Amino acid modified astragalus polysaccharide, and preparation method and antibacterial application thereof
By introducing amino acids into Astragalus polysaccharide molecules, its water solubility and antibacterial activity are improved, solving the problem of limited application of Astragalus polysaccharide and realizing the application of broad-spectrum antibacterial effect and green degradable antibacterial agent.
Patent Information
- Application Number
- CN202311500330.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-11-10
- Publication Date
- 2026-08-25
- Estimated Expiration
- 2043-11-10
AI Technical Summary
Existing Astragalus polysaccharides suffer from poor water solubility, poor acid and alkali resistance, and low biological activity, which limits their application in the food, pharmaceutical, and daily chemical industries.
By introducing amino acids, such as glycine, lysine, and arginine, into Astragalus polysaccharide molecules, amino acid-grafted modified Astragalus polysaccharides are formed, which increase intramolecular electrostatic repulsion, improve water solubility, and enhance antibacterial activity by binding to bacterial surfaces through amino and carboxyl groups.
It significantly improves the water solubility and antibacterial activity of Astragalus polysaccharide, achieving a broad-spectrum antibacterial effect against Gram-positive and Gram-negative bacteria, making it a green, biodegradable, and highly efficient antibacterial agent with the potential to replace antibiotics.
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Figure CN117551215B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of polysaccharide modification technology, and in particular to an amino acid-modified Astragalus polysaccharide, its preparation method, and its antibacterial applications. Background Technology
[0002] Antibiotics are a class of drugs with strong bactericidal effects. However, the continued use of antibiotics increases the concentration of residual antibiotics in the environment, leading to bacterial resistance and posing a serious threat to human health. Polysaccharides are natural macromolecular compounds formed by the condensation of monosaccharide molecules. Their antibacterial activity is achieved by increasing cell membrane permeability, inhibiting the adsorption of pathogenic bacteria to host cells, or blocking the transmembrane transport of nutrients and energy substances. Polysaccharides with antibacterial activity can replace antibiotics, which to some extent alleviates the problems of antibiotic resistance, toxic side effects, and continuously decreasing antibacterial efficiency. Polysaccharide antibacterial agents have high development prospects in the future food, daily chemical, and pharmaceutical fields.
[0003] Astragalus polysaccharide is a dried exudate extracted from the stems and branches of plants in the genus *Astragalus* in Asia. It is a high-molecular-weight anionic natural polysaccharide, primarily composed of D-galacturonic acid monomers linked by α-1,4 linkages, and containing numerous carboxyl, hydroxyl, and methoxy groups. The structure of astragalus polysaccharide contains arabinose, xylose, galactose, rhamnose, fucose, and galacturonic acid residues. Due to its high viscosity, good biocompatibility, biodegradability, and non-toxicity, it is widely used in the food, pharmaceutical, and daily chemical industries. However, its poor water solubility, poor acid and alkali resistance, and low biological activity significantly limit its applications. Therefore, modification of astragalus polysaccharide is crucial for expanding its practical applications. Summary of the Invention
[0004] To address the problems of poor water solubility, poor acid and alkali resistance, and low biological activity of existing astragalus polysaccharides, this invention provides an amino acid-modified astragalus polysaccharide, its preparation method, and its antibacterial application.
[0005] To solve the above-mentioned technical problems, the technical solution provided by the present invention is as follows:
[0006] An amino acid-modified Astragalus polysaccharide has the chemical structural formula shown in Formula I:
[0007]
[0008] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H, respectively.
[0009]
[0010] Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0011] Astragalus polysaccharide molecules have a large number of main chain components and stable glycosidic bonds. They are more likely to form a large number of interlayer close-range hydrogen bonds, resulting in a compact molecular structure and poor hydrophilicity, which greatly limits their practical application. In addition, astragalus polysaccharide has low antibacterial activity and a narrow antibacterial spectrum, making it difficult to achieve the ideal antibacterial effect.
