A composition for brain and eyesight improvement, its preparation method, product and application

Abstract

The present application belongs to the technical field of functional food, and particularly relates to a composition for invigorating brain and improving eyesight, a preparation method, a product and an application thereof. The composition is prepared from algal oil, linseed oil, arachidonic acid, marigold extract, corn xanthophyll suspension oil, NA nervonic acid and PS phosphatidylserine in a specific weight ratio. The marigold extract is prepared by adopting a freezing crushing pretreatment, combining with an enzymatic hydrolysis, an ultrasonic-microwave synergistic extraction technology, being concentrated through a nanofiltration membrane and being vacuum freeze-dried. The composition realizes the high-efficiency invigorating brain and improving eyesight effect through the synergistic effect of the components and the innovative extraction process, and is significantly improved in the component activity reservation and the effective component extraction rate compared with the traditional formula and extraction method.

Description

Technical Field

[0001] This invention belongs to the field of functional food technology, specifically relating to a brain-boosting and vision-improving composition, its preparation method, product, and application. Background Technology

[0002] With the rapid development of technology and significant improvement in productivity, electronic products have become deeply integrated into people's work and lives. Whether it's prolonged screen time for work or immersive entertainment experiences, the amount of time spent using eyes has increased dramatically. At the same time, intensified social competition and the growing pressure of learning and life have led to chronic mental stress and fatigue, placing an excessive burden on the brain. These factors have collectively resulted in an explosive growth in demand for eye care and brain health products.

[0003] Currently, the market is flooded with products claiming to aid memory improvement and relieve eye strain. Among memory-enhancing products, those primarily containing DHA, EPA, vitamins, and zinc are common; traditional Chinese medicine products often use ingredients like Alpinia oxyphylla, walnuts, and Gastrodia elata. For eye strain relief products, lutein esters, zeaxanthin, carotene, and blueberry extract are highly sought after, while traditional Chinese medicine products often include ingredients like goji berries, chrysanthemum, cassia seeds, and bilberries. However, existing technologies face several challenges: First, eye care products often have limited functionality, focusing solely on improving eye problems and failing to meet consumers' diverse health needs for simultaneously enhancing brainpower and memory. Second, existing brain-boosting products often rely on single ingredients or simple formulas, resulting in unsatisfactory effects even with long-term use, hindering comprehensive improvement of brain function. Third, the preparation of product extracts often relies on simple mechanical or solvent soaking methods, which are not only inefficient but also limit the absorption and utilization of the resulting products in the body, significantly reducing their efficacy.

[0004] To address these issues, researchers have conducted extensive studies, resulting in several related invention patents. For example, CN119235021A discloses a composition with both brain-boosting and vision-enhancing effects, along with its applications. This composition comprises DHA algal oil powder, walnut peptides, lutein esters, and other ingredients, and can be formulated into various dosage forms. It effectively improves memory and visual fatigue symptoms, enhances visual acuity, and is scientifically formulated and safe. Another example is CN119453474A, which discloses an eye-protecting and brain-boosting composition. By combining extracts of chokeberry, astaxanthin, and DHA, it achieves multi-pathway improvement of visual problems and simultaneous enhancement of cognitive abilities, playing a positive role in protecting the retina, promoting blood circulation in the eyes, and protecting nerve cells.

[0005] While these patents have met some of the market's demand for eye and brain health products, there is still room for improvement. For example, in terms of raw material selection, the existing patented raw material combinations have not fully explored the components with higher eye and brain health benefits; in terms of extraction technology, there is potential to improve the extraction efficiency and activity retention of the effective components in the raw materials; and in terms of formula optimization, the synergistic effects between the various components need to be further strengthened. Therefore, developing a product with better raw material selection, more efficient extraction technology, and scientific ingredient ratios, which can more effectively achieve multiple benefits such as relieving eye fatigue, preventing myopia, and enhancing memory, has become a pressing technical challenge in the field of eye and brain health products. Summary of the Invention

[0006] To address the aforementioned shortcomings, this invention provides a multi-component, scientifically formulated composition for improving brain function and eyesight. This composition has multiple benefits, including relieving eye fatigue, preventing myopia, and enhancing memory.

