Application of DVM nanoparticles in preparation of products for treating and / or preventing alcoholic liver diseases
By preparing DVM nanoparticles, the problems of insufficient water solubility and liver targeting of gallic acid derivatives in the treatment of alcoholic liver disease were solved, achieving more efficient hepatocellular protection and alcohol detoxification effects.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- KUNMING UNIV OF SCI & TECH
- Filing Date
- 2026-03-10
- Publication Date
- 2026-05-19
AI Technical Summary
Existing natural polyphenolic compounds, such as gallic acid and its derivatives, have limitations in their clinical application in the treatment of alcoholic liver disease due to poor water solubility, rapid metabolism, insufficient liver targeting, and low bioavailability.
DVM nanoparticles were developed by compounding gallic acid derivative VM and dihydromyricetin DM in a 1:3 ratio and adding Pluronic F-127 to prepare nanoparticles. After stirring with ethanol solution, treating with formaldehyde aqueous solution and glycine aqueous solution, the nanoparticles were prepared by water dialysis and freeze drying in MWCO 3500 to form nanoparticles with synergistic liver-protective effects.
DVM nanoparticles significantly reduced hepatocyte apoptosis and ROS production in alcohol-induced HepG2 cells, increased ADH activity, decreased AST and ALT activity, and enhanced the rate of alcohol detoxification in mice, demonstrating excellent hepatoprotective effects.
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Abstract
Description
Technical Field
[0001] This invention relates to the field of biomedical technology, specifically the application of DVM nanoparticles in the preparation of products for the treatment and / or prevention of alcoholic liver disease. Background Technology
[0002] Alcoholic liver disease (ALD) seriously endangers human health and is one of the most common types of liver disease worldwide. It is closely related to oxidative stress, inflammatory response, lipid metabolism disorder and gut microbiota imbalance. Acetaldehyde and reactive oxygen species produced during ethanol metabolism can induce hepatocyte damage and further promote the disease to alcoholic fatty liver, hepatitis, liver fibrosis, cirrhosis and even liver cancer through mechanisms such as damaging the intestinal barrier, activating inflammatory pathways and inhibiting lipid breakdown.
[0003] Currently, clinical treatment strategies for alcoholic liver disease (ALD) are still relatively limited. Commonly used clinical treatments mainly include abstinence from alcohol and nutritional support, but these have limitations such as severe withdrawal symptoms, limited efficacy, and high relapse rates. Moreover, there are currently no specific drugs for the clinical treatment of alcoholic liver disease. Common clinical drugs for the treatment of alcoholic liver disease include silymarin and S-adenosylmethionine.
[0004] Natural polyphenols, with their well-defined antioxidant and anti-inflammatory properties, have shown great potential in the prevention and treatment of ALD (Alzheimer's disease) and have become a research hotspot. However, most natural polyphenols, including gallic acid and some of its derivatives with hepatoprotective activity, generally suffer from inherent defects such as poor water solubility, rapid metabolism, insufficient liver targeting, and low bioavailability, which severely restrict their clinical translation and application. Although existing technologies have disclosed the hepatoprotective activities of some gallic acid derivatives, there is still room for improvement in their efficacy and stability.
[0005] Therefore, in order to overcome the problems existing in the prior art, the problem that this invention needs to solve is: how to develop a novel gallic acid derivative delivery system with higher efficiency and better stability, and provide DVM nanoparticles for the treatment or prevention of alcoholic liver disease. Summary of the Invention
[0006] The purpose of this invention is to provide DVM nanoparticles for the treatment or prevention of alcoholic liver disease. The DVM nanoparticles contain gallic acid derivative VM and dihydromyricetin. When compounded in a mass ratio of 1:3, they have a synergistic effect in the treatment or prevention of alcoholic liver disease.
[0007] To achieve the above objectives, the present invention provides the following technical solution:
[0008] A DVM nanoparticle for treating or preventing alcoholic liver disease, said DVM nanoparticle containing galline. acid derivatives biologyVM and dihydromyricetin; the mass ratio of the gallic acid derivative VM and dihydromyricetin is 1:3; the gallic acid... acid derivatives VM has the structural formula described in Formula I;
[0009]
[0010] Formula I.