[0012] To address the problems of poor water solubility and low bioactivity of existing Astragalus polysaccharides, this invention introduces multiple amino and carboxyl groups into the Astragalus polysaccharide molecule by grafting glycine, lysine, and arginine. This increases the electrostatic repulsion within the Astragalus polysaccharide molecule, making the molecular chain more extended and reducing the close-range hydrogen bonding within the molecule, thus effectively improving the water solubility of Astragalus polysaccharide. Simultaneously, the introduction of amino groups promotes the adsorption of Astragalus polysaccharide on bacterial surfaces, facilitating the full expression of its antibacterial activity. The carboxyl groups can bind to glycoprotein receptors on the bacterial cell surface, inhibiting bacterial metabolic activities and enhancing the bactericidal effect of Astragalus polysaccharide.
[0013] The specific amino acid-modified Astragalus polysaccharide provided by this invention has stable performance, is green and biodegradable, water-soluble and has high antibacterial activity, and is expected to become a new type of green macromolecular antibacterial agent to replace antibiotics, with broad potential application value.
[0014] Preferably, R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0015] During the experiment, the inventors attempted to modify polysaccharides by grafting various amino acids onto them to enhance their antibacterial activity. During the research and development process, they unexpectedly discovered that modifying Astragalus polysaccharides with glycine, arginine, and lysine significantly improved their antibacterial activity. In particular, glycine grafting modification dramatically enhanced the antibacterial activity. A 6 mg / mL aqueous solution of glycine-modified Astragalus polysaccharide showed a 100% inhibition rate against both Escherichia coli and Staphylococcus aureus; a 4 mg / mL aqueous solution showed an inhibition rate of approximately 48% against Escherichia coli and approximately 89% against Staphylococcus aureus. It also exhibited excellent antibacterial activity against both Gram-positive and Gram-negative bacteria, demonstrating a broad antibacterial spectrum. This effect far exceeded the expectations of those skilled in the art, achieving unexpected technical results.
[0016] Preferably, the molar degree of substitution of the amino acid is 0.3 to 1.1.
[0017] It should be noted that the above amino acids include L and D conformations.
[0018] This invention also provides a method for preparing the above-mentioned amino acid-modified Astragalus polysaccharide, comprising at least the following steps:
[0019] Astragalus polysaccharide, formaldehyde solution, amino acids and alkaline solution are added to alcohol solution, mixed evenly, and reacted at 50℃~70℃ for 2h~5h to obtain amino acid modified astragalus polysaccharide.
[0020] Grafting polysaccharides with amino acids can preserve their biodegradability while imparting the bioactivity of amino acids. However, existing methods for grafting polysaccharides with amino acids have the following drawbacks: The N-bromosuccinimide bromination method, using organic solvents, has poor safety. Although the reaction conditions are mild, the condensation reaction between the active carboxyl and hydroxyl groups hinders the improvement of the antibacterial activity of the modified polysaccharides.
[0021] The present invention provides a method for preparing amino acid-modified astragalus polysaccharide. Using astragalus polysaccharide, formaldehyde, and specific amino acids as raw materials, amino acid-grafted modified astragalus polysaccharide is prepared via the Mannich reaction. The amino acid grafting site occurs only on the fifth carbon of the sugar ring, preserving both the amino and carboxyl groups of the amino acids, thus retaining their biological activity. It also preserves the active carboxyl group of the astragalus polysaccharide, significantly improving the antibacterial properties of the synthesized amino acid-modified astragalus polysaccharide. This method achieves broad-spectrum antibacterial activity against both Gram-positive and Gram-negative bacteria. Furthermore, it is biodegradable, capable of degradation in vivo or in the environment. It is a green, novel, and highly efficient antibacterial material, potentially becoming a substitute for antibiotics. This method is of great significance in addressing the current problems of increased bacterial resistance and environmental accumulation caused by antibiotics, and has promising application prospects.
