Method for regulating and controlling cavity size and potential of yeast glucan particle carrier
The method of phosphate buffer and NaCl solution treatment combined with NaOH solution was solved by solving the problem of cavity size and potential regulation of yeast glucan particles, and yeast glucan particles with high permeability and spacious space were prepared, suitable for carrier materials.
Patent Information
- Application Number
- CN202510664764.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-22
- Publication Date
- 2025-09-02
AI Technical Summary
The prior art is difficult to effectively regulate the cavity size and potential of yeast glucan particles, and the preparation process is not safe and controllable enough.
After treating yeast cells with phosphate buffer and NaCl solution, combined with different concentrations of NaOH solution and pH adjustment, the yeast cells were treated by plasmid-solving method and alkaline solvent to regulate the cavity structure and potential of yeast glucan particles.
The regulation of the cavity structure of yeast glucan particles is achieved, and its performance as a carrier material is improved. It has a wider intracellular space and high permeability. The preparation process is safe and controllable.
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Figure CN120574346A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of polysaccharide carrier preparation, and in particular to a method for regulating the cavity size and potential of a yeast glucan particle carrier. Background Art
[0002] Yeast glucan is a natural polysaccharide derived from yeast cell walls. It forms a network structure within the yeast cell wall, forming a tight triple helix structure through hydrogen bonding. This unique molecular structure gives yeast cells unique biological functions. Further research on yeast glucan has revealed a variety of biological activities, including immunomodulatory, anti-inflammatory, anti-tumor, and hypoglycemic effects.
[0003] An extraction method that preserves the yeast glucan structure yields yeast glucan particles that retain the structural morphology of the yeast cell wall. These particles are porous and hollow. Their hollow structure and excellent biological activity make them an excellent natural carrier. Compared to intact yeast, yeast glucan particles possess a more spacious intracellular space and high permeability. Currently, most yeast glucan particle preparation methods rely on organic solvent treatment to remove yeast contents. This method primarily relies on alkaline treatment to remove contents, resulting in a large yeast cell wall pore size that is difficult to control. Summary of the Invention
[0004] In view of the shortcomings of the existing technology, the present invention provides a method for regulating the cavity size and potential of yeast glucan particle carriers, which can effectively obtain the cavity structure of yeast glucan particles and regulate their potential, and the reaction process is safe and controllable.
[0005] To achieve the above objectives, the present invention is implemented through the following technical solutions: A method for regulating the cavity size and potential of a yeast glucan particle carrier, comprising the following steps: S1. Rinse the yeast cells with phosphate buffered saline (PBS), collect the precipitate by centrifugation, and then wash the precipitate with distilled water. S2, suspending the precipitate after washing in the above step S1 in a NaCl solution, stirring and shaking at a constant temperature, and then centrifuging to collect the precipitate, and then continuing to wash the precipitate with distilled water; S3, suspending the precipitate washed in step S2 in NaOH solutions of different concentrations, stirring at a constant temperature of 80°C, adjusting the pH to 4.5, and collecting the precipitate after centrifugation; S4. The precipitate collected in the above step S3 is washed with distilled water and then freeze-dried to obtain yeast glucan particles with different cavity sizes and electrical potentials.
[0006] Preferably, the concentration of the phosphate buffer solution (PBS) in step S1 is 0.01 M and the pH is 6.8.
[0007] Preferably, the concentration of the NaCl solution in step S2 is 100 g / L.
[0008] Preferably, the temperature of the constant temperature oscillation stirring in step S2 is 50-60° C., the stirring speed is 150-220 rpm, and the stirring time is 45-50 h.
[0009] Preferably, the NaOH solution in step S3 is selected from any one of 1M, 2M, and 3M.
[0010] Preferably, the constant temperature stirring time in step S3 is 50-80 minutes.
[0011] Preferably, HCl is used to adjust the pH in step S3.
[0012] Preferably, the centrifugation in steps S1-S4 is performed at a speed of 6000 rpm for 10 minutes.
