Application of composition containing osmanthus fragrans extract in preparation of product for improving liver function and relieving liver injury

Abstract

The invention provides an application of a composition containing a sweet osmanthus extract in preparation of a product for improving liver functions and relieving liver injury. The preparation method comprises the following specific steps: (1) naturally drying sweet osmanthus in the shade, crushing, sieving, soaking in ultrapure water, carrying out suction filtration, and collecting a sweet osmanthus extracting solution; and (2) carrying out rotary evaporation on the osmanthus fragrans extracting solution in the step (1) until yellow syrup-shaped liquid is formed, so as to form an osmanthus fragrans concentrated solution, freezing, and carrying out vacuum drying, so as to obtain the osmanthus fragrans extract. According to the composition containing the osmanthus fragrans extract, ROS and MDA indexes in liver cells damaged by DBP can be reduced, reduction of GSH indexes is relieved, meanwhile, inflammatory indexes NF-kappa B, TNF-alpha and IL-6 are reduced, it is indicated that the composition containing the osmanthus fragrans extract has the capacity of inhibiting oxidative stress and inflammation in the damaged liver cells, and the composition containing the osmanthus fragrans extract has the advantages of being capable of being used for preventing and treating liver injury. Liver function impairment caused by DBP can be remarkably relieved, the application prospect is good, and practical significance is achieved.

Description

Technical Field

[0001] The invention relates to the technical field of plant extraction, and in particular to application of a composition containing osmanthus fragrans extract in preparing a product for improving liver function and relieving liver damage. Background Art

[0002] The liver is the main organ for absorption, distribution, metabolism and elimination in the human body, and plays a key role in sugar metabolism, protein metabolism, lipid metabolism, etc. Factors such as viral infection, bacterial infection, alcohol, drug-induced liver injury, liver ischemia-reperfusion, and pathogenic liver injury will stimulate the body to produce too many highly reactive oxygen free radicals, induce oxidative stress, trigger inflammatory reactions, and then lead to liver cell apoptosis, hepatitis, liver fibrosis, liver injury and other lesions, endangering human health. At present, the treatment of liver injury in clinical practice is mainly based on the removal of the cause, and there are also a large number of drugs for the treatment of liver injury, mainly including Chinese herbal medicines, biological drugs, chemical drugs and trace elements. At present, Chinese medicines and Chinese medicine ingredients such as Flammulina velutipes polysaccharides have a certain protective effect on stress-induced liver injury induced by carbon tetrachloride. It can be seen that Chinese medicines and Chinese medicine active ingredients can be used to protect acute liver injury, which has important practical significance. However, some studies have shown that some anti-liver injury drugs will inevitably produce biological toxicity. It can be seen that the prevention and treatment of liver injury is still a serious issue at present, and the search for effective liver protection substances has always been a hot topic of concern for medical workers. Herbal medicines have the characteristics of multi-link, multi-level and multi-target comprehensive effects, and have unique advantages in the treatment of liver diseases. However, the different ingredients and dosages of herbal medicines have led to very different therapeutic effects, and the prevention and treatment results are not satisfactory. Summary of the invention

[0003] In view of this, the present invention proposes the use of a composition containing osmanthus extract in preparing a product for improving liver function and alleviating liver damage.

[0004] The technical solution of the present invention is achieved in this way:

[0005] A composition containing osmanthus extract is used in the preparation of a product for improving liver function and alleviating liver damage, wherein the osmanthus extract is an aqueous extract of osmanthus, and the preparation method is specifically as follows:

[0006] (1) drying osmanthus naturally in the shade, crushing and sieving, soaking in ultrapure water, filtering, and collecting osmanthus extract;

[0007] (2) subjecting the osmanthus extract in step (1) to rotary evaporation until it is evaporated into a yellow syrupy liquid to form an osmanthus concentrate, freezing, and vacuum drying to obtain a yellow solid, i.e., an osmanthus extract.

[0008] Furthermore, in the preparation method of osmanthus extract, in step (1), the sieving is through a 150-200 mesh sieve; the material-liquid ratio of osmanthus to ultrapure water is 1:8-12 g / mL; and the soaking time is 12-24 h;

[0009] In step (2), the rotary evaporation is carried out at 65-75°C and 20-50rpm for 4-6 hours; the freezing is carried out at -25 to -15°C for 10-12 hours; and the vacuum drying is carried out at 15-25Pa and -60 to -50°C for 8-12 hours.

[0010] Furthermore, the concentration of the osmanthus extract in the product is 2-200 μg / mL.

[0011] Furthermore, the concentration of osmanthus extract in the product is 200 μg / mL.

