A composition containing phlorotannins and a method of preparing and using the same

Abstract

The application belongs to the technical field of biological stimulants, and particularly discloses a composition containing phaeodactosan and a preparation method and application thereof. The composition containing phaeodactosan contains 0.08-0.12 g / L of phaeodactosan and 0.1-0.5 g / L of Lactobacillus pentosus extracellular polysaccharide, and the Lactobacillus pentosus extracellular polysaccharide is prepared by fermentation of Lactobacillus pentosus. The composition can effectively promote fruit color change and sweetness increase of crops.

Description

Technical Field

[0001] This invention relates to the field of biostimulant technology, and in particular to a composition containing brown algae oligosaccharides, its preparation method, and its application. Background Technology

[0002] Alginate oligosaccharides, also known as seaweed oligosaccharides or alginate oligosaccharides, are natural polysaccharides and uronic acids extracted from the cell walls of brown algae. They are the only polysaccharides containing a carboxyl group in their monomer molecules. Alginate oligosaccharides can induce callus formation, significantly increase the content of intracellular hormones (such as indoleacetic acid), and stimulate the expression of auxin-related genes by promoting hormone levels, lipase, amylase, and protease, thereby promoting seed germination, seedling growth, and rooting.

[0003] Microbial extracellular polysaccharides (EPS) are a class of high-molecular-weight compounds secreted by microorganisms during their growth process. Widely found in bacteria, fungi, yeast, and other microorganisms, EPS comprises various polysaccharides and proteins, possessing a complex chemical composition and structure. EPS serves as an interaction medium between microbial cells and their external environment. It not only protects microbial cells from environmental stresses but also promotes cell adhesion, aggregation, and growth, forming microbial biofilms. Furthermore, EPS exhibits certain biological activities, including antioxidant, antibacterial, and antiviral effects, and shows broad application prospects.

[0004] Currently, there are many studies on the application of microbial extracellular polysaccharides or brown algae oligosaccharides in plant or crop cultivation, but there are few reports on the combined application of microbial extracellular polysaccharides and brown algae oligosaccharides. Summary of the Invention

[0005] In view of the above, it is necessary to provide a composition containing fucoidan oligosaccharides, its preparation method, and its application. The composition of the present invention comprises fucoidan oligosaccharides and Lactobacillus pentosolicus extracellular polysaccharides, which can effectively promote the color change and sweetening of crop fruits.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0007] A composition containing fucoidan oligosaccharides, the composition containing 0.08-0.12 g / L fucoidan oligosaccharides and 0.1-0.5 g / L Lactiplantibacillus pentosus extracellular polysaccharide, the Lactiplantibacillus pentosus extracellular polysaccharide being prepared by fermentation of Lactiplantibacillus pentosus HHTMQ4010-1.

[0008] Furthermore, the composition contains 0.1 g / L of fucoidan oligosaccharide and 0.3 g / L of Lactobacillus pentosus extracellular polysaccharide.

[0009] Furthermore, the Lactiplantibacillus pentosus is Lactiplantibacillus pentosus HHTMQ4010-1, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33047, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, on December 12, 2024.

[0010] Furthermore, the method for preparing the extracellular polysaccharide of *Lactobacillus pentosus* includes the following steps:

[0011] (1) Seed culture: The activated Lactobacillus pentosus HHTMQ4010-1 strain was inoculated into the seed culture medium and cultured to obtain the seed culture solution;

[0012] (2) Fermentation culture: The seed culture solution is inoculated into a fermentation medium and cultured to obtain a fermentation broth;

[0013] (3) Removal and drying: The fermentation broth is subjected to sterilization, settling, alcohol precipitation and centrifugation to collect the precipitate. After drying the precipitate, the extracellular polysaccharide of Lactobacillus pentosus is obtained.

[0014] Furthermore, the seed culture medium comprises the following raw material components: glucose 30 g / L, peptone 15 g / L, dipotassium hydrogen phosphate 1.5 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.04 g / L, and Tween 80 1 mL / L.

[0015] Furthermore, the fermentation medium comprises the following raw material components: 10 g / L tryptone, 10 g / L soybean peptone, 8 g / L beef extract, 4 g / L yeast extract, 20 g / L glucose, 20 g / L lactose, 2 g / L dipotassium hydrogen phosphate, 2 g / L triammonium citrate, 5 g / L sodium acetate, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate, and 1 mL / L Tween 80.

