Use of a nutritional composition

Abstract

The present application belongs to the field of food, and particularly relates to a use of a nutritional composition. The present application provides a non-therapeutic use of a nutritional composition in the preparation of a food for assisting in improving memory, the nutritional composition comprising necessary active ingredients as raw materials shown in (I) and (II) as follows: (I) a vitamin D component, (II) lutein, the mass ratio of the total necessary active ingredients shown in (I) to the total necessary active ingredients shown in (II) being 1:(2-110), and the combination of the vitamin D and the lutein in a certain proportion can effectively slow down synaptic plasticity damage, and the two have a synergistic effect.

Description

Technical Field

[0001] This invention belongs to the food field, specifically relating to the non-therapeutic use of a nutritional composition in the preparation of food that helps improve memory. Background Technology

[0002] With economic and social development, people's pace of life has accelerated, and stress has generally increased. People of all ages may experience varying degrees of cognitive decline or memory loss. This stress has varying degrees of impact on physical health, most commonly causing headaches and memory loss. In more severe cases, learning and cognitive functions are impaired, seriously affecting daily life. Furthermore, the probability of developing neurodegenerative diseases (such as Alzheimer's and Parkinson's) increases with age. Currently, the specific pathogenesis of these diseases is not fully understood, but oxidative stress in the central nervous system is one of the important contributing factors. Studies have shown that memory decline or regression is related to damage to the hippocampus in the brain, reduced neurotransmitters, and altered synaptic plasticity.

[0003] The basic building block of the brain is the neuron, which consists of dendrites, axons, and a cell body. Neurons have clearly defined functions, responsible for receiving, transmitting, and processing information. Axon terminals connect to the dendrites of new neurons, eventually forming a vast neural network structure. Signal transmission between axons and dendrites occurs through pulse firing. Synaptic plasticity refers to the reversible alteration of the morphology, structure, and function of synaptic connections between neurons. This alteration can be presynaptic or postsynaptic, such as an increase or decrease in the amount of neurotransmitters released by synaptic vesicles in presynaptic neurons, or changes in receptor density and postsynaptic potential in postsynaptic neurons. Synaptic plasticity is a crucial foundation for the nervous system's adaptation to changes in the external environment and for the formation of learning and memory. Damage to synaptic plasticity may manifest as a decrease in the number of synapses, damage to synaptic structure, impaired neurotransmitter release or receptor function, and abnormal expression of synaptic-related proteins (such as PSD-95 and SYN). Damage to synaptic plasticity is closely related to impaired memory function and the occurrence of depression. Brain-derived neurotrophic factor (BDNF) has a significant impact on hippocampal synaptic plasticity, and memory function can be improved by regulating the BDNF signaling pathway.

[0004] In recent years, vitamin D (VD) deficiency has become a global health problem, affecting nearly one billion people. Besides its crucial role in bone calcium metabolism, vitamin D also plays important roles in other areas such as the cardiovascular, endocrine, and nervous systems. Vitamin D is an essential nutrient for the human body. Its synthesis begins with the conversion of 7-dehydrocholesterol in the skin into inactive VD3 under light exposure. VD3 binds to vitamin D binding protein (VBP) in plasma and is transported to the liver, where it is converted into 25-hydroxycholecalciferol (25(OH)D3) by 25-hydroxylase. This is the main circulating form of VD in the body and a key biomarker for assessing VD levels. 25(OH)D3 is further transported to the kidneys, where it is converted into the active form 1,25-dihydroxycholecalciferol (1,25(OH)2D3) by 25-hydroxy-1α-hydroxylase. This is the main active form of VD in the body. Studies have shown that vitamin D hydroxylase exists in the brain, and regions such as the hippocampus and cortex are rich in vitamin D receptors (VDRs), providing a physiological basis for the role of 1,25(OH)2D3 in the brain. Vitamin D can induce the expression of nerve growth factor (NGF), promoting neuronal neurite growth and differentiation; regulate the expression of L-type calcium channels, which is crucial for synaptic plasticity; and has antioxidant and anti-inflammatory effects, which can reduce the damage of oxidative stress and inflammation to nerve cells. In addition, vitamin D has also shown certain protective effects in neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. For example, the study cited in reference 1 found that vitamin D can inhibit ferroptosis by activating the Nrf2 / HO-1 pathway, thereby alleviating 6-hydroxydopamine (6-OHDA)-induced pheochromocytoma (PC12) cell damage.

[0005] Lutein is a natural carotenoid widely found in nature, especially in green vegetables (such as spinach and kale) and certain flowers. In the human body, it is mainly concentrated in the macula of the retina, hence its name. In recent years, lutein has received widespread attention for its various health benefits, particularly for eye health. Lutein is a powerful antioxidant that neutralizes free radicals and reduces oxidative stress damage to cells. It plays a protective role in the retina, preventing light-induced oxidative damage. Lutein can absorb blue light, reducing its harmful effects on the eyes and thus protecting the retina from light damage. Lutein can cross the blood-brain barrier and has a protective effect on the central nervous system.

[0006] Despite existing research, such as that cited in reference 1 above, there is still room for further discussion regarding the development of synergistic nutritional effects of multiple substances in current technologies.