[0012] Furthermore, the composition also includes a long-stem grape fern extract.

[0013] Furthermore, the mass ratio of the long-stem grape fern extract to the osmanthus extract is 0.1-0.3:1;

[0014] The preparation method of the long-stem grape fern extract comprises:

[0015] (1) subjecting the long-stem grape fern algae to freeze-thaw treatment, freezing it below 0°C for 1-2 days, thawing it naturally to room temperature, repeating this process 2-3 times, crushing it, adding 70%-80% v / v ethanol solution to extract it twice, each time for 30-60 minutes, and collecting the alcohol extract;

[0016] (2) adding the filter residue to the composite enzymatic hydrolysate, extracting at 25-30° C. and 20-30 kHz ultrasonic wave for 20-30 min, combining with the alcohol extract, concentrating under reduced pressure, and then adding glucan and stirring to obtain the long-stem grape fern extract.

[0017] Furthermore, in step (2), the composite enzymatic hydrolysate is obtained by mixing β-glucosidase, cellulase, xylanase, pectinase and water in a mass ratio of 1:2-4:0.4-0.6:1-3:200-300; the solid-liquid ratio of the filter residue to the composite enzymatic hydrolysate is 1:8-12 g / mL; and the mass ratio of the glucan to the filter residue is 0.05-0.08:1.

[0018] Furthermore, the product is a lozenge.

[0019] Furthermore, each 1.0 g of the lozenge contains 0.5-0.7 g of a composition containing osmanthus extract, 0.006-0.008 g of lily powder, 0.005-0.007 g of clove powder, 0.001-0.002 g of peppermint extract, 0.003-0.005 g of corn peptide powder, 0.05-0.10 g of pumpkin powder, 0.05-0.10 g of xylitol, 0.001-0.003 g of stevioside, 0.01-0.03 g of citric acid, 0.001-0.003 g of menthol, 0.001-0.002 g of edible salt, 0.006-0.008 g of magnesium stearate and the remainder of sorbitol.

[0020] Furthermore, the preparation method of the buccal tablet is specifically as follows: a composition containing osmanthus extract, lily powder, clove powder, mint extract, corn peptide powder, pumpkin powder, xylitol, stevioside, citric acid, menthol, edible salt, and sorbitol are uniformly mixed, 50-60% v / v ethanol is added to prepare a soft material, wet granules are prepared through a 20-30 mesh sieve, dried under reduced pressure, granulated through a 16-20 mesh sieve, magnesium stearate is added, mixed, and then tableted to obtain the target buccal tablet.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] The composition containing osmanthus extract of the present invention can significantly improve liver function and alleviate liver damage. According to the test examples, the composition containing osmanthus extract of the present invention can reduce ROS and MDA indexes in liver cells damaged by DBP, alleviate the decline of GSH index, and at the same time, the inflammatory indexes NF-κB, TNF-α and IL-6 all decrease, indicating that the composition containing osmanthus extract of the present invention has the ability to inhibit oxidative stress and inflammation in damaged liver cells, and can significantly alleviate liver function damage caused by DBP. DETAILED DESCRIPTION

[0023] In order to better understand the technical content of the present invention, specific embodiments are provided below to further illustrate the present invention.

[0024] Unless otherwise specified, the experimental methods used in the embodiments of the present invention are all conventional methods.

[0025] Unless otherwise specified, the materials, reagents, etc. used in the embodiments of the present invention can be obtained from commercial sources.

[0026] Example 1

[0027] The preparation method of osmanthus extract comprises:

[0028] (1) Dry the osmanthus naturally in the shade, crush it and pass it through a 200-mesh sieve, add it into ultrapure water at a solid-liquid ratio of 1:10 g / mL, soak it for 12 hours, filter it with suction, and collect the osmanthus extract;

[0029] (2) The osmanthus extract in step (1) is subjected to rotary evaporation at 70° C. and 30 rpm for 5 h, until it evaporates to a yellow syrupy liquid to form an osmanthus concentrate, which is then frozen at -20° C. for 12 h, and vacuum dried at 20 Pa and -57° C. for 12 h to obtain a yellow solid, i.e., an osmanthus extract.

[0030] Example 2

[0031] The preparation method of osmanthus extract comprises:

[0032] (1) Dry the osmanthus naturally in the shade, crush it and pass it through a 200-mesh sieve, add it into ultrapure water at a solid-liquid ratio of 1:8 g / mL, soak it for 24 hours, filter it with suction, and collect the osmanthus extract;

[0033] (2) The osmanthus extract in step (1) is subjected to rotary evaporation at 65° C. and 20 rpm for 4 h, until it evaporates to a yellow syrupy liquid to form an osmanthus concentrate, which is then frozen at -25° C. for 10 h, and vacuum dried at 15 Pa and -60° C. for 8 h to obtain a yellow solid, i.e., an osmanthus extract.

