Sea cucumber processing boiled liquid extract and application thereof

Abstract

The invention provides a sea cucumber processing boiled liquid extract and application thereof. The preparation process of the extract is simple, and the process is easy to operate. According to the preparation process disclosed by the invention, silt and foreign matters in the sea cucumber boiled liquid are removed by adopting a high-speed centrifugal machine, a large amount of salt and moisture in the sea cucumber boiled liquid are removed by adopting a nanofiltration technology, concentration and vacuum drying or freeze drying are performed by adopting a film evaporation technology, and dry powder, namely the extract, is obtained by crushing and sieving the obtained dry matter. The extract retains full nutritional ingredients such as sea cucumber protein, polysaccharide, saponin and the like, has anticoagulation and anti-fatigue functions, has the functions obviously superior to those of traditional dried sea cucumbers, and can be used for preparing food and functional food.

Description

Technical Field

[0001] This invention relates to the food extract and its uses, specifically to a sea cucumber processed boiled extract and its uses. Background Art

[0002] Sea cucumber (Ludeigothurea grisea), belonging to the class Holothuroidea, is most commonly known as the spiny sea cucumber (Apostichopus japonicas). It is a food with high nutritional value and health benefits. Sea cucumber has anticoagulant properties, relieves fatigue, improves immunity, and enhances the body's resistance to disease, making it very suitable for people in a sub-healthy state.

[0003] In the production and processing of sea cucumbers, to prevent autolysis by the sea cucumber's own autolytic enzymes, the sea cucumbers are first boiled in water to inactivate the enzymes, thus generating a large amount of boiling wastewater. The loss rates of total sugar, polysaccharides, and saponins in dried sea cucumbers are between 10% and 50%; the loss rates in salted sea cucumbers are all higher than 60% (Wang Jingyuan, Research on Evaluation Methods of Nutritional Quality of Sea Cucumbers, Master's Thesis, Shanghai Ocean University, 2019). This results in some nutrients from the sea cucumber entering the boiling liquid during the cooking process, such as proteins, polysaccharides, and saponins. Sea cucumber boiling liquid contains a large amount of nutrients, but it is generally discharged as wastewater, causing pollution to the surrounding environment. Utilizing modern scientific technologies and methods to recover and utilize the nutrients in sea cucumber boiling liquid for the development of functional foods is of significant value.

[0004] Current research on sea cucumber cooking liquid includes studies on the content of active ingredients and extraction and separation methods. However, the following problems exist: sea cucumber cooking liquid contains multiple active ingredients, but the content of these active ingredients is low, resulting in high recovery costs; some recycling processes are complex, involving numerous steps and requiring sophisticated equipment, while the product recovery rate is low; or only one component is recovered, lacking a systematic and rational multi-component recycling approach. Chinese invention patent application CN102382198A discloses a process for extracting sea cucumber mucopolysaccharides from sea cucumber processing liquid and preparing instant freeze-dried powder. This process first involves ultrafiltration desalting and concentration of the cooking liquid, followed by enzymatic hydrolysis of the ultrafiltration retentate, ethanol precipitation of the hydrolysate, and vacuum drying of the collected precipitate to obtain crude sea cucumber polysaccharides. Meanwhile, Chinese invention patent CN102406048B discloses a method for extracting sea cucumber glycoproteins from sea cucumber blanching liquid. This method involves concentrating the sea cucumber cooking liquid under reduced pressure, followed by ethanol precipitation. The precipitate is collected, dried, and powdered. The powder is then desalted with ethanol to remove the precipitate. Finally, the precipitate is obtained through protease hydrolysis and ultrafiltration to retain the extracted sea cucumber glycoprotein mixture. Furthermore, Chinese invention patents CN105616327B, CN104262451B, and CN105266075B involve methods for extracting sea cucumber saponins from sea cucumber cooking liquid, often employing molecular weight screening methods such as ethanol precipitation combined with ultrafiltration.

