NRF2-targeted composition capable of efficiently releasing sulforaphane and application of NRF2-targeted composition

Abstract

The invention relates to a nutrition targeting composition which comprises a component 1 for providing glucoraphanin, a component 2 for providing myrosinase, vitamin C and magnesium stearate and is capable of efficiently releasing sulforaphane and activating an NRF2 target spot. The invention also relates to the use of said composition for the preparation of a product for the prevention or treatment of diseases or conditions that can be prevented or treated with sulforaphane.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and relates to a composition that efficiently releases sulforaphane and activates the NRF2 target, and its application in the preparation of products for the prevention or treatment of diseases or conditions that can be prevented or treated by sulforaphane. Background Technology

[0002] Chronic non-communicable diseases, or chronic diseases, have become a major global public health challenge, and their prevention and control strategies are shifting from traditional treatment to early prevention and precise intervention. Nutrition-targeted (or precision nutrition) intervention, with "nutrogenomics" as its core and combined with "clinical evidence-based" technology, constructs a dynamic matching system of genes and active ingredients. By screening dietary active ingredients that can specifically regulate the expression or function of key genes, it promotes the implementation of personalized nutrition intervention and provides an innovative approach for the prevention and control of chronic diseases. Nuclear factor E2-related factor 2 (NRF2) is a transcription factor that regulates the cellular response to antioxidant stress and is also a key transcription factor of the human endogenous antioxidant system. After activation, it can regulate more than 500 genes (such as phase II enzyme NQO1, glutathione thiotransferase, etc.), enhance endogenous defenses such as detoxification, mitochondrial function, and clearance of abnormal proteins (Pall, ML, & Levine, S. Acta Physiologica Sinica., 2015, 67(1), 1-18.), and thus become a core target for chronic disease intervention. Based on the activation effect of the NRF2 pathway, it can play a therapeutic and preventive role in a variety of diseases, including gastric ulcers, Helicobacter pylori infection, cancer, diabetes, cardiovascular disease, autism, schizophrenia, depression, and Alzheimer's disease.

[0003] Sulforaphane (SFN), also known as sulforaphane, is the most potent NRF2 activator found in edible plants to date. Compared to plant polyphenols such as curcumin and resveratrol, which have lower bioavailability, sulforaphane can effectively activate the NRF2 pathway in multiple tissues in the human body at dietary limits (Houghton CA. Curr Top Med Res. 2020;2:123.). Studies have shown that sulforaphane can induce the expression of cell protection genes by efficiently activating the NRF2 signaling pathway, thereby exerting a sustained and more effective antioxidant and chemopreventive effect (Young-Joon S. Science. 2003, 3, 768). Current technology also indicates that sulforaphane and its precursor compound sulforaphane glycoside have clear chemopreventive effects against gastric ulcers and Helicobacter pylori infection (CN1935003A; CN1170472C; CN101208079B). Therefore, developing compositions that can efficiently deliver and stably release sulforaphane is of great value for achieving nutritional intervention targeting NRF2.

[0004] Cruciferous plants are the main source of sulforaphane and its precursor compound, sulforaphane glycosides, with broccoli (Brassica oleracea) being the core source, as its seeds and sprouts contain relatively higher levels of sulforaphane glycosides. However, broccoli contains non-biologically active sulforaphane glycosides, which require the breakdown of myrosinase from cruciferous plants to be converted into active sulforaphane. However, sulforaphane, as an isothiocyanate, is chemically unstable and easily degraded by oxygen and temperature, making it unsuitable for storage and application. To address this limitation, existing technology proposes a solution: pre-mixing myrosinase and sulforaphane glycoside raw materials into solid forms, thus avoiding enzymatic decomposition of sulforaphane glycosides during storage. The enzymatic hydrolysis is then completed in a solution environment after being added to water or taken orally, enabling effective absorption of sulforaphane glycosides (see CN110946996A). However, research has found that for existing products used for chronic disease prevention, especially mixtures of black myrosinase and sulforaphane raw materials (including raw materials in the form of broccoli and / or its extracts) in solid form, the release of sulforaphane in aqueous solution is limited, affecting the utilization of the products.

