Moringa oleifera extract containing nicotinamide ribose as well as preparation method and application of moringa oleifera extract

Abstract

The invention provides a moringa oleifera extract containing an anti-aging functional component nicotinamide ribose as well as a preparation method and application of the moringa oleifera extract. Belongs to the technical field of food and medicine. The invention provides the moringa oleifera extract containing a trace amount of natural anti-aging component nicotinamide ribose (NR) in fresh moringa oleifera seeds, and the moringa oleifera seed extract freeze-dried powder product contains 23.77 mu g / g of nicotinamide ribose, has anti-aging activity, and can be used for preparing new anti-aging drugs or anti-aging cosmetics or prepared into food. The invention also provides a simple and fast preparation technology of the moringa seed extract containing nicotinamide ribose (NR). The preparation method is high in operability, high in efficiency and high in yield.

Description

Technical Field

[0001] This invention belongs to the fields of food and medicine, specifically relating to Moringa extract containing the anti-aging functional ingredient nicotinamide ribose, its preparation method, and its application. Background Technology

[0002] Anti-aging research has become a consistently hot topic. In recent years, NR, NMN, and NAD have been... + Research on the anti-aging mechanisms has yielded a series of breakthroughs: Birkisdóttir's team used a mouse model of premature aging DNA repair deficiency to verify the anti-aging intervention potential of NR; Pérez-Rodríguez revealed that NR induces mitochondrial remodeling in the distal convoluted tubule of the kidney by regulating cytochrome b5 reductase 3 expression; and Elhassan found that NR can significantly increase NAD in skeletal muscle of the elderly. + At the metabolic level, Arslan's team confirmed that NMN and NR reverse ovarian aging by balancing mitochondrial fission-fusion dynamics. In the field of neuroprotection, NR not only delays melanocyte aging but also improves cognitive function in Alzheimer's disease model mice. Liu et al. found that NMN combined with lycopene reduces oxidative damage and prevents D-galactose-induced cognitive decline through the Keap1-Nrf2 pathway. Regarding reproductive aging, Huang's team confirmed that long-term NMN treatment improves ovarian reserve in mice by enhancing mitophagy, and NMN's regulation of the aortic anti-aging miRNA profile in aged mice suggests its epigenetic rejuvenation potential. Zhang's research further elucidates NAD... + Oxidized forms of NAD reverse muscle stem cell aging through mitochondrial unfolded protein responses, and the life-extending effect of NR was demonstrated in a muscular dystrophy model. A Wagner systematic review indicates that NAD... + Decreased NAD levels lead to skeletal muscle aging by affecting the activity of enzymes such as PARP, CD38, and ssirtuins, while nutritional supplementation and exercise intervention can synergistically restore NAD. + Steady state. At the mechanistic level, NAD... + Not only does it participate in the metabolic regulation of Sirtuins and PARPs as a redox coenzyme, its biosynthesis (dependent on NAMPT) synergistically maintains metabolic rhythms with SIRT1. Its age-related decline directly leads to nuclear-mitochondrial communication impairment and induces degenerative diseases. Numerous studies have shown that combined activation of the Sirtuin pathway and exogenous NAD supplementation... +The strategy of using precursors (NR / NMN) has become a cutting-edge research hotspot for reversing multi-organ aging phenotypes, preventing and treating neurodegenerative diseases, and extending healthy lifespan. Studies have reported that trace amounts of nicotinamide riboside are naturally present in milk, at a concentration of approximately 1-2 μM. Additionally, some fruits and vegetables, such as broccoli, cucumber, and cabbage, are speculated to contain trace amounts of nicotinamide riboside, but there is currently no definitive experimental data to support this. NAD+ may be present in some traditional anti-aging herbs (such as ginseng and astragalus). + Related precursors. And Dendrobium ( ) Dendrobium Plants are believed to commonly contain nicotinamide mononucleotide (NMN) and nicotinamide adenine dinucleotide (NAD). + ), including Dendrobium officinale ( Dendrobium officinale The NMN content of Dendrobium officinale is significantly higher than that of other closely related species in the same genus. The mass fraction of NMN in fresh Dendrobium officinale reaches 0.9319 μg·g⁻¹, and it also contains the unique anti-aging component NAD. + Its content is even more outstanding, more than 3 times that of NMN.

