Hemp seed protein peptide for improving mental and physical fatigue and preparation method and application thereof

Abstract

This invention provides a hemp seed protein peptide for improving mental and physical fatigue, its preparation method, and its applications. It belongs to the field of hemp seed protein peptide technology. The method includes extracting hemp seed protein solution using an alkali-soluble acid precipitation method; performing three-step enzymatic hydrolysis steps using Serratia marcescens, thermophilic protease, and flavor protease to obtain the final hydrolysate; and performing post-treatment and drying to obtain the final product. Clinical studies of the hemp seed protein peptide prepared by this invention have shown that: 1. It significantly relieves mental fatigue: promotes nitric oxide production, increases brain oxygen and blood supply; rapidly boosts spirits within 15 minutes, improves concentration and reaction time; and maintains energy levels for a long time, improving lethargy. 2. It significantly relieves physical fatigue: rapidly reduces lactic acid and urea nitrogen accumulation, relieves muscle soreness; increases energy reserves, provides sustained anti-fatigue, extends endurance by 2-4 hours, and maintains physical strength; significantly shortens the recovery period after excessive fatigue, improves postoperative limb heaviness and soreness, and promotes immune recovery.

Description

Technical Field

[0001] This invention relates to the field of hemp seed protein peptide technology, specifically to a hemp seed protein peptide that improves mental and physical fatigue, its preparation method, and its application. Background Technology

[0002] Hemp seeds, belonging to the genus Cannabis in the family Moraceae, are annual herbaceous plants. The fruit is oval, smooth, grayish-green or grayish-yellow, with fine white, brown, or black markings; the leaves are relatively large, light green, with fine lines on the surface; the flowers are small, pale yellow, smooth, and polygonal; the fruit is small, dark brown, and has a hard shell; flowering occurs from May to June, and fruiting from July to August. The *Shennong Bencao Jing* (Shennong's Classic of Materia Medica) records that "hemp seeds are sweet and neutral in nature, rich in lipids, and function to lubricate the intestines and relieve constipation, while also having a slight nourishing effect, making them suitable for the elderly, postpartum women, and those with constipation due to insufficient body fluids and blood."

[0003] Currently, athletes, those engaged in high-intensity mental and physical labor, and post-operative patients commonly experience fatigue accumulation and slow recovery. Developing safe, efficient, and side-effect-free natural anti-fatigue products has become a research hotspot. Hemp seeds, as a food and medicine source, are high in protein and have a well-balanced amino acid composition. Hemp seed protein peptides, prepared through enzymatic hydrolysis, are easily absorbed and have strong bioactivity, exhibiting significant advantages in anti-fatigue, antioxidant, and endurance enhancement, and possess promising development prospects.

[0004] Chinese invention patent CN112410393B, entitled "A Hemp Seed Active Peptide and Its Preparation Method and Application," discloses a hemp seed active peptide and its preparation method and application. The preparation method involves first mixing and pulverizing pre-cooled defatted hemp seed meal with dry ice to obtain hemp seed meal powder. Then, water and calcium carbonate are added to prepare an alkaline suspension, which is then ground and sieved. Next, a two-step enzymatic hydrolysis is performed using a mixed protease and a neutral protease. Finally, the enzymes are inactivated, filtered, centrifuged, decolorized, and sterilized to obtain the hemp seed active peptide.

[0005] The active peptides obtained by the above preparation method are mainly effective in anti-oxidation. Further research is needed on hemp seed protein peptides that have fatigue-relieving and immunity-enhancing effects. Summary of the Invention

[0006] In view of this, the present invention provides a hemp seed protein peptide that improves mental and physical fatigue, its preparation method and application, so as to achieve the purpose of the obtained hemp seed protein peptide having anti-fatigue and improving immunity.

[0007] To achieve the above objectives, the present invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, comprising the following steps: S1. Pulverize defatted hemp seed meal and extract hemp seed protein solution using alkali dissolution and acid precipitation method; S2. Adjust the temperature and pH of the hemp seed protein solution, and hydrolyze it with Serratia marcescens to obtain a hydrolysate; adjust the temperature and pH of the hydrolysate, add thermophilic protease, and continue hydrolysis to obtain a secondary hydrolysate; adjust the temperature and pH of the secondary hydrolysate, add flavor protease, and perform a third hydrolysis to obtain the final hydrolysate. S3. The final enzymatic hydrolysate is post-treated and dried to obtain the anti-fatigue hemp seed protein peptide.