[0016] The present invention also provides a method for preparing the composition containing brown algae oligosaccharides, wherein the preparation method comprises: weighing the brown algae oligosaccharides and the extracellular polysaccharide of Lactobacillus pentosus by weight, adding them to water, and stirring evenly to obtain the composition containing brown algae oligosaccharides.

[0017] The present invention also provides the application of the aforementioned composition containing brown algae oligosaccharides in promoting color change and sweetening of crop fruits.

[0018] The present invention has the following beneficial effects:

[0019] This invention combines brown algae oligosaccharides and extracellular polysaccharides of Lactobacillus pentosus in a certain proportion. The combination of the two can effectively promote the color change and sweetening of crop fruits. The ingredients are simple and easy to use. Detailed Implementation

[0020] To make the technical problems, technical solutions and advantages of the present invention clearer, a detailed description will be given below in conjunction with specific embodiments.

[0021] All features disclosed in this specification, or all steps in all disclosed methods or processes, may be combined in any way, except for mutually exclusive features and / or steps.

[0022] Unless otherwise stated, each feature disclosed in this specification (including any appended claims and abstract) is merely one example of a series of equivalent or similar features.

[0023] Example 1

[0024] This embodiment provides an extracellular polysaccharide from *Lactiplantibacillus pentosus*, strain HHTMQ4010-1. This strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33047, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, on December 12, 2024. Detailed information about this strain is disclosed in Chinese patent application CNCN120866134A.

[0025] The specific preparation method of the above-mentioned extracellular polysaccharide of Lactobacillus pentosus includes the following steps:

[0026] 1. Culture medium preparation

[0027] (1) Seed culture medium: glucose 30g / L, peptone 15g / L, dipotassium hydrogen phosphate 1.5g / L, magnesium sulfate 0.2g / L, manganese sulfate 0.04g / L, Tween 80 1mL / L. pH 6.5, sterilized at 121 ℃ for 20 min.

[0028] (2) Fermentation medium: tryptone 10 g / L, soybean peptone 10 g / L, beef extract 8 g / L, yeast extract 4 g / L, glucose 20 g / L, lactose 20 g / L, dipotassium hydrogen phosphate 2 g / L, triammonium citrate 2 g / L, sodium acetate 5 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.04 g / L, Tween 80 1 mL / L. pH 6.0, sterilized at 121 ℃ for 20 min.

[0029] 2. Seed culture

[0030] The activated Lactobacillus pentosus HHTMQ4010-1 was cultured in the seed culture medium and cultured at 37°C for 12 h to obtain the seed culture solution.

[0031] 3. Fermentation culture

[0032] The seed culture solution was inoculated into the fermentation medium at an inoculation rate of 3% (v / v) and cultured at 30°C for 16 hours to obtain the fermentation broth.

[0033] 4. Remove impurities and dry

[0034] The fermentation broth was centrifuged at 4000 r / min for 15 min to remove bacterial cells. The supernatant was then concentrated to 1 / 5 of its original volume using a rotary evaporator at 80°C. An equal volume of 20% trichloroacetic acid was added, and the mixture was allowed to stand for 30 min to denature and precipitate the proteins. After centrifugation at 4000 r / min for 15 min, the supernatant was collected and the pH was adjusted to 7.0. The mixture was then incubated in a 90°C water bath for 30 min, followed by centrifugation at 4000 r / min for 15 min. The supernatant was collected, and an equal volume of 95% ethanol was added. The mixture was allowed to stand for 12 h. The precipitate obtained after centrifugation at 4000 r / min for 15 min was the extracellular polysaccharide of *Lactobacillus pentosus*.

[0035] Example 2

[0036] This embodiment provides a composition containing fucoidan oligosaccharides. The preparation method of the composition is as follows: Weigh fucoidan oligosaccharides and Lactobacillus pentosaccharide extracellular polysaccharides according to the ratio of 0.08g fucoidan oligosaccharides and 0.1g Lactobacillus pentosaccharide extracellular polysaccharides in 1L of the composition, add them to water, and stir evenly to obtain the composition containing 0.08g / L fucoidan oligosaccharides and 0.1g / L Lactobacillus pentosaccharide extracellular polysaccharides.

