A nutritional composition based on marine shrimp peptides for anti-obesity and improving metabolic syndrome

Abstract

The present application relates to the field of biological medicine and functional food, and discloses a kind of nutrition composition based on marine shrimp peptide for anti-obesity and improving metabolic syndrome;Including: Chinese tube whip shrimp peptide active component and pharmaceutically acceptable adjuvant, the Chinese tube whip shrimp meat protein is used as raw material, and the Chinese tube whip shrimp meat protein is prepared by alkaline protease enzymolysis, and the Chinese tube whip shrimp meat protein contains peptide segment with molecular weight less than 1 kDa, and APPPPP, HPPPP and MPLPP can be detected.The pharmaceutical composition can reduce the weight and fat tissue weight under the state of obesity, improve dyslipidemia, abnormal liver function and liver steatosis, reduce oxidative stress level, improve intestinal mucosal barrier damage and intestinal permeability increase, and regulate intestinal flora structure change.The active component in the present application is clear in origin, and the preparation process is clear, has the synergistic effect from lipid metabolism regulation, liver protection to intestinal barrier improvement, and can be used for preparing medicine for preventing or treating obesity and improving metabolic syndrome.

Description

Technical Field

[0001] This invention relates to the fields of biomedicine and functional foods, specifically to a nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome. Background Technology

[0002] Obesity is a significant contributing factor to metabolic syndrome, often accompanied by dyslipidemia, hepatic steatosis, increased oxidative stress, and impaired intestinal barrier function, thereby affecting metabolic homeostasis. Therefore, developing naturally derived active ingredients with a proven track record and good safety profile for combating obesity and improving metabolic syndrome has significant application value.

[0003] In existing technologies, interventions for obesity mostly involve chemical drugs, nutritional regulators, or ordinary protein hydrolysates. Although some marine-derived bioactive peptides are used for lipid-lowering, anti-oxidation, or nutritional supplementation, these are usually limited to the broad category of marine peptides or proteolytic hydrolysates, with insufficient documentation regarding raw material sources, preparation conditions, key bioactive peptide segments, and their correlation with metabolic improvement effects.

[0004] Existing marine bioactive peptide formulations often exhibit only single lipid-lowering or nutritional supplementation effects due to their broad range of raw materials and unclear compositional boundaries. They struggle to achieve stable and synergistic regulation of lipid metabolism abnormalities, liver damage, oxidative stress, and intestinal barrier imbalances, resulting in unclear effects on improving obesity and metabolic syndrome. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a nutritional composition based on marine shrimp peptides for combating obesity and improving metabolic syndrome, thereby resolving the technical problems existing in the prior art.

[0006] The above-mentioned technical objective of the present invention is achieved through the following technical solution: A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome, comprising: Active peptide components and pharmaceutically acceptable excipients from the Chinese mantis shrimp; The active peptide component of the Chinese spiny shrimp is a polypeptide component obtained by alkaline protease hydrolysis of Chinese spiny shrimp meat protein.

[0007] Preferably, the active peptide component of the Chinese mantis shrimp is obtained by non-cellular biotransformation extraction technology, which is to use Chinese mantis shrimp meat protein as a substrate in a system without the participation of living cells, and to catalyze protein hydrolysis by alkaline protease. The conditions for alkaline protease hydrolysis include: a material-to-liquid ratio of 1:2, a hydrolysis temperature of 55°C, a hydrolysis pH of 8.0, an enzyme dosage of 7000 U / g, a hydrolysis time of 6 h, and a protein hydrolysis degree of 20% or higher in the hydrolysis product.

[0008] Preferably, the active peptide component of the Chinese mantis shrimp contains peptides with a molecular weight of less than 1 kDa.

[0009] Preferably, the active peptide component of the Chinese mantis shrimp contains APPPPP, HPPPP, or MPLPP.

[0010] Preferably, the pharmaceutical composition is used to reduce body weight, epididymal fat weight and perirenal fat weight in obese individuals, and to improve obesity-related lipid metabolism abnormalities. The lipid metabolism abnormalities include elevated triglycerides, elevated total cholesterol, elevated low-density lipoprotein cholesterol, and decreased high-density lipoprotein cholesterol.

[0011] Preferably, the pharmaceutical composition is used to improve obesity-related liver dysfunction, hepatic steatosis, and hepatic lipid accumulation; The abnormal liver function included elevated levels of alanine aminotransferase and aspartate aminotransferase.

