阅读说明：本技术 一种平卧菊三七提取物、防治急性肾损伤制剂及其制备和使用方法 (Gynura procumbens extract, preparation for preventing and treating acute kidney injury and preparation and using methods thereof ) 是由 吴东 龚建平 黄文平 陈新滔 李梦楚 林俊 温晓斌 于 2021-06-29 设计创作，主要内容包括：本发明涉及医药技术,尤其涉及一种平卧菊三七提取物、防治急性肾损伤制剂及其制备和使用方法。所述提取物及制剂包含β-谷甾醇-3-O-龙胆二糖苷、3-吲哚甲酸、毒豆甲酮及正三十二烷醇,具体制备方法为：取平卧菊三七取茎或叶进行提取后真空浓缩和喷雾干燥制得成品。所述提取物及制剂在降低大鼠血尿素水平及降低大鼠肌酐水平两项实验结果为阳性,提示平卧菊三七醇提物及其制剂在治疗急性肾损伤方面具有一定的作用,开发了平卧菊三七的新用途,并且在降低大鼠血尿素水平及降低肌酐水平达到保护肾脏作用,且没有明显的毒副作用；采用提纯方法对不利于直接食用和使用的茎等部位进行处理,使其同样可以作为原料,拓展了原料选用范围。(The invention relates to a pharmaceutical technology, in particular to a gynura procumbens extract, a preparation for preventing and treating acute kidney injury and a preparation and a using method thereof. The extract and the preparation comprise beta-sitosterol-3-O-gentiobioside, 3-indolecarboxylic acid, coumarone and n-dotriacontanol, and the preparation method comprises the following steps: extracting stems or leaves of Gynura procumbens, vacuum concentrating, and spray drying to obtain the final product. The extract and the preparation have positive test results in reducing the blood urea level of rats and reducing the creatinine level of rats, so that the gynura procumbens alcohol extract and the preparation thereof have certain effects in treating acute kidney injury, develop new application of gynura procumbens, achieve the effect of protecting the kidney in reducing the blood urea level of rats and reducing the creatinine level, and have no obvious toxic or side effect; the stem and other parts which are not beneficial to direct eating and use are treated by adopting a purification method, so that the stem and other parts can also be used as raw materials, and the selection range of the raw materials is expanded.)

1. A preparation for preventing and treating acute kidney injury, which is characterized by comprising a compound 1, a compound 2, a compound 3 and n-dotriacontanol;

the compound 1 is

The compound 2 is

Said R2 includes-COCH 3;

the compound 3 is

2. An extract of gynura procumbens comprising compound 4, compound 5, compound 6 and n-dotriacontanol;

the compound 4 is

The compound 5 is

The compound 6 is

3. A preparation method of Gynura procumbens extract is characterized by comprising the following steps:

(1) taking a proper gynura procumbens sample, cleaning, removing impurities, drying, crushing, adding 8-10 times of 70% v/v ethanol, and soaking for 10-60min to obtain a first mixture;

(2) extracting the first mixture for 1-10 times at 80-100 ℃ under reflux, wherein each time of extraction lasts for 0.5-3h, and filtering the extracted first mixture through 100-mesh filter cloth to obtain a first solution;

(3) concentrating the first solution under vacuum to a relative density of 1.05-1.10 to obtain a second solution;

(4) and (3) spray drying the solution II at the temperature of feed liquid materials of 30-40 ℃, the temperature of inlet air of 105-110 ℃, the air compression ratio of 80-150 and the feed liquid speed of 10-60ml/min to obtain a final product.

4. The method according to claim 3, wherein the stems or leaves of Gynura procumbens are extracted.

5. A method for preparing a preparation for preventing and treating acute kidney injury, which comprises the step of preparing the extract according to claim 3 or 4.

6. A method for using an agent for preventing and treating acute kidney injury, which comprises using the agent of claim 1 or the agent produced by the method of claim 5.

