the stability and uniformity of the product are improved.

[0027] 2. The present invention adopts a medium-low temperature reaction system to carry out carboxymethylation preparation at a temperature of 20°C to 65°C, and performs temperature-controlled carboxymethylation in steps to avoid the generation of a large number of by-products due to high-temperature reactions, and better retains the C2 acetyl group of chitin, reducing the shedding of the C2 acetyl group, which is beneficial to improving the safety, effectiveness and biocompatibility of the product.

[0028] 3. The present invention utilizes the strong alkaline conditions after the carboxymethyl reaction to assist in purification. The aqueous solution after the crude product is dissolved is strongly alkaline. Compared with neutral conditions, carboxymethyl chitosan has better solubility and lower viscosity, which can reduce filter element clogging during purification and improve filtration efficiency.

[0029] 4. During purification, the present invention requires less purified water to completely dissolve the crude product, and the concentration of the crude product aqueous solution is increased. Under the dual influence of alkaline conditions and the concentration of the product aqueous solution, the intermolecular electrostatic effect is significantly enhanced, and the amount of alcohol required to induce alcohol precipitation is greatly reduced. The mass ratio of chitosan, purified water and alcohol solution is 1: (40-100): (30-80), and the product is further purified by combining step-by-step elution. The process is simple to operate, low in cost and high in product purity.

[0030] 5. In terms of mass ratio, the amount of ethanol (95%) used in the neutral alcohol precipitation process for preparing carboxymethyl chitosan is 105 times that of chitosan, while the amount of ethanol (95%) used in the alkali-assisted alcohol precipitation process is only 57 times the mass of chitosan, and the latter has obvious cost advantages. Compared with the traditional process, the preparation process of the present invention is short in time, uses less ethanol, and effectively saves ethanol. The traditional preparation process of the heterogeneous reaction system is optimized, and the purification process is carried out by using alkali-assisted alcohol precipitation and step-by-step elution technology, which overcomes the problems of high purification cost and high salt content of carboxymethyl chitosan in the prior art. It is not easy to be blocked during purification and filtration, and has the advantages of simple process, low cost, good purification effect, etc., and is suitable for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a process flow chart of this application.

[0032] Figure 2 This is the infrared spectrum of carboxymethyl chitosan prepared in Example 3. DETAILED DESCRIPTION

[0033] The present invention uses an alcohol solution as a dispersant to perform swelling treatment on chitosan, so that the inside of chitosan molecules can be fully infiltrated, the dispersibility of chitosan raw materials in the system is maintained, the uneven penetration of the alkaline solution during subsequent alkalization is alleviated, and the uniformity of alkalization is improved; during carboxymethylation, a neutralization reaction occurs when the carboxylation reaction liquid contacts the alkali in the system to release a large amount of heat, and when the temperature is too high, the acetyl group in the chitosan is easy to be separated, and the side reaction products increase at the same time, so the carboxylation reaction liquid needs to be added in batches, and the system temperature is controlled at the same time. Heat treatment is avoided throughout the preparation of carboxymethyl chitosan, the deacetylation degree of the product and the by-products are reduced, the C2 acetyl group is effectively protected, and the carboxymethyl substitution is biased towards the C6 and C3 positions.

[0034] Under alkaline conditions, the carboxymethyl group (-COOH) in carboxymethyl chitin is more easily ionized to form carboxymethyl anions (-COO-), increasing its solubility in water. Under neutral conditions, the carboxymethyl group (-COOH) of carboxymethyl chitin is not as ionized as under alkaline conditions, and its solubility is lower. During alcohol precipitation, the amount of alcohol used is related to the solubility of carboxymethyl chitin in the solution and the interaction between molecules. Under alkaline conditions, carboxymethyl chitin has better solubility, stronger electrostatic repulsion between molecules, and molecules are easier to disperse in water, so less ethanol is needed to induce precipitation. Under neutral conditions, molecules tend to aggregate more, so more alcohol solution is needed to break this aggregation state and promote the formation of precipitation.

[0035] Step-by-step elution utilizes the solubility characteristics of soluble impurities and uses a low-concentration alcohol solution with a high water content to elute and dissolve the soluble impurities. However, carboxymethyl chitosan in a low-concentration alcohol system is not easy to dissolve due to its strong electrostatic repulsion. It is then eluted and filtered with high-concentration ethanol to achieve further purification of carboxymethyl chitosan.