[0007] References:

[0008] Reference 1: Mu Qingshuang, Li Peishan, Li Yanxia, ​​et al. Study on the mechanism of vitamin D in alleviating 6-hydroxydopamine-induced PC12 cell damage by inhibiting ferroptosis through Nrf2 / HO-1 [J]. Journal of Xinjiang Medical University, 2023, 46(12):1580-1588. Summary of the Invention

[0009] The problem the invention aims to solve

[0010] While many scholars have conducted research on relieving mental stress and treating memory decline caused by physiological, biochemical, or disease-related factors, drug treatments often have side effects. Therefore, active nutrients that can improve memory are gradually becoming a research hotspot both domestically and internationally, especially in the development of functional foods that can improve memory for daily consumption by various population groups.

[0011] Currently, the existing technologies, as mentioned above, cited reference 1 only studied vitamin D and focused on the mechanism of ferroptosis, paying attention to apoptosis, oxidative stress indicators (MDA, SOD, GSH), iron metabolism-related genes (GPX4, FTH-1, etc.), and the Nrf2 / HO-1 pathway. However, it did not study the mechanism of action on synaptic plasticity, nor did it involve changes in the expression of synaptic plasticity-related proteins or neurotrophic factors. In addition, the study in this reference did not involve the mechanism of its combination with other active ingredients in food, health products, and other fields.

[0012] Furthermore, research on the mechanistic role of lutein in synaptic plasticity is currently lacking. Therefore, further research is needed on how to combine nutritional components in functional food compositions for daily consumption by various population groups to maximize their effectiveness in assisting memory improvement.

[0013] Based on this, the present invention aims to provide a nutritional composition containing vitamin D and lutein as the main active ingredients, and further explores and discovers that the combined use of vitamin D and lutein has a significant effect on slowing down synaptic plasticity damage, and further studies the range of their ratios to achieve the above-mentioned synergistic effect. As a result, a safe and effective nutritional intervention is provided for the prevention and adjuvant improvement of memory decline, depressive behavior, mild cognitive impairment, and neurodevelopmental delay.

[0014] Solution for solving the problem

[0015] To address the aforementioned technical problems, the present invention provides the following technical solution:

[0016] [1]. The non-therapeutic use of a nutritional composition in the preparation of a food that helps improve memory, characterized in that the nutritional composition comprises the essential active ingredients as raw materials as shown in (I) and (II) below:

[0017] (I) A vitamin D component, wherein the vitamin D component is selected from one or more of vitamin D2 and vitamin D3.

[0018] (II) Lutein, said lutein comprising free form and / or esterified form;

[0019] Furthermore, the total mass ratio of the essential active ingredient shown in (I) to the total essential active ingredient shown in (II) is 1:(2-110).

[0020] The aids in mitigating neuronal synaptic plasticity include at least one of the following: increasing the survival rate of damaged neurons, enhancing the ability of damaged neurons to secrete trophic factors, and enhancing the ability of damaged neurons to express synaptic plasticity-related proteins.

[0021] [2]. According to the non-therapeutic use described in [1], the characteristic is that (II) is free lutein.

[0022] [3]. According to the non-therapeutic use described in [1] or [2], the characteristic is that the neuronal synaptic plasticity damage is mediated by oxidative stress.

[0023] [4]. According to the non-therapeutic use described in [1] or [2], the ability of the injured nerve cells to secrete trophic factors includes helping to increase the secretion of nerve growth factor and / or brain-derived neurotrophic factor by neuroendocrine cells.

[0024] [5]. According to the non-therapeutic use described in [1] or [2], the ability of the injured nerve cells to express synaptic plasticity-related proteins includes contributing to the expression of postsynaptic density protein 95 and / or synaptophysin protein.

[0025] [6]. According to the non-therapeutic use described in [1] or [2], the food is characterized in that it includes any one or more of infant food, children's food, adolescent food, maternal food, adult food and food for the middle-aged and elderly.

[0026] [7]. For non-therapeutic use as described in [1] or [2], the food is characterized in that it contains any one or more of the following ingredients: plant-based ingredients, animal dairy ingredients, animal meat ingredients, functional additives and any acceptable excipients.

[0027] [8]. According to the non-therapeutic use described in [1] or [2], the food is characterized in that, based on the total dry matter content of the food, the vitamin D component in the food is 1 to 50 μg / 100g, and the lutein content in the food is 100 to 300 μg / 100g.

[0028] The effects of the invention

[0029] This invention discloses the non-therapeutic application of a nutritional composition containing vitamin D and lutein in the preparation of health supplements or functional foods that aid in memory improvement. The nutritional composition of this invention, by combining vitamin D and lutein in a specific ratio, exhibits a significant synergistic effect in slowing down synaptic plasticity damage in nerve cells.

[0030] Specifically, this invention demonstrates that the nutritional composition can synergistically improve the decreased survival rate of nerve cells caused by oxidative stress. Experimental results show that, compared with the use of vitamin D or lutein alone, the combined use of the two can more effectively improve the survival rate of damaged nerve cells in combating oxidative stress (H2O2)-induced PC12 nerve cell damage.

[0031] This invention confirms that the nutritional composition can synergistically improve the insufficient secretion of neurotrophic factors in neuroendocrine cells caused by oxidative stress. Experimental results show that, compared with the use of vitamin D or lutein alone, the combined use of the two can promote the secretion of NGF and / or BDNF in PC12 nerve cells in the fight against oxidative stress (H2O2)-induced damage, thereby mitigating the damage to synaptic plasticity in nerve cells.