[0034] Example 3

[0035] The preparation method of osmanthus extract comprises:

[0036] (1) Dry the osmanthus naturally in the shade, crush it and pass it through a 200-mesh sieve, add it into ultrapure water at a solid-liquid ratio of 1:12 g / mL, soak it for 24 hours, filter it with suction, and collect the osmanthus extract;

[0037] (2) The osmanthus extract in step (1) is subjected to rotary evaporation at 75° C. and 50 rpm for 6 h, until it evaporates to a yellow syrupy liquid to form an osmanthus concentrate, which is then frozen at -15° C. for 12 h, and vacuum dried at 25 Pa and -50° C. for 12 h to obtain a yellow solid, i.e., an osmanthus extract.

[0038] Example 4

[0039] The composition containing osmanthus extract comprises osmanthus extract and long-stem grape fern extract in a mass ratio of 1:0.2.

[0040] The preparation method of osmanthus extract comprises:

[0041] (1) Dry the osmanthus naturally in the shade, crush it and pass it through a 200-mesh sieve, add it into ultrapure water at a solid-liquid ratio of 1:10 g / mL, soak it for 12 hours, filter it with suction, and collect the osmanthus extract;

[0042] (2) The osmanthus extract in step (1) is subjected to rotary evaporation at 70° C. and 30 rpm for 5 h, until it evaporates to a yellow syrupy liquid to form an osmanthus concentrate, which is then frozen at -20° C. for 12 h, and vacuum dried at 20 Pa and -57° C. for 12 h to obtain a yellow solid, i.e., an osmanthus extract.

[0043] The preparation method of the long-stem grape fern extract comprises:

[0044] (1) The long-stem grape fern algae was subjected to freeze-thaw treatment, frozen below 0°C for 1 day, and thawed naturally to room temperature, repeated 3 times, crushed, and extracted twice with 75% v / v ethanol solution, each time for 45 min, and the ethanol extract was collected;

[0045] (2) A composite enzymatic solution is obtained by mixing β-glucosidase, cellulase, xylanase, pectinase and water in a mass ratio of 1:3:0.5:2:250, and the filter residue is added to the composite enzymatic solution at a solid-liquid ratio of 1:10 g / mL. The mixture is extracted with an ultrasonic wave at 20 kHz for 25 min at a temperature of 30°C, combined with the alcohol extract, and concentrated under reduced pressure. Then, glucan is added at a mass ratio of glucan to filter residue of 0.07:1, and the mixture is stirred to obtain a long-stem grape fern extract.

[0046] Test Example 1

[0047] The osmanthus extract prepared in Example 1 was tested for its physiological activity in improving liver function and alleviating liver tissue damage.

[0048] 1. Experimental methods: (a) Cell culture: HepG2 cells were inoculated in complete culture medium (containing 89% high glucose medium + 10% fetal bovine serum + 1% penicillin-streptomycin) and cultured at 37°C and 5% CO2 saturated humidity.

[0049] (b) Preparation of liver organ chip: 99 μL cells (1×10 6 Cells / mL) were mixed with 20 μL of 15 mg / mL fibrinogen (F8630, Sigma) and 1 μL of 100 U / mL thrombin (T7513, Sigma) in preheated DPBS (C14190500BT, Gibco), and the mixture was then added to the middle channel of the microfluidic chip and incubated for 40 min for gelation.

[0050] (c) Drug induction: The experimental groups were administered with high-sugar culture medium containing different concentrations of the osmanthus extract of Example 1 (low-dose osmanthus extract 2 μg / mL, medium-dose osmanthus extract 20 μg / mL, and high-dose osmanthus extract 200 μg / mL) for 24 h, and the positive group was treated with high-sugar culture medium containing vitamin C (VitC: 50 μg / mL) for 24 h for pre-protection, and then modeled with dibutyl phthalate (DBP: 50 μg / ml).

[0051] (d) Grouping: blank group, model group (DBP: 50 μg / mL), low-dose group (DBP: 50 μg / mL + osmanthus extract 2 μg / mL), medium-dose group (DBP: 50 μg / mL + osmanthus extract 20 μg / mL), high-dose group (DBP: 50 μg / mL + osmanthus extract 200 μg / mL) and positive group (DBP: 50 μg / mL + VitC: 50 μg / mL).

