Brain function enhancing peptide hydrolysate-containing beverage composition and preparation method and application thereof

Abstract

The application provides a brain function improving brain peptide hydrolysate drink composition and a preparation method and application thereof, relates to the technical fields of peptide drinks and medicines. The brain protease hydrolysate concentrate, walnut peptides and intestinal peptides are matched and the dosages thereof are preferably selected, so that the drink composition is fully absorbed, the nervous system is better activated, the excitability of brain cells is improved, the growth and lengthening of nerve synapses are stimulated, the repair of brain nerves is helped, the gastrointestinal movement is enhanced, the gastrointestinal mucosa is better protected, the burden of the spleen and stomach is reduced, and the application is especially suitable for patients with weak spleen and stomach.

Description

Technical Field

[0001] This invention relates to the field of peptide beverages and pharmaceuticals, and in particular to a brain-enhancing beverage composition containing brain peptide hydrolysates, its preparation method, and its application. Background Technology

[0002] Brain injury is damage to brain tissue caused by trauma to the head and face. Common clinical symptoms include headache, altered consciousness, and intracranial hemorrhage. Treatment generally requires conservative or surgical intervention. Mild cases have a relatively good prognosis, while severe cases often result in varying degrees of neurological deficits (limb paralysis, slurred speech, sensory disturbances, etc.) and have a poorer prognosis.

[0003] The prognosis of traumatic brain injury (TBI) is closely related to the severity of the injury and factors such as the patient's age, health status, timely treatment, and rehabilitation measures. Generally speaking, the prognosis for mild TBI is relatively optimistic, while the prognosis for severe TBI is more severe. Mild TBI usually has a good prognosis, and patients can often fully recover after receiving appropriate treatment. Mild TBI typically manifests as brief loss of consciousness or symptoms such as headache and dizziness after head injury, and usually does not leave obvious sequelae. The prognosis for severe TBI is often more complex. Severe TBI can have long-term effects on the patient's physical and cognitive functions. Patients with severe TBI may experience long-term loss of consciousness, memory loss, speech disorders, limb disabilities, and other serious sequelae; in such cases, patients may require comprehensive rehabilitation treatment and long-term care.

[0004] Peptides are important components of brain tissue structure and can provide raw materials for the physiological activities of brain cells. However, most peptides currently used are whole peptides, which often have poor effects and are difficult to digest during use by patients. When whole peptides are hydrolyzed into small molecule peptides, the small molecule peptides are unstable, and the absorption and therapeutic effect are poor when ingested in large quantities. For those with weak spleen and stomach, symptoms such as indigestion, dry stools and constipation may even occur. Summary of the Invention

[0005] This invention provides a brain function-enhancing beverage composition containing hydrolyzed brain peptides, its preparation method, and its applications. On one hand, the provided brain function-enhancing beverage composition contains hydrolyzed brain peptides, providing rich nutrients for the recovery of brain function. Furthermore, the small hydrolyzed brain peptide molecules in the composition are relatively stable, the formulation of the brain peptide-containing composition is reasonable, easily absorbed, and results in rapid recovery of brain function with minimal burden on the gastrointestinal tract. On the other hand, this application provides a brain function-enhancing beverage composition containing hydrolyzed brain peptides, its preparation method, and its applications. The prepared brain peptide beverage composition has good stability, is easily absorbed, and can be used to treat brain injury.

[0006] Technical solution:

[0007] In a first aspect, this application provides a brain function-enhancing beverage composition containing brain peptide hydrolysates:

[0008] A brain function-enhancing beverage composition containing brain peptide hydrolysates, characterized in that, based on the total mass of the beverage composition, it comprises 1-12 wt% brain protein hydrolysate concentrate; 0.1-2 wt% walnut peptides; 0.01-0.1 wt% intestinal peptides; 1-8 wt% stabilizer; and 1-7 wt% concentrated fruit juice.