[0005] Therefore, it is necessary to overcome the problem of low sulforaphane release rate in existing products, especially the limited release of sulforaphane in aqueous solutions of compositions containing sulforaphane and myrosinase. There is an urgent need to provide a composition containing sulforaphane and myrosinase that can efficiently release sulforaphane in aqueous solutions and thus effectively activate the NRF2 target. Summary of the Invention

[0006] While researching the dosage form of the combination of sulforaphane and myrosinase, the inventors unexpectedly discovered that adding magnesium stearate during the formulation process not only enhances the particle flowability and compressibility of the composition containing sulforaphane, myrosinase, and vitamin C, but also promotes the conversion of sulforaphane to sulforaphane, significantly enhances the release of sulforaphane in the aqueous solution, and greatly improves the formation rate of sulforaphane.

[0007] Based on this discovery, in a first aspect of the present invention, a composition for enhancing sulforaphane release is provided, comprising the following components:

[0008] (1) Provides component 1 of glucosinolates;

[0009] (2) Provides component 2 of black mustard enzyme;

[0010] (3) Vitamin C; and

[0011] (4) Magnesium stearate.

[0012] In this invention, the component 1 providing sulforaphane can be any substance or raw material capable of providing a source of sulforaphane compounds. Preferably, the component 1 providing sulforaphane is selected from cruciferous plants, their extracts, and mixtures thereof. The cruciferous plants are preferably selected from broccoli, cauliflower, red cabbage, Brussels sprouts, or cabbage, with broccoli being particularly preferred. The cruciferous plants can be all or part of a plant, such as the whole plant, its above-ground parts, flower heads, sprouts, seeds, or combinations thereof. The term broccoli refers to all or part of a broccoli plant. Preferably, the broccoli is selected from the edible parts in the general sense; more preferably, the broccoli is selected from broccoli flower heads, broccoli seeds, and broccoli sprouts, and combinations thereof.

[0013] In this invention, the component 1 providing sulforaphane can also be an extract of cruciferous plants, such as a solvent extract, preferably a water extract, an alcohol extract, or a hydroalcoholic extract. Besides plant tissues, extracts, and mixtures thereof from cruciferous plants, the component 1 providing sulforaphane in this invention can also include chemically synthesized, semi-chemically synthesized, enzymatically synthesized, or biosynthesized sulforaphane. The broccoli extract refers to an extract of all or part of the broccoli plant, including but not limited to extracts of broccoli, broccoli heads, broccoli seeds, and / or broccoli sprouts. On the other hand, the extract is an extract obtained by solvent extraction, preferably a water extract, an alcohol extract, or a hydroalcoholic extract, particularly preferably a water extract.

[0014] In this invention, the component 1 providing sulforaphane is preferably selected from broccoli florets, broccoli seeds, broccoli sprouts, broccoli extracts and mixtures thereof, as well as sulforaphane in chemically or biosynthetically synthesized forms.

[0015] In this invention, the component 2 providing black myrosinase can be any suitable substance or raw material capable of providing a source of black myrosinase. Preferably, the component 2 providing black myrosinase is selected from cruciferous plants, their extracts, and mixtures thereof. Preferably, the component 2 providing black myrosinase is selected from horseradish, radish, and cabbage. In some preferred embodiments, the component 2 providing black myrosinase is selected from extracts of horseradish, radish, and cabbage; in other preferred embodiments, the component 2 providing black myrosinase is selected from the juice or pulp of horseradish, radish, and cabbage, or a powder obtained by drying the juice or pulp.