[0003] Moringa ( Moringaoleifera Moringa, also known as the drumstick tree or miracle tree, is a deciduous tree belonging to the Moringaceae family, native to India and widely distributed in tropical regions. In Guangdong and Taiwan, my country, it is commonly cultivated as an economic crop, and its medicinal and edible value is considered one of the "Three Treasures of the World," alongside ginseng and Ganoderma lucidum. Studies have reported that Moringa is rich in flavonoids (36 types), fatty acids (22 types), alkaloids (17 types), phenylpropanoids (14 types), and over a hundred secondary metabolites, including steroids, terpenes, and isothiocyanates. Its extracts have exhibited multiple biological activities, including anti-COVID-19, anti-aging, antibacterial, anticancer, antioxidant, and anti-inflammatory effects; however, most of the active ingredients remain unclear. Numerous studies have reported on the anti-aging functions of Moringa extract: Athira's team found that water extract of Moringa leaves can significantly reduce lipid peroxidation levels and lipofuscin deposition in the brain tissue of aged rats, while simultaneously increasing serotonin and antioxidant enzyme activity, suggesting its protective effect against age-related oxidative damage to the brain; Xu et al. demonstrated through in vitro experiments that Moringa leaf extract can effectively inhibit key skin aging enzymes (elastase, collagenase, and hyaluronidase), and screened active ligands using multi-target bioaffinity ultrafiltration-high performance liquid chromatography-mass spectrometry (AUF-HPLC-MS) technology, verifying their specific binding to target enzymes through molecular docking; Lim et al. systematically evaluated the bioactivity of four Moringa products (oil, fermented capsules, powder, and tea), finding that fermented capsules can significantly enhance superoxide dismutase activity, while fermented products (capsules, powder, and tea) have particularly prominent inhibitory effects on elastase and tyrosinase, confirming their anti-aging and whitening effects, but basically did not involve nicotinamide riboside (NR) and its related active substances. Summary of the Invention

[0004] In view of the above-mentioned shortcomings in the existing technology, the purpose of this invention is to provide a Moringa extract containing the anti-aging functional ingredient nicotinamide ribose, its preparation method, and its application in the preparation of food, cosmetics, and pharmaceuticals.

[0005] To achieve the above-mentioned objectives of the present invention, the present invention provides the following technical solution: A moringa seed extract containing nicotinamide ribose, wherein the moringa seed extract containing nicotinamide ribose is prepared by the following method: 1) Take fresh moringa pods, and manually remove the moringa pods to obtain fresh seeds; 2) Grind the moringa seeds, filter the juice through defatted gauze, and further extract the residue three times with ultrasonically using an equal weight of ultrapure water; 3) After combining the juice and extract, centrifuge and collect the supernatant; 4) The supernatant was freeze-dried to obtain moringa seed extract freeze-dried powder.

[0006] The moringa seed extract containing nicotinamide ribose, as described above, wherein the lyophilized moringa extract contains not less than 23.77 μg / g of nicotinamide ribose.

[0007] A method for preparing a Moringa seed extract containing nicotinamide ribose, comprising the following steps: 1) Take fresh moringa pods, and manually remove the moringa pods to obtain fresh seeds; 2) Grind the moringa seeds, filter the juice through defatted gauze, and further extract the residue three times with ultrasonically using an equal weight of ultrapure water; 3) After combining the juice and extract, centrifuge and collect the supernatant; 4) The supernatant was freeze-dried to obtain moringa seed extract freeze-dried powder.

[0008] A pharmaceutical composition comprising, as the active ingredient, a moringa seed extract containing nicotinamide ribose, and a pharmaceutically acceptable carrier.

[0009] Food containing a moringa seed extract containing nicotinamide ribose, and commonly used food additives.

[0010] The cosmetic contains a moringa seed extract containing nicotinamide ribose, and commonly used cosmetic excipients.

[0011] The application of a moringa seed extract containing nicotinamide ribose in food preparation.

[0012] The application of a moringa seed extract containing nicotinamide ribose in the preparation of cosmetics.

[0013] The use of the moringa seed extract containing nicotinamide ribose, or the pharmaceutical composition of claim 4, in the preparation of anti-aging drugs.

[0014] The preparation method of the pharmaceutical composition involves first obtaining a moringa seed extract containing nicotinamide ribose using the aforementioned preparation method, and then adding a pharmaceutically acceptable carrier.