[0036] The main chemical reaction formula involved in the present invention is as follows:

[0037]

[0038] To further understand the present invention, the preparation and purification method of carboxymethyl chitosan provided by the present invention is specifically described below in conjunction with the embodiments, but the present invention is not limited to these embodiments, and non-essential improvements and adjustments made by technicians in this field under the core guiding idea of ​​the present invention still belong to the protection scope of the present invention.

[0039] Example 1

[0040] Preparation of carboxymethyl chitin: Weigh 2.0 kg of chitin powder after 20 mesh sieve, add to 16 kg of isopropanol, stir at room temperature for 2 hours. Add 7.2 kg of 50% sodium hydroxide aqueous solution, react at room temperature for 1.5 hours; under the above alkaline conditions, add 12 kg of 35% chloroacetic acid-isopropanol solution in batches, cool in a water bath, control the solution temperature within 45°C, add within 1 hour, react at room temperature for 1 hour after adding liquid, filter and remove the supernatant.

[0041] Purification of carboxymethyl chitosan: Add 100 kg of purified water to the filtered product, stir and dissolve; filter twice with a 45 μm filter element to remove the filter residue; slowly add 64 kg of 95% ethanol while stirring, let stand, and discard the supernatant; add 12 kg of 70% ethanol to the product, adjust the pH to 6-8 with hydrochloric acid, add 12 kg of 95% ethanol after the pH value stabilizes, stir and filter, and remove the filtrate; add 12 kg of 95% ethanol to the filtered product, stir and filter, and remove the filtrate; dehydrate the product with 16 kg of anhydrous ethanol 1-2 times, filter, and dry in a vacuum at 60°C.

[0042] Example 2

[0043] Preparation of carboxymethyl chitin: Weigh 2.0 kg of chitin powder after 20 mesh sieve, add to 16 kg of isopropanol, stir at room temperature for 2 hours. Add 7.2 kg of 50% sodium hydroxide aqueous solution, react at room temperature for 1.5 hours; under the above alkaline conditions, add 12 kg of 35% chloroacetic acid-isopropanol solution in batches, cool in a water bath, control the solution temperature within 25°C, add within 1 hour, react at room temperature for 1 hour after adding liquid, filter and remove supernatant.

[0044] Purification of carboxymethyl chitosan: Add 200 kg of purified water to the filtered product, stir and dissolve; filter twice with a 45 μm filter element to remove the filter residue; slowly add 160 kg of 95% ethanol while stirring, let stand, and discard the supernatant; add 12 kg of 70% ethanol to the product, adjust the pH to 6-8 with hydrochloric acid, add 12 kg of 95% ethanol after the pH value stabilizes, stir and filter, and remove the filtrate; add 12 kg of 95% ethanol to the filtered product, stir and filter, and remove the filtrate; dehydrate the product with 16 kg of anhydrous ethanol 1-2 times, filter, and dry in a vacuum at 60°C.

[0045] Example 3

[0046] Preparation of carboxymethyl chitin: Weigh 2.0 kg of chitin powder after 20 mesh sieve, add to 16 kg of isopropanol, and react at 65°C for 0.5 h. Add 7.2 kg of 50% sodium hydroxide aqueous solution, and react at 65°C for 1 h. Under the above alkaline conditions, add 12 kg of 35% chloroacetic acid-isopropanol solution in batches, cool in a water bath, control the solution temperature within 65°C, add within 1 h, react at 65°C for 1 h after adding, filter, and remove the supernatant.

[0047] Purification of carboxymethyl chitosan: Add 100 kg of purified water to the filtered product, stir and dissolve; filter twice with a 45 μm filter element to remove the filter residue; slowly add 64 kg of 95% ethanol while stirring, let stand, and discard the supernatant; add 12 kg of 70% ethanol to the product, adjust the pH to 6-8 with hydrochloric acid, add 12 kg of 95% ethanol after the pH value stabilizes, stir and filter, and remove the filtrate; add 12 kg of 95% ethanol to the filtered product, stir and filter, and remove the filtrate; dehydrate the product with 16 kg of anhydrous ethanol 1-2 times, filter, and vacuum dry at 60°C.

[0048] Example 4

[0049] Preparation of carboxymethyl chitin: Weigh 2.0 kg of chitin powder after 20 mesh sieve, add to 16 kg of isopropanol, and react at 65°C for 0.5 h. Add 7.2 kg of 50% sodium hydroxide aqueous solution, and react at 65°C for 1 h. Under the above alkaline conditions, add 12 kg of 35% chloroacetic acid-isopropanol solution in batches, cool in a water bath, control the solution temperature within 65°C, add within 1 h, react at 65°C for 1 h after adding, filter, and remove the supernatant.