[0009] The excipients are 0.1-2 wt%, wherein the excipients include one or more of the following: jujube seed compound powder, γ-aminobutyric acid and taurine;

[0010] Additives 0.03-0.2 wt%, wherein the additives include one or more of the following: preservatives, flavor modifiers, stabilizers, and flavorings;

[0011] And the remaining water.

[0012] Brain peptides can help activate the nervous system, increase the excitability of brain cells, and aid in the repair of brain nerves. Peptides have a certain activating effect on the nervous system, so supplementing with peptides during the repair process after brain nerve damage helps accelerate the recovery of nerve function. At the same time, brain peptides are rich in lipids and unsaturated fatty acids, which are important components of the brain, and peptides can promote the repair of brain nerves.

[0013] Intestinal peptide concentrate is a biological product produced using porcine small intestine as raw material through bio-extraction technology. It regulates gastrointestinal digestion and absorption. Its main components are a mixture of active intestinal peptides, amino acids, and intestinal polysaccharides (glycosaminoglycans, GAGs). It features higher concentrations of active substances and smaller molecular weights, making it easier for the body to ingest and absorb in large quantities. Intestinal peptides can regulate gastrointestinal function. Vasoactive intestinal peptides are neurotransmitters with dual regulatory effects in the body. In the digestive system, they promote the relaxation of intestinal smooth muscle and simultaneously relax muscles such as the lower esophageal sphincter and intestinal smooth muscle. In the nervous system, they dilate blood vessels, increase blood flow, and also dilate the smooth muscle of the trachea and bronchi.

[0014] Walnut peptides can stimulate the growth and elongation of nerve synapses, helping to repair brain nerves; brain cells require a certain amount of energy to carry out physiological activities, and walnut peptides can also provide sufficient energy; the combination of brain peptides can better promote the development of brain cells and help repair brain nerves.

[0015] This application uses a concentrated hydrolyzed brain protein hydrolysate, with small molecule peptides preferred for the brain peptides, which facilitates the full absorption of the beverage composition. In addition, intestinal peptides are added, which can enhance the digestive function of the gastrointestinal tract, better protect and repair damaged gastrointestinal mucosa, enhance gastrointestinal motility, promote the absorption of polypeptides by the intestines, and improve the speed of brain injury recovery, while not increasing the burden on the spleen and stomach, making it especially suitable for those with weak spleen and stomach.

[0016] Furthermore, the beverage composition, based on the total mass of the beverage composition, comprises 3-10 wt% brain protein hydrolysate concentrate; 0.5-1.5 wt% walnut peptide; 0.01-0.05 wt% intestinal peptide; 3-5 wt% stabilizer; and 3-5 wt% concentrated fruit juice.

[0017] Excipients 0.5-1.5wt%,

[0018] Additives 0.08-0.12 wt%,

[0019] And the remaining water.

[0020] Furthermore, the taurine content in the solid content of the beverage composition is less than 1 wt%; the mass ratio of the intestinal peptide to taurine is (0.13-1.67):1.

[0021] Furthermore, the mass ratio of γ-aminobutyric acid to taurine is (10-20):1.

[0022] Furthermore, the mass ratio of the intestinal peptide to taurine is (0.2-1):1.

[0023] Furthermore, the excipients include 0.3-0.5 wt% jujube seed compound powder, 0.4-0.6 wt% γ-aminobutyric acid and 0.02-0.06 wt% taurine; the additives consist of 0.05-0.12 wt% food flavoring, 0.015-0.021 wt% nisin and 0.003-0.009 wt% sucralose.

[0024] Furthermore, the concentrated fruit juice includes one or more of blueberry concentrate, apple concentrate, and lemon concentrate;

[0025] The brain protein hydrolysate concentrate contains amino acids with sequences such as Leu-Phe-Leu-Pro-Arg.

[0026] Furthermore, the stabilizer comprises 2-5 wt% erythritol and 0.06-0.1 wt% xanthan gum.

[0027] Furthermore, the brain protein hydrolysate concentrate includes amino acids with sequences such as Leu-Phe-Leu-Pro-Arg.