[0011] Preferably, the DVM nanoparticles further contain Pluronic F-127, and the mass ratio of the gallic acid derivative VM, dihydromyricetin and Pluronic F-127 is 1:3:4.
[0012] Furthermore, this invention discloses a method for preparing DVM nanoparticles as described above, comprising the following steps:
[0013] Step 1: Weigh gallic acid derivative, dihydromyricetin and Pluronic F-127 according to the mass ratio, then dissolve them in ethanol solution and stir well.
[0014] Step 2: Add a 37 vol% formaldehyde aqueous solution to the product obtained in Step 1 and continue stirring. After 3-6 minutes, add a 10 mg / mL glycine aqueous solution and continue stirring for 2-6 hours.
[0015] Step 3: After dialysis purification of the product from Step 2 with MWCO 3500 water for 46-50 hours, the product is freeze-dried to obtain the DVM nanoparticles.
[0016] Furthermore, this invention discloses the use of DVM nanoparticles as described above in the preparation of products for the treatment and / or prevention of alcoholic liver disease.
[0017] Preferably, the product is a drug for treating and / or preventing alcoholic liver disease.
[0018] Finally, the present invention discloses a medicament for treating and / or preventing alcoholic liver disease, wherein the active ingredient of the medicament is DVM nanoparticles as described above.
[0019] Preferably, the dosage form of the drug is one of liquid, tablet, capsule, powder, or pill.
[0020] Compared with the prior art, the beneficial effects of the present invention are:
[0021] The DVM nanoparticles prepared by co-preparing gallic acid derivatives VM and DM provided by this invention, in alcohol-induced HepG2 cells, can better reduce hepatocyte apoptosis, ROS generation, increase ADH activity, reduce intracellular AST and ALT activity, and increase the alcohol detoxification rate in mice, compared with gallic acid and the positive drug silybin, thus exhibiting superior hepatoprotective effects. Attached Figure Description
[0022] Figure 1 Mass spectra of gallic acid derivative compounds 1-7;
[0023] Figure 2 The results of toxicity tests on HepG2 cells by gallic acid derivative compounds 1-7;
[0024] Figure 3 The results show the aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels in alcohol-induced HepG2 cells, with A representing AST and B representing ALT. ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P < 0.01 indicates a comparison between the sample group and the model group (EtOH);
[0025] Figure 4 The figures show the structural characterization results of DVM nanoparticles; in the figure, A is the transmission electron microscopy image, B is the particle size distribution, C is the dynamic light scattering, D is the X-ray diffraction, E is the Zeta potential, and F is the Fourier transform infrared spectroscopy.
[0026] Figure 5 To test the toxicity of different concentrations of DVM (5~40 μg / mL) on HepG2 cells;
[0027] Figure 6 The figure shows the inhibitory effect of DVM on alcohol-induced apoptosis and ROS in HepG2 cells; the effect of DVM on alcohol-induced acetaldehyde dehydrogenase (ADH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels in HepG2 cells; ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; * P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P < 0.01 indicates a comparison between the sample group and the model group (EtOH); ### P < 0.001 indicates a comparison between the sample group and the model group (EtOH);
[0028] Figure 7The results are from mouse balance tests, whisker response tests, tail suspension tests, and tilt plate tests. ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P < 0.01 indicates a comparison between the sample group and the model group (EtOH);
[0029] Figure 8 The figures show the serum ethanol, liver tissue AST, and ALT levels in mice. ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P < 0.01 indicates a comparison between the sample group and the model group (EtOH);
[0030] Figure 9 H&E staining of liver tissue from mice induced by alcohol (EtOH) using DVM;
[0031] Figure 10 The figure shows the effects of ADH, glutathione (GSH), catalase (CAT), and superoxide dismutase (SOD) levels in mouse liver tissue. ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P<0.01 indicates a comparison between the sample group and the model group (EtOH). Detailed Implementation
[0032] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise specified, the instruments, reagents, and materials involved in the following embodiments are all conventional instruments, reagents, and materials already existing in the prior art and can be obtained through legitimate commercial channels. Unless otherwise specified, the experimental methods and detection methods involved in the following examples are all conventional experimental methods and detection methods already existing in the prior art.
