Use of fucoidan in the preparation of a product for promoting physiological recovery after coral cutting

Abstract

This invention discloses the application of fucoidan in the preparation of products for promoting physiological recovery after coral cutting. The invention utilizes 100 mg / L fucoidan to treat cut corals by immersing them in a solution for 30 min / day for 7 consecutive days. Results show that the treated corals exhibit increased net weight gain, and the levels of redox-related enzymes, symbiotic zooxanthellae density, and chlorophyll a content rapidly recovered to or approached pre-operative normal levels. The increase in zooxanthellae density and chlorophyll a content indicates that fucoidan treatment can increase the photosynthetic activity of corals, providing sufficient energy for coral repair. The decrease in redox-related enzyme levels indicates that fucoidan treatment can directly remove some reactive oxygen species from corals, reducing oxidative damage and creating a favorable internal environment for tissue healing. In short, this invention discloses that fucoidan treatment can promote the survival rate of transplanted corals after cutting and optimize coral reef ecological restoration, which is of great significance for coral transplantation.

Description

Technical Field

[0001] This invention belongs to the field of coral reef repair technology, specifically, it relates to the application of fucoidan in the preparation of products for promoting physiological recovery after coral reef cutting. Background Technology

[0002] Coral reef ecosystems are among the most biodiverse ecosystems on Earth, playing vital ecological and service roles. However, due to multiple factors such as ocean acidification, global warming, and pollution, coral reefs worldwide are experiencing severe degradation. Currently, transplantation restoration based on asexual coral reproduction is one of the main conservation measures, with transplanting coral seedlings being the most common method.

[0003] However, the cutting and transplantation process creates trauma on the coral surface, triggering a strong oxidative stress response and leading to a large accumulation of reactive oxygen species (ROS). Excessive ROS damages coral cells and their symbiotic zooxanthellae, causing a decrease in zooxanthellae density and weakened photosynthetic capacity, which in turn affects energy supply, delays wound healing, and may even lead to transplantation failure. Therefore, promoting rapid postoperative repair and alleviating oxidative damage are key to improving transplantation survival rates.

[0004] Currently, there are no mature methods for post-operative coral treatment, relying mainly on the coral's own healing abilities and lacking effective external interventions. Although some studies have explored the use of antibiotics and probiotics to assist coral recovery, these methods have limitations such as complex operation, high cost, and the potential introduction of exogenous pollution. In recent years, natural polysaccharides have attracted attention in the field of tissue repair due to their good biocompatibility and bioactivity, but the evaluation of their effectiveness in coral post-operative repair remains a blank.

[0005] Fucoidan, also known as fucoidan or fucoidan sulfate (CAS: 9072-19-9), is a natural sulfated polysaccharide mainly found in the cell wall mucus of brown algae (such as kelp, wakame, and fucus vesiculosus), and also abundant in marine animals such as sea cucumbers and sea urchins. It is primarily composed of L-fucose linked by α-1,3 or α-1,4 glycosidic bonds. Its main applications include gastrointestinal protection (inhibiting Helicobacter pylori adhesion, promoting mucosal repair, etc.), immune enhancement (activating macrophages, NK cells, T / B lymphocytes, etc.), anti-tumor (inducing tumor cell apoptosis, inhibiting proliferation, etc.), anticoagulation (inhibiting prothrombin activation, inhibiting platelet aggregation, reducing blood viscosity, etc.), and antioxidant and anti-inflammatory effects (scavenging free radicals, resisting ultraviolet radiation, reducing oxidative damage). It is highly safe and widely used. Currently, fucoidan has not been used for postoperative physiological repair in corals. Summary of the Invention

[0006] The purpose of this invention is to overcome the above-mentioned defects and deficiencies in the prior art and to provide the application of fucoidan in promoting physiological recovery after coral cutting or in the preparation of products for promoting physiological recovery after coral cutting.

[0007] The second objective of this invention is to provide a method for promoting physiological recovery after coral cutting surgery.

[0008] The above-mentioned objective of this invention is achieved through the following technical solution: This invention aims to address the technical problem of the lack of effective methods to promote healing after the cutting and transplantation of reef-building corals. It provides a simple, efficient, and standardized post-operative treatment method for corals that can significantly promote post-operative coral growth, accelerate the recovery of the symbiotic system, and alleviate oxidative stress. Specifically, cutting corals are soaked in a fucoidan solution. Results show that, compared to the control group, *Gyrococcus pluvialis* corals soaked in fucoidan solution exhibit increased net weight gain, and redox-related enzymes, symbiotic zooxanthellae density, and chlorophyll a content can recover to or approach pre-operative normal levels more quickly. The increase in symbiotic zooxanthellae density and chlorophyll a content indicates that fucoidan treatment increases the photosynthetic activity of corals, providing sufficient energy for coral repair. The decrease in redox-related enzyme levels indicates that fucoidan treatment can directly remove some reactive oxygen species from corals, reducing their dependence on their own antioxidant system, thereby mitigating oxidative damage and creating a favorable internal environment for tissue healing. This invention provides a theoretical basis for the application of fucoidan in coral repair and solves the technical problem of the lack of effective treatment methods after the cutting of reef-building corals.