[0022] The reaction equation for preparing amino acid-modified Astragalus polysaccharide according to this invention is as follows:
[0023]
[0024] Under the catalysis of alkali, the active hydrogen on the fifth carbon of the sugar ring of Astragalus polysaccharide, which is attached to the carboxyl group, undergoes a condensation reaction with the aldehyde group and the amino group on the amino acid to obtain amino acid modified Astragalus polysaccharide.
[0025] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H, respectively.
[0026]
[0027] Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0028] Preferably, the alkaline solution is a sodium carbonate solution, a sodium hydroxide solution, or a potassium hydroxide solution.
[0029] Preferably, the mass concentration of the alkaline solution is 30% to 35%.
[0030] More preferably, the molar ratio of alkali to amino acids in the alkaline solution is 1:1 to 1.2:1.
[0031] By controlling the amount of alkali added, the hydrogen bonds in Astragalus polysaccharides can be disrupted, weakening the intermolecular forces and causing the molecular chains to relax, which is beneficial for the modification reaction. Simultaneously, the alkali can protect the carboxyl groups on amino acids, preventing self-condensation reactions and reducing the R-NH3 groups in the amino acids. + It is converted to R-NH2, which makes the reaction easier to proceed.
[0032] Preferably, the formaldehyde solution has a mass concentration of 37% to 40%.
[0033] Preferably, the molar ratio of the amino acid to formaldehyde is 1:1 to 1:1.3.
[0034] Preferably, the molar ratio of the amino acid to the astragalus polysaccharide is 0.5:1 to 1.2:1.
[0035] The optimal ratio of formaldehyde, amino acids, and astragalus polysaccharides ensures that the amino acids have a suitable degree of substitution, thereby improving both water solubility and antibacterial activity.
[0036] Preferably, the alcohol solution is an aqueous solution of isopropanol, an aqueous solution of ethanol, or an aqueous solution of tert-butanol, wherein its mass concentration is 85% to 95%.
[0037] Preferably, the amount of alcohol solution added is 2.5 to 5 times the mass of the astragalus polysaccharide.
[0038] The optimal alcohol solution and the amount of alcohol solution added can promote the full reaction between amino acids and astragalus polysaccharides, obtain a suitable degree of substitution, ensure the effective improvement of antibacterial activity and water solubility, and at the same time, improve the reaction efficiency.
[0039] It should be noted that after the reaction was completed, hydrochloric acid was added to the reaction system to adjust the pH to neutral, and then ethanol was added for precipitation and washing. After filtration and freeze drying, amino acid-modified astragalus polysaccharide was obtained.
[0040] For example, precipitation and washing are performed using an ethanol solution with a mass concentration of 75% to 85%.
[0041] The method for preparing amino acid-modified astragalus polysaccharide provided by this invention is simple, the reaction conditions are mild, and the prepared modified astragalus polysaccharide has stable properties and high antibacterial activity. It can be used as a natural antibacterial agent to replace antibiotics and has potential application value in the preparation of antibacterial health foods or medicines.
[0042] This invention also provides the application of the above-mentioned amino acid-modified Astragalus polysaccharide in the preparation of antibacterial materials.
[0043] This invention uses natural biodegradable astragalus polysaccharide as a matrix and modifies it through the Mannich reaction using specific amino acids. This significantly improves the antibacterial activity and water solubility of the astragalus polysaccharide. The preparation method is simple and achieves the goal of preparing modified astragalus polysaccharide in one pot. It is suitable for industrial-scale production and has broad application prospects in the fields of food, health food, biomedicine and cosmetics. Attached Figure Description
[0044] Figure 1 The infrared spectrum of the amino acid-modified Astragalus polysaccharide prepared in Example 1 of this invention;
[0045] Figure 2 The graph shows the antibacterial effects of Astragalus polysaccharide, the amino acid-modified Astragalus polysaccharide prepared in Examples 1 and 4-5, in the antibacterial activity test of this invention. Detailed Implementation
[0046] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.