[0037] Example 3

[0038] This embodiment provides a composition containing fucoidan oligosaccharides. The preparation method of the composition is as follows: Weigh fucoidan oligosaccharides and Lactobacillus pentosaccharide extracellular polysaccharides according to the ratio of 0.1g fucoidan oligosaccharides and 0.3g Lactobacillus pentosaccharide extracellular polysaccharides in 1L of the composition, add them to water, and stir evenly to obtain the composition containing 0.1g / L fucoidan oligosaccharides and 0.3g / L Lactobacillus pentosaccharide extracellular polysaccharides.

[0039] Example 4

[0040] This embodiment provides a composition containing fucoidan oligosaccharides. The preparation method of the composition is as follows: Weigh fucoidan oligosaccharides and Lactobacillus pentosaccharide extracellular polysaccharides according to the ratio of 0.12g fucoidan oligosaccharides and 0.5g Lactobacillus pentosaccharide extracellular polysaccharides in 1L of the composition, add them to water, and stir evenly to obtain the composition containing 0.12g / L fucoidan oligosaccharides and 0.5g / L Lactobacillus pentosaccharide extracellular polysaccharides.

[0041] Example 5

[0042] This embodiment provides the application of a composition containing fucoidan oligosaccharides in promoting color change and sweetening of crop fruits. The specific application method is as follows:

[0043] 1. Test crop: Potted dwarf tomato.

[0044] 2. Test samples: The composition containing brown algae oligosaccharides described in Example 3, a 0.1 g / L brown algae oligosaccharide solution, a 0.3 g / L Lactobacillus pentosus extracellular polysaccharide solution, and a composition of 0.1 g / L brown algae oligosaccharides and 0.3 g / L Coccidia sacchari extracellular polysaccharides.

[0045] The method for preparing the extracellular polysaccharide of *Coccella korshinskii* is as follows:

[0046] (1) The activated strain of Coccidia sacchariformis HHQGLZ30-1 (for strain information, please refer to Chinese patent application CN120349912A) was inoculated into MRS medium and cultured in a constant temperature shaker for 12 h to obtain seed culture solution.

[0047] (2) Fermentation culture: The seed culture solution was inoculated into MRS medium at an inoculation rate of 3% (v / v) and cultured at 37℃ for 14h to obtain fermentation broth;

[0048] (3) The polysaccharide fermentation broth is subjected to bacterial removal, alcohol precipitation and centrifugation drying to obtain the extracellular polysaccharide of Coccidia sacchari.

[0049] 3. Experimental design: There are 4 treatments in total, with 5 replicates for each treatment, arranged in randomized block design.

[0050] 4. Test treatment

[0051] The details are shown in Table 1 below.

[0052] Table 1 Test Treatment

[0053]

[0054] 5. Survey Indicators and Methods

[0055] The details are shown in Table 2 below.

[0056] Table 2 Survey Indicators and Methods

[0057]

[0058] 6. Test Results

[0059] (1) Effects of different treatments on the number of color-changing fruits and the number of mature fruits in dwarf tomatoes

[0060] Table 3. Effects of different treatments on the number of fruits that have changed color.

[0061]

[0062] As shown in Table 3, except that the treatment with Lactobacillus pentosus extracellular polysaccharide solution alone did not accelerate the color change rate of tomatoes, the treatment with fucoidan oligosaccharide + Lactobacillus pentosus extracellular polysaccharide, fucoidan oligosaccharide, or fucoidan oligosaccharide + Coccidioides saccharide all accelerated the coloring process of dwarf tomatoes, significantly increased the number of color-changing fruits, greatly improved the color change rate, and the effect of promoting fruit color change was: fucoidan oligosaccharide + Lactobacillus pentosus extracellular polysaccharide treatment > fucoidan oligosaccharide treatment > fucoidan oligosaccharide or fucoidan oligosaccharide + Coccidioides saccharide treatment.

[0063] Table 4. Effects of different treatments on the number of mature fruits.

[0064]

[0065] Table 4 shows that, except for tomatoes treated with Lactobacillus pentosate extracellular polysaccharide solution alone, which did not accelerate fruit ripening, all other treatments improved the coloring rate of dwarf tomatoes, accelerated fruit ripening, and increased the number of ripe fruits. The effect of accelerating fruit ripening was: treatment with fucoidan oligosaccharide + Lactobacillus pentosate extracellular polysaccharide > treatment with fucoidan oligosaccharide > treatment with fucoidan oligosaccharide or treatment with fucoidan oligosaccharide + Coxsackie exopolysaccharide.