[0007] The technical solution of this invention is as follows:

[0008] On one hand, the present invention provides a brain-boosting and vision-improving composition comprising the following components in parts by weight: 200-240 parts algal oil, 60-80 parts flaxseed oil, 8-12 parts arachidonic acid, 4-8 parts marigold extract, 0.6-2 parts zeaxanthin suspension oil, 0.02-0.04 parts NA nervonic acid, and 0.4-0.6 parts PS phosphatidylserine;

[0009] The preparation method of the marigold extract includes the following steps:

[0010] S1. Pretreatment: Marigold flowers are frozen, crushed, and then vacuum dried;

[0011] S2, Single extraction: The pretreated material is mixed with the extraction solvent and subjected to ultrasonic-assisted extraction in the presence of a compound enzyme;

[0012] S3. Secondary extraction: The filter residue after the first extraction is mixed with the extraction solvent again and subjected to variable amplitude ultrasonic extraction.

[0013] S4. Post-processing: Combine the extracts and perform microfiltration, nanofiltration concentration and drying sequentially to obtain marigold extract;

[0014] The complex enzyme described in step S2 includes cellulase, β-glucanase, and hemicellulase;

[0015] The ultrasound-assisted extraction in step S2 uses dual-frequency ultrasound, with a low frequency of 20-30kHz and a power of 260-350W, and a high frequency of 100-120kHz and a power of 150-220W.

[0016] The frequency of the variable amplitude ultrasound in step S3 changes periodically between 20kHz and 40kHz, with a change period of 4-6 minutes, and the ultrasound power is 350-450W.

[0017] Preferably, in some embodiments, the low frequency in step S2 is 30kHz and the power is 350W, and the high frequency is 120kHz and the power is 220W.

[0018] In other embodiments, the low frequency is 20kHz with a power of 260W, and the high frequency is 100kHz with a power of 150W.

[0019] Preferably, in some embodiments, the frequency of the variable amplitude ultrasound in step S3 changes periodically between 20 kHz and 40 kHz, with a change period of 4 minutes, and the ultrasound power is 450 W.

[0020] In other embodiments, the frequency of the variable amplitude ultrasound varies periodically between 20 kHz and 40 kHz, with a period of 6 minutes, and the ultrasound power is 350 W.

[0021] Specifically, the mass percentages of cellulase, β-glucanase, and hemicellulase in step S2 are 2-4:1-3:0.5-1.5.

[0022] Preferably, the mass percentages of cellulase, β-glucanase, and hemicellulase in step S2 are 3:2:1.

[0023] Specifically, the extraction time for ultrasound-assisted extraction in step S2 is 1-2 hours, and the extraction temperature is 35-40℃.

[0024] Specifically, the extraction solvent in step S2 is a 30-50% aqueous ethanol solution; the pH value of the extraction solvent is 4.5-5.5.

[0025] Preferably, the extraction solvent in step S2 is a 40% aqueous ethanol solution; the pH value of the extraction solvent is 5.0.

[0026] Specifically, during the ultrasound-assisted extraction process described in step S2, the ultrasound is paused for 1-5 minutes every 10-30 minutes, and the mixture is stirred at a speed of 150-250 r / min.

[0027] Preferably, in some embodiments, during the ultrasound-assisted extraction process described in step S2, the ultrasound is paused for 4 minutes every 25 minutes, and the mixture is stirred at a speed of 220 r / min.

[0028] Preferably, in some other embodiments, during the ultrasound-assisted extraction process described in step S2, the ultrasound is paused for 2 minutes every 15 minutes, and the mixture is stirred at a speed of 180 r / min.

[0029] Specifically, the extraction time for the variable amplitude ultrasound in step S3 is 0.8-1.2 hours, and the extraction temperature is 32-38℃.

[0030] Specifically, the freezing process in step S1 is carried out at a temperature of -30 to -50°C for 8 to 16 hours.

[0031] The average particle size of the pulverized particles is 0.5-1.0 mm;

[0032] The vacuum drying process involves a vacuum level of 5-15 Pa, a temperature of 20-30°C, and a time of 3-5 hours.

[0033] Specifically, the microfiltration process in step S4 uses a filter membrane with a pore size of 0.2-0.8 μm;

[0034] The nanofiltration membrane used has a molecular weight cutoff of 500-1000 Da;

[0035] The drying process is vacuum freeze drying, and the moisture content after drying is ≤3.0%.

[0036] Preferably, in some embodiments, the composition comprises the following components in parts by weight: 240 parts algal oil, 60 parts linseed oil, 8 parts arachidonic acid, 8 parts marigold extract, 0.6 parts zeaxanthin suspension oil, 0.02 parts NA nervonic acid, and 0.6 parts PS phosphatidylserine;

[0037] In other embodiments, the ingredients include the following components in parts by weight: 200 parts algal oil, 80 parts linseed oil, 12 parts arachidonic acid, 4 parts marigold extract, 2 parts zeaxanthin suspension oil, 0.04 parts NA nervonic acid, and 0.4 parts PS phosphatidylserine.