[0009] Therefore, the present invention provides the use of fucoidan in promoting physiological recovery after coral cutting or in the preparation of products for promoting physiological recovery after coral cutting.

[0010] Furthermore, the coral is a reef-building coral.

[0011] Furthermore, the reef-building coral is a shield-shaped gyro coral.

[0012] Furthermore, the physiological recovery includes increased net coral weight gain, decreased redox-related enzyme activity, and increased symbiotic zooxanthellae density and chlorophyll a content. The increase in symbiotic zooxanthellae density and chlorophyll a content indicates that fucoidan treatment increases the photosynthetic activity of corals, providing sufficient energy for coral repair; the decrease in redox-related enzyme levels indicates that fucoidan treatment can directly remove some reactive oxygen species from corals, reducing their dependence on their own antioxidant system, thereby mitigating oxidative damage and creating a favorable internal environment for tissue healing.

[0013] The present invention also provides a method for promoting physiological recovery after coral cutting, wherein the method involves soaking the cut coral seedlings in a fucoidan solution.

[0014] This invention, through systematic screening of fucoidan treatment concentration and soaking time, determined the optimal process parameters. The results showed that a fucoidan treatment regimen of 100 mg / L concentration, soaking for 30 min / day for 7 consecutive days, significantly promoted postoperative growth of *Gyrococcus pluvialis*, accelerated the recovery of symbiotic zooxanthellae, and alleviated oxidative stress, representing the optimal treatment conditions. This regimen is repeatable, exhibits stable effects, and shows promising application prospects.

[0015] Furthermore, the concentration of the fucoidan solution is 50–150 mg / L.

[0016] Preferably, the concentration of the fucoidan solution is 100 mg / L.

[0017] Furthermore, the solvent for the fucoidan is water with a salinity of 33.

[0018] Furthermore, the soaking treatment time is 10–60 min / day.

[0019] Preferably, the soaking treatment time is 30 min / day.

[0020] Furthermore, the soaking treatment lasts for 5 to 10 days.

[0021] Preferably, the soaking treatment lasts for 7 days.

[0022] Furthermore, the soaking conditions for the coral seedlings are a temperature of 27°C and a light cycle of 12h:12h.

[0023] Specifically, the coral seedlings were soaked in a 100 mg / L fucoidan solution for 30 min / day for 7 days at a temperature of 27°C and a photoperiod of 12h:12h.

[0024] The present invention also provides a preferred and feasible method for treating fucoidan to promote physiological recovery after coral reef cutting, the method comprising the following steps: S1. Coral seedling preparation: Prepare shield-shaped gyroscope corals (… Turbinaria peltata Cut into pieces with dimensions of 2×2×2cm 3 Cube-shaped coral seedlings, ready for use; S2. Preparation of Fucoidan Solution: Take fucoidan powder, dissolve it in coral culture water (salinity 33, pH 8.1, temperature 27℃) to prepare a solution with a concentration of 100 mg / L; S3. Soaking treatment: The coral seedlings prepared in step S1 are completely immersed in the fucoidan solution prepared in step S2 and soaked for 30 minutes under normal aquaculture conditions (temperature 27℃, photoperiod 12h:12h). S4. Recovery and Culture: After soaking, the coral seedlings are removed and returned to the conventional coral culture system for recovery and culture. S5. Repeat the process: Repeat S3 and S4 once a day for 7 consecutive days.

[0025] Compared with the prior art, the present invention has the following beneficial effects: This invention provides the application of fucoidan in the preparation of products to promote physiological recovery after coral cutting. The invention utilizes 100 mg / L fucoidan to treat cut corals by immersing them in a solution for 30 minutes / day for 7 consecutive days. Results show that the treated corals exhibit increased net weight gain, and redox-related enzymes, symbiotic zooxanthellae density, and chlorophyll a content rapidly recovered to or approached pre-operative normal levels. The increase in zooxanthellae density and chlorophyll a content indicates that fucoidan treatment increases the photosynthetic activity of corals, providing sufficient energy for coral repair. The decrease in redox-related enzyme levels indicates that fucoidan treatment can directly remove some reactive oxygen species from corals, reducing their dependence on their own antioxidant systems, thereby mitigating oxidative damage and creating a favorable internal environment for tissue healing. In other words, fucoidan treatment can promote rapid coral growth after cutting, restore the health of the symbiotic system, alleviate oxidative stress, improve coral transplant survival rate, and optimize coral reef ecological restoration technology. This invention provides a standardized operating procedure for coral transplantation, with clear process parameters, simple operation, and easy implementation, showing broad application prospects. Attached Figure Description