[0033] Example 1: Isolation and purification of gallic acid derivatives
[0034] 1. After air-drying, 5 kg of rose petals from the golden-edged rose (Rosa chinensis cv. 'JinBian') collected from Anning, Yunnan Province were crushed using a pulverizer. The crushed petals were then ultrasonically extracted with a methanol aqueous solution (90%) at a ratio of 1:8 (m / v, g / mL) for 0.5 hours each time, for a total of three extractions. All extracts were collected, the solvent was removed by rotary evaporation, and the extract was freeze-dried to obtain 0.5 kg of crude extract, which was stored at 4℃.
[0035] 2. Dissolve 0.5 kg of crude extract completely in 4 L of pure water, and then extract it four times in sequence with dichloromethane, ethyl acetate and n-butanol at a volume ratio of 1:1 (aqueous phase: organic phase). Collect each phase extract and concentrate it under vacuum in a rotary evaporator to dryness. Finally, dichloromethane phase (46.75 g), ethyl acetate phase (160.38 g), n-butanol phase (132.54 g) and aqueous phase (151.93 g) are obtained.
[0036] 3. The ethyl acetate phase rich in VM was chromatographically analyzed using a medium-pressure column loaded with RP-C18 packing material. Eluents were obtained using methanol-water solutions of different concentrations (10%~100%), with 6 column volumes eluted in each solvent system. Fractions containing the same compounds were combined by thin-layer chromatography to obtain a total of 6 fractions (JF1~JF6).
[0037] 4. Fragment JF2 (20 g) was purified by medium-pressure preparative chromatography using a gradient elution of 10 vol%, 30 vol%, 50 vol%, 70 vol%, and 90 vol% methanol-water solutions. The eluent was collected and rotary evaporated to obtain five subfractions (JF2-Fr.A~E). JF2-Fr. C (8.2 g) was separated by silica gel column chromatography and eluted with chloroform-methanol (7:1) to obtain compounds 1 (1.5 g) and 2 (35 mg). Fragment JF4 (20 g) was purified by medium-pressure preparative chromatography using a gradient elution of 10 vol%, 30 vol%, 50 vol%, 70 vol%, and 90 vol% methanol-water solutions. The eluent was collected and rotary evaporated to obtain five fractions (JF4-Fr.AE). JF4-Fr. A (7 g) was separated by silica gel column chromatography and eluted with chloroform-methanol (10:1) to obtain compounds 5 (10 mg) and 7 (1.1 g). Fragment JF4 (10 g) was separated and purified by medium-pressure preparative chromatography, using a gradient elution of 10 vol%, 30 vol%, 50 vol%, 70 vol%, and 90 vol% methanol-water solutions. The eluent was collected and rotary evaporated to obtain five fractions (JF4-Fr.AE). JF4-Fr. B (2.3 g) was separated by silica gel column chromatography and eluted with chloroform-methanol (15:1, 10:1, and 5:1) to obtain compounds 3 (95 mg), 4 (23 mg), and 6 (13 mg).
[0038] Compound 1 is gallic acid;
[0039] Compound 2 is methyl gallate;
[0040] Compound 3 is ethyl gallate;
[0041] Compound 4 is Urolithin;
[0042] Compound 5 is Corilagin;
[0043] Compound 6 is ellagic acid;
[0044] Compound 7 is VM;
[0045] The relevant mass spectrometry data of compounds 1-7 are as follows: Figure 1 As shown in the figure, A to G correspond to compounds 1 to 7, respectively.
[0046] Example 2: Hepatoprotective effect of gallic acid derivatives in alcohol-induced HepG2 cells
[0047] (1) Cell culture
[0048] The experiment was conducted using the human liver-derived cell line (HepG2, purchased from the Kunming Institute of Zoology, Chinese Academy of Sciences). The cells were cultured in DMEM medium containing 10% fetal bovine serum (FBS) and 1% penicillin / streptomycin (Gibco, Big Island, New York, USA) and incubated at 37°C in a 5 vol% CO2 incubator.