[0016] In this invention, the magnesium stearate can be a product refined from a mixture of oxidase and edible-grade solid beverage fatty acids (mainly stearic acid). In the composition of this invention, the magnesium stearate can be "pharmaceutical-grade magnesium stearate," "food-grade magnesium stearate," "magnesium stearate conforming to the standards of the Chinese Pharmacopoeia," or "magnesium stearate conforming to GB 1886.91 'National Food Safety Standard for Food Additives: Magnesium Stearate'." Preferably, the magnesium stearate in this invention can be in the form of dried magnesium stearate powder, or in the form of its hydrate, alcohol, or water-alcohol mixture. The alcohol can be selected from ethanol, methanol, propanol, or butanol. Dried magnesium stearate powder is particularly preferred in this invention.

[0017] In this invention, the mass ratio of component 1, component 2, vitamin C and magnesium stearate is (10-90):(1-90):(0.1-10):(1-10), preferably (70-90):(10-30):(0.5-1.5):(1-3), and most preferably 80:12:1:2.

[0018] In some embodiments, the composition of the present invention for improving sulforaphane release may consist of the following components:

[0019] (1) Provides component 1 of glucosinolates;

[0020] (2) Provides component 2 of black mustard enzyme;

[0021] (3) Vitamin C; and

[0022] (4) Magnesium stearate.

[0023] The compositions of the present invention are preferably in solid form, for example, in the form of powder, granules, capsules, or tablets. More preferably, in the compositions of the present invention, components 1, 2, and magnesium stearate are all present in solid form. For example, component 1 may be in the form of an extract, a sprout, or a seed powder (including lyophilized powder); component 2 may be in the form of a dried powder or a dried extract (including lyophilized powder); and magnesium stearate may be in the form of magnesium stearate powder. In some preferred embodiments, component 1 of the present invention is selected from broccoli seed extract, broccoli sprout powder, broccoli floret lyophilized powder, and mixtures thereof.

[0024] In another aspect, the present invention provides the use of the nutritionally targeted composition for improving sulforaphane release as described herein in the preparation of products for the prevention and / or treatment of diseases or conditions that can be prevented and / or treated using sulforaphane. The products include pharmaceuticals or food, preferably pharmaceuticals. In some preferred embodiments, the diseases or conditions associated with the prevention and / or treatment using sulforaphane are selected from cancer, diabetes, cardiovascular disease, Helicobacter pylori infection, autism, schizophrenia, depression, Alzheimer's disease (AD), and pulmonary fibrosis.

[0025] In another aspect, the present invention provides a method for converting sulforaphane into sulforaphane in vitro, comprising the following steps:

[0026] 1) Provide a composition for increasing sulforaphane release as described in this invention.

[0027] 2) Mix the composition with water or an aqueous solution.

[0028] In another aspect, the present invention also provides a method for supplementing sulforaphane to a subject in need, comprising administering to the subject a nutrient-targeting composition as described in the present invention for improving sulforaphane release.

[0029] In another aspect, the present invention also provides a method for promoting the release of sulforaphane in an aqueous solution, comprising the step of mixing a mixture containing component 1 providing sulforaphane, component 2 providing myrosinase and vitamin C with magnesium stearate.

[0030] In another aspect, the present invention also provides the use of magnesium stearate in the preparation of products for improving the release of sulforaphane in water or aqueous solutions or for promoting the conversion of sulforaphane to sulforaphane in water or aqueous solutions.

[0031] The inventors unexpectedly discovered that by adding magnesium stearate to the composition, the conversion rate of sulforaphane released from the composition containing sulforaphane, myrosinase and vitamin C in water can be effectively improved, thereby achieving enhanced sulforaphane release and effectively overcoming the problem of low sulforaphane release in the prior art. Attached Figure Description

[0032] Figure 1 This is a bar chart showing the release of sulforaphane from the composition (powder) and the control composition in aqueous solution.