[0052] 2. Determination of oxidative stress and inflammatory markers

[0053] The reactive oxygen species (ROS) assay kit (BL714A) of Biosharp, the malondialdehyde (MDA) test kit (A003-1) of Nanjing Jiancheng Bioengineering Institute, the trace reduced glutathione (GSH) assay kit (A006-2-1) and the ELISA kit of Shanghai ELISA Biotechnology Co., Ltd. were used to measure the inflammatory indexes, including human tumor necrosis factor-α (TNF-α) (YJ077385), human interleukin-6 (IL-6) (YJ058097), and human nuclear factor-κB (NF-κB) (YJ026330). The assay operation was carried out strictly in accordance with the instructions of the kit. The results are shown in Table 1.

[0054] Table 1

[0055]

[0056] As can be seen from Table 1, different concentrations of osmanthus extracts (2-200 μg / mL) in Example 1 of the present invention can reduce ROS and MDA indicators in liver cells damaged by DBP, alleviate the decline of GSH indicators, and at the same time, the inflammatory indicators NF-κB, TNF-α and IL-6 all decrease. When the concentration of osmanthus extract is 200 μg / mL, the effect is most obvious, indicating that it has the ability to inhibit oxidative stress and inflammation in damaged liver cells, and is better than the positive drug vitamin C.

[0057] 3. Determination of the effects on functional indicators of damaged liver tissue

[0058] The results are shown in Table 2 using the alanine aminotransferase (ALT / GPT) test kit (C009-2-1), aspartate aminotransferase (AST / GOT) test kit (C010-2-1), total bile acid (TBA) test kit (E003-2-1), lactate dehydrogenase (LDH) test kit (A020-2) and the ELISA human albumin (ALB) test kit (YJ038466) of Shanghai ELISA Biotechnology Co., Ltd. in strict accordance with the instructions for use of the kit.

[0059] Table 2

[0060]

[0061]

[0062] As can be seen from Table 2, different concentrations of osmanthus extract (2-200 μg / mL) in Example 1 of the present invention can alleviate the liver function damage caused by DBP to varying degrees, especially when the concentration of osmanthus extract is 200 μg / mL, it can significantly alleviate the liver function damage caused by DBP, and can be closest to the indicators of the blank group.

[0063] Comparative Example 1

[0064] The difference from Example 4 is that the mass ratio of the long-stem grape fern extract and the osmanthus extract is 1:0.2, and the rest is consistent with Example 4.

[0065] Comparative Example 2

[0066] The osmanthus extract obtained by patent CN102085223A is used.

[0067] Test Example 2

[0068] The substances obtained in Example 4 and Comparative Examples 1-2 were compared in terms of improving liver function and alleviating liver damage. The specific methods were the same as those in Experimental Example 1 (the concentration of osmanthus extract was 200 μg / mL). The results are shown in Tables 3-4.

[0069] Table 3

[0070]

[0071]

[0072] Table 4

[0073] sample ALB ALT AST LDH TBA Blank Group 154.11±0.84 1.09±0.039 0.15±0.12 0.49±0.02 3.70±1.26 Model Group 124.81±11.21 1.93±0.036 0.84±0.08 1.10±0.02 5.95±0.16 Example 4 153.22±12.45 1.10±0.025 0.19±0.07 0.65±0.05 3.72±1.28 Comparative Example 1 138.14±14.51 1.58±0.078 0.55±0.09 0.94±0.06 4.57±1.34 Comparative Example 2 142.16±13.87 1.48±0.067 0.48±0.08 0.86±0.05 4.24±1.36

[0074] As can be seen from Table 3, compared with Example 4, Comparative Example 1-2 is not as good as Example 4 in reducing ROS and MDA indicators in liver cells damaged by DBP, and is not as good as Example 4 in alleviating the decline of GSH indicators, while the decline of inflammatory indicators NF-κB, TNF-α and IL-6 is not as good as Example 4. As can be seen from Table 4, compared with Example 4, Comparative Example 1-2 is not as good as Example 4 in alleviating liver function damage caused by DBP.

[0075] Application Examples

[0076] A lozenge, each 1.0 g of which contains 0.6 g of a composition containing osmanthus extract, 0.007 g of lily powder, 0.006 g of clove powder, 0.002 g of mint extract, 0.004 g of corn peptide powder, 0.10 g of pumpkin powder, 0.10 g of xylitol, 0.002 g of stevioside, 0.02 g of citric acid, 0.002 g of menthol, 0.002 g of edible salt, 0.007 g of magnesium stearate and the remainder of sorbitol.