[0038] In another aspect, the present invention provides a method for preparing the aforementioned composition, comprising the following steps:

[0039] (1) Mix the algal oil, linseed oil and arachidonic acid in the specified amounts to obtain a uniform oil phase mixture;

[0040] (2) Add marigold extract to the oil phase mixture obtained in step (1), mix well, and obtain mixture A;

[0041] (3) Add zeaxanthin suspension oil to mixture A obtained in step (2), mix well, and obtain mixture B;

[0042] (4) Add NA nervonic acid and PS phosphatidylserine to the mixture B obtained in step (3), mix well, and the composition is obtained.

[0043] In another aspect, the present invention provides a functional food, characterized in that it comprises the aforementioned composition.

[0044] Specifically, the health products also include excipients.

[0045] The excipients include, but are not limited to, one or more of the following: fillers, disintegrants, lubricants, and flavoring agents.

[0046] The filler includes, but is not limited to, one or more of lactose, microcrystalline cellulose, starch, and mannitol.

[0047] The disintegrants include, but are not limited to, one or more of the following: sodium carboxymethyl starch, crospovidone, and low-substituted hydroxypropyl cellulose.

[0048] The lubricant includes, but is not limited to, one or more of magnesium stearate, talc, silica, and polyethylene glycol.

[0049] The flavoring agents include, but are not limited to, one or more of the following: steviol glycosides, aspartame, sucrose, and flavorings.

[0050] In another aspect, the present invention provides the application of the aforementioned composition or functional food in the preparation of brain-boosting and vision-improving products.

[0051] Specifically, the product is a functional food or a medicine.

[0052] The beneficial effects of this invention are as follows:

[0053] (1) This invention achieves an organic combination of multiple effects such as relieving eye fatigue, preventing myopia, and enhancing memory through the scientific combination of various functional ingredients, overcoming the shortcomings of existing eye care products with single effects, and meeting the needs of consumers to improve brainpower and enhance memory at the same time.

[0054] (2) The present invention uses multiple extraction methods to prepare marigold extract, which improves the extraction efficiency. Furthermore, through scientific concentration and drying processes, high-quality active ingredients are obtained, which improves the absorption and utilization efficiency of the product in the body.

[0055] (3) The formula of the present invention is scientifically and rationally designed, and the components work synergistically to effectively improve memory and visual fatigue symptoms, enhance the duration of clear vision, and have significant brain-boosting and vision-enhancing effects.

[0056] (4) The preparation method of the present invention is simple and feasible, the process flow is clear, it is easy to industrialize and produce, and the stability and consistency of the product are guaranteed through scientific quality control. Attached Figure Description

[0057] Figure 1Typical images of the average fluorescence intensity of apoptotic cells in the eyes of zebrafish in each group are shown. A represents the normal control group; B represents the model control group; C represents coenzyme Q; D represents Example 1; E represents Example 2; F represents Comparative Example 1; G represents Comparative Example 2; H represents Comparative Example 3; I represents Comparative Example 4; and the green fluorescent particles indicated by the red arrows represent apoptotic cells.

[0058] Figure 2 The diagram shows the ratio of retinal pigment epithelium diameter to scleral diameter in zebrafish from each group. A represents the normal control group; B represents the MO standard control group; C represents the model control group; E represents Example 1; D represents Atropine sulfate monohydrate; F represents Example 2; G represents Comparative Example 1; H represents Comparative Example 2; I represents Comparative Example 3; and J represents Comparative Example 4.

[0059] Figure 3 Typical zebrafish movement trajectories for each group are shown below. A represents the normal control group; B represents Ginkgo biloba and Cistanche deserticola tablets; C represents Example 1; D represents Example 2; E represents Comparative Example 1; F represents Comparative Example 2; G represents Comparative Example 3; H represents Comparative Example 4; and the blue boxes represent the blue areas of the cross maze, which are the quantitative areas. Detailed Implementation

[0060] The present invention will be further clearly and completely illustrated below through embodiments. These embodiments are only some examples of the present invention and are not intended to limit the present invention, but are only for illustrating the present invention. Unless otherwise specified, the experimental methods used in the following embodiments are all conventional experiments, and the materials and reagents used in the following embodiments are commercially available unless otherwise specified.