[0005] The extraction and preparation processes described above are cumbersome and complex, and the resulting nutrients are mostly single-category components such as polysaccharides, saponins, and polypeptides, while sea cucumber nutrients are combinations of these categories. This invention's patented preparation process uses a high-speed centrifuge to remove mud and foreign matter from the sea cucumber boiling liquid, employs nanofiltration technology to remove a large amount of salt and water, uses thin-film evaporation technology for concentration, and freeze-drying or spray drying to obtain a dried powder, i.e., the extract. This extract retains all the nutrients of sea cucumber, including protein, polysaccharides, and saponins, and compared with traditional dried sea cucumber, it has significant anticoagulant and anti-fatigue effects, making it suitable for preparing food and functional foods. Summary of the Invention

[0006] This invention provides a sea cucumber boiled extract, its composition, and its uses. The preparation process of the extract is as follows:

[0007] 1. Place the cleaned fresh sea cucumbers into water at 70-105℃ and heat for 10-30 minutes to obtain a decoction;

[0008] 2. The boiling liquid is centrifuged at high speed (12,000-16,000 rpm or higher) to purify it and obtain a clear filtrate.

[0009] 3. The clarified filtrate is passed through a 200-600 Dalton nanofiltration membrane for desalination and dehydration to obtain a concentrated desalinated solution;

[0010] 4. Desalting and dewatering solution is concentrated under reduced pressure using a thin-film membrane to obtain a concentrated solution with a solid content in the range of 10-40%.

[0011] 5. The concentrate is dried under vacuum at 60-80℃, or spray-dried, or freeze-dried to obtain a dried product with a water content of less than 6%;

[0012] 6. The dried material is pulverized and sieved to obtain a dry powder finer than a 40-mesh sieve, which is the sea cucumber extract.

[0013] The extract contains 2-4% total saponins from sea cucumber; 2.5-4.5% polysaccharides from sea cucumber; and more than 20% protein.

[0014] The total saponin content of the sea cucumber decoction extract is 2-4%, the polysaccharide content is 2.5-4.5%, and the protein content is greater than 20%. The sodium chloride content should be less than 8%, and the loss on drying should be less than 6%.

[0015] The extract is used in the preparation of food and functional food; the extract can be made into powders, solid beverages, liquid beverages, gel candies, compressed candies, tablets, soft capsules, hard capsules, and granules.

[0016] The extract and the food and functional food prepared therefrom, as well as the powders, solid beverages, liquid beverages, gel candies, compressed candies, tablets, soft capsules, hard capsules, and granules made therefrom, can be used by sub-healthy individuals and have the effects of delaying aging, relieving fatigue, improving immunity, improving memory, assisting in lowering blood lipids, assisting in lowering blood pressure, improving blood viscosity, relieving arteriosclerosis, and assisting in anti-tumor treatment.

[0017] The preparation process of the above-mentioned sea cucumber extract can be further described as follows:

[0018] 1. Remove the internal organs from fresh sea cucumbers and wash away the mud and sand. Because sea cucumbers live in shallow seas and feed on seaweed and other organisms, their bodies and surfaces contain a lot of mud, sand and other foreign objects. Therefore, after removing the internal organs, they still need to be thoroughly rinsed with clean water.

[0019] 2. Dehydrate the cleaned, live sea cucumbers in water at 70-105℃ for 10-30 minutes. Sea cucumbers with their internal organs removed have a water content of over 90% and contain abundant autolytic enzymes. Boiling the sea cucumbers removes about 80% of the water and deactivates the autolytic enzymes. Saponins and polysaccharides in sea cucumbers are easily soluble in water, and some water-soluble proteins, polypeptides, and amino acids also dissolve into the boiling liquid. Under heating conditions, sea cucumber fats, mostly unsaturated fatty acids, can also be transferred into the water. Traditional boiling of sea cucumbers is a necessary step in the preparation of dried and ready-to-eat sea cucumbers, but it also results in a significant amount of nutrients being released into the boiling liquid. This boiling liquid is discharged as waste, causing environmental pollution and the loss of valuable nutrients. Experimental analysis shows that the boiling process results in the loss of 20-40% of nutrients, especially saponins, which are more soluble in water and are lost more readily.