[0032] This invention confirms that the nutritional composition can synergistically improve the abnormal expression of postsynaptic density protein 95 and / or synaptophysin protein induced by oxidative stress. Experimental results show that, compared with the use of vitamin D or lutein alone, the combined use of the two can effectively upregulate the expression of PSD-95 and / or SYN in combating oxidative stress (H2O2)-induced damage to PC12 neurons, thus alleviating synaptic plasticity damage caused by oxidative stress in PC12 cells.

[0033] Based on the above mechanisms and effects, the vitamin D and lutein composition of the present invention is particularly suitable for preparing non-therapeutic foods that help improve memory, providing a safe and effective nutritional intervention for the prevention and assistance in improving memory decline, depressive behavior, mild cognitive impairment, and neurodevelopmental delay. Detailed Implementation

[0034] The following describes embodiments of the present invention, but the present invention is not limited thereto. The present invention is not limited to the various configurations described below, and various modifications can be made within the scope of the claims. Embodiments and examples obtained by appropriately combining the technical means disclosed in different embodiments are also included in the technical scope of the present invention.

[0035] In this invention, "comprising," "having," "including," or "containing" can mean included or open-ended, and does not exclude additional, uncited elements or method steps. At the same time, "comprising," "having," "including," or "containing" can also mean closed-ended, excluding additional, uncited elements or method steps.

[0036] In this invention, the meaning of "may" includes both performing a certain process and not performing a certain process.

[0037] In this invention, "optional" or "optionally" means that certain substances, components, execution steps, application conditions, etc., are used or not used.

[0038] In this invention, the numerical ranges represented by "value A ~ value B", "value A - value B", and "value A above / below" refer to the ranges including the endpoint values ​​A and B.

[0039] In this invention, the term "about" is used to define that the numerical ranges and parameters of this invention are approximate values, and the relevant values ​​in the specific embodiments have been presented as precisely as possible. Unless otherwise explicitly stated, it should be understood that all ranges, quantities, values, and percentages used in this invention are modified by the term "about". Here, "about" generally means that the actual value is within ±1%, ±0.8%, ±0.5% of a specific value or range. Furthermore, the values ​​and ranges appearing in this invention should be understood to include systematic errors that are unavoidable in industrial production.

[0040] In this invention, terms such as "some specific / preferred embodiments," "other specific / preferred embodiments," and "implementation" refer to specific elements (e.g., features, structures, properties, and / or characteristics) related to a particular embodiment that are included in at least one of the embodiments described herein, and may or may not be present in other embodiments. Furthermore, it should be understood that these elements may be combined in any suitable manner in various embodiments.

[0041] In this invention, all unit names used are international standard unit names.

[0042] In this invention, "infants and toddlers" refers to the human group under the age of 3 years, including infants aged 0-6 months, older infants aged 6-12 months, and toddlers aged 12-36 months.

[0043] In this invention, "children" refers to the human group aged 3-6 years.

[0044] In this invention, "teenagers" refers to the human group aged 7-18.

[0045] In this invention, "pregnant women" includes women who are pregnant and women who are breastfeeding.

[0046] In this invention, "middle-aged and elderly" refers to the human group aged 41 and above.

[0047] Unless otherwise defined, other technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0048] <Nutritional Combinations>

[0049] The nutritional composition provided by this invention comprises the essential active ingredients shown in (I) to (II) below:

[0050] (I) Vitamin D components,

[0051] (II) Lutein;

[0052] The total mass ratio of the essential active ingredient shown in (I) to the total essential active ingredient shown in (II) is 1:(2-110), for example, it can be 1:3, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:11, 1:12, 1:13, 1:14, 1:15, 1:16, 1:17, 1:18, 1:19, 1:20, 1:21, 1:22, 1:23, 1:24, 1:25, 1:26, 1:27, 1:28, 1:29, 1:30, 1:31, 1:32, 1:33, 1:34, 1:35, 1:36, 1:37, 1:38, 1:39, 1:40, 1:41, 1:42, 1:43, 1:44, 1:45, 1:46, 1:47, 1:48, 1:49, 1:50, 1:51, 1:52, 1:53, 1:54, 1:55, 1:56, 1:57, 1:58, 1:59, 1:60, 1:61, 1:62, 1:63, 1:64, 1:65, 1:66, 1:67 1:68, 1:69, 1:70, 1:71, 1:72, 1:73, 1:74, 1:75, 1:76, 1:77, 1:78, 1:79, 1:80, 1:81, 1:82, 1:83, 1:84, 1:85, 1:86, 1:87, 1:88, 1:89, 1:90, 1:91, 1:92, 1:93, 1:94, 1:95, 1:96, 1:97, 1:98, 1:99, or 1:105, etc.; preferably, the necessary activity shown in (I) The mass ratio of the active ingredient to the essential active ingredient shown in (II) is 1:(5-50); more preferably, the mass ratio of the essential active ingredient shown in (I) to the essential active ingredient shown in (II) is 1:(5-40); even more preferably, the mass ratio of the essential active ingredient shown in (I) to the essential active ingredient shown in (II) is 1:(5-30); and even more preferably, the mass ratio of the essential active ingredient shown in (I) to the essential active ingredient shown in (II) is 1:(5-25).

[0053] The combination of the essential active ingredient shown in (I) and the essential active ingredient shown in (II) within the above-mentioned mass ratio range exhibits a significant synergistic effect in mitigating neuronal synaptic plasticity damage.