[0015] Compared with existing technologies, this invention has the following advantages: This invention uses UPLC-MS / MS technology to collect mass spectrometry data from the pericarp, pulp, and fresh seed extracts of Moringa pods, and preliminarily analyzes their chemical composition characteristics by constructing a molecular network system, identifying the NR compound and annotating its structure. Simultaneously, for the screened potential anti-aging active ingredients, structural characterization is completed by combining mass spectrometry fragmentation patterns or standard comparison, and the content is determined using the external standard method. Furthermore, the anti-aging activity of the target components is systematically evaluated through in vitro cell models (such as SIRT1 activity detection and ROS scavenging experiments) and molecular docking simulations, resulting in a novel anti-aging product. Based on this, a simple and efficient preparation process for Moringa seed extract containing nicotinamide ribose (NR) has been improved, producing a freeze-dried Moringa seed extract product containing 23.77 μg / g of nicotinamide ribose. Attached Figure Description

[0016] Figure 1 This is a schematic diagram of the nicotinamide riboside (NR) structure. Figure 2 TIC diagram for nicotinamide ribose (NR) standard; Figure 3 TIC and EIC plots of nicotinamide ribose (NR) in fresh Moringa seeds; Figure 4 Secondary mass spectrometry of nicotinamide ribose (NR) in fresh Moringa seeds; Figure 5 Qualitative analysis of nicotinamide ribose in freeze-dried Moringa seed powder; Figure 6 The external standard curve for nicotinamide ribose (NR) in Moringa seed extract is given by lnΔε = 0.66268lnC +11.55609. Detailed Implementation

[0017] The following description, in conjunction with the accompanying drawings, uses embodiments of the present invention to further illustrate the substantive content of the invention, but does not limit the invention thereto.

[0018] Example 1 1. Preparation of Moringa seed extract containing nicotinamide ribose (NR).

[0019] The main preparation process includes: 1) 2205.1 g (PWMo) of fresh Moringa pods were accurately weighed, and 229.7 g (YSWMo) of fresh seeds were obtained after manually removing the Moringa pods. 2) After grinding the moringa seeds in a mortar and pestle, filter the juice through defatted gauze and then further extract the residue three times with ultrasonically using an equal weight of ultrapure water. 3) After combining the juice and extract, centrifuge and collect the supernatant; 4) The supernatant was freeze-dried to obtain 14.9 g of lyophilized Moringa seed extract powder (FPDMo).

[0020] Structural annotation of nicotinamide riboside (NR) in Moringa seeds.

[0021] Comparison of primary and secondary mass spectra and retention times of lyophilized Moringa seed extract powder (FPDMo) and nicotinamide ribose standards indicates that Moringa seeds may contain nicotinamide ribose (NR), with the structural formula shown below. Figure 1 .

[0022] By comparing nicotinamide ribose standards ( Figure 2 Retention time and UV spectrum of fresh Moringa seed extract freeze-dried powder (FPDMo) Figure 3 ) and secondary mass spectrometry (MS) 2 )feature( Figure 4 ),Discover m / z The compound at 255.0973 Dalton had the same retention time as the standard, and the secondary mass spectrometry fragments (53.0397, 55.0194, 57.0343, 69.0342, 80.0527, 123.0555) were a perfect match. Figure 3 and Figure 4 Preliminary evidence confirms the presence of nicotinamide ribose (NMR) in fresh Moringa seeds. Figure 5 This result confirms the presence of natural nicotinamide ribose (NR) in moringa seeds.

[0023] Qualitative analysis of nicotinamide ribose (NR) in Moringa seeds.

[0024] Qualitative analysis methods.

[0025] 1) Accurately weigh a certain amount of freeze-dried Moringa seed powder and dissolve it in a 10 ml volumetric flask to prepare an aqueous solution of a certain concentration; 2) After filtering the sample through a syringe microporous membrane, it was analyzed using an Agilent 6545 LC / Q-TOF liquid chromatography-mass spectrometry system. The analytical method is as follows: Mobile phase: Solvent A [0.1% formic acid aqueous solution (v / v)] and solvent B (acetonitrile); Time program: 0 min, 95% B; 7 min, 95% B; Detection wavelengths: 210, 230, 260, 280, and 365 nm. Injection volume: 2 μL; Column temperature: 30℃; Flow rate: 1 mL / min; Q-TOF-MS detection in positive ion mode; Mass spectrometry parameters: Dry gas temperature 300℃, flow rate 8 L / min, nebulizer gas pressure 35 psi, upper voltage limit 3500 V, fragmentation voltage 175 V, skimmer voltage 65 V. Mass range: 50–1000 Da. Reference ions: m / z 121.050873 and 922.009798 Da. Mass spectrometry mode: AUTO ms / ms.