[0020] 3. Centrifugal purification of the boiling liquid: Because sea cucumbers still contain a significant amount of mud, sand, and foreign matter after harvesting and during processing, these substances gradually dissolve due to the sea cucumber's autolytic enzymes before cooking. During boiling, these substances detach as flaky particles and fall into the boiling liquid. Therefore, after removing the cooked sea cucumbers, the boiling liquid needs to be centrifuged using a high-speed centrifuge at 12,000 rpm or higher to remove impurities and obtain a clear filtrate.

[0021] 4. The clarified filtrate undergoes nanofiltration for desalination and dehydration. Since approximately 40% of the solids in the sea cucumber boiled liquid are salts, primarily sodium chloride, it's crucial to remove as much salt as possible to achieve acceptable levels when extracting nutrients from the boiled liquid. Ion exchange resin desalination is an option, but it's cumbersome, inefficient, and costly. Given the high salt content and large volume of the boiled liquid, ion exchange resin is unsuitable. Nanofiltration is the optimal choice, offering compact equipment, high efficiency, low cost, and continuous, automated operation. Nanofiltration uses 400-600 Da (Dalton) membranes, which can retain nutrients such as polysaccharides, saponins, and proteins while removing small molecules like sodium chloride and water, achieving both desalination and dehydration.

[0022] 5. Thin-film vacuum concentration: Nanofiltration solution is concentrated using a thin-film method to obtain a solids content of 10-40%. Because the nutrient content in sea cucumber boiled liquid is 1-3%, the concentration after nanofiltration is less than 10%. If conventional drying is used, the high drying temperature and long heating time can easily lead to nutrient degradation, affecting product quality. The thin-film vacuum concentration process uses a low solution heating temperature and short heating time, resulting in high efficiency and further increasing the solids content.

[0023] 6. Vacuum drying, freeze drying, or spray drying at 60-80℃ yields solids with a moisture content of less than 6%. Vacuum drying, freeze drying, or spray drying result in shorter heating times, higher drying efficiency, and easier pulverization and sieving of the dried material, thus ensuring product quality.

[0024] 7. The dried material is pulverized and sieved to obtain a dry powder finer than a 40-mesh sieve, which is the sea cucumber extract.

[0025] Methods for component analysis of sea cucumber extract

[0026] 1. The determination of total saponins in sea cucumber was performed using ultraviolet spectrophotometry.

[0027] The reference standard is ginsenoside Rb1, and the reagent is 5% vanillin-glacial acetic acid solution. Prepare the reference standard, sample, and reagents to appropriate concentrations. First, measure the reference standard to plot a standard curve. See [link to standard curve for ginsenoside Rb1] for details. Figure 1 Next, accurately pipette 0.2 mL of the above solution into 10 mL stoppered colorimetric tubes. After evaporating the reagents under reduced pressure, add 0.2 mL of 5% vanillin-glacial acetic acid solution to each test solution, followed by 0.8 mL of perchloric acid. Shake well and react in a 60°C water bath for 15 min, then cool in an ice-water bath. Dilute the cooled stoppered tubes with 5 mL of glacial acetic acid, mix well, and let stand at room temperature for 10 min. Measure the absorbance of an appropriate amount of the reaction solution at 546 nm and calculate the result from the standard curve.

[0028] 2. The determination of total polysaccharides in sea cucumber was performed using ultraviolet spectrophotometry.

[0029] Reference standard D(+) anhydrous glucose, reagents phenol, sulfuric acid. Prepare the reference standard, sample, and reagents to appropriate concentrations. First, measure the reference standard to plot a standard curve. Then, pipette 0.2 mL of the above reaction solution into each test tube and add distilled water to a volume of 2.0 mL. Add 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid (analytical grade). Let stand for 10 min, then vortex to mix thoroughly. Place the test tubes in a 40℃ water bath for 20 min. After cooling to room temperature, measure the absorbance of an appropriate amount of the reaction solution at 490 nm. Calculate the absorbance from the standard curve.

[0030] 3. Protein content was determined using the Kjeldahl method.

[0031] Instruction manual illustrations

[0032] Figure 1 Reference standard curve of ginsenoside Rb1

[0033] Figure 2 Comparison chart of rotor dwell time for related products Detailed Implementation

[0034] The following are specific implementation examples of the present invention, used to further illustrate the advantages and features of the present invention, but the present invention is not limited to the following implementation examples.