[0054] In some embodiments, the nutritional composition comprises an active ingredient (a component that performs a specific physiological function, i.e., a component that has a non-therapeutic effect that helps alleviate synaptic plasticity damage) and an inactive ingredient (a substance that does not have an effect that helps alleviate synaptic plasticity damage). Exemplarily, the inactive ingredient may be other nutrients, any food-acceptable excipient, and / or substances that are generated during the production or acquisition of the active ingredient and cannot be effectively separated from the active ingredient or do not need to be separated. In some embodiments, the nutritional composition consists of the active ingredient and the inactive ingredient.

[0055] In some embodiments, the vitamin D component used as a raw material may be selected from one or more of vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). For the purpose of better achieving the improved effects of the present invention, it is preferred that part or all of the vitamin D is derived from cholecalciferol, for example, all of it.

[0056] In some implementations, the vitamin D may be of natural origin, such as extracted from cod liver oil, lanolin, yeast, or lichen.

[0057] In other embodiments, the vitamin D may be obtained by chemical synthesis or bio-fermentation techniques.

[0058] In some embodiments, the lutein includes free and / or esterified forms (and these substances collectively constitute the total lutein composition of the present invention), for example, one or more selected from lutein crystals, lutein esters, lutein oil suspensions, or lutein microparticles.

[0059] In some embodiments, the lutein may be of natural origin, such as plant extracts, typically such as marigold (Tagetes erecta) flower extract, etc.

[0060] In other embodiments, the lutein may be derived from raw materials prepared through artificial synthesis.

[0061] The present invention does not impose any particular limitation on the form of the nutritional composition; typically, it can be a liquid or a solid. From the perspective of ease of production, transportation, storage, and use, the nutritional composition of the present invention is preferably a solid.

[0062] <Uses of Nutritional Combinations>

[0063] This invention proposes that combining vitamin D and lutein in a certain ratio can help improve conditions related to synaptic plasticity damage (non-therapeutic purposes), and that the two substances have a synergistic effect.

[0064] In principle, this invention does not specifically limit the causes of synaptic plasticity damage. Such causes may include abnormal protein aggregation and toxicity, neuroinflammation and abnormal activation of glial cells, mitochondrial dysfunction, hormonal or metabolic disorders, chronic stress or environmental factors, etc. However, it has been found that the vitamin D and lutein of this invention are particularly effective in improving synaptic plasticity damage caused by oxidative stress.

[0065] In some implementations, the improvement of neuronal synaptic plasticity includes improving neurotransmitter release disorders, improving neurotransmitter receptor dysfunction, and improving abnormal expression of synaptic-related proteins.

[0066] In some embodiments, the improvement in neuronal synaptic plasticity includes at least one of increasing the survival rate of damaged neurons, enhancing the ability of damaged neurons to secrete trophic factors, and enhancing the ability of damaged neurons to express synaptic plasticity-related proteins.

[0067] In some embodiments, the enhancement of the ability of damaged nerve cells to secrete trophic factors includes helping to increase the secretion of nerve growth factor and / or brain-derived neurotrophic factor from neuroendocrine cells.

[0068] In some implementations, the enhancement of the ability of damaged nerve cells to secrete trophic factors includes helping to increase the secretion of nerve growth factor and brain-derived neurotrophic factor from neuroendocrine cells.

[0069] In some embodiments, the ability of damaged nerve cells to express synaptic plasticity-related proteins includes contributing to increased expression of postsynaptic density protein 95 and / or synaptophysin protein.

[0070] In some implementations, the ability of damaged nerve cells to express synaptic plasticity-related proteins includes contributing to increased expression of postsynaptic density protein 95 and synaptophysin.

[0071] The methods described in this invention to help alleviate synaptic plasticity damage are not intended to treat or prevent disease, and the conditions related to synaptic plasticity damage described in this invention do not reach the level that can be identified as a disease.

[0072] By improving synaptic plasticity damage as described above, this invention can macroscopically improve memory decline, depressive behavior, cognitive dysfunction, and neurodevelopmental delay, among any one or more of these.

[0073] In some implementations, the cognitive impairment includes non-pathological anxiety, depression, migraines, stress, etc.

[0074] In some embodiments, the neurodevelopmental delay includes non-pathological levels of hyperactivity, inattention, attention deficit, ADHD, sleep disorders, etc. The present invention does not specifically limit the specific non-therapeutic foods containing or prepared using the above-described nutritional composition.

[0075] In some embodiments, the food described in this invention is in the form of a liquid or a solid under normal temperature conditions.

[0076] In some implementations, the food is infant food, children's food, adolescent food, pregnant and postpartum food, adult food, or food for the middle-aged and elderly.

[0077] In some embodiments, the food described in this invention is a confectionery, such as hard candy, gel candy, shortbread candy, compressed candy, and aerated candy.

[0078] In some embodiments, the food described in this invention is a beverage, such as carbonated beverages, tea beverages, coffee beverages, fruit and vegetable juice beverages, and lactic acid bacteria beverages.

[0079] In some embodiments, the food described in this invention is a dairy product, such as milk powder, cheese, yogurt, liquid milk, etc.

[0080] In some embodiments, the food described in this invention is a baked food, such as bread, cake, and biscuits.

[0081] In some embodiments, the food products described in this invention are dietary supplements, such as hard capsules, soft capsules, tablets, oral liquids, pills, granules, and powders.