[0012] Preferably, the pharmaceutical composition is used to improve oxidative stress levels; The improvement in oxidative stress levels includes reducing malondialdehyde (MDA) levels and increasing glutathione peroxidase (GLP) activity, catalase activity, and superoxide dismutase (SOD) activity.

[0013] Preferably, the pharmaceutical composition is used to improve intestinal mucosal barrier damage and increased intestinal permeability; The improvement in intestinal permeability includes reducing serum diamine oxidase levels and reducing lipopolysaccharide levels in the liver.

[0014] Preferably, the pharmaceutical composition is used to regulate obesity-related gut microbiota imbalance to improve gut microbiota structure changes induced by a high-fat diet.

[0015] In summary, the present invention has the following main beneficial effects: This application uses shrimp meat protein from the Chinese mantis shrimp (Sinapis sinensis) as raw material and prepares active peptide components from it using alkaline protease under specific enzymatic hydrolysis conditions. This achieves the goal of obtaining active ingredients with a clearly defined composition, clear origin, and low molecular weight short peptide characteristics. The active components not only contain peptides with a molecular weight less than 1 kDa, but also contain representative short peptides such as APPPPP, HPPPP, and MPLPP. The compositional characteristics of this enrichment of short peptides provide a stable material basis for subsequent antioxidant activity, metabolic regulation, and organ protection effects, avoiding the problems of vague sources of marine protein hydrolysates, unclear boundaries of active components, and difficulty in establishing a correspondence with specific functions in existing technologies.

[0016] By intervening in a high-fat diet-induced obesity model using the active peptide components of the Chinese whip shrimp, this application achieves the effects of reducing weight, decreasing epididymal and perirenal fat accumulation, and improving abnormalities in triglycerides, total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol. Simultaneously, by improving abnormalities in alanine aminotransferase and aspartate aminotransferase, and alleviating hepatic steatosis and lipid accumulation, it achieves a synchronous regulatory effect on obesity-related liver damage. In other words, this application does not only target a single weight or lipid-lowering indicator, but rather achieves a holistic intervention effect on metabolic syndrome-related abnormalities through the coordinated improvement of lipid metabolism abnormalities and liver pathological damage.

[0017] By enhancing the activities of glutathione peroxidase, catalase, and superoxide dismutase and reducing malondialdehyde (MDA) content through the active peptide components of the Chinese whip shrimp, this approach alleviates oxidative stress in the liver and intestines. Furthermore, by improving the structure of the ileal and colonic mucosa, reducing serum diamine oxidase and liver lipopolysaccharide levels, and regulating high-fat diet-induced changes in gut microbiota structure, this approach repairs the intestinal barrier, reduces endotoxin transport, and maintains gut-hepatic axis homeostasis. Therefore, this application achieves a continuous regulatory effect from weight control and metabolic improvement to organ protection through the synergistic effect of antioxidant regulation, intestinal barrier protection, and microbiota structure improvement, demonstrating a comprehensive technical advantage that distinguishes it from existing schemes that only have single nutritional supplementation or single lipid-lowering effects. Attached Figure Description

[0018] Figure 1 This invention relates to the effect of the dual-enzyme combination method on the degree of hydrolysis and DPPH free radical scavenging rate.

[0019] Figure 2 This is a prediction of the antioxidant activity of the peptides in this invention.

[0020] Figure 3 This invention relates to the molecular docking of peptides (A) APPPPP, (B) HPPPP and (C) MPLPP with Keap1.

[0021] Figure 4 This refers to the total antioxidant capacity and reducing power of the SCPs of this invention.

[0022] Figure 5 This refers to the change in mouse body weight during the modeling process of this invention.

[0023] Figure 6 This invention relates to the effects of SCPs on the morphology of mouse adipose tissue.

[0024] Figure 7 The effects of the SCPs of this invention on the levels of oxidative stress (MDA, GSH-Px, CAT, SOD) in the liver of mice.

[0025] Figure 8 The present invention describes the effects of SCPs on intestinal permeability (DAO, LPS) and oxidative stress levels (SOD, MDA) in mice.

[0026] Figure 9 The effects of the SCPs of this invention on liver function indicators (ALT, AST) in mice.

[0027] Figure 10 This invention relates to the effects of SCPs on mouse liver structure.