[0028] Furthermore, the beverage composition, based on the total mass of the beverage composition, includes 6 wt% of a concentrated brain protein hydrolysate.

[0029] Walnut peptide 1.04 wt%; intestinal peptide 0.02 wt%; stabilizer: the stabilizer consists of erythritol 4 wt% and xanthan gum 0.08 wt%; concentrated fruit juice: blueberry concentrated juice 4 wt%; excipients: the excipients consist of jujube seed compound powder 0.4 wt%, γ-aminobutyric acid 0.5 wt%, and taurine 0.05 wt%; additives: the additives consist of food flavoring 0.08 wt%, nisin 0.019 wt%, and sucralose 0.006 wt%; balance water. Secondly, this application provides a method for preparing a brain-enhancing peptide hydrolysate beverage composition.

[0030] Thirdly, this application provides the use of a brain-enhancing beverage composition containing brain peptide hydrolysate in functional beverages and / or pharmaceuticals for the treatment of brain injury.

[0031] Beneficial effects:

[0032] 1. This application uses a combination of brain protein hydrolysate concentrate, walnut peptides and intestinal peptides, and optimizes their dosage to ensure that the beverage composition is fully absorbed, effectively helping to activate the nervous system, increase the excitability of brain cells, stimulate the growth and elongation of nerve synapses, help repair brain nerves, enhance gastrointestinal motility, better protect and repair the gastrointestinal mucosa, and reduce the burden on the spleen and stomach, making it especially suitable for patients with weak spleen and stomach.

[0033] 2. Furthermore, taurine can rapidly improve brain cell activity and enhance memory; however, excessive intake can also damage the gastrointestinal mucosa, leading to insomnia and high blood pressure. Gamma-aminobutyric acid (GABA) can promote sleep, relieve stress, improve cognitive function, enhance immunity, and improve overall health. The excipients in this application use a synergistic combination of GABA and taurine, along with brain protein hydrolysate and walnut peptides to effectively repair brain nerves. The optimal dosage helps repair brain nerves without increasing the burden on the spleen and stomach, and has minimal irritation to the gastrointestinal mucosa, making it suitable for patients with weak spleen and stomach.

[0034] 3. Furthermore, the stabilizer uses a combination of erythritol (a small molecule alcohol) and xanthan gum (a large molecule polysaccharide) in optimal amounts. The small and large molecules form a good compatibility effect with minimal interaction between them, which effectively stabilizes the small peptides in the brain protein hydrolysate concentrate, thereby improving the stability of the small peptide chain segments and enhancing the effect of the beverage composition on repairing brain nerves. Detailed Implementation

[0035] To make the technical solution of the present invention clearer, the present invention will be further described in detail below with reference to specific embodiments.

[0036] Preparation Example 1: A concentrated solution of brain protein hydrolysate, prepared as follows:

[0037] Step 1: Take out fresh pig brain or thawed frozen pig brain, wash it with purified water, and remove the fascia and blood vessels; mix the pig brain with purified water at a ratio of 1:1.5, then grind it into a homogenate using a colloid mill. After the tissue is basically broken down, adjust the fineness to a finer level, grind and homogenize for 5 minutes per batch, then add 1.3 times the amount of purified water and continue homogenizing for 5 minutes. Mix each batch of homogenized brain fluid evenly, and adjust the pH to 9.5 with potassium hydroxide solution; the coarsest mesh is 100 mesh; the finest mesh is 150 mesh.

[0038] Step 2: Place the adjusted brain homogenate in a microwave-safe container equipped with a stirrer, control the temperature at 60℃, microwave for 20 minutes, and then keep warm for 15 minutes; the microwave container has a frequency of 3300MHz and a power of 25KW; drain the microwave-treated homogenate and separate the protein and lipids twice using a disc centrifuge at 38℃. The first centrifugation should yield 42% of the lighter liquid. After draining the lighter liquid, microwave for another 20 minutes and then centrifuge a second time. The lighter liquid obtained from the second centrifugation is the phospholipid solution. Mix the concentrated solutions obtained from the two centrifugations; the first centrifugation speed is 9000r / min; the second centrifugation speed is 12000r / min; the temperature of the homogenate should not be lower than 38℃ during the second centrifugation.