[0082] In some embodiments, in the food product of the present invention, the mass ratio of vitamin D to lutein is 1:(2-100); preferably, the mass ratio of vitamin D to lutein is 1:(2-50); more preferably, the mass ratio of vitamin D to lutein is 1:(5-40); even more preferably, the mass ratio of vitamin D to lutein is 1:(5-30); and even more preferably, the mass ratio of vitamin D to lutein is 1:(5-25).

[0083] This invention does not impose any specific absolute limits on the content of vitamin D and lutein in food, as long as the requirements of local food-related laws and regulations are met.

[0084] For example, in the food product, the vitamin D content, based on the total dry matter content, is 1-50 μg / 100g, preferably 3-50 μg / 100g, and more preferably 7-30 μg / 100g. The lutein content in the food product is 100-300 μg / 100g, preferably 120-300 μg / 100g, and more preferably 150-300 μg / 100g.

[0085] In addition to the components described above in the nutritional composition, the food may also contain other ingredients, such as common food ingredients like proteins / amino acids, carbohydrates, fats, vitamins, and minerals.

[0086] Furthermore, depending on the type of food and the end needs of the target audience, in some embodiments, the food contains any one or more of the following ingredients: plant-based ingredients, animal dairy ingredients, animal meat ingredients, functional additives, and any acceptable excipients.

[0087] Examples of plant-based ingredients include fruits such as figs, pomegranates, kiwis, oranges, tangerines, pineapples, strawberries, apples, bananas, grapes, pears, cherries, blueberries, blackberries, blackcurrants, cranberries, raspberries, melons, amla, and bilberries, or their extracts; fruits and vegetables such as onions, cucumbers, tomatoes, cauliflower, carrots, spinach, kale, Brussels sprouts, garlic, basil, and oregano, or their extracts; grains such as rice (indica, japonica, glutinous rice), cereals (wheat, barley, oats, rye), corn, sorghum, millet, sorghum, yellow millet, buckwheat, soybeans, broad beans, peas, mung beans, red beans, and kidney beans, or their extracts; nuts such as walnuts, pistachios, cashews, hazelnuts, almonds, apricot kernels, pine nuts, peanuts, sunflower seeds, chestnuts, macadamia nuts, and ginkgo nuts, or their extracts; and coffee or its extracts.

[0088] Animal dairy product ingredients can include fresh milk from cows and sheep, as well as reprocessed dairy products such as whole milk powder, skim milk powder, whey protein concentrate, desalted whey powder, whey protein powder, hydrolyzed whey protein powder, and casein powder.

[0089] Examples of animal meat product ingredients include pork, beef, mutton, seafood, and poultry.

[0090] Examples of functional additives include other vitamin supplements (such as vitamin A, beta-carotene, vitamin E, vitamin K1, vitamin B1, vitamin B2, vitamin B6, vitamin B12, vitamin C, pantothenic acid, folic acid, niacin, biotin, etc.), mineral supplements (such as iron, copper, manganese, zinc, cobalt, molybdenum, chromium, nickel, vanadium, fluorine, selenium, iodine, silicon, tin, etc.), nucleotide supplements (e.g.), dietary fiber (e.g., inulin, konjac flour, galactooligosaccharides, fructooligosaccharides, isomaltooligosaccharides, soybean polysaccharides, cyclodextrin, resistant dextrin, soybean fiber, etc.), and functional polyunsaturated fatty acid supplements (e.g., arachidonic acid oil powder, docosahexaenoic acid oil powder, etc.).

[0091] Any acceptable excipients may include solvents, antioxidants, antibacterial agents, thickeners, diluents, cosolvents, stabilizers, emulsifiers, fillers, disintegrants, lubricants, coating materials, anti-caking agents, flavoring agents, sweeteners, food flavorings, and food colorings.

[0092] Example

[0093] The embodiments of the present invention will be described in detail below with reference to experimental examples. However, those skilled in the art will understand that the following experimental examples are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified, the experimental examples were conducted under conventional conditions or conditions recommended by the manufacturer. Unless otherwise specified, all materials and instruments used are commercially available conventional products.

[0094] The raw materials used in the experimental examples of this invention include: Vitamin D3 (cholecalciferol) (purchased from MCE, USA) and lutein (purchased from Sigma, USA).

[0095] The main reagents used in the experimental examples of this invention include: NGF kit and BDNF kit (purchased from Shanghai Jianglai Biotechnology Co., Ltd.), and BCA protein concentration assay kit (purchased from Beyotime Biotechnology Co., Ltd.)

[0096] The main instruments used in the experimental examples of this invention include: the SynergyHTX fully automated microplate reader (purchased from BioTek) and a carbon dioxide incubator (purchased from Thermo Fisher Scientific China Co., Ltd.)