7. The method of use of claim 6, wherein the renal injury is acute renal injury caused by an external infection.

8. The method of use of claim 6, wherein the renal injury is a treatment-inappropriate renal injury.

9. The method of use of claim 6, wherein the renal injury is a drug side effect induced renal injury.

Technical Field

The invention relates to a pharmaceutical technology, in particular to a gynura procumbens extract, a preparation for preventing and treating acute kidney injury and a preparation and a using method thereof.

Background

Gynura procumbens (Lour.) Merr is a plant of genus Gynura of Compositae, is distributed in Vietnam, Thailand, Indonesia and Africa as well as Guangdong, Hainan, Guizhou, Yunnan, Jiangxi and other places of China, is a traditional food and medicine, is used for treating diabetes, cancer and hypertension, has the effects of clearing away heat and toxic materials, stopping bleeding and stopping cough, promoting blood circulation and removing obstruction in channels, improving the immunity and antiviral ability of human bodies, supplementing calcium, resisting cold, reducing hypertension, hyperlipidemia, improving immunity and the like, and is a non-toxic plant which can be used as both medicine and food.

The inventor of the invention finds that the existing gynura procumbens application has the following problems: 1. the application field needs to be further expanded, and the effects of small toxic and side effects, low price and small environmental pollution of the Chinese herbal medicine are fully exerted; the 2 horizontal chrysanthemum notoginseng leaf is the traditional medicinal and using part, and the stem is usually thrown away in habit and is not used as food or medicine, which causes great resource waste.

Disclosure of Invention

In order to solve at least one problem mentioned in the background art, the embodiments of the present invention aim to provide an extract of gynura procumbens, a preparation for preventing and treating acute kidney injury, and methods for preparing and using the same, wherein gynura procumbens is used for treating acute kidney injury, a new use of gynura procumbens is developed, and in good effect, gynura procumbens can be used for preventing and treating acute kidney injury, and good economic and social benefits are generated; and simultaneously, the parts such as stems can be treated to be used as raw materials, so that the selection range of the raw materials is expanded.

A preparation for preventing and treating acute kidney injury comprises compound 1, compound 2, compound 3 and n-dotriacontanol;

the compound 1 is

The compound 2 is

Said R2 includes-COCH 3 and-COCH 2CH 3;

the compound 3 is

An extract of Gynura procumbens comprises compound 4, compound 5, compound 6 and n-dotriacontanol;

the compound 4 is

The compound 5 is

The compound 6 is

A preparation method of Gynura procumbens extract comprises the following steps:

(1) taking a proper gynura procumbens sample, cleaning, removing impurities, drying, crushing, adding 8-10 times of 70% v/v ethanol, and soaking for 10-60min to obtain a first mixture;

(2) extracting the first mixture for 1-10 times at 80-100 ℃ under reflux, wherein each time of extraction lasts for 0.5-3h, and filtering the extracted first mixture through 100-mesh filter cloth to obtain a first solution;

(3) concentrating the first solution under vacuum to a relative density of 1.05-1.10 to obtain a second solution;

(4) and (3) spray drying the solution II at the temperature of feed liquid materials of 30-40 ℃, the temperature of inlet air of 105-110 ℃, the air compression ratio of 80-150 and the feed liquid speed of 10-60ml/min to obtain a final product.

Preferably, the gynura procumbens is extracted from stems or leaves.

A preparation method of a preparation for preventing and treating acute kidney injury comprises the preparation step of the gynura procumbens extract.

An application method of a preparation for preventing and treating acute kidney injury, the preparation for preventing and treating acute kidney injury or the preparation for preventing and treating acute kidney injury prepared by the method.

Preferably, the kidney injury is acute kidney injury caused by external infection.

Preferably, the kidney injury is caused by an inappropriate treatment.

Preferably, the kidney injury is a kidney injury caused by a side effect of a drug.

In order to better verify the treatment effects of the gynura procumbens extract and the preparation for preventing and treating acute kidney injury, the effect verification test of 180-220g male Wistar rats obtains remarkable effect, and the gynura procumbens extract has definite curative effect, no toxic or side effect and wide application prospect.