[0013] The present invention provides a method for regulating the cavity size and potential of a yeast glucan particle carrier, which has the following advantages over the prior art: (1) The present invention combines a mass-solution method with an alkaline solvent to treat yeast cells to obtain yeast glucan particles. This method can effectively obtain the cavity structure of yeast glucan particles and simultaneously regulate their electrical potential. This solves the problem of narrow intracellular space and low cell wall porosity in yeast and lays a good foundation for the development of yeast glucan particles as carrier materials.
[0014] (2) The present invention can obtain yeast glucan particles that maintain the structural morphology of yeast cell walls, and the yeast glucan particles have a porous structure and a hollow structure, and have a more spacious intracellular space and high permeability.
[0015] (3) The present invention first uses a mass-solution method to treat the yeast. The mass-solution method is relatively mild compared to the solvent method and causes less damage to the cell wall. Then, the yeast is treated with an organic solvent, which can better remove the contents in the yeast and obtain yeast glucan particles with larger pores, thereby realizing the regulation of the cavity structure and potential of the yeast glucan particles.
[0016] (4) The preparation process of the present invention is simple, the raw materials are cheap and easily available, and the preparation conditions of the yeast glucan particles are controllable and convenient. The present invention has obvious advantages as a carrier material. BRIEF DESCRIPTION OF THE DRAWINGS
[0017] Figure 1 Schematic diagram of the sugar content of different yeast glucan particles in the examples of the present invention; Figure 2 Schematic diagram of the protein content of different yeast glucan particles in the examples of the present invention; Figure 3 Schematic diagram of the Zeta potential of different yeast glucan particles in the examples of the present invention; Figure 4 The morphology of different yeast glucan particles in the examples of the present invention is characterized, wherein the scale bar in A is 1 μm, and the scale bar in B is 5 μm. DETAILED DESCRIPTION
[0018] To make the objectives, technical solutions, and advantages of the embodiments of the present invention more clear, the technical solutions in the embodiments of the present invention are clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.
[0019] Example 1: Yeast glucan particle carrier preparation: (1) Rinse 100 g of yeast cells with phosphate buffered saline (PBS) (0.01 M, pH 6.8), then centrifuge (6000 rpm, 10 min), discard the supernatant to collect the precipitate, and wash the collected precipitate with distilled water; (2) The washed precipitate was suspended in a 10% (100 g / L) NaCl solution in a beaker, and the beaker was placed in a shaking incubator at 55°C and 180 rpm for 48 hours. The precipitate was collected by centrifugation (6000 rpm, 10 min), and the collected precipitate was washed with distilled water; (3) The precipitate was suspended in NaOH solutions of different concentrations (1 M, 2 M, 3 M) and stirred continuously at 80 °C for 1 hour. The pH was adjusted to 4.5 with HCl and the precipitate was collected by centrifugation (6000 rpm, 10 min) and the supernatant was discarded. (4) The collected precipitate is washed with distilled water and finally freeze-dried to obtain yeast glucan particles with different cavity sizes and potentials.
[0020] The yeast glucan particles obtained by using the NaOH solution of different concentrations are shown in Table 1 below: Table 1
[0021] Comparative Example 1: Yeast glucan particle carrier preparation: 100 g of yeast cells were washed with phosphate buffered saline (PBS) (0.01 M, pH 6.8), and then centrifuged (6000 rpm, 10 min) to discard the supernatant to collect the precipitate. The collected precipitate was washed with distilled water and freeze-dried to obtain the yeast glucan particle carrier NPYC.
[0022] Comparative Example 2: Yeast glucan particle carrier preparation: (1) Rinse 100 g of yeast cells with phosphate buffered saline (PBS) (0.01 M, pH 6.8), then centrifuge (6000 rpm, 10 min), discard the supernatant to collect the precipitate, and wash the collected precipitate with distilled water; (2) The washed precipitate was suspended in a 10% (100 g / L) NaCl solution in a beaker, and the beaker was placed in an oscillating incubator at 55°C and 180 rpm for 48 hours. The precipitate was collected by centrifugation (6000 rpm, 10 min), washed with distilled water, and freeze-dried to obtain the yeast glucan particle carrier PYC.
