Adsorption extraction method of chitooligosaccharide

Abstract

The application discloses an adsorption extraction method of chitosan oligosaccharide, and comprises the following steps: (1) zeolite modification: placing natural clinoptilolite in a sodium chloride solution with a certain concentration, boiling, filtering, and obtaining modified zeolite; (2) adsorption: adjusting the pH of a chitosan oligosaccharide solution to 5.5-6.5, then adding the modified zeolite, fully oscillating, and filtering to collect the modified zeolite; (3) desorption: placing the collected modified zeolite in a hydrochloric acid solution, fully oscillating, filtering, and obtaining a purified chitosan oligosaccharide solution. The application uses cheap and easily available clinoptilolite as an adsorption material to realize the extraction and purification of chitosan oligosaccharide in a solution, and has the advantages of simple operation, low cost, and the like, and has important significance for expanding the application of zeolite in the preparation and separation process of chitosan oligosaccharide and improving the preparation process of chitosan oligosaccharide.

Description

Technical Field

[0001] This invention relates to the field of marine bioengineering, and in particular to an adsorption and extraction method for chitosan oligosaccharides. Background Technology

[0002] Chitosan oligosaccharides are linear oligosaccharides composed of D-glucosamine linked by β-1,4 glycosidic bonds, containing some acetylglucosamine residues, with a degree of polymerization between 2 and 20. Chitosan oligosaccharides possess various biological activities, including antibacterial, antioxidant, antitumor, immune-enhancing, and plant growth-stimulating effects, and are widely used in biostimulants, animal feed, and health foods.

[0003] Currently, chitosan oligosaccharides are mainly obtained through chitosan degradation, primarily through chemical, enzymatic, and physical methods. During the extraction and degradation of chitosan oligosaccharides, a series of chitosan oligosaccharides with different degrees of polymerization, as well as neutral sugar molecules such as glucose, acetylglucosamine, and chitosan oligosaccharides, and other inorganic salts and impurities are generated. Separating and purifying chitosan oligosaccharides to obtain high-purity or even mono-degree-of-polymerization chitosan oligosaccharides is crucial for elucidating the structure-activity relationship between their chemical structure and biological activity. Activated carbon can adsorb and extract chitosan oligosaccharides from solution, removing some inorganic salts; however, activated carbon also adsorbs other neutral sugar molecules, thus failing to effectively purify chitosan oligosaccharides. Currently, there is no effective method for purifying and extracting chitosan oligosaccharides. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention provides an adsorption extraction method for chitosan oligosaccharides, which can adsorb and extract chitosan oligosaccharides from the solution, separating them from neutral sugars and other molecules in the solution, and achieving a high yield.

[0005] To achieve the above objectives, the technical solution of the present invention is as follows:

[0006] An adsorption extraction method for chitosan oligosaccharides includes the following steps:

[0007] (1) Zeolite modification: Natural clinoptilolite is placed in a sodium chloride solution of a certain concentration, boiled, filtered, and modified zeolite is obtained.

[0008] (2) Adsorption: Adjust the pH of the chitosan oligosaccharide solution to 5.5-6.5, then add modified zeolite, shake thoroughly, filter, and collect the modified zeolite;

[0009] (3) Desorption: The collected modified zeolite was placed in hydrochloric acid solution, shaken thoroughly, and then filtered to obtain the purified chitosan oligosaccharide solution.

[0010] In the above scheme, in step (1), the mass concentration of sodium chloride solution is 15% to 25%.

[0011] In the above scheme, in step (1), the natural clinoptilolite is modified three times using sodium chloride solution.

[0012] In the above scheme, in step (2), the amount of modified zeolite used is 10-15 times the mass of chitosan oligosaccharide in the chitosan oligosaccharide solution.

[0013] In the above scheme, the sufficient oscillation time in step (2) is 3-6 hours.

[0014] In the above scheme, in step (3), the concentration of hydrochloric acid solution is 0.5-3 mol / L.

[0015] In the above scheme, the sufficient oscillation time in step (3) is 2-6 hours.

[0016] In the above scheme, the chitosan oligosaccharide solution used in step (2) is an aqueous solution containing neutral sugar molecules.