[0008] Optionally, the temperature and pH of the hemp seed protein solution are adjusted to 45-50℃ and 8-9; the amount of serrata peptide enzyme added is 0.5-1% of the hemp seed protein mass, and the enzymatic hydrolysis reaction time is 2.5-3.5h.

[0009] Optionally, the temperature and pH of the hydrolysate are adjusted to 55-60°C and 7-8; the amount of thermophilic protease added is 1.5-2.5% of the protein content of hemp seed, and the hydrolysis reaction time is 1.5-2.5 h.

[0010] Optionally, the temperature and pH of the secondary enzymatic hydrolysate are adjusted to 45-55°C and 6.5-7.5; the amount of flavor protease added is 1-2% of the protein content of hemp seeds, and the enzymatic hydrolysis reaction time is 1-2 hours.

[0011] Optionally, the alkali dissolution and acid precipitation method includes adding an alkali solution to defatted hemp seed meal, stirring and extracting, centrifuging and taking the supernatant, adjusting the pH of the supernatant with an acid solution to precipitate the protein, centrifuging and collecting the hemp seed protein precipitate; redissolving the protein precipitate in water to obtain a hemp seed protein solution.

[0012] Optionally, the alkaline solution is a sodium hydroxide solution with a pH of 9-10.0; the material-to-liquid ratio during stirring extraction is 1:(12-17), the stirring extraction time is 1-2 hours, and the temperature is 45-55℃; the acid solution is a 1-1.5 mol / L hydrochloric acid solution, and the pH of the supernatant is adjusted to 4.5-6.5.

[0013] Optionally, the post-processing includes sterilizing the final enzymatic hydrolysate, centrifuging and taking the supernatant, passing the supernatant through an ultrafiltration membrane, and then freeze-drying the filtrate under vacuum to obtain the anti-fatigue hemp seed protein peptide.

[0014] Optionally, the sterilization is performed by water bath sterilization at 90~95℃ for 10~15 minutes, and the centrifugation is performed at a speed of 7000~9000 rpm for 15~25 minutes.

[0015] To achieve the above objectives, the present invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, and the resulting anti-fatigue hemp seed protein peptides include the following amino acid sequences: SEQ ID NO.1: MARNF, SEQ ID NO.2: MDTRLL.

[0016] To achieve the above objectives, the present invention provides the application of hemp seed protein peptides, which improve mental and physical fatigue, in the preparation of anti-fatigue foods or health products.

[0017] The above-described technical solution of the present invention has at least the following beneficial effects: 1. The technical solution provided by the present invention uses Serratia peptides for internal digestion, and the enzymatic hydrolysis efficiency is high. On the basis of Serratia peptides hydrolysis, thermophilic protease is used for further enzymatic hydrolysis, and then flavor protease is used for external digestion and debittering. The three different hydrolytic enzymes work together to efficiently obtain specific hemp seed protein peptides.

[0018] 2. As demonstrated in the experimental examples, analysis of serum samples from subjects using immune cell markers CD3+ and CD3+ / CD4+ showed that the hemp seed protein peptides prepared in this invention can promote the improvement of the subjects' immune capacity, i.e., they have an immune recovery effect. Assessment of postoperative fatigue and sleep quality indices in subjects showed that the hemp seed protein peptides prepared in this invention significantly reduced fatigue scores and significantly improved sleep quality indices. In conclusion, the hemp seed protein peptides prepared in this invention have anti-fatigue and immune-enhancing effects. Attached Figure Description

[0019] Figure 1 This is a graph showing the difference in the percentage of CD3+ cells in each group of subjects before and after treatment in the experimental examples of this invention; Figure 2 This is a graph showing the difference in the percentage of CD3+ / CD4+ cells in different groups of subjects before and after treatment in the experimental examples of this invention; Figure 3 This is a graph showing the difference in the percentage of CD3+ / CD8+ cells in different groups of subjects before and after treatment in the experimental examples of this invention; Figure 4 This is a graph showing the differences in fatigue scores among subjects in different groups before and after treatment in the experimental examples of this invention. Figure 5 This is a graph showing the differences in sleep volume scores among subjects in different groups before and after treatment in the experimental examples of this invention. Detailed Implementation

[0020] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the following will be described in conjunction with the accompanying drawings of the embodiments of the present invention. Figures 1-5The technical solutions of the embodiments of the present invention will be clearly and completely described herein. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the described embodiments of the present invention are within the scope of protection of the present invention.