[0023] Detection: 1. Sugar content determination: The sugar content of each yeast glucan particle carrier was determined using the phenol-sulfuric acid method. The glucose standard was dried overnight in a vacuum oven and dissolved in deionized water to prepare a standard solution with a concentration gradient of 0.01 mg / mL to 0.1 mg / mL. Separately, the vacuum-dried sample was dispersed in deionized water to prepare a 0.1 mg / mL suspension. 1 mL of phenol solution (5 w / w%) was added to 1 mL of the glucose standard solution and sample suspension, and the mixture was thoroughly mixed. 5 mL of concentrated sulfuric acid was added to the mixture in an ice bath, and the mixture was thoroughly mixed. After cooling, the absorbance of the solution at 490 nm was measured using a UV-visible spectrophotometer. The sugar content of the sample was calculated from the glucose standard curve.
[0024] Specific results can be found in Figure 1 , PYC-A3 has the highest sugar content.
[0025] 2. Protein content determination: The nitrogen content of the yeast glucan particle carriers in each group was measured by an elemental analyzer, and the protein content was calculated based on the nitrogen content. Specific results can be found in Figure 2 As shown, NPYC has the highest protein content.
[0026] 3. Potential measurement The zeta potential of each group of yeast glucan particle carriers was measured using a dynamic light scattering particle size analyzer. Prior to measurement, an aliquot of the yeast glucan particle carrier was diluted with deionized water to avoid multiple scattering caused by particle interactions. Specific results such as Figure 3 As shown in the figure, the zeta potential of yeast glucan particle carriers obtained by different treatments were different, and the zeta potential of PYC-A1, PYC-A2, and PYC-A3 was higher than that of PYC and NPYC.
[0027] 4. Transmission electron microscopy morphology characterization: Each group of yeast glucan particle carrier samples was evenly dispersed in deionized water at a concentration of 50 μg / mL. 2.5 μL of the dispersion was dropped on the surface of a single-polished single-crystal silicon wafer and naturally dried. After the sample surface was gold-plated, the sample morphology was observed using a scanning electron microscope. Another 2.5 μL of the above dispersion was dropped on the surface of a carbon support film and naturally dried. The sample morphology was observed using a transmission electron microscope.
[0028] Specific results such as Figure 4 As shown, PYC-A1, PYC-A2, and PYC-A3 formed obvious cavity structures.
[0029] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the various embodiments of the present invention.
Claims
1. A method for regulating the cavity size and potential of a yeast glucan particle carrier, characterized in that: The method comprises the following steps: S1. Rinse the yeast cells with phosphate buffered saline (PBS), collect the precipitate by centrifugation, and then wash the precipitate with distilled water. S2, suspending the precipitate after washing in the above step S1 in a NaCl solution, stirring and shaking at a constant temperature, and then centrifuging to collect the precipitate, and then continuing to wash the precipitate with distilled water; S3, suspending the precipitate washed in step S2 in NaOH solutions of different concentrations, stirring at a constant temperature of 80°C, adjusting the pH to 4.5, and collecting the precipitate after centrifugation; S4. The precipitate collected in the above step S3 is washed with distilled water and then freeze-dried to obtain yeast glucan particles with different cavity sizes and electrical potentials.
2. The method for controlling cavity size and potential according to claim 1, wherein: The concentration of the phosphate buffer solution (PBS) in step S1 is 0.01 M and the pH is 6.
8.
3. The method for controlling cavity size and potential according to claim 1, wherein: The concentration of the NaCl solution in step S2 is 100 g / L.
4. The method for controlling cavity size and potential according to claim 1, wherein: The constant temperature oscillating stirring in step S2 is performed at a temperature of 50-60° C., a stirring speed of 150-220 rpm, and a stirring time of 45-50 h.
5. The method for controlling cavity size and potential according to claim 1, wherein: In step S3, the NaOH solution is selected to be any one of 1M, 2M, and 3M.
6. The method for controlling cavity size and potential according to claim 1, wherein: The constant temperature stirring time in step S3 is 50-80 minutes.
7. The method for controlling cavity size and potential according to claim 1, wherein: In step S3, HCl is used to adjust the pH.
8. The method for controlling cavity size and potential according to claim 1, wherein: The centrifugation in steps S1-S4 is performed at a speed of 6000 rpm for 10 minutes.