[0028] Figure 11 This invention relates to the effects of SCPs on the structure of the mouse ileum.

[0029] Figure 12 This invention relates to the effects of SCPs on the colonic structure of mice. Detailed Implementation

[0030] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0031] Example 1 refer to Figure 1-12 A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome, comprising: Shrimp meat from *Sinapis sinensis* was selected as the raw material. After pretreatment, a feed-liquid system was established, and protease was added for enzymatic hydrolysis to obtain the active peptide components of *Sinapis sinensis*. During the enzymatic screening process, the hydrolytic effects of trypsin, alkaline protease, papain, pepsin, and neutral protease under different conditions were compared, with the degree of hydrolysis and free radical scavenging rate used as screening indicators. The screening results showed that alkaline protease and trypsin had relatively better hydrolytic effects. Further investigation of dual-enzyme combinations revealed that while the combination of alkaline protease and trypsin improved some hydrolysis indicators, the DPPH free radical scavenging rate was still lower than that of alkaline protease alone. Considering both the hydrolytic effect and cost, alkaline protease was ultimately selected to prepare the active peptide components of *Sinapis sinensis*.

[0032] The degree of hydrolysis was determined using the OPA method, and the degree of protein hydrolysis was calculated by detecting the change in free amino content before and after enzymatic hydrolysis. The degree of hydrolysis is used to characterize the extent to which the shrimp meat protein of *Sinapis sinensis* is cleaved into low molecular weight peptides by alkaline protease.

[0033] The alkaline protease hydrolysis conditions were: a material-to-liquid ratio of 1:2, a temperature of 55°C, a pH of 8.0, an enzyme dosage of 7000 U / g, and a hydrolysis time of 6 hours. Under these conditions, the degree of hydrolysis and free radical scavenging rate of the alkaline protease hydrolysis products were both at optimal levels. Specifically, the free radical scavenging rate reached a high level when the enzyme dosage was 7000 U / g and the hydrolysis time was 6 hours. Further increasing the enzyme dosage or extending the hydrolysis time did not improve the hydrolysis effect; instead, it may have caused damage to the active site or resulted in enzyme waste.

[0034] After enzymatic hydrolysis, the hydrolysis products were collected, and the active component of *Sinapis sinensis* peptide was prepared in lyophilized powder form and stored at low temperature for later use. Finally, an alkaline protease was selected for the large-scale preparation of *Sinapis sinensis* peptide, and the lyophilized powder was collected and stored at -20°C for subsequent experimental use. Therefore, those skilled in the art can reproduce the *Sinapis sinensis* peptide active component identical to that of this application.

[0035] In the preparation process, the active peptide components of the Chinese mantis shrimp are obtained using a non-cellular biotransformation extraction technique. Specifically, using Chinese mantis shrimp meat protein as a substrate, in a cell-free system, alkaline protease catalyzes the directed hydrolysis of the protein, gradually breaking down large protein molecules into low-molecular-weight peptide components. By controlling the enzymatic hydrolysis time, enzyme dosage, and pH conditions, the enzymatic hydrolysis reaction is kept in a stable state, thereby obtaining an active product mainly composed of short peptides. Combined with the enzymatic hydrolysis conditions described in Example 1, a high degree of hydrolysis can be achieved within the preferred process range, thus ensuring the release efficiency of the target peptide fragments.

[0036] In this embodiment, the acquisition of the active peptide components of *Sinapis sinensis* is a non-cellular biotransformation extraction process. Specifically, in a cell-free system, using *Sinapis sinensis* meat protein as a substrate, alkaline protease catalyzes protein hydrolysis, gradually breaking down large protein molecules into low-molecular-weight peptide components. After enzymatic hydrolysis, the degree of hydrolysis of the hydrolysate is measured. The results show that under the conditions of a material-to-liquid ratio of 1:2, a temperature of 55℃, a pH of 8.0, an enzyme dosage of 7000 U / g, and a hydrolysis time of 6 h, the degree of protein hydrolysis reaches 20% or higher, and can even exceed 40%. This demonstrates that the above-mentioned non-cellular biotransformation process can effectively release short peptide components from *Sinapis sinensis* protein, providing a basis for the subsequent preparation of peptides with a molecular weight less than 1 kDa and representative peptides such as APPPPP, HPPPP, and MPLPP.