[0047] To better illustrate the present invention, further examples are provided below.
[0048] Example 1
[0049] This embodiment provides a method for preparing glycine-modified Astragalus polysaccharide, including the following steps:
[0050] 128 g of 90% isopropanol aqueous solution was added to a three-necked flask. Then, 32.4 g (0.2 mol) of Astragalus polysaccharide, 12.76 g of 40% formaldehyde solution, 12.75 g (0.17 mol) of glycine and 22.44 g of 30.3% sodium hydroxide solution were added sequentially and mixed thoroughly. The mixture was heated to 60 °C and reacted for 4 h. After the reaction was completed, hydrochloric acid was added to the reaction solution to adjust the pH to 6.8. Then, 80% ethanol solution was added for washing and precipitation. The filter cake was dried to constant weight to obtain glycine-modified Astragalus polysaccharide (TG-Gly) as shown in Formula I.
[0051]
[0052] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0053] The content of carboxylate ions in glycine-modified Astragalus polysaccharide was determined by acid-base titration. The molar degree of substitution of glycine was found to be 0.61, and the apparent viscosity of a 1 wt% glycine-modified Astragalus polysaccharide aqueous solution was 2100 mPa·s.
[0054] Example 2
[0055] This embodiment provides a method for preparing glycine-modified Astragalus polysaccharide, including the following steps:
[0056] Add 81 g of 85% tert-butanol solution to a three-necked flask, then add 32.4 g (0.2 mol) of Astragalus polysaccharide, 9.76 g of 37% formaldehyde solution, 7.51 g (0.1 mol) of glycine, and 20.8 g of 32.5% potassium hydroxide solution in sequence. Mix well and heat to 60 °C for 4 h. After the reaction is complete, add hydrochloric acid to the reaction solution to adjust the pH to 6.8, then add 80% ethanol solution for washing and precipitation. Dry the filter cake to constant weight to obtain glycine-modified Astragalus polysaccharide (TG-Gly) as shown in Formula I.
[0057]
[0058] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0059] The content of carboxylate ions in glycine-modified Astragalus polysaccharide was determined by acid-base titration. The molar degree of substitution of glycine was found to be 0.44, and the apparent viscosity of a 1 wt% glycine-modified Astragalus polysaccharide aqueous solution was 3650 mPa·s.
[0060] Example 3
[0061] This embodiment provides a method for preparing glycine-modified Astragalus polysaccharide, including the following steps:
[0062] 162 g of 95% ethanol solution was added to a three-necked flask. Then, 32.4 g (0.2 mol) of Astragalus polysaccharide, 24.67 g of 38% formaldehyde solution, 18.02 g (0.24 mol) of glycine and 80.0 g of 35% sodium carbonate solution were added sequentially and mixed thoroughly. The mixture was heated to 60 °C and reacted for 4 h. After the reaction was completed, hydrochloric acid was added to the reaction solution to adjust the pH to 6.8. Then, 80% ethanol solution was added for washing and precipitation. The filter cake was dried to constant weight to obtain glycine-modified Astragalus polysaccharide (TG-Gly) as shown in Formula I.
[0063]
[0064] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0065] The content of carboxylate ions in glycine-modified Astragalus polysaccharide was determined by acid-base titration. The molar degree of substitution of glycine was found to be 0.79, and the apparent viscosity of a 1 wt% glycine-modified Astragalus polysaccharide aqueous solution was 3750 mPa·s.
[0066] Example 4
[0067] This embodiment provides a method for preparing arginine-modified astragalus polysaccharide, including the following steps:
[0068] 128 g of 90% isopropanol aqueous solution was added to a three-necked flask. Then, 32.4 g (0.2 mol) of Astragalus polysaccharide, 12.76 g of 40% formaldehyde solution, 29.61 g (0.17 mol) of arginine and 22.44 g of 30.3% sodium hydroxide solution were added sequentially and mixed thoroughly. The mixture was heated to 60 °C and reacted for 4 h. After the reaction was completed, the pH of the reaction solution was adjusted to 6.9 with hydrochloric acid. Then, 80% ethanol solution was added for washing and precipitation. The filter cake was dried to constant weight to obtain arginine-modified Astragalus polysaccharide (TG-Arg) as shown in Formula I.