[0061] The sources of the raw materials for this invention are shown in Table 1:

[0062] Table 1. Sources of raw materials

[0063]

[0064] Example 1

[0065] 1.1 Formula

[0066] A brain-boosting and vision-improving composition, the formula of which is shown in Table 2:

[0067] Table 2 Formulation of Example 1

[0068]

[0069] 1.2 Preparation method

[0070] 1.2.1 Preparation method of marigold extract

[0071] (1) Pretreatment: Select fresh marigold flowers free from pests and diseases, remove the stems and sepals, and freeze them at -40℃ for 12 hours to allow the water in the cells to fully crystallize. Then, use an ultra-low temperature pulverizer to pulverize the frozen flowers into particles with an average particle size of about 0.8 mm. Next, place these particles in a vacuum drying oven and dry them at a vacuum degree of 10 Pa and a temperature of 25℃ for 4 hours to remove some of the bound water, destroy the cell structure, and improve the subsequent extraction efficiency.

[0072] (2) Single-stage ultrasonic extraction: The pretreated marigold granules were added to a mixture of 45% ethanol aqueous solution, 0.3% (m / m) cellulase, 0.2% (m / m) β-glucanase, and 0.1% (m / m) hemicellulase in an acetate-sodium acetate buffer solution (pH 5.0) at a material-to-liquid ratio of 1:12 (mass of marigold granules: total volume of mixed liquid). The mixture was then transferred to an ultrasonic extraction device for dual-frequency ultrasonic synergistic extraction. The low-frequency ultrasonic frequency was set at 30 kHz with a power range of 350 W; the high-frequency ultrasonic frequency was set at 120 kHz with a power range of 220 W. Extraction was performed at 30°C for 2 hours. During extraction, the ultrasonic treatment was paused for 4 minutes every 25 minutes, and the mixture was stirred at 220 r / min to ensure full contact between the materials and promote the release of intracellular active ingredients.

[0073] (3) Solid-liquid separation and secondary ultrasonic extraction: After the first ultrasonic extraction, the mixture was separated into solid and liquid components using a plate and frame filter press to obtain filter residue and the first extract. The filter residue was added to a 45% ethanol aqueous solution at a material-to-liquid ratio of 1:8 and then transferred to the ultrasonic extraction device again. Variable amplitude ultrasonic extraction was used, with the ultrasonic frequency varying periodically between 20kHz and 40kHz for 4 minutes. The ultrasonic power was kept constant at 450W, and extraction was performed at 38℃ for 0.8 hours. Within this parameter range, low-frequency ultrasound could deeply break down the stubborn cell structures remaining after the first extraction, while high-frequency ultrasound accelerated the diffusion of residual active ingredients.

[0074] (4) Combining and Concentration: The secondary extract and the primary extract are combined. Larger particulate impurities are first removed by microfiltration (pore size 0.45μm), and then concentrated by nanofiltration. The molecular weight cutoff is set to 800Da. The concentration is carried out at 0.4MPa pressure to 1 / 4 of the original volume to enrich the target components.

[0075] (5) Drying: Transfer the concentrate to a vacuum freeze dryer and pre-freeze it at a pre-freezing temperature of -50°C for 3 hours to completely freeze the concentrate. Then, sublime dry it under a vacuum of 2Pa and a temperature of 32°C until the moisture content drops below 2.5% to obtain high-quality marigold extract powder.

[0076] 1.2.2 Preparation method of brain-boosting and vision-improving composition

[0077] The prescribed amounts of algal oil, linseed oil, and arachidonic acid were mixed and stirred at 50°C to form a homogeneous oil phase mixture. Marigold extract powder was then slowly added to the oil phase mixture, and stirring was continued for 30 minutes to ensure thorough dispersion. Next, zeaxanthin suspension was added, and stirring was continued for 20 minutes. Finally, NA nervonic acid and PS phosphatidylserine were added, and the mixture was stirred at 30°C for 15 minutes to obtain a homogeneous and stable brain-boosting and vision-improving composition.

[0078] Example 2

[0079] 2.1 Formula

[0080] A brain-boosting and vision-improving composition, the formula of which is shown in Table 3:

[0081] Table 3 Formulation of Example 2

[0082]

[0083] 2.1 Preparation method

[0084] The difference from Example 1 lies in steps (2) and (3) of the preparation method of marigold extract; the rest is the same as in Example 1.