[0037] In the aforementioned non-cellular biotransformation process, by controlling temperature, pH, and enzyme dosage, the protein hydrolysis process is carried out under mild conditions, without introducing organic solvents or highly corrosive reagents, thus forming a key technology path for green manufacturing. Specifically, the enzymatic hydrolysis process uses water as the reaction medium, and the reaction conditions are moderate temperature and a weakly alkaline environment. After enzymatic hydrolysis, the active peptide components of *Triplophysa chinensis* can be obtained through simple separation and drying steps, thereby reducing the generation of harmful byproducts and lowering the environmental burden.

[0038] After obtaining the active peptide components from the Chinese mantis shrimp, their functional properties were systematically evaluated, forming a functional evaluation technology system integrated with the preparation process. This evaluation system includes in vitro antioxidant capacity testing, cellular-level oxidative stress protection experiments, and animal model validation. In in vitro evaluation, the active components' free radical scavenging ability was confirmed by measuring total antioxidant capacity and reducing power. In cell models, its protective effect on cell viability was verified through H2O2-induced oxidative damage experiments. In animal experiments, its effects on body weight, adipose tissue, blood lipids, liver function, oxidative stress indicators, and intestinal barrier function were evaluated using a high-fat diet-induced obesity model. This forms a continuous technical pathway from preparation to functional validation.

[0039] In vitro antioxidant capacity tests correspond to total antioxidant capacity and reducing power indicators; cellular level oxidative stress protection experiments correspond to LS174t cell survival rate indicators; animal model validation corresponds to indicators of changes in body weight, epididymal fat, perirenal fat, TC, TG, LDL-C, HDL-C, ALT, AST, MDA, GSH-Px, CAT, SOD, DAO, LPS, and gut microbiota structure.

[0040] Through the synergistic application of the above-mentioned non-cellular biotransformation extraction technology, key green manufacturing technology and functional evaluation technology, the active components of the Chinese whip shrimp peptide in this application not only have a clear source and a controllable preparation process, but also their functions can be verified through multi-level evaluation results, thereby ensuring that those skilled in the art can implement the technical solution of this application based on the contents of the specification.

[0041] The preparation process uses water as the reaction medium and is carried out under mild temperature and weak alkaline conditions. No organic solvents or strong corrosive reagents are introduced. After enzymatic hydrolysis, the active peptide components of Chinese whip shrimp are obtained by separation and drying.

[0042] Example 2 The free amino acid composition of the active peptide component of *Triplophysa chinensis* prepared in Example 1 was analyzed. Seventeen amino acids were detected in the active component, including seven essential amino acids and six flavor amino acids. The amino acids with higher content were arginine (34.627 mg / g), phenylalanine (25.455 mg / g), glycine (24.451 mg / g), and tyrosine (23.467 mg / g). Essential amino acids accounted for 39.36% of the total amino acid mass, while non-essential amino acids accounted for 60.64%. The relevant results are shown in Table 1.

[0043] Table 1. Amino acid composition of SCPs

[0044] Note: * indicates essential amino acids, # indicates flavor amino acids. Therefore, it can be seen that the active peptide component of the Chinese mantis shrimp not only has the characteristics of polypeptide components, but also has a clear amino acid composition basis.

[0045] The active components of the *Sinapis sinensis* peptides obtained in Example 1 were detected using ultra-high performance liquid chromatography-tandem Q-TOF mass spectrometry (UHPLC-QTOF). De novo sequencing was used to analyze the mass spectrometry data to identify its peptide composition. Under ALC ≥ 95%, 17 high-confidence peptides were identified, with lengths ranging from 4 to 8 amino acids and molecular weights all less than 1 kDa, indicating that the active components belong to a low-molecular-weight, short-peptide enrichment group. The relevant results are shown in Table 2.

[0046] Table 2. Peptides identified in SCPs

[0047] Among the aforementioned high-confidence peptides, APPPPP, HPPPP, and MPLPP were detected. Specifically, APPPPP had an ALC of 98%, a length of 6 amino acids, and a molecular weight of 574.3115; HPPPP had an ALC of 98%, a length of 5 amino acids, and a molecular weight of 543.2805; and MPLPP had an ALC of 97%, a length of 5 amino acids, and a molecular weight of 553.2934.