[0066] Tables 3 and 4 together show that fucoidan alone accelerates the coloring and ripening process of dwarf tomato fruits, while *Lactobacillus pentosus* extracellular polysaccharide alone does not. However, the combination of fucoidan and *Lactobacillus pentosus* extracellular polysaccharide exhibits a synergistic effect, significantly accelerating the coloring and ripening process of dwarf tomato fruits. This may be because *Lactobacillus pentosus* extracellular polysaccharide enhances the effect of fucoidan on dwarf tomatoes, or there may be some interaction between certain monosaccharides in *Lactobacillus pentosus* extracellular polysaccharide and fucoidan, thereby promoting the coloring and ripening process of dwarf tomato fruits. However, there is a certain antagonistic effect between fucoidan and *Coccidioidomyces coccidioidomyces* extracellular polysaccharide.

[0067] Table 5 Effects of different treatments on Brix (%) in fruit

[0068]

[0069] As shown in Table 5, all treatment groups improved the Brix of tomato fruits compared to the CK group, but the differences between groups D1-D3 were not significant, while the difference in group S1 was more significant. This indicates that the treatment of brown algae oligosaccharide + pentosyl lactobacillus extracellular polysaccharide can synergistically increase the sweetness of tomato fruits.

Claims

1. A composition containing fucoidan oligosaccharides, characterized in that, The composition contains 0.08-0.12 g / L of fucoidan oligosaccharide and 0.1-0.5 g / L of extracellular polysaccharide of Lactobacillus pentosus, wherein the extracellular polysaccharide of Lactobacillus pentosus is prepared by fermentation of Lactobacillus pentosus.

2. The composition containing fucoidan oligosaccharides according to claim 1, characterized in that, The composition contains 0.1 g / L of brown algae oligosaccharides and 0.3 g / L of Pentosaccharide Lactobacillus extracellular polysaccharides.

3. The composition containing fucoidan oligosaccharides according to claim 1, characterized in that, The Lactiplantibacillus pentosus is Lactiplantibacillus pentosus HHTMQ4010-1, and the preservation number of Lactiplantibacillus pentosus HHTMQ4010-1 is CGMCC No.33047.

4. The composition containing fucoidan oligosaccharides according to claim 1, characterized in that, The method for preparing the extracellular polysaccharide of Lactobacillus pentosus includes the following steps: (1) Seed culture: The activated Lactobacillus pentosus HHTMQ4010-1 strain was inoculated into the seed culture medium and cultured to obtain the seed culture solution; (2) Fermentation culture: The seed culture solution is inoculated into a fermentation medium and cultured to obtain a fermentation broth; (3) Removal and drying: The fermentation broth is subjected to sterilization, settling, alcohol precipitation and centrifugation to collect the precipitate. After drying the precipitate, the extracellular polysaccharide of Lactobacillus pentosus is obtained.

5. The composition containing fucoidan oligosaccharides according to claim 4, characterized in that, The seed culture medium comprises the following raw material components: glucose 30 g / L, peptone 15 g / L, dipotassium hydrogen phosphate 1.5 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.04 g / L, and Tween 80 1 mL / L.

6. The composition containing fucoidan oligosaccharides according to claim 4, characterized in that, The fermentation medium comprises the following raw material components: 10 g / L tryptone, 10 g / L soybean peptone, 8 g / L beef extract, 4 g / L yeast extract, 20 g / L glucose, 20 g / L lactose, 2 g / L dipotassium hydrogen phosphate, 2 g / L triammonium citrate, 5 g / L sodium acetate, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate, and 1 mL / L Tween 80.

7. The method for preparing a composition containing fucoidan oligosaccharides according to claim 1, characterized in that, The preparation method is as follows: weigh the brown algae oligosaccharide and the extracellular polysaccharide of Lactobacillus pentosus according to weight, add them to water, and stir evenly to obtain the composition containing brown algae oligosaccharide.

8. The use of the composition containing fucoidan as described in any one of claims 1-6 in promoting color change and sweetening of crop fruits.

Images