[0050] Purification of carboxymethyl chitosan: Add 200 kg of purified water to the filtered product, stir and dissolve; filter twice with a 45 μm filter element to remove the filter residue; slowly add 160 kg of 95% ethanol while stirring, let stand, and discard the supernatant; add 12 kg of 70% ethanol to the product, adjust the pH to 6-8 with hydrochloric acid, add 12 kg of 95% ethanol after the pH value stabilizes, stir and filter, and remove the filtrate; add 12 kg of 95% ethanol to the filtered product, stir and filter, and remove the filtrate; dehydrate the product with 16 kg of anhydrous ethanol 1-2 times, filter, and dry in a vacuum at 60°C.

[0051] Comparative Example 1

[0052] This comparative example uses the preparation and purification methods of Example 1 and Example 3, except that: the carboxymethyl chitosan in Example 1 is prepared at room temperature, the carboxymethyl chitosan in Example 3 is prepared at 65° C., and the carboxymethyl chitosan in this comparative example is prepared at 90° C. The remaining technical features are the same as those of Example 1 and Example 3.

[0053] Comparative Example 2

[0054] This comparative example uses the preparation and purification method of Example 1, except that the chitosan raw material in this comparative example is not subjected to swelling pretreatment after being crushed. The remaining technical features are the same as those of Example 1.

[0055] Comparative Example 3

[0056] This comparative example uses the preparation method of Example 2. When purifying carboxymethyl chitosan, the pH is first adjusted to neutral and then filtered and precipitated with alcohol, that is, the alkali-assisted alcohol precipitation of the technical solution of the present invention is not used. Specifically:

[0057] Purification of carboxymethyl chitosan: Add 12kg of 95% ethanol to the filtered product to elute and remove the supernatant, repeat the above operation twice; add 200kg of purified water, stir and dissolve; adjust the pH to 6-8 with hydrochloric acid, filter twice with a 45μm filter element, and remove the filter residue; slowly add 480kg of 95% ethanol under stirring, let it stand, and discard the supernatant; add 12kg of 95% ethanol to the product, stir and filter, and remove the filtrate; add 12kg of 95% ethanol again, stir and filter, and remove the filtrate; dehydrate the product with 16kg of anhydrous ethanol 1-2 times, filter, and vacuum dry at 60°C.

[0058] Comparative Example 4

[0059] This comparative example uses the preparation method of Example 3. When purifying carboxymethyl chitosan, the pH is first adjusted to neutral and then filtered and precipitated with alcohol, that is, the alkali-assisted alcohol precipitation of the technical solution of the present invention is not used. Specifically:

[0060] Purification of carboxymethyl chitosan: Add 12kg of 95% ethanol to the filtered product to elute and remove the supernatant, repeat the above operation twice; add 100kg of purified water, stir and dissolve; adjust the pH to 6-8 with hydrochloric acid, filter twice with a 45μm filter element, and remove the filter residue; slowly add 240kg of 95% ethanol under stirring, let it stand, and discard the supernatant; add 12kg of 95% ethanol to the product, stir and filter, and remove the filtrate; add 12kg of 95% ethanol again, stir and filter, and remove the filtrate; dehydrate the product with 16kg of anhydrous ethanol 1-2 times, filter, and vacuum dry at 60°C.

[0061]

[0062] The physical and chemical performance indicators of the products obtained in the examples were tested.

[0063] (1) Molecular weight index detection

[0064] Dissolve the dry sample in 0.1mol / L sodium chloride solution, adjust the dilution multiple to ensure that the outflow time exceeds 200 seconds, and shake the solution; filter the solution using a No. 3 vertical fritted glass funnel and collect the filtrate; inject the test solution into tube 2 of the Ubbelohde viscometer, fix the viscometer in a constant temperature water bath at 25℃±0.1℃, and ensure that the water bath liquid level is higher than point C; after standing for 15 minutes, connect the latex tube to tubes 1 and 3, clamp tube 3, and draw air from tube 1 to raise the liquid level to the middle of ball C, open tube 3 first, then open tube 1, and record the outflow time T when the liquid level drops from line m1 to line m2. Repeat the measurement twice to ensure that the difference between the two times is within ±0.5%, and take the average value as the outflow time T of the test solution. The outflow time is T0. Calculate as follows:

[0065] Intrinsic viscosity η=lnηr / C

[0066] Where ηr is T / T0

[0067] C is the concentration of the test solution, g / mL

[0068] Calculate the molecular weight according to the following formula:

[0069] [η] = 1.81 × 10 ～3 M 0.93

[0070] (2) Insoluble matter index detection

[0071] According to the test method specified in 6.15 of YY / T 0953-2020, weigh 1.0 g of carboxymethyl chitosan (calculated as dry product) and stir it in 100 ml of purified water until it is completely dissolved; transfer the solution to a 1-liter beaker, add 900 ml of water, heat to a slight boil and keep for 0.5 hours. During the heating process, the beaker mouth needs to be covered. Use a constant-weight sand core funnel (No. 3) for filtration and wash the residue with water. And dry it to constant weight in an electric hot air drying oven at 100°C to 105°C.

[0072] (3) Degree of substitution test

[0073] The test shall be carried out according to the test method specified in Appendix B of YY / T 0953~2020.

[0074] (4) Deacetylation degree detection

[0075] Weigh 0.2g of sample, add 25mL of 0.3mol / L hydrochloric acid solution, and stir until the sample is completely dissolved. Titrate with 0.1mol / L sodium hydroxide titration solution. Use glass electrode as indicator and calomel electrode as reference, and record the corresponding relationship between the volume (mL) of 0.1mol / L sodium hydroxide titration solution consumed and the pH value.

[0076] The jump on the titration curve was obtained by the quadratic differential method. The 0.1 mol / L sodium hydroxide titration solution consumed between the jump points pH2 (V2) 4.2 and pH3 (V3) 8.6 was the titration of -NH4+, so the deacetylation degree of carboxymethyl chitosan could be obtained.

[0077] The calculation formula is as follows:

[0078]

[0079] Where: V3 and V2 are the volumes (mL) of standard NaOH solution consumed during the third and second jumps, respectively;

[0080] C NaOH : is the concentration of standard NaOH solution (mol / L);

[0081] W: sample weight (g).

[0082] 219: Average molecular weight of sugar group.

[0083] The test results are shown in the table below.

[0084]

[0085]

[0086] The comparison of the amount of ethanol (calculated as 95%) used in each embodiment is shown in the table below.

[0087] Ethanol dosage / kg Chitosan: ethanol (mass ratio) Example 1 114 1：57 Example 2 210 1：105 Example 3 114 1：57 Example 4 210 1：105 Comparative Example 1 114 1：57 Comparative Example 2 114 1：57 Comparative Example 3 520 1：260 Comparative Example 4 280 1：140

[0088] There is no significant difference in the molecular weight, insoluble matter and degree of substitution among the embodiments. The reaction temperature of Embodiment 3, Embodiment 4 and Comparative Example 4 is 65°C, and the deacetylation degree thereof is higher than that of the other embodiments reacted at room temperature. The reaction temperature of Comparative Example 1 is 90°C. The high reaction temperature leads to a high degree of deacetylation of the product, indicating that lowering the reaction temperature is beneficial to retaining the acetyl group in the chitin molecule.

[0089] The insoluble matter index of Comparative Example 2 is significantly higher than that of the other embodiments and comparative examples, and the degree of substitution index is significantly lower than that of the other embodiments and comparative examples, indicating that the purity of carboxymethyl chitosan prepared by this scheme is lower than that of other embodiments or comparative examples, that is, the lack of swelling pretreatment will affect the subsequent reaction efficiency.

[0090] In Comparative Example 3, after 200 kg of purified water chitosan crude product was adjusted to neutrality, the viscosity became higher, the filter pores were easily blocked during filtration, and the difficulty of purification increased. In addition, 480 kg of 95% ethanol was required for neutral alcohol precipitation, compared with the usage of 160 kg of 95% ethanol for alkali-assisted alcohol precipitation in Example 2, the process difficulty was high and the amount of ethanol used was large. Therefore, Example 2 had similar test indicators to Comparative Example 3, but was easy to operate and low in cost.

[0091] Similarly, in Example 3, only 64 kg of 95% ethanol is needed to achieve alcohol precipitation of the product during purification, which has process and cost advantages compared to 240 kg of 95% ethanol used during alcohol precipitation in Comparative Example 4.