[0048] Furthermore, among the 17 peptides with ALC ≥ 95%, 15 contain proline, 9 contain alanine, and 9 contain leucine, indicating a high proportion of hydrophobic amino acid residues in this active component. Combined with the known pattern that hydrophobic residues are beneficial for bioactive peptides to exert antioxidant or metabolic regulatory effects, it can be concluded that the active component of the Chinese whip shrimp peptide in this application already possesses the structural basis for subsequently exerting antioxidant and anti-obesity effects.

[0049] Example 3 Bioinformatics analysis was performed on the high-confidence peptides identified in Example 2 to screen for potential key active peptides. Among the 17 peptides with ALC ≥ 95%, 7 peptides had an activity score ≥ 0.5, including 3 peptides with a free radical scavenging ability score ≥ 0.5: HPPPP, APPPPP, and MPLPP. This result indicates that these three peptides were not only detected but also scored highly in activity prediction, and can be considered representative key peptides among the active peptide components of *Sinapis sinensis*.

[0050] Molecular docking analysis was performed on HPPPP, APPPPP, and MPLPP with Keap1 protein. APPPPP forms four hydrogen bonds with Keap1, HPPPP forms two hydrogen bonds, and MPLPP forms three hydrogen bonds. Furthermore, APPPPP interacts with residues such as VAL-467, VAL-420, CYS-368, THR-560, VAL-465, ASN-381, THR-388, and SER-391. Since the Keap1-Nrf2-ARE pathway is closely related to cellular antioxidant defense, these docking results demonstrate a clear correlation between the key short peptides in the active peptide components of *C. sinensis* and their antioxidant capacity.

[0051] This embodiment describes HPPPP, APPPPP, and MPLPP as representative key peptide segments in the active components of Chinese prawn peptides, rather than completely separating them from the overall active components and defining them as the sole active ingredients.

[0052] Example 4 The total antioxidant capacity and reducing power of the active peptide component of *Triplophysa chinensis* prepared in Example 1 were tested in vitro. When the concentration of the active component was 10 mg / mL, its reducing power was 0.59 and its total antioxidant capacity was 0.86. Moreover, the total antioxidant capacity and reducing power showed an increasing trend with the increase of sample concentration.

[0053] The cellular safety of the active peptide components of *Viburnum chinense* was investigated using LS174t cells as an in vitro cell model. Within a concentration range of 200 μg / mL to 1400 μg / mL, the active components showed no cytotoxicity to LS174t cells. Within a concentration range of 400 μg / mL to 1000 μg / mL, the relative cell proliferation rate showed an increasing trend, reaching a high level at 1000 μg / mL. Based on these results, 200 μg / mL, 600 μg / mL, and 1000 μg / mL were selected as subsequent evaluation concentrations.

[0054] The viability of LS174t cells pretreated with active peptides from *C. sinensis* under H2O2-induced oxidative damage conditions was evaluated. Compared with the oxidative damage group, low, medium, and high concentrations of active peptides from *C. sinensis* significantly improved the viability of LS174t cells, with no significant difference between 600 μg / mL and 1000 μg / mL. These results indicate that the active component not only possesses antioxidant capacity in in vitro chemical evaluation but also exerts a protective effect against oxidative stress damage at the cellular level.

[0055] The above in vitro test results corroborate the activity prediction and molecular docking results of the key peptides in Example 3, jointly demonstrating that the low molecular weight short peptides, especially HPPPP, APPPPP and MPLPP peptides, in the active components of Chinese prawn peptides are intrinsically related to the antioxidant properties of the active components.

[0056] Example 5 A high-fat diet-induced obesity model was established using C57BL / 6J mice. The mice were first induced with a high-fat diet for 7 weeks to establish the obesity model, followed by continuous gavage administration of the drug for 4 weeks. Experimental groups included a normal diet group (ND), a model group (MOD), a positive control group (PC), a low-dose group of Chinese prawn peptide (SCPs-L), and a high-dose group of Chinese prawn peptide (SCPs-H), with 10 mice in each group.

[0057] Weight changes during modeling, such as Figure 5 As shown in the table, the average weight of mice in the high-fat diet group reached approximately 32g, which was 1.23 times that of mice in the normal diet group, indicating that the obesity model was successfully established. The weight results of mice in each group after 4 weeks of drug administration are shown in Table 4-3. The final weight of the ND group was 27.150±1.25g, the MOD group was 29.892±1.11g, the PC group was 28.353±2.00g, the SCPs-L group was 27.438±1.35g, and the SCPs-H group was 27.204±1.82g. It can be seen that the final weights of the SCPs-L and SCPs-H groups were lower than those of the MOD group and close to those of the ND group, indicating that the active components of the Chinese whip shrimp peptide can reduce body weight in obese individuals.