[0017] The present invention provides an adsorption and extraction method for chitosan oligosaccharides using modified zeolite. While natural zeolite is a commonly used adsorbent, its application in the adsorption and extraction of chitosan oligosaccharides from solution has not been reported. Modified zeolite enhances ion exchange performance and also possesses the adsorption characteristics of porous molecular sieves. The inventors investigated the effects of different modification methods, including NaCl, HCl, NaOH, and cetyltrimethylammonium bromide (CTAB), on the adsorption capacity of zeolite for chitosan oligosaccharides. They found that NaCl-modified zeolite exhibited the highest yield after both adsorption and desorption of chitosan oligosaccharides. NaCl modification induces an ion exchange reaction between Na ions and cations in the zeolite, thereby activating some cations on the framework. Furthermore, the zeolite framework carries a negative charge, leaving some vacancies unfilled by metal cations. Salt modification allows metal cations to enter the zeolite channels and fill these vacancies, thus increasing the ion exchange capacity of the zeolite.

[0018] The inventors systematically studied the pH conditions and kinetic data of modified zeolite adsorption of chitosan oligosaccharides, discovering that chitosan oligosaccharides, as the only alkaline oligosaccharide in nature, exhibit a unique interaction mechanism with zeolite. Due to the presence of numerous free amino groups in the chitosan molecule, the pH requirement for adsorption with zeolite is specific. Although zeolite exhibits the highest adsorption capacity for chitosan oligosaccharides under strongly alkaline conditions, the adsorption process is irreversible, and the chitosan oligosaccharides cannot be desorbed and recovered, thus limiting practical applications. Under neutral and acidic conditions, the inventors found that zeolite achieves maximum adsorption of chitosan oligosaccharide solutions in a slightly acidic range of pH 5.5-6.5. This is related to the presence of amino groups (NH2 or NH3) in the solution carried by the chitosan oligosaccharides. + The presence of NH3 is relevant, indicating that it carries NH3. +Amino oligosaccharides with weak cationic properties are most easily adsorbed by ion exchange interaction with zeolite, while other neutral sugars generally cannot under these conditions.

[0019] This invention also limits the desorption solution because using sodium chloride solutions of different concentrations as desorbents cannot fully displace chitosan oligosaccharides from zeolites, resulting in a recovery rate of only about 40%. However, when this invention uses 0.5-3 mol / L hydrochloric acid solution for elution, the recovery rate reaches at least 75%, which is a significant improvement.

[0020] This invention targets the only naturally occurring basic amino oligosaccharide. Based on the adsorption kinetics of modified zeolite and the unique amino acid base theory of chitosan oligosaccharide, it discovers the optimal conditions for the adsorption and desorption of chitosan oligosaccharide in solution after modification of natural zeolite. A new method for efficient extraction and purification of chitosan oligosaccharide is proposed, which has a high extraction yield, is simple to operate, and has low cost. It is of great significance for optimizing and improving the preparation process of high-purity chitosan oligosaccharide and saving production costs. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings used in the description of the embodiments or the prior art will be briefly introduced below.

[0022] Figure 1 This is the high-performance liquid chromatography (HPLC) chromatogram of chitosan oligosaccharides extracted in Example 1.

[0023] Figure 2 This is the mass spectrometry analysis spectrum of the chitosan oligosaccharide extracted in Example 1. Detailed Implementation

[0024] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention.

[0025] This invention provides an adsorption and extraction method for chitosan oligosaccharides, and specific embodiments are as follows:

[0026] Example 1

[0027] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0028] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles;

[0029] (3) Desorption: The zeolite filter residue was placed in 30 ml of 2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 81%.

[0030] Example 2

[0031] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 15%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0032] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 5.5 with NaOH solution, then add 1 g of modified zeolite, shake for 3 hours and then filter quickly to collect the modified zeolite particles.

[0033] (3) Desorption: The zeolite filter residue was placed in 30 ml of 0.5 mol / L hydrochloric acid solution for desorption. After shaking for 2 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 75%.

[0034] Example 3

[0035] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 25%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0036] (2) Adsorption: Take 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.5 with NaOH solution, then add 1.5 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles.

[0037] (3) Desorption: The zeolite filter residue was placed in 30 ml of 2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 85%.

[0038] Example 4

[0039] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0040] (2) Adsorption: Take 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1 g of modified zeolite, shake for 4 hours and then filter quickly to collect the modified zeolite particles.

[0041] (3) Desorption: The zeolite filter residue was placed in 30 ml of 1 mol / L hydrochloric acid solution for desorption. After shaking for 4 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 79%.

[0042] Example 5

[0043] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 25%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0044] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1.2 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles;

[0045] (3) Desorption: The zeolite filter residue was placed in 30 ml of 3 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 88%.