[0021] Example 1 This invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, comprising the following steps: S1. Crush defatted hemp seed meal and pass it through a 60-mesh sieve. Then, add sodium hydroxide solution with a pH of 10.0 to the defatted hemp seed meal and stir to extract. The ratio of sodium hydroxide solution to defatted hemp seed meal is 1:15. The stirring and extraction temperature is 50℃ and the stirring and extraction time is 1.5h. After extraction, centrifuge and collect the supernatant. Adjust the pH of the supernatant to 4.5 with 1.5 mol / L hydrochloric acid solution to precipitate the protein. Centrifuge and collect the hemp seed protein precipitate. Redissolve the protein precipitate in deionized water to obtain the hemp seed protein solution. S2. Adjust the temperature of the hemp seed protein solution to 50℃ and the pH to 8. Add 0.8% (by weight of hemp seed protein) of Serratia marcescens enzyme and hydrolyze for 3 hours to obtain the hydrolysate. Adjust the temperature of the hydrolysate to 60℃ and the pH to 7.5. Add 2% (by weight of hemp seed protein) of thermophilic protease and continue hydrolysis for 2 hours to obtain the secondary hydrolysate. Adjust the temperature of the secondary hydrolysate to 50℃ and the pH to 7. Add 1.5% (by weight of hemp seed protein) of flavor protease and perform a third hydrolysis for 1.5 hours to obtain the final hydrolysate. S3. The final enzymatic hydrolysate was sterilized in a water bath at 90°C for 15 min, and then centrifuged at 8000 rpm for 20 min to obtain the supernatant. The supernatant was then passed through ultrafiltration membranes with molecular weight cutoffs of 10 kDa and 3 kDa to obtain a filtrate containing components with molecular weights less than 3 kDa. The filtrate was then freeze-dried under vacuum to obtain anti-fatigue hemp seed protein peptides.

[0022] Example 2 This invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, comprising the following steps: S1. Crush defatted hemp seed meal and pass it through a 60-mesh sieve. Then, add sodium hydroxide solution with a pH of 9.5 to the defatted hemp seed meal and stir to extract. The ratio of sodium hydroxide solution to defatted hemp seed meal is 1:17. The stirring and extraction temperature is 55℃ and the stirring and extraction time is 1 hour. After extraction, centrifuge and collect the supernatant. Adjust the pH of the supernatant to 5 with 1.2 mol / L hydrochloric acid solution to precipitate the protein. Centrifuge and collect the hemp seed protein precipitate. Redissolve the protein precipitate in deionized water to obtain hemp seed protein solution. S2. Adjust the temperature of the hemp seed protein solution to 45℃ and the pH to 9, add 1% of the hemp seed protein weight of Serratia proteinase and hydrolyze for 3.5 hours to obtain the enzymatic hydrolysate; adjust the temperature of the enzymatic hydrolysate to 58℃ and the pH to 8, add 1.5% of the hemp seed protein weight of thermophilic protease and continue enzymatic hydrolysis for 2.5 hours to obtain the secondary enzymatic hydrolysate; adjust the temperature of the secondary enzymatic hydrolysate to 55℃ and the pH to 7.5, add 1% of the hemp seed protein weight of flavor protease and perform a third enzymatic hydrolysis for 2 hours to obtain the final enzymatic hydrolysate; S3. The final enzymatic hydrolysate was sterilized in a water bath at 92°C for 13 min, and then centrifuged at 7000 rpm for 25 min to obtain the supernatant. The supernatant was then passed through ultrafiltration membranes with molecular weight cutoffs of 10 kDa and 3 kDa to obtain a filtrate containing components with molecular weights less than 3 kDa. The filtrate was then freeze-dried under vacuum to obtain anti-fatigue hemp seed protein peptides.