[0039] Step 3: Adjust the pH of the concentrated solution obtained in Step 2 to 5.0 with dilute hydrochloric acid, causing the protein to precipitate out in large cloud-like flakes and separate into layers. Then, remove the liquid using a centrifuge with a 250-mesh filter cloth and collect the filter residue to obtain brain protein isolate. Dissolve the collected filter residue in twice the amount of purified water to obtain a 5% solution. Adjust the pH to 9.0, then add trypsin (5000u / g) at 4.5% of the amount of isolated protein and continue homogenization and circulation. When the pH drops to around 7.5, add alkali to adjust it to 8.6, circulate for another 3 minutes, and then drain the slurry.

[0040] Step 4: Before microwave digestion, check the pH value of the slurry obtained in Step 3. If the pH value is less than 7, add alkali to adjust the pH value to 8.2 and perform microwave-assisted enzymatic hydrolysis. Keep warm for 15 minutes to inactivate the enzyme. Discharge the enzyme hydrolysate after inactivation. When the temperature of the enzyme hydrolysate drops to less than 40°C, add a 1.5% chitosan solution dissolved in 15% acetic acid under stirring conditions to adjust the pH value to 4.8. Precipitate undigested brain proteins, phospholipids and other macromolecular impurities. Centrifuge the precipitate through a 200-mesh filter cloth to remove the precipitate. Filter the centrifuged liquid through a 0.25µm hollow fiber membrane. Filter the filtrate through an ultrafiltration membrane that can intercept molecules with a molecular weight of 10,000 Da. The permeate is the pig brain polypeptide solution. If only polypeptides are to be processed, concentrate this solution under reduced pressure and then freeze-dry to obtain polypeptide powder.

[0041] Step 5: Filter the polypeptide solution using a nanofiltration membrane with a molecular weight cutoff of 1200 Da. Then concentrate the filtrate using a reverse osmosis membrane. Freeze and dry the concentrate to obtain a brain protein hydrolysate concentrate with a solid content of 55 wt%. The brain protein hydrolysate contains 80 wt% small molecule peptides, 14.5% nitrogen in dry matter, and 0.6% fat.

[0042] LC-MS / MS separation and identification

[0043] The amino acid sequence and molecular weight of the concentrated brain protein hydrolysate were determined using LC-MS / MS. Samples were first subjected to reductive alkylation and desalting before chromatography. The capillary liquid chromatography column used was a 300 μm × 5 mm Acclaim PepMap RPLCC18 pre-column (5 μm) and an analytical column of 150 μm × 150 mm (1.9 μm). Mobile phase A consisted of 0.1% (v / v) formic acid and 2% (v / v) acetonitrile, and mobile phase B consisted of 0.1% (v / v) formic acid and 80% (v / v) acetonitrile. The flow rate was 600 nL / min, and the analysis time for each component was 60 min.

[0044] The MS and MS / MS parameters are as follows:

[0045] (1) MS parameters: resolution is 70000; maximum injection time is 40ms; scan range is 300-1400m / z.

[0046] (2) MS / MS parameters: resolution 175000; maximum injection time 60ms; scan range 300-1400m / z;

[0047] TopN=20;NCE / steepedNCE=27.

[0048] The amino acid sequence of the peptide was determined by analyzing the raw MS / MS file with Mascot software based on the sample type and comparing it with the Uniprot database (https: / / www.uniprot.org / ) Susscrofa (Pig).

[0049] After mass spectrometry data retrieval, the R4 fraction was identified using PSMFDR≤0.01 and ProteinFDR≤0.01 as screening criteria for peptide, site, and protein identification, respectively. The amino-terminal sequence of the peptide was obtained, and the concentrated brain protein hydrolysate contained amino acids with sequences such as Leu-Phe-Leu-Pro-Arg.