[0069]
[0070] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0071] The content of carboxylate ions in arginine-modified Astragalus polysaccharide was determined by acid-base titration. The molar degree of substitution of arginine was found to be 0.67, and the apparent viscosity of a 1 wt% arginine-modified Astragalus polysaccharide aqueous solution was 2510 mPa·s.
[0072] By replacing the addition ratio and reaction conditions of each raw material in this embodiment with other conditions specified in this invention, the arginine-modified astragalus polysaccharides prepared can achieve technical effects that are basically equivalent to the modified astragalus polysaccharides described above.
[0073] Example 5
[0074] This embodiment provides a method for preparing lysine-modified Astragalus polysaccharide, including the following steps:
[0075] 128 g of 90% isopropanol aqueous solution was added to a three-necked flask. Then, 32.4 g (0.2 mol) of Astragalus polysaccharide, 12.76 g of 40% formaldehyde solution, 24.85 g (0.17 mol) of lysine and 22.44 g of 30.3% sodium hydroxide solution were added sequentially and mixed thoroughly. The mixture was heated to 60 °C and reacted for 4 h. After the reaction was completed, the pH of the reaction solution was adjusted to 6.8 with hydrochloric acid. Then, 80% ethanol solution was added for washing and precipitation. The filter cake was dried to constant weight to obtain lysine-modified Astragalus polysaccharide (TG-Lys) as shown in Formula I.
[0076]
[0077] Where n is 200 to 600, and R1, R2, R3, and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
[0078] The content of carboxylate ions in lysine-modified Astragalus polysaccharide was determined by acid-base titration. The molar degree of substitution of lysine was found to be 0.64, and the apparent viscosity of a 1 wt% glycine-modified Astragalus polysaccharide aqueous solution was 2720 mPa·s.
[0079] By replacing the addition ratio and reaction conditions of each raw material in this embodiment with other conditions specified in this invention, the lysine-modified astragalus polysaccharides prepared can achieve technical effects that are basically equivalent to the modified astragalus polysaccharides described above.
[0080] The infrared spectra of Astragalus polysaccharide and the amino acid-modified Astragalus polysaccharide prepared in Examples 1 and 4-5 are shown below. Figure 1 As shown in the graph, at 3410 cm -1 The broad absorption peak appearing nearby is caused by the stretching vibration of the hydroxyl group, at 2913 cm⁻¹. -1 Stretching vibration peaks belonging to the methylene group were observed nearby, at 1651 cm⁻¹. -1 and 1456cm -1 This corresponds to the symmetrical and asymmetrical stretching vibration peaks of the carboxyl group. In amino acid-modified Astragalus polysaccharide, the peak at 1317 cm⁻¹ is... -1The appearance of the stretching vibration peak of NH nearby proves the successful grafting of amino acid onto the Astragalus polysaccharide molecule.
[0081] Performance testing
[0082] 1. Substitutability Test
[0083] The degree of substitution of glutamic acid and glycine-modified Astragalus polysaccharides was determined by acid-base titration. The specific steps are as follows:
[0084] Take a 732 type cation exchange resin and place it in a beaker. Soak and stir it with 3 mol / L hydrochloric acid solution for 0.5 h, then wash it repeatedly with distilled water until Cl cannot be detected. - Then, the 732 type cation exchange resin was immersed and stirred in a 2.2 mol / L sulfuric acid solution for 3 hours, and washed repeatedly with distilled water until SO4 was undetectable in the aqueous solution. 2- This yields an activated cation exchange resin.