[0033] Figure 2 This is a bar chart showing the release of sulforaphane from the composition (tablet) of the present invention and the control composition in aqueous solution. Detailed Implementation

[0034] Preparation Example 1 Preparation of the composition (powder) and control composition of the present invention

[0035] Preparation of the composition of the present invention: 400 g of broccoli seed water extract (containing 15.1% sulforaphane, from Brassica Protection Products LLC, USA, hereinafter the same) was mixed evenly with 60 g of horseradish powder, 5 g of vitamin C, and 10 g of magnesium stearate to obtain 475 g of composition 1, wherein sulforaphane accounts for 12.72%. The above composition was packaged into small bags of 5 g each to obtain the corresponding powder product (the composition of the present invention).

[0036] Preparation of the control composition: 400 g of broccoli seed water extract (containing 15.1% sulforaphane) was mixed evenly with 60 g of horseradish powder and 5 g of vitamin C to obtain 465 g of the control composition, in which sulforaphane accounted for 12.99%. The above control composition was packaged into small bags of 5 g each to obtain the corresponding powder product (control composition).

[0037] Take 1 g of each of the two powder products and add them to 200 ml of artificial gastric fluid. Use 37 ℃ to simulate brewing conditions. Take a sample at 90 min and determine the content of sulforaphane in the aqueous solution by HPLC. Calculate the sulforaphane formation rate.

[0038] HPLC determination of sulforaphane: The sample solution was filtered through a 0.45 μm filter membrane and analyzed by HPLC. HPLC conditions: Column: Huapu Company Unitary C18 (4.6 mm × 250 mm, 5 μm); Column temperature: 30 ℃; Mobile phase: 70% water-30% acetonitrile; Flow rate: 0.8 mL / min; Injection volume: 10 μL; UV detection wavelength: 245 nm.

[0039] The experimental results are shown in Table 1 below.

[0040] Table 1. Comparison of sulforaphane formation rates between the powder products of the present invention and the control composition (n=2)

[0041]

[0042] The experimental results show that, compared to the control composition without magnesium stearate, the composition of the present invention containing magnesium stearate increased the formation rate of sulforaphane in aqueous solution by 60%, a significant difference, which is a very surprising result. This indicates that the added magnesium stearate significantly improved the conversion of sulforaphane to sulforaphane in aqueous solution. Such a result is unexpected.

[0043] Preparation Example 2Preparation of the composition (tablet) of the present invention and its control tablet

[0044] Preparation of the composition (tablets) of the present invention: 400 g of broccoli seed water extract (containing 15.1% sulforaphane), 60 g of horseradish powder, 5 g of vitamin C, 10 g of magnesium stearate, and 500 g of other tableting excipients (specifically starch, maltodextrin, and hydroxypropyl methylcellulose in a w / w ratio of 5:80:2, the same below) were mixed evenly, compressed into tablets at a weight of 1 g / tablet, and coated with a film to obtain approximately 975 g of tablets of the present invention, wherein sulforaphane accounts for 6.19%. The above-mentioned tablets of the present invention were dispensed into bottles of 60 tablets each, and after adding a desiccant, the bottle openings were sealed to obtain the corresponding tablet product.

[0045] Preparation of the control tablet: 400 g of broccoli seed water extract (containing 15.1% sulforaphane), 60 g of horseradish powder, 5 g of vitamin C, and 500 g of other tableting excipients were mixed evenly and compressed into tablets at a weight of 1 g / tablet. The tablets were then film-coated to obtain approximately 965 g of the tablets of this invention, wherein sulforaphane accounted for 6.26%. The tablets of this invention were dispensed into bottles of 60 tablets each, and after adding a desiccant, the bottles were sealed to obtain the corresponding tablet product.

[0046] In Example 2, 30 g of each of the two tablet products were ground and 1 g of powder was added to 200 ml of artificial gastric fluid and heated at 37 °C to simulate brewing conditions. At 90 min, a sample was taken and the content of sulforaphane in the aqueous solution was determined by HPLC, and the formation rate of sulforaphane was calculated.