[0085] (2) Single-stage ultrasonic extraction: The pretreated marigold granules were added to a mixture of 45% ethanol aqueous solution, 0.3% (m / m) cellulase, 0.2% (m / m) β-glucanase, and 0.1% (m / m) hemicellulase in an acetate-sodium acetate buffer solution (pH 5.0) at a material-to-liquid ratio of 1:12 (mass of marigold granules: total volume of mixed liquid). The mixture was then transferred to an ultrasonic extraction device for dual-frequency ultrasonic synergistic extraction. The low-frequency ultrasonic frequency was set at 20 kHz with a power range of 260 W; the high-frequency ultrasonic frequency was set at 100 kHz with a power range of 150 W. Extraction was performed at 35℃ for 1 hour. During extraction, the ultrasonic treatment was paused for 2 minutes every 15 minutes, and the mixture was stirred at 180 r / min to ensure full contact between the materials and promote the release of intracellular active ingredients.

[0086] (3)Solid-liquid separation and secondary ultrasonic extraction: After the first ultrasonic extraction, the mixed solution is subjected to solid-liquid separation by a plate and frame filter press to obtain filter residue and the first extract. The filter residue is added to an aqueous ethanol solution with a volume fraction of 45% at a solid-liquid ratio of 1:8, and then transferred to the ultrasonic extraction device again. This time, variable amplitude ultrasonic extraction is used, where the ultrasonic frequency varies periodically between 20 kHz and 40 kHz with a variation period of 6 minutes, the ultrasonic power is kept constant at 350 W, and extraction is carried out at 32 °C for 1.2 hours. Within this parameter range, the low-frequency ultrasonic waves can deeply break the stubborn cell structures remaining after the first extraction, and the high-frequency ultrasonic waves accelerate the diffusion of the remaining active ingredients.

[0087] Comparative example

[0088] Set the comparative example by referring to Example 1. The differences between the comparative example and Example 1 are shown in Table 4:

[0089] Table 4

[0090]

[0091] Effect example 1 Evaluation of the efficacy of anti-blue light eye damage

[0092] 1.1 Detection materials

[0093] 1.1.1 Sample information

[0094] The brain-strengthening and eye-clearing compositions prepared in Examples 1 - 2 and Comparative Examples 1 - 4 all use DMSO as the solvent.

[0095] Positive control: Coenzyme Q10, yellow powder, batch number K2211223, Shanghai Aladdin Biochemical Technology Co., Ltd., with DMSO as the solvent.

[0096] [[ID=​​​​​​​​Dissecting microscope (SZX7, OLYMPUS, Japan); CCD camera (VertA1, Shanghai Tusen Vision Technology Co., Ltd., China); Precision electronic balance (CP214, OHAUS, USA); 6-well plate (Zhejiang Beilanbo Biotechnology Co., Ltd., China); Blue light meter (50W 450nm, Huiheng Light Source Technology Mall, China); Motorized focusing continuous zoom fluorescence microscope (AZ100, Nikon, Japan); Ultrasonic cleaner (JP-010T, Shenzhen Jiemeng Cleaning Equipment Co., Ltd., China).

[0100] Methylcellulose (batch number C2004046, Shanghai Aladdin Biochemical Technology Co., Ltd., China); dimethyl sulfoxide (DMSO, batch number BCCD8942, Sigma, Switzerland); acridine orange (AO, batch number C15109250, Shanghai Maclean Biochemical Technology Co., Ltd., China); streptase E (batch number G12511Y118034, Shanghai Yuanye Biotechnology Co., Ltd., China).

[0101] 1.2 Detection Method

[0102] Wild-type AB strain zebrafish at 1 dpf were randomly selected and irradiated with blue light to establish a zebrafish blue light-induced eye damage model. At 3 dpf, well-developed model zebrafish were randomly assigned to 6-well plates, with 30 zebrafish treated in each well (experimental group). The corresponding samples (concentrations shown in Table 5) were dissolved in water and administered, along with a positive control of coenzyme Q10 at a concentration of 62.5 μg / mL. A normal control group (no blue light irradiation, given water) and a model control group (irradiated with blue light, given water) were also established, with a well volume of 3 mL. After treatment at 28℃ for 1 day, zebrafish in each experimental group were stained with AO light-protected material for 30 min, washed three times with standard dilution water, and then 10 zebrafish were randomly selected from each experimental group and photographed under a fluorescence microscope. ImageJ software was used to analyze and collect data. The average fluorescence intensity of apoptotic cells in the zebrafish eyes was analyzed and statistically analyzed to evaluate the anti-blue light-induced eye damage efficacy of the samples. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software, and p < 0.05 indicated that the difference was statistically significant.

[0103] 1.3 Test Results

[0104] Under the experimental conditions, Examples 1-2 and Comparative Examples 1-4 all demonstrated anti-blue light eye damage efficacy, specifically by reducing the average fluorescence intensity of apoptotic cells in the eye. Furthermore, Examples 1-2 showed superior anti-blue light eye damage efficacy compared to Comparative Examples 1-4. See Table 5 for details. Figure 1 .