[0026] Figure 1 The effects of different concentrations and soaking times of fucoidan treatment on the net weight gain of *Gyrococcus pluvialis* seedlings after surgery were investigated. Figure 1 In the figures, 'a' represents the net weight gain on day 7; 'b' represents the net weight gain on day 15; and 'c' represents the net weight gain on day 30. Note: Different lowercase letters indicate significant differences between different concentration treatment groups within the same soaking time. P <0.05%, different capital letters indicate significant differences in soaking time within the same concentration ( P <0.05).

[0027] Figure 2 The effects of different concentrations and soaking times of fucoidan treatment on zooxanthellae density and chlorophyll a content in *Gymnodon salina* seedlings after surgery were investigated. Figure 2 In the figure, 'a' represents the change in zooxanthellae density on day 7; 'b' represents the change in zooxanthellae density on day 15; 'c' represents the change in zooxanthellae density on day 30; 'd' represents the change in chlorophyll a content on day 7; 'e' represents the change in chlorophyll a content on day 15; and 'f' represents the change in chlorophyll a content on day 30. Note: The dashed lines in the figure represent the zooxanthellae density and chlorophyll a content of the shield-shaped gyroscope coral when it is normal before surgery. Different lowercase letters indicate significant differences between different concentration treatment groups within the same immersion time.P <0.05%, different capital letters indicate significant differences in soaking time within the same concentration ( P <0.05).

[0028] Figure 3 The effects of different concentrations and soaking times of fucoidan treatment on the antioxidant enzyme activity and lipid peroxidation level of *Gyrococcus pluvialis* seedlings after surgery were investigated. Figure 3 In the figure, a represents SOD; b represents CAT; c represents GSH; and d represents LPO. The horizontal axis represents the concentration treatment groups at different immersion times; the dashed line in the figure represents the antioxidant enzyme activity and lipid peroxidation level of *Gyrococcus pluvialis* before surgery when it was normal. * indicates significant differences within the group (…). P <0.05), ** indicates extremely significant difference within the group (P<0.01); an asterisk without a line indicates significant difference within the group, and a line indicates significant difference between specific groups. Detailed Implementation

[0029] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0030] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0031] Example 1: Effect of polysaccharide treatment on net weight gain of corals I. Experimental Methods 1. Experimental materials Shield-shaped gyroscope coral ( Turbinaria peltata The samples were collected from the Xuwen Coral Reef National Nature Reserve in Guangdong Province (109°96′E, 20°23′N). After collection, they were temporarily held in the laboratory for two months under the following conditions: 300 L of water, water temperature 27℃, pH 8.1, salinity 33, and light intensity 200 μmol / m³. -2 s -1 Artificial seawater was prepared using coral salt (Instant Ocean) with a light-dark ratio of 12 h:12 h.

[0032] Fucoidan was purchased from Maclean Company (F885876), CAS: 9072-19-9.

[0033] 2. Experimental Design Cut the healthy coral into 2×2×2 cm pieces. 3Coral seedlings were randomly divided into nine treatment groups. A two-factor design was used: factor one was the concentration of fucoidan (50, 100, 150 mg / L), and factor two was the soaking time (10, 30, 60 min). A control group (without fucoidan treatment) was also included. Each group contained 14 coral seedlings. Soaking at the corresponding concentration and time was performed daily at 14:00 for 7 consecutive days, with routine maintenance during the remaining time. Three coral seedlings from each group were randomly selected before treatment (day 0), and at 7, 15, and 30 days after treatment to measure various indicators.

[0034] 3. Net weight gain determination: Three coral seedlings were selected from each group and weighed on days 0, 7, 15, and 30 using a hydrostatic balance (TP5002, Xiniu Technology). The weight gain relative to day 0 was calculated.

[0035] II. Experimental Results Net weight gain results as follows Figure 1 As shown, on postoperative day 7, the net weight gain in all treatment groups with 100 mg / L was significantly higher than that in the control group. P <0.05)( Figure 1 a). 15 days post-surgery, the net weight gain in the 100 mg / L × 30 min treatment group was significantly increased (0.257 ± 0.035 g). Figure 1 b). At 30 days post-surgery, the net weight gain in the 100 mg / L × 30 min treatment group (0.353 ± 0.032 g) was still significantly higher than that in the control group (0.240 ± 0.036 g), an increase of 47.1%. Figure 1 c).