[0077] The preparation method of the above-mentioned buccal tablets is specifically as follows: a composition containing osmanthus extract, lily powder, clove powder, mint extract, corn peptide powder, pumpkin powder, xylitol, stevioside, citric acid, menthol, edible salt, and sorbitol are mixed evenly, 50% v / v ethanol is added to make a soft material, wet granules are made through a 20-mesh sieve, dried under reduced pressure, granulated through a 16-mesh sieve, magnesium stearate is added, mixed evenly, and then tableted to obtain the target buccal tablets.

[0078] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. Use of a composition containing osmanthus extract in preparing a product for improving liver function and alleviating liver damage, characterized in that: Osmanthus fragrans extract is used as an effective ingredient, and the osmanthus fragrans extract is a water extract of osmanthus fragrans. The specific preparation method is as follows: (1) drying osmanthus naturally in the shade, crushing and sieving, soaking in ultrapure water, filtering, and collecting osmanthus extract; (2) subjecting the osmanthus extract in step (1) to rotary evaporation until it is evaporated into a yellow syrupy liquid to form an osmanthus concentrate, freezing, and vacuum drying to obtain a yellow solid, i.e., an osmanthus extract.

2. The use according to claim 1, characterized in that In the step (1) of the preparation method of osmanthus extract, the sieving is through a 150-200 mesh sieve; the material-liquid ratio of the osmanthus to ultrapure water is 1:8-12 g / mL; the soaking time is 12-24 h; In step (2), the rotary evaporation is carried out at 65-75°C and 20-50rpm for 4-6 hours; the freezing is carried out at -25 to -15°C for 10-12 hours; and the vacuum drying is carried out at 15-25Pa and -60 to -50°C for 8-12 hours.

3. The use according to claim 1, characterized in that: The concentration of the osmanthus extract in the product is 2-200 μg / mL.

4. The use according to claim 3, characterized in that: The concentration of the osmanthus extract in the product is 200 μg / mL.

5. The use according to claim 1, characterized in that: The composition also includes an extract of Pteris longifolia.

6. The use according to claim 5, characterized in that The mass ratio of the long-stem grape fern extract to the osmanthus extract is 0.1-0.3:1; The preparation method of the long-stem grape fern extract comprises: (1) subjecting the long-stem grape fern algae to freeze-thaw treatment, freezing it below 0°C for 1-2 days, thawing it naturally to room temperature, repeating this process 2-3 times, crushing it, adding 70%-80% v / v ethanol solution to extract it twice, each time for 30-60 minutes, and collecting the alcohol extract; (2) adding the filter residue to the composite enzymatic hydrolysate, extracting at 25-30° C. and 20-30 kHz ultrasonic wave for 20-30 min, combining with the alcohol extract, concentrating under reduced pressure, and then adding glucan and stirring to obtain the long-stem grape fern extract.

7. The use according to claim 6, characterized in that In step (2), the composite enzymatic hydrolysis solution is obtained by mixing β-glucosidase, cellulase, xylanase, pectinase and water in a mass ratio of 1:2-4:0.4-0.6:1-3:200-300; the solid-liquid ratio of the filter residue to the composite enzymatic hydrolysis solution is 1:8-12 g / mL; and the mass ratio of the glucan to the filter residue is 0.05-0.08:

1.

8. The use according to claim 1, characterized in that The product is a buccal tablet.

9. The use according to claim 8, characterized in that Each 1.0 g of the lozenge contains 0.5-0.7 g of a composition containing osmanthus extract, 0.006-0.008 g of lily powder, 0.005-0.007 g of clove powder, 0.001-0.002 g of mint extract, 0.003-0.005 g of corn peptide powder, 0.05-0.10 g of pumpkin powder, 0.05-0.10 g of xylitol, 0.001-0.003 g of stevioside, 0.01-0.03 g of citric acid, 0.001-0.003 g of menthol, 0.001-0.002 g of edible salt, 0.006-0.008 g of magnesium stearate and the remainder of sorbitol.

10. The use according to claim 9, characterized in that: The preparation method of the buccal tablet is specifically as follows: uniformly mixing a composition containing osmanthus extract, lily powder, clove powder, mint extract, corn peptide powder, pumpkin powder, xylitol, stevioside, citric acid, menthol, edible salt and sorbitol, adding 50-60% v / v ethanol to prepare a soft material, sieving through a 20-30 mesh sieve to prepare wet granules, drying under reduced pressure, sieving through a 16-20 mesh sieve to size granules, adding magnesium stearate to mix well and then tableting to obtain the target buccal tablet.

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