[0085] Weigh 2g each of Astragalus polysaccharide, glycine-modified Astragalus polysaccharide, arginine-modified Astragalus polysaccharide, and lysine-modified Astragalus polysaccharide, and disperse them separately in 80mL of 80% ethanol aqueous solution. Exchange the polysaccharides using an activated cation exchange resin (3 times the mass of the polysaccharide sample) for 3 hours to ensure complete exchange of sodium ions into hydrogen ions in the amino acid-modified Astragalus polysaccharide. Separate the activated cation exchange resin and dry the polysaccharides to constant weight. Accurately weigh mg of the exchanged sample and dissolve it in 15mL of 0.08mol / L sodium hydroxide solution. Back-titrate the sodium hydroxide solution with 0.08mol / L hydrochloric acid solution, record the amount of hydrochloric acid used, and subtract the amount of sodium hydroxide solution consumed by the uronic acid in the Astragalus polysaccharide itself. Calculate the amount of sodium hydroxide consumed by the grafted amino acids.
[0086] DS = M1A / (1 - M2A)
[0087] A=(C1V1-C2V2) / m
[0088] In the formula: DS is the degree of carboxyl substitution in the sample; M1 is the molar mass of a single astragalus polysaccharide unit, g / mol; M2 is the molar mass of the amino acid monomer reacting with astragalus polysaccharide, g / mol; C1 is the concentration of sodium hydroxide, mol / L; V1 is the volume of sodium hydroxide consumed, L; C2 is the concentration of hydrochloric acid, mol / L; V2 is the volume of hydrochloric acid consumed, L; m is the mass of the titrated sample, g.
[0089] 2. Antibacterial activity test
[0090] (1) Bacterial culture methods
[0091] The bacterial strains used in this experiment were *Escherichia coli* and *Staphylococcus aureus*. The lyophilized bacterial powders of *E. coli* and *Staphylococcus aureus* were dissolved in water to obtain a bacterial suspension (15 mg / mL). A small amount of the bacterial suspension was inoculated into a solid culture medium using a sterile toothpick, and the medium was incubated upside down in a 37°C incubator for 12 hours. A single colony was picked and placed into a centrifuge tube containing 1 mL of liquid culture medium, and incubated with shaking for 12 hours. This process was repeated twice to obtain a stable third-generation strain. The bacterial culture was then transferred from the centrifuge tube to a petri dish containing 50 mL of liquid culture medium and incubated at 37°C for 12 hours. The culture was then stored at 4°C for later use.
[0092] Solid culture medium: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L sodium chloride, 20 g / L agar powder, autoclaved at 120°C for 30 min.
[0093] Liquid culture medium: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L sodium chloride, pH 7.40 ± 0.01, autoclaved at 120°C for 30 min.
[0094] (2) Antibacterial test
[0095] Astragalus polysaccharide (TG), amino acid-modified astragalus polysaccharide prepared in Examples 1 (TG-Gly), and Examples 4 (TG-Arg) to 5 (TG-Lys) were added to deionized water to prepare polysaccharide sample solutions with concentrations of 2 mg / mL, 4 mg / mL, 6 mg / mL, 8 mg / mL, 10 mg / mL, and 12 mg / mL. Phosphate buffer (pH = 7.4) was used as a control.
[0096] The Escherichia coli and Staphylococcus aureus cultures were diluted to a final concentration of 10 μL with phosphate buffer at pH 7.4. 6 CFU / mL, under aseptic conditions, take 1 mL of the above bacterial solution and 9 mL of polysaccharide sample solution, mix them with a vortex mixer, incubate at 35℃ for 24 h, measure the absorbance value at 600 nm, and calculate the inhibition rate. Each group of experiments is repeated three times.
[0097] Antibacterial rate = A1 - (A - A0) / A1 * 100%
[0098] In the formula: A1 is the absorbance of sterile water and bacterial solution;
[0099] A represents the absorbance of the polysaccharide sample solution with added bacterial solution;
[0100] A0 represents the absorbance of the polysaccharide sample solution.
[0101] The results are shown in Table 1.