[0058] During the administration period, the food intake in the ND group was significantly higher than that in other groups, while the food intake in the MOD group was significantly lower than that in the ND and PC groups. The food intake in the SCPs-L and SCPs-H groups was not significantly different from that in the PC group. This suggests that the weight loss induced by the active peptide components of *Sinapis sinensis* is not simply achieved through appetite suppression, but is more likely related to its regulation of lipid metabolism, liver function, and intestinal function.

[0059] The results of the weight of mouse organs and adipose tissue are shown in Table 3.

[0060] Table 3 Weight of mouse organs and adipose tissue

[0061] The epididymal fat weight in the MOD group was 1.119±0.34g, and the perirenal fat weight was 0.309±0.10g; in the SCPs-L group, the epididymal fat weight was 0.776±0.17g, and the perirenal fat weight was 0.211±0.09g; and in the SCPs-H group, the epididymal fat weight was 0.775±0.19g, and the perirenal fat weight was 0.195±0.07g. Compared with the MOD group, treatment with the active component of *Sinapis sinensis* peptide significantly reduced both epididymal and perirenal fat, indicating that this active component can reduce fat formation and adipose tissue accumulation.

[0062] Example 6 Serum lipid metabolism and liver function parameters were measured in mice of each group. Compared with the ND group, the MOD group showed significantly increased serum TC, TG, LDL-C, ALT, and AST levels, and significantly decreased HDL-C levels. Compared with the MOD group, the SCPs-H group significantly improved TC, TG, LDL-C, ALT, AST, and HDL-C levels, while the SCPs-L group showed no significant improvement in ALT, TG, and HDL-C levels. There were no significant differences in AST, ALT, LDL-C, and HDL-C levels between the SCPs-H group and the PC group.

[0063] The above results indicate that the active peptide components of *Sinapis sinensis* can not only improve lipid metabolism abnormalities induced by a high-fat diet in mice, but also improve abnormal liver function. Specifically, changes in TC, TG, LDL-C, and HDL-C directly correspond to the lipid metabolism abnormalities described in the claims, while changes in ALT and AST directly correspond to the abnormal liver function described in the claims. These results form a complete logical chain of model establishment, abnormal indicators, and intervention improvement.

[0064] H&E staining results of epididymal adipose tissue are as follows Figure 6 As shown in the figure, compared with the MOD group, the volume of adipocytes was significantly reduced after treatment with the active component of Chinese whip shrimp peptide, indicating that the active component can alleviate fat accumulation, which is corroborated by the results of body weight and adipose tissue weight.

[0065] Example 7 The appearance, Oil Red O staining, and H&E staining of the livers of mice in each group were observed. The livers of mice in the ND group were dark red with no obvious fat spots on the surface and sharp liver margins. The livers of mice in the MOD group were light red with an oily feel on the cut surface, indistinct outlines, and a softer texture. The livers of the SCPs-treated groups, especially the SCPs-H group and the PC group, were closer in appearance to the ND group. Oil Red O staining showed a significant increase in lipid droplets in the MOD group, while a significant decrease in lipid droplets was observed in the SCPs-treated groups, especially the SCPs-H group. H&E staining showed that the liver tissue structure of the MOD group was disordered, with diffuse fat vesicles of different sizes and loose cytoplasm, while the liver tissue structure of the SCPs-H group was significantly restored, and the number of fat vesicles was significantly reduced.

[0066] The active peptide components of *Trichoderma sinense* can improve hepatic steatosis and lipid accumulation induced by a high-fat diet. This result, together with the improvements in blood lipids and liver function reported in Example 6, indicates that the active components in this application do not merely cause changes in a single biochemical indicator, but rather have a practical ameliorative effect on obesity-related liver pathological changes.