[0023] Example 3 This invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, comprising the following steps: S1. Crush defatted hemp seed meal and pass it through a 100-mesh sieve. Then, add sodium hydroxide solution with a pH of 9 to the defatted hemp seed meal and stir to extract. The ratio of sodium hydroxide solution to defatted hemp seed meal is 1:14. The stirring and extraction temperature is 52℃ and the stirring and extraction time is 1.8h. After extraction, centrifuge and collect the supernatant. Adjust the pH of the supernatant to 4.8 with 1.1mol / L hydrochloric acid solution to precipitate the protein. Centrifuge and collect the hemp seed protein precipitate. Redissolve the protein precipitate in deionized water to obtain the hemp seed protein solution. S2. Adjust the temperature of the hemp seed protein solution to 48℃ and the pH to 8.5, add 0.5% (by weight of hemp seed protein) of Serratia proteinase and hydrolyze for 3 hours to obtain the enzymatic hydrolysate; adjust the temperature of the enzymatic hydrolysate to 56℃ and the pH to 7.6, add 1.9% (by weight of hemp seed protein) of thermophilic protease and continue enzymatic hydrolysis for 2.2 hours to obtain the secondary enzymatic hydrolysate; adjust the temperature of the secondary enzymatic hydrolysate to 48℃ and the pH to 7.5, add 1.6% (by weight of hemp seed protein) of flavor protease and perform a third enzymatic hydrolysis for 1.7 hours to obtain the final enzymatic hydrolysate; S3. The final enzymatic hydrolysate was sterilized in a water bath at 93°C for 12 min, and then centrifuged at 7500 rpm for 23 min to obtain the supernatant. The supernatant was then passed through ultrafiltration membranes with molecular weight cutoffs of 10 kDa and 3 kDa to obtain a filtrate containing components with molecular weights less than 3 kDa. The filtrate was then freeze-dried under vacuum to obtain anti-fatigue hemp seed protein peptides.

[0024] Example 4 This invention provides a method for preparing hemp seed protein peptides that improve mental and physical fatigue, comprising the following steps: S1. Crush defatted hemp seed meal and pass it through an 80-mesh sieve. Then, add sodium hydroxide solution with a pH of 9.5 to the defatted hemp seed meal and stir to extract. The ratio of sodium hydroxide solution to defatted hemp seed meal is 1:12. The stirring and extraction temperature is 45℃ and the stirring and extraction time is 2 hours. After extraction, centrifuge and collect the supernatant. Adjust the pH of the supernatant to 4.5 with 1 mol / L hydrochloric acid solution to precipitate the protein. Centrifuge and collect the hemp seed protein precipitate. Redissolve the protein precipitate in deionized water to obtain the hemp seed protein solution. S2. Adjust the temperature of the hemp seed protein solution to 45℃ and the pH to 8, add 0.7% (by weight of hemp seed protein) of Serratia marcescens enzyme and hydrolyze for 2.5 hours to obtain the hydrolysate; adjust the temperature of the hydrolysate to 55℃ and the pH to 7, add 2.5% (by weight of hemp seed protein) of thermophilic protease and continue hydrolysis for 1.5 hours to obtain the secondary hydrolysate; adjust the temperature of the secondary hydrolysate to 45℃ and the pH to 6.5, add 2% (by weight of hemp seed protein) of flavor protease and perform a third hydrolysis for 1 hour to obtain the final hydrolysate; S3. The final enzymatic hydrolysate was sterilized in a water bath at 95°C for 10 min, and then centrifuged at 9000 rpm for 15 min to obtain the supernatant. The supernatant was then passed through ultrafiltration membranes with molecular weight cutoffs of 10 kDa and 3 kDa to obtain a filtrate containing components with molecular weights less than 3 kDa. The filtrate was then freeze-dried under vacuum to obtain anti-fatigue hemp seed protein peptides.

[0025] The hemp seed protein peptides prepared in Examples 1-4 were analyzed by mass spectrometry using HPLC-M, and the amino acid sequences of the peptides are as follows: SEQ ID NO.1: MARNF (Met-Ala-Arg-Gln-Phe), SEQ ID NO.2: MDTRLL (Met-Asp-Thr-Arg-Leu-Leu).