[0050] Brain protein hydrolysate concentrate is also available for purchase, such as the one prepared by Shaoxing Jiayun Biotechnology Co., Ltd., with a small molecule peptide content of 40-90wt%.

[0051] Example 1: A brain function-enhancing beverage composition containing brain peptide hydrolysate, using the raw materials listed in Table 1, and prepared as follows:

[0052] After mixing the raw materials, the mixture is stirred at 300 r / min for 40 min at 25°C (or 30°C in other embodiments) (or 30-500 r / min in other embodiments) to prepare a beverage composition containing brain peptide hydrolysate.

[0053] The brain protein hydrolysate concentrate was prepared using the brain protein hydrolysate concentrate from Preparation Example 1; walnut peptide: a small molecule peptide of walnut with a molecular weight less than 1000 Da, solubility > 90%, and moisture content < 5%; intestinal peptide: a molecular weight less than 1000 Da, solubility > 90%, and moisture content < 6%.

[0054] The jujube seed compound powder is 120 mesh with a purity of 98%; xanthan gum has an average molecular weight of 2 million and a purity of 98%.

[0055] Examples 2 to 5 describe a brain function-enhancing beverage composition containing brain peptide hydrolysate, which differs from Example 1 in the types and weight settings of the raw materials used, as shown in Table 1.

[0056] Table 1. List of raw materials and their weights used in the beverage compositions of Examples 1 to 5.

[0057]

[0058] Examples 5 to 10 describe a brain function-enhancing beverage composition containing brain peptide hydrolysate, which differs from Example 1 in that the weight settings of intestinal peptides and taurine are different, as shown in Table 2.

[0059] Table 2. Amounts of intestinal peptides and taurine used in the beverage compositions of Examples 5 to 10 and Example 1.

[0060]

[0061] Example 11, a brain function-enhancing beverage composition containing brain peptide hydrolysate, differs from Example 1 in that jujube seed compound powder replaces γ-aminobutyric acid and taurine in equal amounts.

[0062] Example 12, a brain function-enhancing beverage composition containing brain peptide hydrolysate, differs from Example 1 in that erythritol replaces xanthan gum in an equal amount.

[0063] Example 13, a brain function-enhancing beverage composition containing brain peptide hydrolysate, differs from Example 1 in that xanthan gum replaces erythritol in an equal amount.

[0064] Comparative Example 1, a brain function-enhancing beverage composition containing brain peptide hydrolysate, differs from Example 1 in that it uses intestinal peptides in an equal amount to replace walnut peptides.

[0065] Comparative Example 2, a brain function-enhancing beverage composition containing brain peptide hydrolysate, differs from Example 1 in that it uses walnut peptides in an equal amount to replace intestinal peptides.

[0066] Experimental performance:

[0067] C57 mice (18-24g), male, 10 mice per group, were administered the drug at a dose of 20mg / kg each time. The drug was administered by gavage after modeling (using the examples and comparative examples respectively). The normal group and the control group were administered the same volume of physiological saline by gavage. The administration was once daily for 2 consecutive weeks.

[0068] Preparation of traumatic brain injury model:

[0069] Mice were anesthetized with chloral hydrate, fixed on a stereotaxic apparatus, shaved, and their scalps were cut to expose the skull. The skull was then examined in the left hemisphere.

[0070] A 4mm diameter circular hole was made in a ball to expose the brain tissue. The ball was then struck using a free-fall impactor to establish a craniocerebral injury model. The normal group underwent craniotomy without any manipulation, the control group received impact without medication, and the drug-treated group received the corresponding drug via gavage after impact. After administration, the mice were placed on cages lined with filter paper, and the number and quality of feces were observed and recorded daily.

[0071] Cerebral edema level test:

[0072] After the animal is euthanized, its brain tissue is collected, weighed, and recorded as wet weight (WW). The brain tissue is then dried in an oven at 110°C for 24 hours, weighed, and recorded as dry weight (DW). Moisture content (%) = (WW - DW) / WW × 100%.