[0035] Example 1: Sea cucumber extracts obtained at different temperatures and rotation speeds

[0036] Fresh sea cucumbers, simulating wild sea cucumber growth of 6-8 years, were harvested, their internal organs removed, and the mud and sand rinsed off. They were then transferred to a boiling tank and dynamically boiled in water at 85℃ for 20 minutes. The boiling liquid was continuously introduced into a tubular centrifuge at 16,000 rpm to obtain a clear filtrate. The clear filtrate was then desalted and dehydrated using nanofiltration. The desalted and dehydrated liquid was then transferred to a membrane equipment for vacuum concentration; distillable liquids were removed, resulting in a concentrated liquid with a solids content of 25.3%. The concentrated liquid was then transferred to a vacuum drying device at 75℃ and a vacuum degree less than 0.09 MPa for 8 hours. The dried material was pulverized through a 40-mesh sieve to obtain a dry powder, which is the sea cucumber extract.

[0037] Example 2: Sea cucumber extracts obtained at different temperatures and rotation speeds

[0038] Fresh sea cucumbers, simulating wild sea cucumber growth of 3-5 years, were harvested, their internal organs removed, and the mud and sand rinsed off. They were then transferred to a boiling tank and dynamically boiled in water at 95℃ for 10 minutes. The boiling liquid was continuously introduced into a tubular centrifuge at 12000 rpm to obtain a clear filtrate. The clear filtrate was then desalted and dehydrated using nanofiltration. The desalted liquid was then transferred to a membrane equipment for vacuum concentration; the distillate was removed, yielding a concentrated liquid with a solid content of 34.3%. The concentrated liquid was then transferred to a vacuum drying device at 80℃ and a vacuum degree less than 0.09 MPa for 5 hours. The dried material was pulverized through a 40-mesh sieve to obtain a dry powder, which is the sea cucumber extract.

[0039] Example 3 Sea cucumber extract obtained by freeze-drying

[0040] Fresh sea cucumbers, typically farmed in shallow seas for two years, were harvested, eviscerated, rinsed of mud and sand, and then transferred to a boiling tank. They were dynamically boiled in water at 75°C for 25 minutes. The boiling liquid was continuously fed into a tubular centrifuge at 13,000 rpm to obtain a clear filtrate. The clear filtrate was then desalted and dehydrated using nanofiltration. The desalted and dehydrated liquid was then concentrated under reduced pressure using a membrane apparatus; the distillate was removed, yielding a concentrated liquid with a solid content of 19.7%. The concentrated liquid was then freeze-dried under the following conditions to obtain a dried product. The dried product was pulverized and passed through a 40-mesh sieve to obtain a dry powder, which is the sea cucumber extract.

[0041]

[0042] Example 4: Analysis of saponins, polysaccharides, and other components in sea cucumber extracts from Examples 1-3.

[0043] The sea cucumber extracts from the three samples were analyzed, and the results are as follows:

[0044] Example 1 Example 2 Example 3 Total saponins in sea cucumber (%) 3.52 3.10 2.21 sea ​​cucumber polysaccharide % 4.13 3.35 2.73 protein% 27.33 25.38 30.15 Sodium chloride % 5.8 4.7 6.5 Loss on drying % 5.5 5.3 5.8

[0045] Example 5: Anticoagulant effect of sea cucumber decoction extract

[0046] Experimental animal: SD rat.

[0047] Sea cucumber decoction extract powder, dried sea cucumber ultrafine powder, and fresh sea cucumber powder were dissolved in physiological saline, and the in vitro anticoagulant activity of each sample was observed. The method is as follows:

[0048] (1) Take 0.2g of each sample and dilute to a 10mL centrifuge tube (concentration of 20mg / mL), centrifuge at 4500r / min for 20min, and collect the supernatant for later use (this is the original solution); (2) Dilute 10, 100, and 1000 times according to the ratio for later use; (3) Add whole blood at a ratio of 1:9, mix well, and observe the blood coagulation state. In addition, add heparin sodium as a positive control.