[0067] The level of oxidative stress in liver tissue was measured, and the results are shown in [the table below]. Figure 7 Compared with the ND group, the MOD group showed a significant increase in MDA content in the liver of mice, while the activities of GSH-Px, CAT, and SOD were significantly decreased. Compared with the MOD group, both the SCPs-H group and the PC group showed significant increases in GSH-Px, CAT, and SOD activities, and decreased MDA content. These results indicate that the active components of the Chinese whip shrimp peptides can enhance the liver's antioxidant defense system, reduce lipid peroxidation damage, and thus alleviate obesity-related liver oxidative stress.

[0068] The results of Examples 3 and 4 further illustrate that the active component of the Chinese whip shrimp peptide in this application can improve liver oxidative stress because it contains low molecular weight short peptides and has in vitro antioxidant and cell protection capabilities.

[0069] Example 8 Morphological observations were performed on the ileum and colon tissues of mice in each group. In the ileum tissue, compared with the ND group, the MOD group had shallower crypt depth, incomplete and shortened intestinal villi; after treatment in the SCPs-H group and PC group, the length of intestinal villi increased, the number of goblet cells increased, and the pathological damage to the intestine was reduced.

[0070] In the colon tissue, the ND group showed intact crypt structure with a large number of goblet cells; the MOD group showed altered crypt structure with branching, destruction of goblet cells and part of the mucosal layer, and abnormal inflammatory infiltration in some areas; after SCPs treatment, the integrity of the crypt structure was restored, the number of goblet cells increased, and mucin secretion and mucosal barrier formation were improved, with the SCPs-H group showing better results.

[0071] Further testing of intestinal permeability-related indicators and intestinal local oxidative stress indicators was conducted, and the results are shown in [the table below]. Figure 8 Compared with the ND group, the MOD group showed significantly increased serum DAO levels, significantly increased liver LPS content, increased MDA content in colon tissue, and decreased SOD activity. Compared with the MOD group, the active components of the Chinese mantis shrimp peptide significantly reduced serum DAO and liver LPS, increased colonic SOD activity, and decreased colonic MDA content. This indicates that the active components can reduce endotoxin transport caused by increased intestinal permeability and alleviate local oxidative stress in the intestine, thereby improving intestinal mucosal barrier damage.

[0072] The above results indicate that the improvement of the intestinal barrier by the active component of the Chinese prawn peptide in this application is not only reflected in the restoration of tissue morphology, but also in the decrease of functional indicators DAO and LPS, as well as the improvement of colonic oxidative stress. Therefore, it can directly correspond to the technical features of improving intestinal mucosal barrier damage and improving intestinal permeability in the claims.

[0073] Example 9 Diversity and composition analysis of the gut microbiota in each group of mice was performed. In the α-diversity and β-diversity analyses, intervention with SCPs partially reversed the changes in gut microbiota structure induced by a high-fat diet, indicating that the active components of *Sinapis sinensis* peptides have a regulatory effect on high-fat diet-induced gut microbiota dysbiosis.

[0074] At the phylum level, the mouse gut microbiota mainly consisted of Firmicutes, Verrucomicrobiota, and Actinobacteria. Compared with the ND group, the MOD group showed an increased relative abundance of Firmicutes, while the relative abundance of Verrucomicrobiota and Actinobacteria decreased. Administration of the active peptide component of *Sinapis sinensis* reversed these changes to some extent. At the family level, the relative abundance of bacteria such as *Eubacterium coprostanoligenes*, *Akkermansiaceae*, and *Lactobacillaceae* was increased in the SCPs group.

[0075] At the genus level, compared with the ND group, the relative abundance of Faecalibaculum and Eubacterium_nodatum_group decreased in the MOD group, while the relative abundance of Turicibacter and Coriobacteriaceae_UCG-002 increased; compared with the MOD group, the relative abundance of norank_f_Eubacterium_coprostanoligenes_group and Turicibacter increased in the SCPs group, while the abundance of Coriobacteriaceae_UCG-002 and Negativibacillus decreased.

[0076] Clinical factor association analysis further revealed correlations between lipid metabolism, liver function, and intestinal permeability-related indicators and the abundance of various specific bacterial communities. This indicates that the active components of the Chinese spiny shrimp peptide improve obesity and metabolic syndrome-related abnormalities not only through changes in host weight, blood lipids, and liver indicators, but also through the regulation of changes in gut microbiota structure, thus forming a continuous chain of action involving microbiota regulation, intestinal barrier protection, LPS reduction, and liver improvement.