[0105] Table 5. Experimental results evaluating the efficacy of the samples in preventing blue light-induced eye damage (n = 10)

[0106]

[0107] Note: Compared with the model control group, *** indicates p<0.001, * indicates p<0.05; compared with the normal control group, no hash indicates p>0.05. ## This indicates that p < 0.01. ### This indicates that p < 0.001.

[0108] As shown in Table 5, Examples 1-2 significantly reduced the fluorescence intensity of blue light-induced apoptotic cells in zebrafish eyes. Example 2 was more effective than the positive control Coenzyme Q10. Although Example 1 was slightly less effective than Coenzyme Q10, both were better than Comparative Examples 1-4, and the effect of Example 2 was closest to the normal state. The improvement effects of Comparative Examples 1-4 were weak, and Comparative Example 4 was even not statistically significant, further highlighting the significant advantages of the examples in preventing blue light eye damage. Overall, the data show that Examples 1-2 exhibit excellent potential for preventing blue light eye damage by inhibiting ocular cell apoptosis and can be used as a preferred solution for anti-blue light eye protection products.

[0109] Example 2: Evaluation of the efficacy of myopia prevention

[0110] 2.1 Testing Materials

[0111] 2.1.1 Sample Information

[0112] The brain-boosting and vision-improving compositions prepared in Examples 1-2 and Comparative Examples 1-4 were prepared using standard dilution water as the solvent.

[0113] Positive control: Atropine sulfate monohydrate, white crystals, batch number I2111034, Shanghai Aladdin Biochemical Technology Co., Ltd., solvent is standard dilution water.

[0114] 2.1.2 Laboratory Animals

[0115] Zebrafish were raised in aquarium water at 28℃ (water quality: 200 mg of readily soluble sea salt added per 1 L of reverse osmosis water; conductivity 450~550 μS / cm; pH 6.5~8.5; hardness 50~100 mg / L CaCO3), bred and provided by our company's aquarium. The laboratory animal use license number is: SYXK (Zhejiang) 2022-0004. The husbandry and management met the requirements of international AAALAC certification (certification number: 001458), and the IACUC ethics review number is: IACUC-2025-12427-01.

[0116] 2.1.3 Instruments, Consumables and Reagents

[0117] Dissecting microscope (SZX7, OLYMPUS, Japan); CCD camera (VertA1, Shanghai Tusen Vision Technology Co., Ltd., China); microinjection apparatus (IM300, Narishige, Japan); needle puller (PC-10, Narishige, Japan); precision electronic balance (CP214, OHAUS, USA); 6-well plate (Zhejiang Beilanbo Biotechnology Co., Ltd., China).

[0118] Methylcellulose (batch number C2004046, Shanghai Aladdin Biochemical Technology Co., Ltd., China).

[0119] 2.2 Evaluation of the efficacy in preventing myopia

[0120] Wild-type AB strain zebrafish at the single-cell stage were randomly selected and placed in 6-well plates, with 30 zebrafish treated in each well. Except for the normal control group and the MO standard control group, all other experimental groups were injected with lumican-MO to establish a zebrafish myopia model. After treatment at 28℃ for 2 days, samples were administered in water (concentrations shown in Table 6), with a positive control of 2500 μg / mL atropine sulfate monohydrate. Normal control, MO standard control, and model control groups were also set up, with a volume of 3 mL per well. After further treatment at 28℃ for 3 days, 10 zebrafish from each experimental group were randomly selected and photographed under a dissecting microscope. Images were saved, and data were analyzed and collected using NIS-Elements D 3.20 advanced image processing software. The ratio of retinal pigment epithelium diameter to scleral diameter was analyzed, and the statistical analysis results of this index were used to evaluate the myopia prevention efficacy of the samples. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS software, and p < 0.05 was considered statistically significant.

[0121] 2.3 Test Results

[0122] Under the experimental conditions, Examples 1-2 and Comparative Examples 1-4 all demonstrated myopia prevention efficacy, specifically by increasing the ratio of retinal pigment epithelium diameter to scleral diameter. Examples 1-2 showed superior myopia prevention efficacy compared to Comparative Examples 1-4. See Table 6 for details. Figure 2 .

[0123] Table 6. Experimental results evaluating the efficacy of the samples in preventing myopia (n = 10)

[0124]

[0125] Note: Compared with the model control group, *** indicates p<0.001, * indicates p<0.05; compared with the normal control group, ### This indicates that p < 0.001.