[0097] The cells used in the experimental examples of this invention are pheochromocytoma (PC12) cells, purchased from the Cell Bank of the Chinese Academy of Sciences. PC12 cells are tumor cells of the sympathetic nervous system, possessing the general characteristics of neuroendocrine cells. They exhibit a neuronal phenotypic response to nerve growth factor (NGF), accompanied by physiological and biochemical changes. They share many structural and functional similarities with dopaminergic neurons and are characterized by rapid growth and proliferation, short culture cycles, and easily controllable culture conditions. Currently, they are frequently used internationally to study the physiology, pathology, and pharmacology of neurons. The neuronal oxidative damage model established based on PC12 cells is an important pharmacological model, often used to screen for active ingredients in vitro. Oxidative stress is one of the common mechanisms leading to nerve cell damage. Treating PC12 cells with oxidants (such as H2O2) can induce synaptic damage, manifested as decreased expression of synaptic-related proteins, altered cell morphology, and neurotransmitter imbalance. Therefore, in the experimental examples of this invention, H2O2 is used as an inducer of oxidative damage to construct the PC12 cell synaptic damage model. In the comparative and examples, vitamin D and lutein were added to observe whether they had a protective effect in slowing down synaptic plasticity damage in PC12 cells.

[0098] Experimental Example 1: Effect of Nutritional Composition on PC12 Cell Viability

[0099] This experiment investigated the effects of vitamin D and lutein at different doses, alone or in combination, on the survival rate of PC12 cells.

[0100] 1. Experimental Methods

[0101] 1.1 Construction of PC12 cell synaptic injury model and experimental grouping

[0102] After PC12 cells were revived, they were cultured in DMEM medium with 5% fetal bovine serum, 10% heat-inactivated horse serum and 1% penicillin-streptomycin added. The culture was then placed in a 37°C, 5% CO2 humidified incubator. The samples were divided into 14 groups and treated differently: ① Blank control group; ② Positive control group (hydrogen peroxide stimulation group): hydrogen peroxide was added to the culture medium to a final concentration of 100 μmol / L and treated for 6 h; ③ Comparative and example groups: as shown in Table 1, each well of each group was filled with 100 μL of vitamin D (Comparative Example 1: 1 μg / mL; Comparative Example 2: 0.5 μg / mL; Comparative Example 3: 0.1 μg / mL), 100 μL of lutein (Comparative Example 4: 20 μg / mL; Comparative Example 5: 10 μg / mL; Comparative Example 6: 5 μg / mL), or 100 μL of a combination of the two (Example 1: 0.5 μg / mL vitamin D, 20 μg / mL lutein; Example 2: 1 μg / mL vitamin D, 10 μg / mL lutein; Example 3: 0.5 μg / mL vitamin D, 10 μg / mL lutein; Example 4: 0.1 μg / mL vitamin D, 10 μg / mL lutein). Lutein; Example 5: 0.1 μg / mL vitamin D, 5 μg / mL lutein; Example 6: 1 μg / mL vitamin D, 5 μg / mL lutein) After treatment for 24 h, hydrogen peroxide was added to a final concentration of 100 μmol / L, and treatment was carried out for 6 h.

[0103] Table 1. Design schemes for mitigating synaptic plasticity damage using different nutrient ratios.

[0104]

[0105] 1.2 Detection of the effect of nutrients on cell viability:

[0106] To investigate the effects of vitamin D and lutein, and their combined use, on PC12 cell viability, this study used the Cell Counting Kit-8 (CCK-8) assay to determine PC12 cell viability. Following the manufacturer's instructions, PC12 cells were seeded into 96-well plates at a density of 1 × 10⁶ cells per well. 4 Cells were divided into groups in 96-well plates. After grouping and processing, 10 μL of CCK-8 reagent kit solution was added to each well of the plate, and the plates were incubated at 37°C for 1 h. The absorbance at 450 nm was measured using a microplate reader, and cell viability was calculated. The results were statistically analyzed and expressed as mean ± standard deviation (x ± SD). P > 0.05 indicated no significant difference, and P < 0.05 indicated a significant difference.

[0107] 2. Experimental Results

[0108] The experimental results are shown in Table 2. After H2O2 treatment, the cell survival rate of PC12 cells in the positive control group was approximately 50%. Compared with the positive control group, both vitamin D and lutein treatments improved cell survival rate. Furthermore, the combined use of vitamin D and lutein significantly improved the survival rate of H2O2-induced PC12 cells, with an even higher cell survival rate.

[0109] Table 2. Effects of Vitamin D and Lutein on the Viability of PC12 Cells

[0110]

[0111] As shown in Table 2, there are limits to the improvement in cell viability when vitamin D or lutein are used alone, and these limits are reached at medium doses of each substance. However, when the two substances are used in combination, effects significantly exceeding these limits are achieved. Generally, if there is no synergistic effect between the two substances, even when combined, the effect is unlikely to exceed the individual limits for improving cell viability.

[0112] Furthermore, as further evidence of the synergistic effect of the two, a comparison between Examples 2-4 and Comparative Examples 1-3 shows that the latter's effect on increasing cell activity reaches a peak with the amount of vitamin D used, while the former, due to the addition of lutein, changes the form of such dosage and effect.

[0113] Experimental Example 2: Effect of Nutritional Composition on the Concentration of Neurotrophic Factor in PC12 Cells

[0114] 1. Experimental Methods

[0115] 1.1 Construction of PC12 cell synaptic injury model and experimental grouping

[0116] Same as Experiment 1.

[0117] 1.2 Measurement of neurotrophic factor-related indicators

[0118] Neurotrophic factors are a class of proteins that play a crucial role in the survival, development, and functional maintenance of neurons. Among them, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are two of the most important neurotrophic factors, playing multiple key roles in the nervous system. NGF and BDNF have complementary and synergistic effects in synaptic plasticity. NGF primarily provides the basis for synaptic plasticity by supporting neuronal survival and growth, while BDNF directly promotes the occurrence and development of synaptic plasticity by regulating neurotransmitter release, enhancing synaptic transmission efficiency, and inducing long-term potentiation (LTP). Together, they maintain the normal function of the nervous system and play important roles in learning, memory, and neuronal injury repair.