[0073] The results of the brain tissue water content measurement are shown in Table 3.

[0074] Behavioral test results:

[0075] The Rotard test is used to determine the latency time for mice to fall off a rotating rod, i.e., the time it takes for them to fall.

[0076] The duration of time spent on the rotating bar; the balance beam test records the number of missteps a mouse makes on a balance beam, and the experiment is mainly used to evaluate the sensorimotor ability of animals after injury.

[0077] Inflammation level measurement:

[0078] The levels of inflammatory factors (TNF-α and IL-1β) in brain tissue were measured using an ELISA enzyme-linked immunosorbent assay kit, and the experimental procedure was performed in accordance with the kit instructions.

[0079] Table 3. Experimental results of Examples 1 to 13, Comparative Example 1, Comparative Example 2, Normal Group and Control Group.

[0080]

[0081] As can be seen from Examples 1, 5 to 10, with the increase of taurine content, the average number of fecal pellets per day in mice decreased, and cerebral edema and inflammation in mice tended to increase. However, the rotating bar test and balance beam test showed good experimental results, indicating that the repair effect on brain nerves was good. In summary, when the taurine content in the solid content of the beverage composition is less than 1 wt%; the mass ratio of γ-aminobutyric acid to taurine is (10-20):1; and the mass ratio of intestinal peptide to taurine is (0.2-1):1, the overall effect is better, the repair effect on brain nerves is better, and at the same time, it does not increase the burden on the spleen and stomach and has little irritation to the gastrointestinal mucosa.

[0082] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention. Therefore, the scope of protection of this patent should be determined by the appended claims.

Claims

1. A drink composition comprising a brain peptide hydrolysate, characterized in that, The beverage composition comprises 1-12 wt% of the brain proteolysate concentrate, 0.1-2 wt% of the walnut peptide, 0.01-0.1 wt% of the intestinal peptide, 1-8 wt% of the stabilizer, 1-7 wt% of the concentrated fruit juice, 0.1-2 wt% of the auxiliary material, 0.03-0.2 wt% of the additive, and the balance of water, based on the total mass of the beverage composition; The brain proteolysate concentrate comprises amino acids with the sequence of Leu-Phe-Leu-Pro-Arg; the auxiliary material comprises Zizyphus jujuba Mill. compound powder, gamma-aminobutyric acid, and taurine; the mass ratio of the gamma-aminobutyric acid to the taurine is (10-20):1; the mass ratio of the intestinal peptide to the taurine is (0.2-1):1; The additive comprises one or a combination of several of preservatives, taste regulators, stabilizers, and essences.

2. The brain peptide hydrolysate-containing beverage composition according to claim 1, characterized by, The beverage composition comprises 3-10 wt% of the brain proteolysate concentrate, 0.5-1.5 wt% of the walnut peptide, 0.01-0.05 wt% of the intestinal peptide, 3-5 wt% of the stabilizer, 3-5 wt% of the concentrated fruit juice, 0.5-1.9 wt% of the auxiliary material, 0.08-0.12 wt% of the additive, and the balance of water, based on the total mass of the beverage composition.

3. The brain peptide hydrolysate-containing beverage composition according to claim 1, wherein The auxiliary material comprises 0.3-0.5 wt% of Zizyphus jujuba Mill. compound powder, 0.4-0.6 wt% of gamma-aminobutyric acid, and 0.02-0.06 wt% of taurine; the stabilizer comprises 2-5 wt% of erythritol and 0.06-0.1 wt% of xanthan gum; The additive comprises 0.05-012 wt% of food essence, 0.015-0.021 wt% of nisin, and 0.003-0.009 wt% of sucralose.

4. The brain peptide hydrolysate-containing beverage composition according to claim 1, characterized by, The concentrated fruit juice comprises one or a combination of several of blueberry concentrate, apple concentrate, and lemon concentrate.

5. Use of the beverage composition containing brain peptide hydrolysate according to any one of claims 1-4 in the preparation of a medicament for the treatment of brain injury.

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