[0026] Experimental Example Objective: To investigate whether hemp seed protein peptides have anti-fatigue and immunity-enhancing effects.

[0027] 1. Selection of subjects: The subjects were individuals who underwent surgery (including laparoscopic, open, and benign / malignant tumor surgeries); The patient's surgery lasted between 1 and 4 hours. The main symptoms include: easy fatigue (such as weakness, loss of appetite, poor concentration, depression, tension, anxiety, etc., evaluated according to the fatigue scale), drowsiness / insomnia (evaluated according to the sleep scale), and weakened immunity. Age between 35 and 75 years old, gender not limited; Those who agree to participate in this research.

[0028] 2. Testing Methods Subjects were randomly assigned to two groups: a placebo group (Group A) and an experimental group (Group E). The placebo group (Group A) received a placebo capsule, while the experimental group (Group E) received capsules containing hemp seed protein peptides. The placebo was a residual food additive without hemp seed protein peptides, identical in appearance, shape, odor, and size to the capsules containing hemp seed protein peptides. Dosage: Twice daily, 30 minutes after breakfast and dinner.

[0029] The trial treatment period was 30 days. Subjects were randomly assigned to groups. After enrollment, drug management personnel distributed trial capsules to the subjects, who then consumed the capsules according to the recommended dosage and method. The trial set two monitoring points: day 1 before treatment (T1) and day 30 of treatment (T2). Subjects were followed up and their indicators were monitored during these monitoring periods. During these visits, data such as laboratory tests, fatigue scales, sleep quality assessments, and immune function indicators were recorded. The trial product was distributed and collected, and the subject took the product continuously for 30 days.

[0030] Laboratory test indicators include immune markers: T lymphocyte subsets: CD3+, CD3+CD4+, CD3+CD8+; CD3+% represents the proportion of all T lymphocytes. CD3+ T cells are an important component of the body's immune system and participate in cellular immune responses. It is usually used to assess overall T cell immune function, mainly for immune status assessment. A decrease in the number of CD3+ T cells may reflect impaired immune system function, while an increase in the number of CD3+ T cells can, to some extent, reflect enhanced immune system function. (CD3+ / CD4+)% is an important indicator of T lymphocyte subsets in the immune system, used to assess cellular immune function. It represents the proportion of helper T cells (CD4+) among all T cells (CD3+). CD4+ T cells play a central role in immune regulation, anti-infection, and tumor surveillance; therefore, this indicator is widely used in immune function assessment and nutritional status monitoring. A decrease in the number of CD4+ T cells is usually associated with immunosuppression, infection (such as HIV), or weakened immune function, while an increase in the number of CD4+ T cells can, to some extent, reflect enhanced immune system function. (CD3+ / CD8+)% is an important indicator for assessing T lymphocyte subsets in the immune system, commonly used to reflect cellular immune function, especially immune surveillance against viruses and tumors. It represents the proportion of cytotoxic T cells (CD8+) among all T cells (CD3+). CD8+ T cells can recognize and kill virus-infected or mutated tumor cells, thus playing a crucial role in anti-infection, anti-tumor, and immune surveillance. An increase in this proportion can, to some extent, reflect enhanced immune system function.

[0031] The fatigue scale was assessed using the Identity-Consequence Fatigue Scale (ICFS). The scale scoring criteria were calculated according to the scale assessment rules. The scale and scale assessment rules are as follows:

[0032] Reference description: Score (1): 1. Not at all; 2. Almost never; 3. Sometimes; 4. Often; 5. Very frequently; 6. Always.

[0033] Score (2): 1. Not at all; 2. Occasionally; 3. Sometimes but less than usual; 4. Almost the same as usual; 5. The same as usual; 6. Does not apply to me.

[0034] The above indicators are quantitative measures of postoperative fatigue in patients; a higher score indicates more severe fatigue. This score is commonly used to assess the impact of perioperative care, treatment, or nutritional interventions on patient recovery. Improvement in perioperative fatigue is crucial for postoperative rehabilitation. Reduced fatigue signifies better recovery and a faster return to normal activities.

[0035] The sleep quality assessment scale uses the Pittsburgh Sleep Quality Index. The scale scoring criteria are calculated according to the scale's assessment rules, as follows: 1. In the past month, have you woken up during the night or woken up early (have difficulty falling back asleep after waking up in the early morning)? (A) Not in the past month. (C) Less than one night on average per week.