[0049] (2) Cytotoxicity evaluation
[0050] 200 μL of HepG2 cells were seeded into 96-well cell culture plates (2 × 10⁶ cells per well). 4 (1 cell), incubated at 37℃ for 24 h, then 200 μL of different concentrations of compounds 1-7 (20 and 40 μM) were added to 96-well plates. After 24 h of incubation, the culture medium was removed, and 200 μL of PBS-dissolved MTT (0.5 mg / mL) solution was added to the 96-well cell culture plate. The plates were then incubated at 37℃ in a 5% CO2 incubator for another 4 hours. 200 μL of DMSO was added and reacted for 10 min. The absorbance was measured at 490 nm using a microplate reader, and cell viability was calculated. The toxicity of the compounds to HepG2 cells was evaluated by the MTT assay. Cell viability (%) = (A 样品 / A 空白 ) × 100%, where A 样品 The absorbance values are shown in Figure 1. The A blank represents the absorbance value of the experimental group with the added compound, and the B blank represents the absorbance value of the blank control group without the added compound. Results are shown in Figure 2. Figure 2 The results in the figure show that compounds 1-7 did not exhibit toxicity to HepG2 cells at concentrations of 20 and 40 μM. Subsequent experiments were conducted using 20 μM.
[0051] Figure 2 The results show the toxicity test results of compounds 1-7 on HepG2 cells; in the figure, 1-7 correspond to compounds 1-7 respectively.
[0052] (3) Evaluation of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels
[0053] HepG2 cells in the logarithmic growth phase were seeded into 6-well cell culture plates (4 × 10⁶ cells per well). 5 Cells were cultured at 37°C for 24 hours (2 mL per cell), and then compounds 1-7 (20 μM) and silybin (20 μM) were added to the culture medium. The cells were then co-incubated with 800 mM ethanol (EtOH) for another 24 hours. Cells were washed with PBS, and complete culture medium containing 800 mM ethanol was added. AST and ALT accumulation in the cells was measured using an AST and ALT assay kit. Results are shown in [Figure number missing]. Figure 3 Compared to other compounds, VM (compound 7) showed significant inhibitory effects on the accumulation of AST and ALT at 20 μM, and was superior to the positive control drug silymarin, demonstrating good protective activity against alcoholic liver injury.
[0054] Figure 3 The figures show the results of measuring the aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels of compounds 1-7 in alcohol-induced HepG2 cells; in the figure, A represents the AST results and B represents the ALT results. ** P < 0.01 indicates a comparison between the model group (EtOH) and the blank group; ## P<0.01 indicates a comparison between the sample group and the model group (EtOH).
[0055] Example 3: Co-assembly of DVM nanoparticles with VM and dihydromyricetin (DM)
[0056] DM (150 mg), Pluronic F-127 (100, 200, 400, and 600 mg), and VM (50 mg) were dissolved in 40 mL of ethanol and stirred magnetically at 25 °C for 15 min. Then, 24.5 μL of formaldehyde solution (37 vol% formaldehyde aqueous solution) was added to the solution; after 5 min, 3 mL of glycine aqueous solution (10 mg / mL) was added, and the mixture was stirred for 4 h. The solution was purified by dialysis with water (MWCO3500) for 48 h and then lyophilized to obtain the sample (DVM). The structure of DVM was characterized as follows. Figure 4 As shown.
[0057] (1) Transmission electron microscopy image of DVM particles ( Figure 4 A) and particle size distribution ( Figure 4(B) TEM images show the morphology and corresponding particle size distribution of the DVM after the proportion of F127 in the DVM increases. Scale bar: 50 nm (n=3). The particles exhibit a uniform spherical morphology, with a smooth and integrated surface, and the size increases from 32 nm to 45 nm.