[0049] The results showed that the original dry powder of sea cucumber boiled extract and its 10-fold dilution had significant anticoagulant effects. The original ultrafine powder of fresh sea cucumber also had significant anticoagulant effects. The results are shown in Table 3.

[0050] Table 3 Results of in vitro anticoagulation pretest of sea cucumber extract

[0051]

[0052] Note: √: Has anticoagulant effect; ×: Has no anticoagulant effect.

[0053] Compared with commercial dried sea cucumber and fresh sea cucumber ultrafine powder, sea cucumber boiled liquid extract has a stronger anticoagulant effect.

[0054] Example 6: Anti-fatigue effect of sea cucumber decoction extract

[0055] Fatigue rotundus test. Male Balb / c mice, 10 mice per group. The fatigue rotundus test measured the rotundus dwell time of each group of mice. Mice were placed on a rotating rod in the center of a cylinder. The mice were required to maintain their balance and follow the rotation of the rod to avoid slipping. When the mice fell, the instrument automatically recorded the rotundus dwell time to assess their fatigue tolerance. Thirty minutes after the last drug administration, the mice were placed on the fatigue rotundus apparatus, with the rotation speed set to 30 rpm. －1 After 3 days of continuous training, the formal experiment began. The time the mice remained on the rotundus was recorded, with the standard being 180 seconds without falling.

[0056] Animal rotata dwell time

[0057]

[0058]

[0059] GH, commercial dried sea cucumber ultrafine powder; HS, dried sea cucumber decoction extract powder; XH, fresh sea cucumber powder.

[0060] Compared with the normal control group, *P<0.05.

[0061] Comparison of rotor dwell time for related products (see) Figure 2

[0062] Commercially available dried sea cucumber ultrafine powder, dried sea cucumber decoction extract powder, and fresh sea cucumber powder all exhibited certain anti-fatigue effects. The sea cucumber decoction extract showed a stronger anti-fatigue effect than commercially available dried sea cucumber.

Claims

1. A sea cucumber decoction extract, wherein the preparation process of the extract is as follows: Step 1: Place the cleaned fresh sea cucumbers into water at 70-105℃ and heat for 10-30 minutes to obtain a decoction. Step 2: Centrifuge the boiling liquid at high speed (12000-16000 rpm) to purify it and obtain a clear filtrate; Step 3: The clarified filtrate is passed through a 200-600 Dalton nanofiltration membrane to obtain a desalted and dehydrated solution; Step 4: The desalted and dehydrated water solution is concentrated under reduced pressure via a thin-film membrane to obtain a concentrated solution with a solid content in the range of 10-40%. Step 5: The concentrate is vacuum dried at 60-80℃, or spray-dried or freeze-dried to obtain a dried product with a water content of less than 6%. Step 6: Crush the dried material and sieve it to obtain a dry powder finer than a 40-mesh sieve, which is the sea cucumber extract.

2. The sea cucumber decoction extract according to claim 1, characterized in that... The total saponin content of the extract is 2-4%.

3. The sea cucumber decoction extract according to claim 1, characterized in that... The extract contains 2.5-4.5% sea cucumber polysaccharides.

4. The sea cucumber decoction extract according to claim 1, characterized in that... The extract contains more than 20% protein.

5. The sea cucumber decoction extract according to claim 1, characterized in that... The sodium chloride content of the extract should be less than 8% and the loss on drying should be less than 6%.

6. The sea cucumber decoction extract according to claim 1, characterized in that... The extract contains 2-4% total saponins from sea cucumber, 2.5-4.5% polysaccharides from sea cucumber, and more than 20% protein. The sodium chloride content should be less than 8%, and the loss on drying should be less than 6%.

7. The sea cucumber decoction extract according to claims 1-6, for use in the preparation of food and functional food.

8. Preparations of powders, solid beverages, liquid beverages, gel candies, compressed candies, tablets, soft capsules, hard capsules, granules, and related formulations according to the extracts described in claims 1-6.

9. The sea cucumber decoction extract according to claims 1-6 is used for delaying aging, relieving fatigue, improving immunity, improving memory, assisting in lowering blood lipids, assisting in lowering blood pressure, improving blood viscosity, relieving arteriosclerosis, and assisting in anti-tumor treatment.

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