[0036] (B) On average, there are one or two nights per week. (D) On average, there are three or more nights per week.

[0037] 2. In the past month, have you experienced any pain or discomfort while sleeping at night? (A) Not in the past month. (C) On average, one or two nights per week.

[0038] (B) Less than one night per week on average. (D) Three or more nights per week on average.

[0039] 3. In the past month, how would you rate your overall sleep quality? (A) Very good (B) Poor (C) Fairly good (D) Very poor 4. In the past month, have you frequently felt drowsy while driving, eating, or participating in social activities? (A) Not in the past month. (C) On average, one or two nights per week.

[0040] (B) Less than one night per week on average. (D) Three or more nights per week on average.

[0041] 5. In the past month, how long does it usually take you to fall asleep each night (from getting into bed to falling asleep): (A) Within 5 minutes. (B) 15-30 minutes. (C) 15-45 minutes. (D) More than 30 minutes.

[0042] Choose A for 20 points, B for 15 points, C for 10 points, and D for 5 points. A score of 76-100 indicates excellent sleep quality. A score of 61-75 indicates good sleep quality. A score of 41-60 indicates average sleep quality. A score of 25-40 indicates poor sleep quality.

[0043] The Sleep Quality Index (SQI) is a tool for assessing patient sleep quality. It is used to evaluate the impact of perioperative or other interventions on sleep quality; a higher score indicates better sleep quality. Improved sleep quality is crucial for postoperative recovery. Good sleep after surgery helps accelerate wound healing, reduce postoperative complications, and improve overall recovery outcomes.

[0044] 3. Experimental Results 3.1 Relevant Results of Immune Indicators 3.1.1 Immune indicators of subjects in each group before treatment

[0045] Placebo group, n=38; Experimental group, n=36 As shown in Table 1, there were no significant differences in immune indicators such as CD3+ and CD3+ / CD4+ among the subjects in each group before treatment.

[0046] 3.1.2 Comparison of immune indicators among subjects before and after treatment

[0047] Placebo group, n=38; Experimental group, n=36 The graph showing the differences in the percentage of CD3+ cells in each group before and after treatment is shown below. Figure 1 The difference in the percentage of CD3+ / CD4+ cells among the subjects before and after treatment is shown in the figure below. Figure 2 The difference in the percentage of CD3+ / CD8+ cells among the subjects before and after treatment is shown in the figure below. Figure 3 .

[0048] Depend on Figure 1 ,2 As shown in Table 2, compared with the baseline data before treatment, the percentage of CD3+, CD3+ / CD4+, and CD3+ / CD8+ in groups A and E increased significantly, indicating that the T lymphocytes and T lymphocyte subsets in each group recovered well within 30 days.

[0049]

[0050] Placebo group, n=38; Experimental group, n=36 As shown in Table 3, after 30 days of treatment, compared with group A, the percentage of CD3+ in group E was significantly increased (P<0.05), and the percentage of CD3+CD8+ was significantly increased (P<0.05). That is, compared with natural recovery, the hemp seed protein peptide provided by the present invention can more effectively promote the recovery of T lymphocytes and T lymphocyte subsets in subjects and improve immunity.

[0051] 3.2 Comparison of quantitative indicators of fatigue and sleep Fatigue assessment was performed using the perioperative fatigue assessment scale, with a Likert scale of 1-6, where 25-35 points indicated mild fatigue, 36-40 points indicated moderate fatigue, and 41-45 points indicated severe fatigue. The sleep quality index scale contained 5 items, with a score range of 0-100.

[0052] 3.2.1 Quantitative indicators of fatigue and sleep in each group of subjects before treatment

[0053] Placebo group, n=38; Experimental group, n=36 As shown in Table 4, there were no significant differences in the two indicators among the subjects in each group before treatment.

[0054] 3.2.2 Comparison of quantitative indicators of fatigue and sleep in different groups of subjects before and after treatment

[0055] Placebo group, n=38; Experimental group, n=36 The graph showing the differences in fatigue scores among the subjects before and after treatment is shown below. Figure 4 The difference in sleep scores among the subjects before and after treatment is shown in the graph. Figure 5 .