[0126] As shown in Table 6, both Example 1 and Example 2 significantly improved the ratio of retinal pigment epithelium diameter to scleral diameter. Example 1 was more effective than the positive control, Atropine sulfate monohydrate, while Example 2 was slightly less effective than the positive control. However, both were significantly better than Comparative Examples 1-4. The improvement effect of Comparative Examples 1-4 was weaker, further highlighting the significant advantages of the examples in preventing myopia.

[0127] Example 3: Evaluation of the effect of promoting brain development

[0128] 3.1 Testing Materials

[0129] 3.1.1 Sample Information

[0130] The brain-boosting and vision-improving compositions prepared in Examples 1-2 and Comparative Examples 1-4 were prepared using standard dilution water as the solvent.

[0131] Positive control: Ginkgo biloba and Cistanche deserticola tablets, brown tablets, batch number 3313N903, Amway (China) Daily Necessities Co., Ltd., solvent is standard dilution water.

[0132] 3.1.2 Laboratory Animals

[0133] Zebrafish were raised in aquarium water at 28℃ (water quality: 200 mg of readily soluble sea salt added per 1 L of reverse osmosis water; conductivity 450~550 μS / cm; pH 6.5~8.5; hardness 50~100 mg / L CaCO3), bred and provided by our company's aquarium. The laboratory animal use license number is: SYXK (Zhejiang) 2022-0004. The husbandry and management met the requirements of international AAALAC certification (certification number: 001458), and the IACUC ethics review number is: IACUC-2025-12427-01.

[0134] 3.1.3 Instruments, Consumables and Reagents

[0135] Dissecting microscope (SZX7, OLYMPUS, Japan); CCD camera (VertA1, Shanghai Tusen Vision Technology Co., Ltd., China); Precision electronic balance (CP214, OHAUS, USA); Zebrafish behavior analysis system (Zebra Lab3.22.3.31, Viewpoint, France); 6-well plate (Zhejiang Beilanbo Biotechnology Co., Ltd., China).

[0136] 3.2 Detection Method

[0137] Five dpf wild-type AB strain zebrafish were randomly selected and placed in beakers, with 30 zebrafish treated in each beaker (experimental group). Water-soluble samples (concentrations shown in Table 7) were administered, along with a positive control of 125 μg / mL Ginkgo biloba and Cistanche deserticola tablets. A normal control group was also included. Each beaker had a volume of 20 mL. After treatment at 28℃ for one day, five zebrafish from each experimental group were randomly placed into a cross-shaped module. The module was divided into four regions: yellow, blue, red, and green. Six modules were placed in each group. Data were collected using a behavior analyzer, and the percentage (%) of the total movement distance of the zebrafish within the blue region relative to the total movement distance of the entire region within 10 minutes was analyzed. The statistical analysis results of this index were used to evaluate the brain development promotion effect of the samples. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software, and p < 0.05 indicated statistical significance.

[0138] 3.3 Test Results

[0139] Under the experimental conditions, Examples 1-2 and Comparative Examples 1-4 all demonstrated the effect of promoting brain development. Furthermore, Examples 1-2 showed a superior effect in promoting brain development compared to Comparative Examples 1-4. See Table 7 for details. Figure 3 .

[0140] Table 7. Experimental results evaluating the efficacy of the samples in promoting brain development (n = 6)

[0141]

[0142] Note: Compared with the normal control group, *p<0.05, ***p<0.001.

[0143] As shown in Table 7, Examples 1-2 significantly increased the proportion of movement in the blue area of ​​zebrafish. Among them, Example 1 was more effective than the positive control Ginkgo biloba and Cistanche deserticola tablets, Example 2 was comparable to the positive control, and both were significantly better than Comparative Examples 1-4. Although Comparative Examples 1-4 had a certain promoting effect, the effect was weak, further highlighting the significant advantages of the Examples in promoting brain development.

[0144] The above detailed description is a specific illustration of one feasible embodiment of the present invention, and this embodiment is not intended to limit the patent scope of the present invention. It should be noted that all equivalent implementations or modifications made without departing from the present invention should be included within the scope of the technical solution of the present invention. Therefore, the protection scope of the present invention should be determined by the appended claims.