[0077] Example 10 Examples 1 to 9 show that the active peptide component of *Sinapis sinensis* in this application has a clear source. It is a polypeptide component prepared by alkaline protease under the conditions of a material-to-liquid ratio of 1:2, a temperature of 55 ℃, a pH of 8.0, an enzyme dosage of 7000 U / g, and an enzymatic hydrolysis time of 6 h, using *Sinapis sinensis* meat protein as raw material. Its composition is clear, containing short peptides with a molecular weight of less than 1 kDa, and key peptide segments such as APPPPP, HPPPP, and MPLPP were detected.

[0078] The active components of the Chinese whip shrimp peptides described herein possess total antioxidant capacity, reducing power, and protective effects against oxidative damage to cells in vitro. In vivo, they can reduce body weight, epididymal fat weight, and perirenal fat weight in high-fat diet-induced obese mice; improve abnormalities in TC, TG, LDL-C, HDL-C, ALT, and AST; alleviate hepatic steatosis and hepatic lipid accumulation; increase GSH-Px, CAT, and SOD activities and reduce MDA content; improve the ileal and colonic mucosal barrier; reduce DAO and LPS; and regulate changes in intestinal flora structure. Therefore, those skilled in the art can realize the active components of the Chinese whip shrimp peptides described in the claims and their use in the preparation of drugs for the prevention or treatment of obesity and the improvement of metabolic syndrome.

[0079] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome, characterized in that, include: Active peptide components and pharmaceutically acceptable excipients from the Chinese mantis shrimp; The active peptide component of the Chinese spiny shrimp is a polypeptide component obtained by alkaline protease hydrolysis of Chinese spiny shrimp meat protein.

2. The nutritional composition for anti-obesity and improvement of metabolic syndrome based on marine shrimp peptides according to claim 1, characterized in that, The active peptide components of the Chinese spiny shrimp were obtained using a non-cellular biotransformation extraction technology. This non-cellular biotransformation extraction technology involves using Chinese spiny shrimp meat protein as a substrate in a system without the participation of living cells, and catalyzing protein hydrolysis through alkaline protease. The conditions for alkaline protease hydrolysis include: a material-to-liquid ratio of 1:2, a hydrolysis temperature of 55°C, a hydrolysis pH of 8.0, an enzyme dosage of 7000 U / g, a hydrolysis time of 6 h, and a protein hydrolysis degree of 20% or higher in the hydrolysis product.

3. The nutritional composition for anti-obesity and improvement of metabolic syndrome based on marine shrimp peptides according to claim 2, characterized in that, The active peptide component of the Chinese whip shrimp contains peptides with a molecular weight of less than 1 kDa.

4. The nutritional composition for anti-obesity and improvement of metabolic syndrome based on marine shrimp peptides according to claim 3, characterized in that, The active peptide components of the Chinese prawn contain APPPPP, HPPPP, or MPLPP.

5. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome according to claim 4, characterized in that, The pharmaceutical composition is used to reduce body weight, epididymal fat weight and perirenal fat weight in obese individuals, and to improve obesity-related lipid metabolism abnormalities. The lipid metabolism abnormalities include elevated triglycerides, elevated total cholesterol, elevated low-density lipoprotein cholesterol, and decreased high-density lipoprotein cholesterol.

6. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome according to claim 5, characterized in that, The pharmaceutical composition is used to improve obesity-related liver dysfunction, hepatic steatosis, and hepatic lipid accumulation. The abnormal liver function included elevated levels of alanine aminotransferase and aspartate aminotransferase.

7. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome according to claim 6, characterized in that, The pharmaceutical composition is used to improve oxidative stress levels; The improvement in oxidative stress levels includes reducing malondialdehyde (MDA) levels and increasing glutathione peroxidase (GLP) activity, catalase activity, and superoxide dismutase (SOD) activity.

8. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome according to claim 7, characterized in that, The pharmaceutical composition is used to improve intestinal mucosal barrier damage and increased intestinal permeability; The improvement in intestinal permeability includes reducing serum diamine oxidase levels and reducing lipopolysaccharide levels in the liver.

9. A nutritional composition based on marine shrimp peptides for anti-obesity and improvement of metabolic syndrome according to claim 8, characterized in that, The pharmaceutical composition is used to regulate obesity-related gut microbiota imbalance to improve gut microbiota structure changes induced by a high-fat diet.

Images