[0056] Depend on Figure 4 , Figure 5 As shown in Table 5, compared with the baseline data before treatment, the fatigue scores of subjects in groups A and E were significantly reduced and the sleep quality index was significantly improved, indicating that the fatigue status of subjects in each group was improved within 30 days, and mental fatigue was improved.

[0057]

[0058] Placebo group, n=38; Experimental group, n=36 As shown in Table 6, after 30 days of treatment, compared with group A, the fatigue score of group E subjects decreased significantly (P<0.05) and the sleep quality index increased significantly (P<0.05). That is, compared with natural recovery, the hemp seed protein peptide provided by the present invention has a more significant effect on improving the fatigue status and sleep quality of the subjects.

[0059] In summary, the hemp seed protein peptides provided by this invention have the effects of anti-fatigue and improving immunity.

[0060] The above are preferred embodiments of the present invention. It should be noted that, for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for preparing hemp seed protein peptides that improve mental and physical fatigue, characterized in that, Includes the following steps: S1. Pulverize defatted hemp seed meal and extract hemp seed protein solution using alkali dissolution and acid precipitation method; S2. Adjust the temperature and pH of the hemp seed protein solution, and hydrolyze it with Serratia marcescens to obtain a hydrolysate; adjust the temperature and pH of the hydrolysate, add thermophilic protease, and continue hydrolysis to obtain a secondary hydrolysate; adjust the temperature and pH of the secondary hydrolysate, add flavor protease, and perform a third hydrolysis to obtain the final hydrolysate. S3. The final enzymatic hydrolysate is post-treated and dried to obtain anti-fatigue hemp seed protein peptides.

2. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 1, characterized in that, The temperature and pH of the hemp seed protein solution are adjusted to 45-50℃ and 8-9; the amount of serrata peptide enzyme added is 0.5-1% of the hemp seed protein mass, and the enzymatic hydrolysis reaction time is 2.5-3.5h.

3. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 1, characterized in that, The temperature and pH of the hydrolysate are adjusted to 55-60℃ and 7-8; the amount of thermophilic protease added is 1.5-2.5% of the protein content of hemp seed, and the hydrolysis reaction time is 1.5-2.5h.

4. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 1, characterized in that, The temperature and pH of the secondary enzymatic hydrolysate are adjusted to 45-55℃ and 6.5-7.5; the amount of flavor protease added is 1-2% of the protein content of hemp seeds, and the enzymatic hydrolysis reaction time is 1-2 hours.

5. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 1, characterized in that, The alkaline dissolution and acid precipitation method includes adding an alkaline solution to defatted hemp seed meal, stirring and extracting, centrifuging and taking the supernatant, adjusting the pH of the supernatant with an acid solution to precipitate the protein, centrifuging and collecting the hemp seed protein precipitate; redissolving the protein precipitate in water to obtain a hemp seed protein solution.

6. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 5, characterized in that, The alkaline solution is a sodium hydroxide solution with a pH of 9-10.0; the material-to-liquid ratio during stirring extraction is 1:(12-17), the stirring extraction time is 1-2 hours, and the temperature is 45-55℃; the acid solution is a 1-1.5 mol / L hydrochloric acid solution, and the pH of the supernatant is adjusted to 4.5-6.

5.

7. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 1, characterized in that, The post-processing includes sterilizing the final enzymatic hydrolysate, centrifuging and taking the supernatant, passing the supernatant through an ultrafiltration membrane, and then freeze-drying the filtrate under vacuum to obtain the anti-fatigue hemp seed protein peptide.

8. The method for preparing hemp seed protein peptides for improving mental and physical fatigue according to claim 7, characterized in that, The sterilization is performed by water bath at 90-95℃ for 10-15 minutes, and the centrifugation is performed at 7000-9000 rpm for 15-25 minutes.

9. A method for preparing hemp seed protein peptides for improving mental and physical fatigue as described in any one of claims 1-8, characterized in that, It includes the following amino acid sequences: SEQ ID NO.1: MARNF, SEQ ID NO.2: MDTRLL.

10. The use of the hemp seed protein peptide of claim 9, which improves mental and physical fatigue, in the preparation of anti-fatigue food or health products.

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