[0046] Example 1: The high-performance liquid chromatogram of purified chitosan oligosaccharide is shown below. Figure 1 As shown, absorption peak A is the solvent peak, and peaks B-F correspond to the chitobiose and chitohexaose components, respectively. Mass spectrometry analysis is as follows: Figure 2 As shown, 341.16, 502.23, 663.30, and 824.37 correspond to the molecular ion peaks [M+H] of chitobiose-chitopentose, respectively. + 544.24 and 705.31 are the molecular ion peaks [M+H] of chitosan trisaccharide and chitosan tetrasaccharide containing one acetyl group, respectively. + 251.62, 332.15, 412.69, and 493.22 correspond to the [M+2H] of chitosan-chitosan respectively. 2+ 353.16 and 433.69 are the [M+2H] values ​​of chitosan tetrasaccharide and chitosan pentasaccharide, respectively, containing one acetyl group. 2+ No molecular ion peak of neutral sugar was observed in the mass spectrometry. The high-performance liquid chromatograms and mass spectra of the purified chitosan oligosaccharides in Examples 2-5 are the same as those in Example 1.

[0047] Comparative Example 1

[0048] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0049] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 7.0 with NaOH solution, then add 1 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles;

[0050] (3) Desorption: The zeolite filter residue was placed in 30 ml of 2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 54%.

[0051] Comparative Example 2

[0052] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0053] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 5.0 with NaOH solution, then add 1 g of modified zeolite, shake for 4 hours and then filter quickly to collect the modified zeolite particles;

[0054] (3) Desorption: The zeolite filter residue was placed in 30 ml of 1 mol / L hydrochloric acid solution for desorption. After shaking for 4 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 52%.

[0055] Comparative Example 3

[0056] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0057] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles;

[0058] (3) Desorption: The zeolite filter residue was placed in 30 ml of 0.2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 41%.

[0059] Comparative Example 4

[0060] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0061] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1 g of modified zeolite, shake for 6 hours and then filter quickly to collect the modified zeolite particles;

[0062] (3) Desorption: The zeolite filter residue was placed in 30 ml of 3.2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered. The resulting chitosan oligosaccharide underwent partial degradation, mainly because the hydrochloric acid concentration was too high, and the glycosidic bonds in the chitosan oligosaccharide chain were unstable and underwent hydrolysis.

[0063] Comparative Example 5

[0064] (1) Zeolite modification: Natural clinoptilolite was placed in a sodium chloride solution with a mass concentration of 20%, boiled for 20 minutes, filtered, and the sodium chloride solution was reused to modify the zeolite three times to obtain modified zeolite.

[0065] (2) Adsorption: Measure 25 ml of 4 g / L chitosan oligosaccharide solution, adjust its pH to 6.0 with NaOH solution, then add 1 g of modified zeolite, shake for 2 hours and then filter quickly to collect the modified zeolite particles;

[0066] (3) Desorption: The zeolite filter residue was placed in 30 ml of 2 mol / L hydrochloric acid solution for desorption. After shaking for 6 hours, it was quickly filtered to obtain a purified chitosan oligosaccharide solution with an extraction recovery rate of 38%.

[0067] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for adsorption and extraction of chitosan oligosaccharides, characterized in that, Includes the following steps: (1) Zeolite modification: Natural clinoptilolite is placed in a sodium chloride solution of a certain concentration, boiled, filtered, and modified zeolite is obtained; (2) Adsorption: Adjust the pH of the chitosan oligosaccharide solution to 5.5-6.5, then add modified zeolite, shake thoroughly, filter, and collect the modified zeolite; (3) Desorption: The collected modified zeolite was placed in hydrochloric acid solution, shaken thoroughly and filtered to obtain the purified chitosan oligosaccharide solution; In step (2), the sufficient oscillation time is 3-6 hours; In step (3), the concentration of the hydrochloric acid solution is 0.5-3 mol / L.

2. The adsorption and extraction method for chitosan oligosaccharides according to claim 1, characterized in that, In step (1), the mass concentration of the sodium chloride solution is 15%~25%.

3. The adsorption and extraction method for chitosan oligosaccharides according to claim 1, characterized in that, In step (1), natural clinoptilolite is modified three times using sodium chloride solution.

4. The adsorption and extraction method for chitosan oligosaccharides according to claim 1, characterized in that, In step (2), the amount of modified zeolite used is 10-15 times the mass of chitosan oligosaccharide in the chitosan oligosaccharide solution.

5. The adsorption and extraction method for chitosan oligosaccharides according to claim 1, characterized in that, In step (3), the oscillation time is 2-6 hours.

6. The adsorption and extraction method for chitosan oligosaccharides according to claim 1, characterized in that, In step (2), the chitosan oligosaccharide solution used is an aqueous solution containing neutral sugar molecules.

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