[0058] (2) Dynamic light scattering (DLS) Figure 4 DLS (Density Spectroscopy) is a technique used to measure the size and distribution of particles in a suspension or solution. Figure 4 The results of B) are consistent.
[0059] (3) X-ray diffraction (XRD) Figure 4 XRD (di-molecular dynamics) is a technique that utilizes the interaction of X-rays with crystalline materials to analyze their crystal structure and phase composition. XRD measures the intermolecular distances within the di-molecular dynamics membrane (DMVM) and reveals the crystal peak structures of the DM and VM. The DVM primarily produces characteristic crystal peaks at 19.1° and 22.3°.
[0060] (4) Zeta potential ( Figure 4 The zeta potential (E) in DVM is a key parameter characterizing the stability of colloidal dispersions, reflecting the potential difference between the particle surface and the surrounding liquid, and affecting particle interaction and stability. The results show that the zeta potential of DVM increases slightly with increasing F127 proportion, but there is no significant difference between DVM 200 and 600.
[0061] (5) Fourier transform infrared (FT-IR) spectroscopy Figure 4 The F in the figure is an infrared spectroscopy analysis method based on Fourier transform mathematical methods and computer technology, which can identify the functional groups of DVM nanoparticles. Fourier transform infrared (FT-IR) spectroscopy confirmed that DVM contains a para-substituted benzene ring (842 cm⁻¹). -1 ), carbonyl (1735 cm) -1 ), oxygen-hydrogen bond (3431 cm) -1 Spectral absorptions of functional groups derived from DM and VM compounds, etc.
[0062] F127:DM+VM with a ratio of 1:1 (m:m) has relatively small particle size and a large zeta potential, which allows it to exist more stably in liquids, making it a sample for subsequent research.
[0063] Figure 4 The figures show the structural characterization results of DVM nanoparticles; in the figure, A is the transmission electron microscopy image, B is the particle size distribution, C is the dynamic light scattering, D is the X-ray diffraction, E is the Zeta potential, and F is the Fourier transform infrared spectroscopy.
[0064] Example 4: Hepatoprotective effect of DVM nanoparticles in alcohol-induced HepG2 cells
[0065] (1) Cell culture
[0066] The experiment was conducted using the human liver-derived cell line (HepG2, purchased from Kunming Institute of Zoology, Chinese Academy of Sciences). The cells were cultured in DMEM medium containing 10 vol% fetal bovine serum (FBS) and 1 vol% penicillin / streptomycin (Gibco, Big Island, New York, USA) and incubated at 37°C in a 5 vol% CO2 incubator.
[0067] (2) Cytotoxicity evaluation
[0068] 200 μL of HepG2 cells were seeded into 96-well cell culture plates (2 × 10⁶ cells per well). 4 Cells were incubated at 37°C for 24 h, and then 200 μL of different concentrations of DVM (5~40 μg / mL) were added to 96-well plates. After 24 h of incubation, the culture medium was removed, and 200 μL of MTT (0.5 mg / mL) solution dissolved in PBS was added to the 96-well cell culture plates. The plates were then incubated at 37°C and 5 vol% CO2 for another 4 hours. 200 μL of DMSO was added and reacted for 10 min. The absorbance was measured at 490 nm using a microplate reader, and cell viability was calculated. The toxicity of DVM to HepG2 cells was evaluated by the MTT assay. Cell viability (%) = (A... 样品 / A 空白 ) × 100%, where A 样品 The absorbance values are shown for the experimental group with added DVM, and the A blank is the absorbance value of the blank control group without added DVM; the results are shown in […]. Figure 5 The results in the figure show that DVM exhibits toxicity to HepG2 cells within the concentration range of 5–40 μg / mL, with concentrations of 40 and 30 μg / mL. Therefore, a concentration of 20 μg / mL was subsequently selected for subsequent experiments.
