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Beta-1,3/1,6-glucan, preparation method therefor, and application thereof in preparing immune enhancement and anti-tumor medicine and functional food

A functional food and immune-enhancing technology, applied in the fields of antineoplastic drugs, applications, food preparation, etc., can solve the problems of large differences in molecular weight, connection mode and branching degree, no tumor growth and metastasis, and unclear structure, etc. Achieve the effect of enhancing immunity and inhibiting tumor growth and metastasis, making up for insufficient market supply and rich raw material resources

Active Publication Date: 2015-10-28
MARINE BIOMEDICAL RES INST OF QINGDAO CO LTD
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0003] At present, most of the β-1,3-glucans on the market come from barley, oats, edible fungi (lentinus edodes, Grifola frondosa, Schizophyllum), yeast and other terrestrial organisms. Due to different sources of raw materials, the obtained β-1, The molecular weight, connection mode and branching degree of 3-glucan vary greatly, and the quality is difficult to control. For example, β-glucan for injection mainly comes from shiitake mushrooms, which is a kind of β-1,6-branched β-1,3-glucan, due to its high molecular weight of 400-800kDa, has poor water solubility
With the continuous expansion of the application field of β-glucan and the continuous growth of market demand, the existing shiitake mushroom β-glucan can no longer meet the needs of the market
[0004] In terms of the preparation of seaweed-derived glucans, a Chinese invention patent (Patent No. 201010169739.7) was found to involve the preparation of seaweed low-molecular-weight glucans. The raw materials are kelp, wakame and hijiki. The method used It adopts the method of 80% ethanol fractional precipitation and hydrogen peroxide oxidation. The molecular weight of the obtained product is only 800-1000 Daltons (Da), the structure is not clear, and it has no activity of inhibiting tumor growth and metastasis.

Method used

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  • Beta-1,3/1,6-glucan, preparation method therefor, and application thereof in preparing immune enhancement and anti-tumor medicine and functional food
  • Beta-1,3/1,6-glucan, preparation method therefor, and application thereof in preparing immune enhancement and anti-tumor medicine and functional food
  • Beta-1,3/1,6-glucan, preparation method therefor, and application thereof in preparing immune enhancement and anti-tumor medicine and functional food

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] 1. Dried and crushed Antarctic brown algae (such as sea velvet, sea bamboo shoots, and Leisong algae), took 400g of algae powder, added 4000mL of 95% ethanol to the algae powder to soak, stirred intermittently for 4 hours, centrifuged to discard the ethanol, and used ethanol to repeatedly extract 1 time, dried to obtain 300g of defatted algae powder.

[0034] 2. Take 30 g of the defatted algae powder, add 600 mL of double distilled water, stir and extract at room temperature for 2-4 hours to obtain a water extract.

[0035] 3. Take the supernatant by centrifugation; add 600mL of 3mol / L calcium chloride aqueous solution to the supernatant under stirring, collect the supernatant after centrifugation, then desalt with a 3000Da dialysis bag or a 1000Da nanofiltration membrane, concentrate under reduced pressure and freeze Dry to obtain 1.6g of crude polysaccharide.

[0036] 4. Dissolve 0.5 g of the crude polysaccharide with 2 mL of water, use distilled water and aqueous so...

Embodiment 2

[0048] Macrophages (Raw264.7) were cultured in high-glucose DMEM medium containing 10% fetal bovine serum, the cell concentration was adjusted, and they were inoculated in 96-well cell culture plates so that the cell concentration was 2×10 4 samples / well, and then add different concentrations of samples, so that the final concentration of β-1,3 / 1,6-glucose is 5 μg / mL, 25 μg / mL, 50 μg / mL, 100 μg / mL, and the blank medium is used as the control. After culturing for 24 hours, the medium was aspirated, the excess medium was washed with PBS, and 0.075% neutral red was added, at 37°C, 5% CO 2 Incubate for half an hour, wash off excess neutral red with PBS, add cell lysate (absolute ethanol: glacial acetic acid = 1:1, volume ratio), mix well and measure the absorbance value at 540nm on a microplate reader. The size evaluates the ability of samples to promote macrophage phagocytosis of neutral red. The above experiment was repeated three times, and the average value was taken for stat...

Embodiment 3

[0050] The macrophage (Raw264.7) cells were revived, subcultured, cultured, plated and cultured for 24 hours after adding drugs. Take a 96-well plate and add 50 μL culture solution and 50 μL diluted standard to each well, then add 50 μL Griess Reagent I to each well, then add 50 μL Griess Reagent II, and measure the absorbance at 540 nm. The above experiment was repeated three times, and the average value was taken for statistical analysis of the results. Such as Figure 5 As shown, when the brown algae β-1,3 / 1,6-glucan (DAG) of the present invention is in the range of 5-100 μg / mL, the release of macrophage NO can be significantly promoted with the increase of the concentration.

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Abstract

The present invention provide a beta-1,3 / 1,6-glucan, a preparation method therefor, and application thereof in preparing immune enhancement and anti-tumor medicine and functional food. The preparation method comprises: drying and milling antarctic brown algae; after degreasing, water extracting, calcium chloride precipitating and anion exchanging resin purification, obtaining glucan with a beta-1,3-glucose main chain and a beta-1,6-glucose branched chain, wherein a molecular weight thereof is between 5-50 kDa. The beta-glucan prepared by the present invention has an immune enhancement activity, which is indicated by swallow of neutral red by in vitro macrophages and an NO release promoting experiment, and has a tumor growth inhibiting effect and a tumor metastasis inhibiting effect which are indicated by an in vivo animal experiment. According to the present invention, beta-1,3 / 1,6-glucan is prepared by using the antarctic brown algae; is characterized in richness and readily availability of raw material sources, simple preparation process, high product purity, strong bioactivity and easiness for industrialization; and has a prospect of being developed into immune enhancement and anti-tumor medicine.

Description

technical field [0001] The invention belongs to the field of marine pharmaceuticals, and relates to a method for preparing high-purity low-molecular-weight glucan from seaweed and its application, in particular to a β-1,3 / 1,6-glucan and its preparation method and its preparation for immune enhancement. And anti-tumor drugs and functional food applications. Background technique [0002] Glucan (Glucan) is a polysaccharide substance with glucose as the only constituent unit, which widely exists in animals, plants, microorganisms and brown algae. There are many kinds of glucans in nature, including linear or branched α / β-glucans, and the connection methods between glucose include 1,2-linkage, 1,3-linkage, 1,4-linkage and 1, 6-connection and other forms, among which β-1,3 and β-1,6-glucan have better biological activities, such as anti-tumor, enhancing immunity, promoting wound healing, anti-radiation and preventing hyperlipidemia, etc. According to the statistics of the World...

Claims

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Application Information

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IPC IPC(8): C08B37/02A61K31/716A23L1/29A61P37/04A61P35/00
CPCA61K31/716
Inventor 于广利郝杰杰丛大鹏管华诗赵峡吕友晶胡婷
Owner MARINE BIOMEDICAL RES INST OF QINGDAO CO LTD
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