[0047] HPLC determination of sulforaphane: The above sample solution was filtered through a 0.45 μm filter membrane and analyzed by HPLC. HPLC conditions: Column: Huapu Company Unitary C18 (4.6 mm × 250 mm, 5 μm); Column temperature: 30 ℃; Mobile phase: 70% water-30% acetonitrile; Flow rate: 0.8 mL / min; Injection volume: 10 μL; UV detection wavelength: 245 nm.

[0048] The experimental results are shown in Table 2 below.

[0049] Table 2. Comparison of sulforaphane formation rates between the tablets containing the composition of the present invention and the control tablets (n=2)

[0050]

[0051] The experimental results show that, compared to the control tablet without magnesium stearate, the tablets of this invention containing magnesium stearate increased the formation rate of sulforaphane in aqueous solution by 60%, a significant difference, which is a very surprising result. This indicates that the conversion of sulforaphane to sulforaphane is significantly improved after the addition of magnesium stearate. Such results are unexpected.

[0052] It should be understood that although the invention has been specifically disclosed through preferred embodiments, those skilled in the art can utilize the optional features, modifications, improvements, and variations of the invention embodied in the disclosure herein, and consider such modifications, improvements, and variations to be within the scope of the invention. The materials, methods, and embodiments provided herein as exemplary preferred examples are exemplary and are not intended to limit the scope of the invention.

Claims

1. A composition comprising the following components: 1) Provides component 1 of glucosinolates; 2) Provides component 2 of black mycosesacylase; 3) Vitamin C; and 4) Magnesium stearate.

2. The composition of claim 1, wherein component 1 is selected from cruciferous plants, their extracts, and mixtures thereof.

3. The composition according to any one of claims 1 to 2, wherein the cruciferous plant is broccoli.

4. The composition according to any one of claims 1 or 3, wherein the cruciferous plant is selected from all, a portion of, or a combination thereof of plants.

5. The composition according to any one of claims 1 to 4, wherein the cruciferous plant is selected from the flower heads, seeds, seedlings, and combinations thereof.

6. The composition according to any one of claims 1 to 5, wherein the mass ratio of component 1, component 2, vitamin C and magnesium stearate is (10-90):(1-90):(0.1-10):(1-10).

7. The composition according to any one of claims 1 to 6, wherein it is in solid form.

8. The composition according to any one of claims 1 to 7, wherein it is in the form of a powder, granules, capsule or tablet.

9. The composition according to any one of claims 1 to 8, wherein component 1 is selected from broccoli seed extract, broccoli sprout powder, broccoli floret freeze-dried powder and combinations thereof, and sulforaphane in chemically or biosynthetically synthesized forms.

10. The composition according to any one of claims 1 to 9, wherein the component 2 is selected from horseradish, radish, cabbage; their juice or slurry; and their extracts.

11. Use of the composition of any one of claims 1 to 10 in the preparation of a product for the prevention or treatment of a disease or condition that can be prevented or treated with sulforaphane.

12. The use as claimed in claim 11, wherein the disease or condition that can be prevented or treated by sulforaphane is selected from cancer, diabetes, cardiovascular disease, Helicobacter pylori infection, autism, schizophrenia, depression, Alzheimer's disease, and pulmonary fibrosis.

13. A method for converting sulforaphane into sulforaphane in vitro, comprising the following steps: 1) Provide a composition as described in any one of claims 1 to 12, 2) Mix the composition with water or an aqueous solution.

14. A method of supplementing a subject with sulforaphane, comprising administering to the subject the composition as described in any one of claims 1 to 13.

15. A method for promoting the release of sulforaphane in an aqueous solution, comprising the step of mixing a sample containing a component 1 providing sulforaphane, a component 2 providing myrosinase and vitamin C with magnesium stearate.

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