[0069] (3) Effects of DVM on intracellular damage biomarkers in HepG2 cells
[0070] HepG2 cells in the logarithmic growth phase were seeded into 12-well cell culture plates (2 × 10⁶ cells per well). 5 Cells were cultured at 37°C for 24 hours (1 mL), followed by the addition of DVM (10 and 20 μg / mL) and silybin (10 μg / mL) to the culture medium, and co-incubated with 800 mM ethanol (EtOH) for another 24 hours. Cells were then washed twice with PBS and digested with trypsin without EDTA. The degree of apoptosis and ROS production were measured by flow cytometry. Results are shown below. Figure 6 As shown in A~C. After undergoing the same cell manipulation procedures described above, HepG2 cells were digested with trypsin without EDTA, sonicated, and the cell suspension was used for further studies to determine acetaldehyde dehydrogenase (ADH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT). Results are shown in [Figure Number]. Figure 6 DVM significantly inhibited the accumulation of AST and ALT at 20 μg / mL and significantly increased ADH activity, showing superiority over the positive control drug silymarin.
[0071] (1) Apoptosis analysis: Compared with the blank control group, alcohol-induced apoptosis of HepG2 cells was significantly increased (P < 0.01). Compared with the model group, DVM at 10 and 20 μg / mL significantly reduced cell apoptosis (P < 0.01), see [link to relevant data]. Figure 6 A and B in the example.
[0072] (2) Intracellular ROS generation: Compared with the blank control group, alcohol-induced ROS generation in HepG2 cells was significantly increased (P < 0.01). Compared with the model group, DVM at 10 and 20 μg / mL significantly reduced ROS generation in cells (P < 0.01), see [link to relevant documentation]. Figure 6 C in the middle.
[0073] (3) Acetaldehyde dehydrogenase (ADH) analysis: Compared with the blank control group, the ADH activity of alcohol-induced HepG2 cells was significantly reduced (P<0.01). Compared with the model group, DVM at 10 and 20 μg / mL significantly increased ADH activity (P < 0.01), see [link to relevant documentation]. Figure 6 D in the middle.
[0074] (4) Aspartate aminotransferase (AST) analysis: Compared with the blank control group, the AST activity of alcohol-induced HepG2 cells was significantly increased (P < 0.01). Compared with the model group, DVM at 10 and 20 μg / mL significantly reduced AST activity (P < 0.01), see [link to relevant data]. Figure 6 E in the middle.
[0075] (5) Alanine aminotransferase (ALT) analysis: Compared with the blank control group, the ALT activity of alcohol-induced HepG2 cells was significantly increased (P < 0.01). Compared with the model group, DVM at 10 and 20 μg / mL significantly reduced ALT activity (P < 0.01), see [link to relevant documentation]. Figure 6 F in the middle.
[0076] Example 5: Hepatoprotective and hangover-relieving activities of DVM nanoparticles
[0077] (1) Animal grouping and treatment: Forty mice that had been fasted for 12 hours but not restricted to water were randomly divided into 5 groups: blank group, model group, silybin group, and the low-dose (DVL) and high-dose (DVH) groups of DVM nanoparticles used in the examples, with 8 mice in each group. Each experimental group was administered the corresponding test substance by gavage for 1 week. During the experimental period (1 week), except for the first day when the mice were given 30 vol% alcohol, they were administered 52 vol% Niulanshan strong-aroma baijiu by gavage every day at a dose of 5 mL / kg. The positive control was silybin (10 mg / kg), the sample low-dose (DVL, 10 mg / kg), and the sample high-dose (DVH, 20 mg / kg). The positive control and the sample were injected into the mice intraperitoneally, and alcohol was administered by gavage half an hour later.
[0078] (2) Erguotou-induced alcohol intoxication experiment in mice: Each experimental group was continuously administered the corresponding test substance by gavage for 1 week. The time for balance test, whisker response test, tail suspension test, and tilt plate test was recorded. The results are shown in […]. Figure 7 .
[0079] (3) Balance test: Compared with the blank control group, the time spent on the rotarod in the model group was significantly shorter (P<0.01). Compared with the model group, the time spent on the rotarod in the DVM group was significantly longer (P<0.01), and the number of rotarod drops in groups 3-5 of Examples all showed a significant regression (P<0.01, see Example 3). Figure 7 (A in the middle).