[0026] Qualitative analysis results.

[0027] By comparing nicotinamide ribose standards ( Figure 2 Retention time and UV spectrum of Moringa seed extract freeze-dried powder (FPDMo) Figure 3 ) and secondary mass spectrometry (MS) 2 )feature( Figure 4 ),Discover m / z The compound at 255.0973 Dalton had the same retention time as the standard, and the secondary mass spectrometry fragments (53.0397, 55.0194, 57.0343, 69.0342, 80.0527, 123.0555) were a perfect match. Figure 3 and Figure 4 Preliminary evidence confirms the presence of nicotinamide ribose (NMR) in Moringa seeds. Figure 5 ).

[0028] Note: Secondary mass spectrometry (A) of nicotinamide ribose standard; secondary mass spectrometry (B) of nicotinamide ribose in lyophilized powder of Moringa seed extract at various collision energies (labeled peaks are characteristic fragment peaks of nicotinamide ribose).

[0029] Quantitative analysis of nicotinamide ribose in fresh Moringa seeds.

[0030] External standard method for quantitative analysis.

[0031] 1) The external standard method was used to quantitatively analyze nicotinamide ribose in Moringa seeds. Specifically, six concentration gradients of nicotinamide ribose standard aqueous solutions (5240 mg / L, 1048 mg / L, 209.6 mg / L, 41.92 mg / L, 8.384 mg / L, and 1.677 mg / L) were precisely prepared, and a standard curve between concentration and EIC peak area was established using mass spectrometry. 2) Construct a linear relationship to calculate the nicotinamide ribose content in Moringa seeds.

[0032] Quantitative analysis results.

[0033] Quantitative analysis showed that the EIC peak area of ​​nicotinamide ribose (NR) standard did not exhibit a direct linear relationship with its concentration. Figure 6 By transforming the peak area with the natural logarithm of the concentration, a linear equation y = 0.0098x + 11.5561 (R² = 0.99956) was established (Table 1).

[0034] In this experiment, 2205.1 g of fresh Moringa pods were processed, and 229.7 g of seeds were isolated. After extraction, purification, and freeze-drying, 14.9 g of Moringa seed extract freeze-dried powder was obtained. Final quantitative analysis showed that the nicotinamide ribose content in the freeze-dried powder of fresh Moringa seed extract was 23.77 ± 0.36 μg / g. Through conversion, the nicotinamide ribose content in fresh Moringa seeds was determined to be 1.542 ± 0.023 μg / g. This result verifies the presence of trace amounts of natural nicotinamide ribose (NR) in Moringa seeds.

[0035] Table 1. Raw data for quantitative analysis of nicotinamide ribose (NR) in fresh Moringa seeds.

[0036] Example 2 Take the Moringa seed extract obtained by the same method as in Example 1, add the excipient at a weight ratio of 1:1-5, and granulate and compress into tablets.

[0037] Example 3 Moringa seed extract obtained by the same method as in Example 1 was made into capsules using conventional capsule formulation methods.

[0038] Example 4 The Moringa seed extract obtained by the same method as in Example 1 was then prepared into tablets according to the following method:

[0039] Example 5 Capsules: Take 100 mg of Moringa seed extract obtained in the same manner as in Example 1, along with an appropriate amount of starch and stearic acid mold. Mix the compound and excipients, sieve, and mix evenly in a suitable container. Fill the resulting mixture into hard gelatin capsules.

[0040] Example 6 Take 1 part of Moringa seed extract obtained by the same method as in Example 1, 10 parts of vegetable fat powder, mix them well, and make them into a solid beverage according to conventional methods.

[0041] Example 7 The Moringa seed extract obtained by the same method as in Example 1, along with an appropriate amount of food additives, was prepared into a food product according to conventional food preparation methods.

[0042] Example 8 Preparation of Moringa Seed Extract Facial Mask.

[0043] Material The moringa seed extract obtained by the same method as in Example 1 was used, while other materials were cosmetic-grade or food-grade raw materials.

[0044] formula Moringa seed extract 12 g, xanthan gum 3.5 g, hydrolyzed hyaluronic acid 2.0 g, sodium hyaluronate 2.0 g, propylene glycol 0.3 g, carbomer 0.5 g, parabens 0.1 g, PPG-10 methyl glucoether 0.25 g, glyceryl glucoside 1.0 g, deionized water 45 g.