[0092] The above are only preferred embodiments of the present invention. Although the present invention has been disclosed as a preferred embodiment, it is not intended to limit the present invention. Any technician familiar with the art can use the above disclosed methods and technical contents to make many possible changes and modifications to the technical solution of the present invention without departing from the scope of the technical solution of the present invention, or modify it into an equivalent embodiment of equivalent changes. Therefore, any simple modification, equivalent change and modification made to the above embodiments according to the technical essence of the present invention without departing from the content of the technical solution of the present invention still falls within the scope of protection of the technical solution of the present invention.

Claims

1. A method for preparing and purifying carboxymethyl chitosan, characterized in that: The method comprises the following steps: 1) Pretreatment: Take the blocky chitosan raw material, crush it, add it into the alcohol solution and stir it thoroughly to make the chitosan powder fully swell; 2) Alkalization: adding a pre-prepared alkaline solution to the above solution to carry out an alkalization reaction; 3) Carboxymethylation: After the alkalization reaction is completed, the pre-prepared carboxylation reaction solution is added in batches to fully carry out the carboxymethylation reaction. After standing, the supernatant is removed and filtered to obtain a crude carboxymethyl chitosan product; 4) Alkali-assisted alcohol precipitation: add an appropriate amount of purified water to the crude carboxymethyl chitosan under stirring, and after it is completely dissolved, purify and filter 1 to 3 times, remove the filter residue, slowly add alcohol solution under stirring, and filter and collect the precipitate after the alcohol precipitation is complete; 5) Stepwise elution: add the above precipitate to a low concentration alcohol solution, adjust the pH to 6-8 with acid under stirring, then add a high concentration alcohol solution, stir evenly and filter, remove the supernatant; add a high concentration alcohol solution again, stir evenly and filter, remove the supernatant; 6) Post-treatment: After elution, add anhydrous ethanol for dehydration, filter, and dry to obtain the pure product.

2. The method for preparing and purifying carboxymethyl chitosan according to claim 1, wherein: In step 1), the chitosan is crushed to a mesh size of 20-60 mesh; the mass ratio of chitosan to alcohol solution is 1:(6-12); the pretreatment stirring time is 0.5h-2.5h, and the reaction temperature is 20°C-65°C.

3. The method for preparing and purifying carboxymethyl chitosan according to claim 1, wherein: In step 1), the alcohol solution is one or both of isopropanol and ethanol, and the alcohol concentration is 85% to 100%.

4. The method for preparing and purifying carboxymethyl chitosan according to claim 1, 2 or 3, characterized in that: In step 2), the mass ratio of chitosan to alkaline solution is 1:(2.5-5); the alkalization reaction time is 1h-2.5h, and the reaction temperature is 20°C-65°C.

5. The method for preparing and purifying carboxymethyl chitosan according to claim 4, wherein: In step 2), the alkaline solution is composed of an alkali and water, the alkali is one or both of sodium hydroxide and potassium hydroxide, and the concentration is 40% to 60%.

6. The method for preparing and purifying carboxymethyl chitosan according to claim 5, wherein: In step 3), the mass ratio of chitosan to carboxylation reaction liquid is 1: (4.5-8); when adding the carboxylation reaction liquid, the temperature needs to be appropriately lowered to control the reaction system temperature not higher than 65°C. After the carboxylation reaction liquid is added, the reaction is stirred for 0.5h-2h, and the reaction temperature is 20°C-65°C.

7. The method for preparing and purifying carboxymethyl chitosan according to claim 6, wherein: In step 3), in the carboxylation reaction solution, the solute is one or both of chloroacetic acid and sodium chloroacetate, and the concentration is 15% to 35%; the solvent is one or both of isopropanol and anhydrous ethanol.

8. The method for preparing and purifying carboxymethyl chitosan according to claim 1, 2, 3, 5, 6 or 7, characterized in that: In step 4), the mass ratio of chitosan, purified water and alcohol solution is 1: (40-100): (30-80); the alcohol solution is one or two of isopropanol and ethanol, and the alcohol concentration is 85%-100%.

9. The method for preparing and purifying carboxymethyl chitosan according to claim 8, characterized in that: In step 4), the pore size of the filter element or filter disc used for purification filtration is 25 μm to 75 μm.

10. The method for preparing and purifying carboxymethyl chitosan according to claim 1, characterized in that: In step 5), the alcohol solution is one or both of isopropanol and ethanol, and can be divided into low concentration 50% to 80% and high concentration 85% to 100% according to the alcohol concentration. The mass ratio of chitosan to the low concentration alcohol solution and the high concentration alcohol solution is 1:(6 to 10):(6 to 10); in step 6), the mass ratio of chitosan to anhydrous ethanol is 1:(6 to 10).

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