[0080] (4) Tentacle response: Compared with the blank control group, the reaction time of the model group after being touched with a cotton swab was significantly increased (P<0.01). Compared with the model group, the reaction time of the DVM group mice after being touched with a cotton swab was significantly decreased (P<0.01, see...). Figure 7 (B in the middle).
[0081] (5) Tail suspension test: Compared with the blank control group, the time to start struggling in the tail suspension state was significantly increased in the model group (P < 0.01). Compared with the model group, the reaction time to start struggling in the DVM group was significantly decreased (P < 0.01, see Figure 7 (C in the middle).
[0082] (6) Inclined board test: Compared with the blank control group, the model group had a significantly shorter time remaining on a 30° inclined cardboard and slid off the cardboard quickly (P < 0.01). Compared with the model group, the DVM group mice were able to stay stably on the cardboard at a higher position and had a significantly longer time on the cardboard (P < 0.01, see...). Figure 7 (D in the middle).
[0083] (7) Effects on serum ethanol concentration, ALT and AST activity in mice: Compared with the normal group, the model group showed significantly increased serum ethanol concentration, ALT and AST activity (P < 0.01); compared with the model group, the DVL and DVH groups and the silymarin group significantly reduced serum ethanol concentration, ALT and AST activity in intoxicated mice (P < 0.01), with the DVH group showing a more significant reduction in serum ethanol concentration, ALT and AST activity. See results below. Figure 8 .
[0084] (8) Effects on mouse liver tissue: In the alcohol-induced acute liver injury model, the hepatocyte structure of mice was disordered, with obvious cell swelling, necrosis, and vacuolar degeneration. The hepatocyte structure in the control group was normal. DVM alleviated liver injury and reduced structural disorder. The results are shown in the figure. Figure 9 .
[0085] (9) Effects on ADH, GSH, CAT, and SOD levels in mouse liver tissue: Compared with the normal group, the activities of ADH, GSH, CAT, and SOD in the model group were significantly decreased (P < 0.01); compared with the model group, both the DVL and DVH groups significantly increased the levels of ADH, GSH, CAT, and SOD in the liver tissue of intoxicated mice (P < 0.01), with the DVH group showing a more significant effect. See results below. Figure 10 .
[0086] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.
Claims
1. A DVM nanoparticle for treating or preventing alcoholic liver disease, characterized in that, The DVM nanoparticles contain gallic acid derivative VM and dihydromyricetin; the mass ratio of gallic acid derivative VM to dihydromyricetin is 1:3; the gallic acid derivative VM has the structural formula described in Formula I; Formula I.
2. The DVM nanoparticles according to claim 1, characterized in that, The DVM nanoparticles also contain Pluronic F-127, and the mass ratio of the gallic acid derivative VM, dihydromyricetin and Pluronic F-127 is 1:3:
4.
3. A method for preparing DVM nanoparticles as described in any one of claims 1 to 2, characterized in that, Includes the following steps: Step 1: Weigh gallic acid derivative, dihydromyricetin and Pluronic F-127 according to the mass ratio, then dissolve them in ethanol solution and stir well. Step 2: Add a 37 vol% formaldehyde aqueous solution to the product obtained in Step 1 and continue stirring. After 3-6 min, add a 10 mg / mL glycine aqueous solution and continue stirring for 2-6 h. Step 3: After dialysis purification of the product from Step 2 with MWCO 3500 water for 46-50 hours, the product is freeze-dried to obtain the DVM nanoparticles.
4. The use of DVM nanoparticles as described in any one of claims 1 to 2 in the preparation of products for the treatment and / or prevention of alcoholic liver disease.
5. The application according to claim 4, characterized in that, The product described is a medication for the treatment and / or prevention of alcoholic liver disease.
6. A medicament for treating and / or preventing alcoholic liver disease, characterized in that, The active ingredient of the drug is the DVM nanoparticle as described in any one of claims 1 to 2.
7. The drug according to claim 6, characterized in that, The dosage form of the drug is one of the following: liquid, tablet, capsule, powder, or pill.