[0119] After cell culture and grouping, cells were collected in centrifuge tubes and stored at -80℃ for later use. The NGF and BDNF levels in PC12 cells were measured according to the instructions of the NGF and BDNF kits. The results were statistically analyzed and expressed as mean ± standard deviation (x ± SD). P > 0.05 indicated no significant difference, and P < 0.05 indicated a significant difference.

[0120] 2. Experimental Results

[0121] NGF and BDNF are two important neurotrophic factors that play crucial roles in neuronal survival, differentiation, synaptic plasticity, and functional regulation. NGF supports neuronal development and maintains neuronal survival, thus providing a basis for synapse formation and plasticity. After nerve injury, NGF can promote the regeneration and repair of damaged neurons, helping to restore neural function and synaptic connections.

[0122] This study verified the effects of nutrients on the secretion of neurotrophic factor (NGF) in PC12 cells. The results are shown in Table 3. Compared with the blank control group, the positive control group treated with H2O2 significantly reduced the intracellular NGF content (P<0.05). Compared with the positive control group, the NGF content in the vitamin D alone treatment group and the vitamin D and lutein combination treatment group were significantly increased (P<0.05). Although the NGF content in the lutein alone treatment group was higher than that in the positive control group, the difference was not statistically significant (P>0.05). Compared with single nutrient treatments, the NGF content in the vitamin D and lutein combination treatment group was significantly increased. Taking Comparative Examples 2, 5, and 3 as examples, the increase in NGF content in Example 3 relative to the positive control group (43.94) was greater than the sum of the increases in NGF content in Comparative Examples 2 and 5 relative to the positive control group (21.25+11.25). The results indicate that the combined use of vitamin D and lutein can synergistically promote the secretion of NGF in cells and play a role in mitigating synaptic plasticity damage.

[0123] Table 3. Effects of nutrients on the secretion of neurotrophic factor (NGF) by PC12 cells.

[0124]

[0125] Note: * indicates a significant difference from the control group (P<0.05), # indicates a significant difference from the positive control group (P<0.05).

[0126] BDNF is a key neurotrophic factor involved in most forms of synaptic plasticity. BDNF promotes the release of neurotransmitters and enhances receptor function, thereby improving the efficiency of synaptic transmission. BDNF is a key molecule inducing long-term potentiation (LTP), one of the main forms of synaptic plasticity closely related to learning and memory. BDNF can activate a series of signaling pathways through its receptor TrkB, promoting the formation and maintenance of LTP.

[0127] This study verified the effects of nutrients on the secretion of neurotrophic factor BDNF in PC12 cells. The results are shown in Table 4. Compared with the blank control group, the positive control group treated with H2O2 significantly reduced the intracellular BDNF content (P<0.05). Compared with the positive control group, the BDNF content in the vitamin D alone treatment group and the vitamin D and lutein combination treatment group were significantly increased (P<0.05), while the BDNF content in the lutein alone treatment group was higher than that in the positive control group, but the difference was not significant (P>0.05). Compared with single nutrient treatment, the vitamin D and lutein combination group showed a greater increase in BDNF. Taking Comparative Examples 1, 5, and 2 as examples, the increase in BDNF content in Example 2 relative to the positive control group (29.16) was greater than the sum of the increases in BDNF content in Comparative Examples 1 and 5 relative to the positive control group (10.07+6.7). In addition, the BDNF content in Examples 2 and 3 was close to that in the blank group (P>0.05). The results showed that the combined use of vitamin D and lutein could synergistically promote the secretion of BDNF in cells and promote synaptic plasticity.

[0128] Table 4. Effects of nutrients on the secretion of neurotrophic factor BDNF by PC12 cells.

[0129]

[0130] Note: * indicates a significant difference from the control group (P<0.05), # indicates a significant difference from the positive control group (P<0.05).

[0131] Experimental Example 3: Effects of Nutritional Compositions on Synaptic Plasticity-Related Proteins

[0132] 1. Experimental Methods

[0133] 1.1 Construction of PC12 cell synaptic injury model and experimental grouping

[0134] Same as Experiment 1.

[0135] 1.2 Determination of the content of synaptic plasticity-related proteins

[0136] Synaptic plasticity-related proteins play a crucial role in the function and adaptation of the nervous system. These proteins influence processes such as nerve signal transmission, learning, and memory by regulating the structure and function of synapses. Among them, postsynaptic density-95 (PSD-95) and synaptophysin (SYN), located at the two ends of neuronal connections, are the main markers of synaptic plasticity. After cell culture and grouping, cells were collected in centrifuge tubes and stored at -80℃ for later use. The levels of SYN and PSD-95 in PC12 cells were measured according to the instructions of the BCA protein quantification kit. The results were statistically analyzed and expressed as mean ± standard deviation (x ± SD). P > 0.05 indicated no significant difference, and P < 0.05 indicated a significant difference.