Claims

1. A composition for improving brain function and eyesight, characterized in that, It includes the following ingredients in parts by weight: 200-240 parts algal oil, 60-80 parts linseed oil, 8-12 parts arachidonic acid, 4-8 parts marigold extract, 0.6-2 parts zeaxanthin suspension oil, 0.02-0.04 parts NA nervonic acid, and 0.4-0.6 parts PS phosphatidylserine; The preparation method of the marigold extract includes the following steps: S1. Pretreatment: Marigold flowers are frozen, crushed, and then vacuum dried; S2, Single extraction: The pretreated material is mixed with the extraction solvent and subjected to ultrasonic-assisted extraction in the presence of a compound enzyme; S3. Secondary extraction: The filter residue after the first extraction is mixed with the extraction solvent again and subjected to variable amplitude ultrasonic extraction. S4. Post-processing: Combine the extracts and perform microfiltration, nanofiltration concentration and drying sequentially to obtain marigold extract; The complex enzyme described in step S2 includes cellulase, β-glucanase, and hemicellulase; The ultrasound-assisted extraction in step S2 uses dual-frequency ultrasound, with a low frequency of 20-30kHz and a power of 260-350W, and a high frequency of 100-120kHz and a power of 150-220W. The frequency of the variable amplitude ultrasound in step S3 changes periodically between 20kHz and 40kHz, with a change period of 4-6 minutes, and the ultrasound power is 350-450W.

2. The composition according to claim 1, characterized in that, The low frequency in step S2 is 30kHz with a power of 350W, and the high frequency is 120kHz with a power of 220W. Or the low frequency is 20kHz, the power is 260W, the high frequency is 100kHz, and the power is 150W; The frequency of the variable amplitude ultrasound in step S3 changes periodically between 20kHz and 40kHz, with a change period of 4 minutes and an ultrasound power of 450W. Alternatively, the frequency of the variable amplitude ultrasound changes periodically between 20kHz and 40kHz, with a period of 6 minutes, and the ultrasound power is 350W.

3. The composition according to claim 1, characterized in that, The mass percentages of cellulase, β-glucanase, and hemicellulase in step S2 are 2-4: 1-3: 0.5-1.

5.

4. The composition according to claim 3, characterized in that, The mass percentages of cellulase, β-glucanase, and hemicellulase in step S2 are 3:2:

1.

5. The composition according to claim 1, characterized in that, The freezing process described in step S1 is carried out at a temperature of -30 to -50°C for 8 to 16 hours. The average particle size of the pulverized particles is 0.5-1.0 mm; The vacuum drying process involves a vacuum degree of 5-15 Pa, a temperature of 20-30℃, and a time of 3-5 hours. The extraction time for ultrasound-assisted extraction in step S2 is 1-2 hours, and the extraction temperature is 35-40℃. In step S2, during the ultrasound-assisted extraction process, the ultrasound is paused for 1-5 minutes every 10-30 minutes, and the mixture is stirred at a speed of 150-250 r / min. The extraction solvent in step S2 is a 30-50% aqueous ethanol solution; the pH value of the extraction solvent is 4.5-5.

5. The extraction time for the variable amplitude ultrasound in step S3 is 0.8-1.2 hours, and the extraction temperature is 32-38℃. The microfiltration process described in step S4 uses a filter membrane with a pore size of 0.2-0.8 μm. The nanofiltration membrane used has a molecular weight cutoff of 500-1000 Da; The drying process is vacuum freeze drying, and the moisture content after drying is ≤3.0%.

6. The composition according to any one of claims 1-5, characterized in that, The ingredients include the following components by weight: 240 parts algal oil, 60 parts linseed oil, 8 parts arachidonic acid, 8 parts marigold extract, 0.6 parts zeaxanthin suspension oil, 0.02 parts NA nervonic acid, and 0.6 parts PS phosphatidylserine; It may include the following ingredients in parts by weight: 200 parts algal oil, 80 parts linseed oil, 12 parts arachidonic acid, 4 parts marigold extract, 2 parts zeaxanthin suspension oil, 0.04 parts NA nervonic acid and 0.4 parts PS phosphatidylserine.

7. A method for preparing the composition according to any one of claims 1-6, comprising the following steps: (1) Mix the algal oil, linseed oil and arachidonic acid in the specified amounts to obtain a uniform oil phase mixture; (2) Add marigold extract to the oil phase mixture obtained in step (1), mix well, and obtain mixture A; (3) Add zeaxanthin suspension oil to mixture A obtained in step (2), mix well, and obtain mixture B; (4) Add NA nervonic acid and PS phosphatidylserine to the mixture B obtained in step (3), mix well, and the composition is obtained.

8. A functional food, characterized in that, Includes the composition according to any one of claims 1-6.

9. The use of the composition according to any one of claims 1-6 or the functional food according to claim 8 in the preparation of brain-boosting and vision-improving products.

10. The application according to claim 9, characterized in that, The product is a functional food or a medicine.

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