[0045] Facial mask manufacturing process Step 1: Add deionized water, xanthan gum, hydrolyzed hyaluronic acid, sodium hyaluronate, propylene glycol, carbomer, and parabens to an emulsifying pot. Heat to 75°C while stirring. Once the temperature is reached, keep warm and continue stirring until the solution becomes clear and homogeneous. Then keep warm for 20 minutes and cool down to 42°C.

[0046] Step 2: Adjust the pH to 7.6 ± 0.2.

[0047] Step 3: Continue cooling to 35℃, add PPG-10 methyl glucoether, Moringa seed extract, and glyceryl glucoside, and continue stirring until homogeneous. Once the mixture passes inspection, it is ready for discharge. Coat the product onto a non-woven fabric substrate at a coating amount of 2.1 g per square centimeter.

[0048] Example 9 Preparation of Moringa Seed Extract Facial Cleanser

[0049] Material The moringa seed extract obtained by the same method as in Example 1 uses cosmetic-grade or food-grade raw materials.

[0050] formula Moringa seed extract 7 g, glycerin 2.0 g, butylene glycol 4.0 g, propylene glycol 0.5 g, sodium EDTA 0.1 g, guar gum 0.3 g, zinc dioxide 2.0 g, C12-15 benzoate 4.0 g, C12-20 alkyl glucoside 3.0 g, C14-22 alcohol 0.5 g, cetearyl alcohol 1.2 g, parabens 0.2 g, sodium stearate 0.3 g, polydimethylsiloxane alcohol 0.5 g, polysorbate 0.2 g, deionized water 55 g.

[0051] Preparation process Weigh the prescribed amount of Moringa seed extract and dissolve it completely in purified water. Dissolve glycerin, butylene glycol, propylene glycol, sodium EDTA, guar gum, zinc dioxide, sodium stearate, C12-15 benzoate, C12-20 alkyl glucoside, C14-22 alcohol, and cetearyl alcohol completely in purified water. Dissolve parabens, polydimethylsiloxane, and polysorbate completely in purified water. Mix the prepared solutions thoroughly and filter through a 0.22 μm microporous membrane. Bottle the filtrate in 30 mL containers, seal, and sterilize.

[0052] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0053] The embodiments described above are merely examples of several implementations of the present invention, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention.

Claims

1. A Moringa seed extract containing nicotinamide ribose, characterized in that: The moringa seed extract containing nicotinamide ribose is prepared by the following method: 1) Take fresh moringa pods, and manually remove the moringa pods to obtain fresh seeds; 2) Grind the moringa seeds, filter the juice through defatted gauze, and further extract the residue three times with ultrasonically using an equal weight of ultrapure water; 3) After combining the juice and extract, centrifuge and collect the supernatant; 4) The supernatant was freeze-dried to obtain moringa seed extract freeze-dried powder.

2. The Moringa seed extract containing nicotinamide ribose as described in claim 1, characterized in that: The lyophilized moringa extract powder contains no less than 23.77 μg / g of nicotinamide ribose.

3. A method for preparing a Moringa seed extract containing nicotinamide ribose, characterized in that: The method includes the following steps: 1) Take fresh moringa pods, and manually remove the moringa pods to obtain fresh seeds; 2) Grind the moringa seeds, filter the juice through defatted gauze, and further extract the residue three times with ultrasonically using an equal weight of ultrapure water; 3) After combining the juice and extract, centrifuge and collect the supernatant; 4) The supernatant was freeze-dried to obtain moringa seed extract freeze-dried powder.

4. A pharmaceutical composition comprising, as an active ingredient, a moringa seed extract containing nicotinamide ribose as described in claim 1 or 2, and an additive pharmaceutically acceptable carrier.

5. A food product containing a moringa seed extract containing nicotinamide ribose as described in claim 1 or 2, and a commonly used food additive.

6. A cosmetic product containing a moringa seed extract containing nicotinamide ribose as described in claim 1 or 2, and commonly used cosmetic excipients.

7. The application of the Moringa seed extract containing nicotinamide ribose as described in claim 1 or 2 in the preparation of food.

8. The use of the moringa seed extract containing nicotinamide ribose as described in claim 1 or 2 in the preparation of cosmetics.

9. The use of a moringa seed extract containing nicotinamide ribose as described in claim 1 or 2, or the pharmaceutical composition as described in claim 4, in the preparation of an anti-aging drug.

10. The method for preparing the pharmaceutical composition according to claim 4, characterized in that: First, a moringa seed extract containing nicotinamide ribose is obtained using the preparation method described in claim 3, and then a pharmaceutically acceptable carrier is added.

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