[0137] 2. Experimental Results

[0138] Synaptic plasticity-related proteins play crucial roles in the function and adaptation of the nervous system. These proteins influence processes such as neural signal transmission, learning, and memory by regulating synaptic structure and function. PSD-95, a key protein in the postsynaptic compactum, participates in synapse formation and maintenance and is a major marker of synaptic plasticity. It regulates synaptic transmission and plasticity through interactions with NMDA and AMPA receptors. PSD-95 plays a vital role in LTP (synaptic transduction), enhancing synaptic transmission by stabilizing receptor expression at the postsynaptic membrane. PSD-95 is essential for maintaining and promoting synaptic plasticity.

[0139] The expression results of PSD-95 are shown in Table 5. Compared with the blank control group, the expression of PSD-95 in the positive control group treated with H2O2 was significantly decreased (P<0.05). Compared with the positive control group, the expression of PSD-95 in the medium-dose vitamin D alone treatment group and the vitamin D and lutein combination treatment group was significantly increased (P<0.05). Although the expression of PSD-95 in the high-dose vitamin D alone treatment group and the lutein treatment group was higher than that in the positive control group, the difference was not significant (P>0.05). Compared with single nutrient treatment, the increase in PSD-95 content in the vitamin D and lutein combination was more significant. Taking Comparative Example 2, Comparative Example 4, and Example 1 as examples, the increase in PSD-95 content in Example 1 relative to the positive control group (0.38) was greater than the sum of the increases in PSD-95 content in Comparative Example 2 and Comparative Example 4 relative to the positive control group (0.19+0.07). The above results indicate that the combination of vitamin D and lutein can synergistically and effectively upregulate PSD-95 expression and alleviate synaptic plasticity damage caused by oxidative stress in PC12 cells.

[0140] Table 5. Effects of nutrients on PSD-95 expression in PC12 cells

[0141]

[0142] Note: * indicates a significant difference from the control group (P<0.05), # indicates a significant difference from the positive control group (P<0.05).

[0143] SYN is a synaptic vesicle membrane protein that primarily participates in the docking and fusion of synaptic vesicles and regulates neurotransmitter release. SYN expression levels reflect synaptic activity and quantity, and are a key marker of synaptic plasticity. SYN participates in neurotransmitter transmission and axonal growth, playing a crucial role in synaptic formation and maintaining plasticity.

[0144] The SYN expression results are shown in Table 6. Compared with the blank control group, the SYN expression in the H2O2-treated positive control group was significantly decreased (P<0.05). Compared with the positive control group, the SYN expression in the vitamin D-treated group and the lutein-treated group was increased, but the change was not significant (P<0.05), while the SYN expression in the vitamin D and lutein combination treatment group was significantly increased (P<0.05). Meanwhile, compared with single nutrient treatments, the increase in SYN content in the vitamin D and lutein combination was more significant. Taking Comparative Examples 3, 6, and 5 as examples, the increase in SYN content in Example 5 relative to the positive control group (4.74) was greater than the sum of the increases in SYN content in Comparative Examples 3 and 6 relative to the positive control group (1.45+1). Therefore, the combination is more synergistically effective in upregulating SYN expression than single treatments, and has a stronger function in maintaining synaptic plasticity.

[0145] Table 6. Effects of nutrients on SYN expression in PC12 cells

[0146]

[0147] Note: * indicates a significant difference from the blank group (P<0.05), # indicates a significant difference from the positive control group (P<0.05).

Claims

1. Non-therapeutic use of a nutritional composition for the manufacture of a foodstuff for assisting the improvement of memory, characterized in that, The aforementioned memory enhancement is achieved by mitigating synaptic plasticity damage mediated by oxidative stress, and the nutritional composition comprises the essential active ingredients as raw materials as shown in (I) and (II) below: (I) Vitamin D component, wherein the vitamin D component is vitamin D3, (II) Lutein, said lutein comprising free form and / or esterified form; Furthermore, the total mass ratio of the essential active ingredient shown in (I) to the total essential active ingredient shown in (II) is 1:(2-110). The mitigation of oxidative stress-mediated neuronal synaptic plasticity damage includes at least one of the following: increasing the survival rate of damaged neurons, enhancing the ability of damaged neurons to secrete trophic factors, and enhancing the ability of damaged neurons to express synaptic plasticity-related proteins. The ability of damaged nerve cells to secrete trophic factors includes helping to increase the secretion of nerve growth factor and / or brain-derived neurotrophic factor by neuroendocrine cells. The ability to enhance the expression of synaptic plasticity-related proteins in damaged nerve cells includes helping to increase the expression of postsynaptic density protein 95 and / or synaptophysin protein.

2. The non-therapeutic use according to claim 1, characterized in that, (II) is free lutein.

3. The non-therapeutic use according to claim 1 or 2, characterized in that, The food products include any one or more of the following: infant food, children's food, adolescent food, and food for pregnant and postpartum women.

4. The non-therapeutic use according to claim 1 or 2, characterized in that, The food products mentioned also include adult food products.

5. The non-therapeutic use according to claim 1 or 2, characterized in that, The food products mentioned also include those for middle-aged and elderly people.

6. The non-therapeutic use according to claim 1 or 2, characterized in that, The food contains any one or more of the following ingredients: plant-based ingredients, animal dairy ingredients, and animal meat ingredients.

7. The non-therapeutic use according to claim 1 or 2, characterized in that, The food also contains functional additives.

8. The non-therapeutic use according to claim 1 or 2, characterized in that, In the food, based on the total dry matter content, the vitamin D component in the food is 1~50 μg / 100g, and the lutein content in the food is 100~300 μg / 100g.

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