[0102] Table 1 Antibacterial rate
[0103]
[0104] (3) Colony count
[0105] Dissolve 10.0 g tryptone, 5.0 g yeast extract, 10.0 g NaCl, and 20.0 g agar powder in 100 mL distilled water and sterilize at 120 °C for 20 min to obtain sterile culture medium. Under ultra-clean air conditions, transfer the sterile culture medium to sterile petri dishes and allow it to solidify. Dilute the bacterial suspension to 10⁻⁶ pH with phosphate buffer (pH 7.4). 6 CFU / mL, then Astragalus polysaccharide, and the amino acid-modified Astragalus polysaccharide prepared in Examples 1 and 4-5 were added to deionized water to prepare a polysaccharide solution with a concentration of 10 mg / mL. The above bacterial culture and polysaccharide solution were mixed separately and incubated at 37.5℃ for 3 h. 50 μL was extracted and transferred to a culture dish containing solid culture medium. The culture dish was inverted and incubated at 37.5℃ for 24 h. Bacterial production was observed, and the results are as follows: Figure 2 As stated above.
[0106] The above results demonstrate that the amino acid-modified astragalus polysaccharide prepared in the embodiments of the present invention has excellent antibacterial activity against Escherichia coli and Staphylococcus aureus. In particular, the glycine-modified astragalus polysaccharide has the highest antibacterial activity, and the antibacterial rate against Escherichia coli and Staphylococcus aureus can reach 100% at a concentration of 6 mg / mL. It is a green, biodegradable, and highly active antibacterial agent with high potential for future applications. It is expected to become a natural macromolecular antibacterial agent that can replace antibiotics and has high practical value.
[0107] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions or improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.
Claims
1. An amino acid-modified Astragalus polysaccharide, characterized in that, Its chemical structural formula is shown in Formula I: Formula I Where n is 200~600, and R1, R2, R3 and R4 are selected from -H or Furthermore, R1, R2, R3, and R4 cannot all be -H at the same time.
2. The amino acid-modified Astragalus polysaccharide as described in claim 1, characterized in that, The molar degree of substitution of the amino acid is 0.3 to 1.
1.
3. A method for preparing amino acid-modified Astragalus polysaccharide according to claim 1 or 2, characterized in that, It should include at least the following steps: Astragalus polysaccharide, formaldehyde solution, glycine and alkaline solution are added to alcohol solution, mixed evenly, and reacted at 50℃~70℃ for 2h~5h to obtain amino acid modified astragalus polysaccharide.
4. The method for preparing amino acid-modified Astragalus polysaccharide as described in claim 3, characterized in that, The alkaline solution is a sodium carbonate solution, a sodium hydroxide solution, or a potassium hydroxide solution.
5. The method for preparing amino acid-modified Astragalus polysaccharide as described in claim 3, characterized in that, The alkaline solution has a mass concentration of 30% to 35%; and / or The formaldehyde solution has a mass concentration of 37% to 40%.
6. The method for preparing amino acid-modified Astragalus polysaccharide as described in claim 3, characterized in that, The molar ratio of glycine to astragalus polysaccharide is 0.5:1 to 1.2:1; and / or The molar ratio of alkali to amino acids in the alkaline solution is 1:1 to 1.2:1; and / or The molar ratio of glycine to formaldehyde is 1:1 to 1:1.
3.
7. The method for preparing amino acid-modified Astragalus polysaccharide as described in claim 3, characterized in that, The alcohol solution is an aqueous solution of isopropanol, an aqueous solution of ethanol, or an aqueous solution of tert-butanol, wherein its mass concentration is 85% to 95%.
8. The method for preparing amino acid-modified Astragalus polysaccharide as described in claim 7, characterized in that, The amount of alcohol solution added is 2.5 to 5 times the mass of the Astragalus polysaccharide.
9. The application of the amino acid-modified Astragalus polysaccharide according to claim 1 or 2 in the preparation of antibacterial materials.