[0036] Example 2: Effects of polysaccharide treatment on coral polyp and xanthophyll density and chlorophyll a content I. Experimental Methods 1. The experimental design is the same as in Example 1.

[0037] 2. Zooxanthellae density: Rinse coral seedlings with 100 mL of sterile seawater using a dental scaler and collect tissue fluid. Take 30 mL, centrifuge at 4000 rpm for 10 min, resuspend the precipitate in 5 mL of formaldehyde, count the cells using a hemocytometer, and divide the result by the coral surface area (aluminum foil method).

[0038] 3. Chlorophyll a content: Take another 30 mL of tissue fluid, centrifuge at 4000 rpm for 10 min, add 8 mL of methanol to the precipitate, extract overnight at 4℃, centrifuge again, take the supernatant, and measure the absorbance at 664, 630, and 750 nm using a spectrophotometer. Calculate the chlorophyll a content according to the formula Chl a (µg / mL) = 13.6849 × (A664 - A750) - 3.4551 × (A630 - A750), and normalize the result to the surface area.

[0039] II. Experimental Results Zooxanthindella density results as follows Figure 2 As shown in AC, 15 days after surgery, the zooxanthellae density in the 100 mg / L × 30 min treatment group reached its peak, significantly higher than that in the control group. P <0.05), and recovered to preoperative levels. 30 days postoperatively, the zooxanthellae density in this group was still significantly higher than that in the control group, approaching preoperative levels.

[0040] The results of chlorophyll a content are as follows: Figure 2 As shown in the data, 15 days post-surgery, the chlorophyll a content was highest in the 100 mg / L × 30 min treatment group, significantly higher than that in the control group (P < 0.05). This indicates that fucoidan not only promotes the recovery of zooxanthellae populations but also enhances their photosynthetic activity, providing sufficient energy for coral restoration.

[0041] Example 3: Effect of polysaccharide treatment on net weight gain of corals I. Experimental Methods 1. The experimental design is the same as in Example 1.

[0042] 2. Antioxidant enzymes and lipid peroxidation indicators: The tissue fluid supernatant was collected, and the activities of superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) content and lipid peroxidation (LPO) level were measured using a kit (Beijing Box, AKAO001M, etc.). At the same time, the total protein content was measured using the Bradford method, and the specific activity of enzymes was calculated.

[0043] II. Experimental Results Antioxidant indicators such as Figure 3 As shown, 7 days post-surgery, SOD, CAT, GSH, and LPO levels in the control group were significantly elevated; while in the 100 mg / L × 30 min treatment group, SOD, CAT, GSH, and LPO levels were significantly lower than in the control group, approaching pre-operative levels. This demonstrates that fucoidan can directly scavenge some reactive oxygen species, reduce corals' dependence on their own antioxidant system, thereby mitigating oxidative damage and creating a favorable internal environment for tissue healing.

[0044] In summary, the fucoidan treatment regimen of 100 mg / L concentration, 30 minutes of immersion daily for 7 consecutive days, significantly promotes postoperative growth of *Gyrococcus pluvialis*, accelerates the recovery of symbiotic zooxanthellae, and alleviates oxidative stress, representing the optimal treatment conditions. This regimen is repeatable, has stable effects, and shows promising application prospects. It provides a clear technical solution for the efficient transplantation of *Gyrococcus pluvialis*, ultimately realizing the application value of improving coral transplant survival rates and optimizing coral reef ecological restoration technology.

Claims

1. Application of fucoidan in promoting physiological recovery after coral cutting or in the preparation of products for promoting physiological recovery after coral cutting.

2. The application according to claim 1, characterized in that, The coral in question is a reef-building coral.

3. The application according to claim 2, characterized in that, The reef-building coral is a shield-shaped gyro coral.

4. The application according to claim 1, characterized in that, The physiological recovery includes increased net weight gain in corals, decreased activity of redox-related enzymes, and increased density of symbiotic zooxanthellae and chlorophyll a content.

5. A method for promoting physiological recovery after coral cutting surgery, characterized in that, The method involves soaking the cut coral seedlings in a fucoidan solution.

6. The method according to claim 5, characterized in that, The concentration of the fucoidan solution is 50–150 mg / L.

7. The method according to claim 5, characterized in that, The solvent for the fucoidan is water with a salinity of 33.

8. The method according to claim 5, characterized in that, The soaking treatment time is 10 to 60 minutes per day.

9. The method according to claim 5, characterized in that, The soaking treatment lasts for 5 to 10 days.

10. The method according to claim 5, characterized in that, The coral seedlings were soaked under the following conditions: temperature 27°C, light cycle 12h:12h.

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