Advantageous effects

The invention provides gynura procumbens extract, a preparation for preventing and treating acute kidney injury and a preparation method and a using method thereof, and the gynura procumbens extract has the following characteristics: 1. the gynura procumbens is used for treating acute kidney injury, the new application of gynura procumbens is developed, the effect of protecting the kidney is achieved by reducing the blood urea level and the creatinine level of a rat, and no obvious toxic or side effect exists; 2. the stem and other parts which are not beneficial to direct eating and use are treated by adopting a purification method, so that the stem and other parts can also be used as raw materials, and the selection range of the raw materials is expanded.

Drawings

FIG. 1 is a flow chart of a process for preparing Gynura procumbens extract.

Detailed Description

The technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the scope of the present invention.

A preparation for preventing and treating acute kidney injury comprises compound 1, compound 2, compound 3 and n-dotriacontanol;

the compound 1 is

The compound 2 is

Said R2 includes-COCH 3;

the compound 3 is

An extract of Gynura procumbens comprises compound 4, compound 5, compound 6 and n-dotriacontanol;

the compound 4 is

The compound 5 is

The compound 6 is

A preparation method of Gynura procumbens extract comprises the following steps:

(1) taking a proper gynura procumbens sample, cleaning, removing impurities, drying, crushing, adding 8-10 times of 70% v/v ethanol, and soaking for 10-60min to obtain a first mixture;

(2) extracting the first mixture for 1-10 times at 80-100 ℃ under reflux, wherein each time of extraction lasts for 0.5-3h, and filtering the extracted first mixture through 100-mesh filter cloth to obtain a first solution;

(3) concentrating the first solution under vacuum to a relative density of 1.05-1.10 to obtain a second solution;

(4) and (3) spray drying the solution II at the temperature of feed liquid materials of 30-40 ℃, the temperature of inlet air of 105-110 ℃, the air compression ratio of 80-150 and the feed liquid speed of 10-60ml/min to obtain a final product.

Preferably, the gynura procumbens is extracted from stems or leaves.

A preparation method of a preparation for preventing and treating acute kidney injury comprises the preparation step of the gynura procumbens extract.

An application method of a preparation for preventing and treating acute kidney injury, the preparation for preventing and treating acute kidney injury or the preparation for preventing and treating acute kidney injury prepared by the method.

Preferably, the kidney injury is acute kidney injury caused by external infection.

Preferably, the kidney injury is caused by an inappropriate treatment.

Preferably, the kidney injury is a kidney injury caused by a side effect of a drug.

In some alternative embodiments, gynura procumbens is used for treating acute kidney injury, a new application of gynura procumbens is developed, and the gynura procumbens has the effect of protecting the kidney by reducing the blood urea level and the creatinine level of rats without obvious toxic or side effect;

in some optional embodiments, parts such as stems which are not beneficial to direct eating and use are treated by a purification method, so that the parts can also be used as raw materials, and the selection range of the raw materials is expanded.

Study on efficacy of gynura procumbens for treating acute kidney injury

TABLE 1 results of blood urea nitrogen (BUN, U/L) data

Note: the t value is less than 0.05 and has significant difference

TABLE 2 Creatinine (CRE, μmol/L) data results

Note: the t value is less than 0.05 and has significant difference

As a result: as can be seen from table 1, compared with the model control group, the gynura procumbens alcohol extract low-dose group, the gynura procumbens alcohol extract medium-dose group and the gynura procumbens alcohol extract high-dose group can reduce the level of the haematuria of rats and have significant difference (t < 0.05); as can be seen from table 2, compared with the model control group, the low dose group, the medium dose group and the high dose group of gynura procumbens alcohol extract can reduce the creatinine level of rats to a certain extent, and the medium dose group has significant difference (t <0.05) in reducing the creatinine level; in conclusion, through experimental study, the results of two experiments of reducing the blood urea level and the creatinine level of a rat are positive, and the gynura procumbens alcohol extract and the preparation thereof have a certain effect on treating acute kidney injury.

NMR analysis of Gynura procumbens extract

Based on the disclosed embodiments, the preparation method comprises the following steps: drying 3.0kg of gynura procumbens, refluxing and extracting with methanol, concentrating the extract under reduced pressure, dispersing in water, and sequentially extracting with petroleum ether and chloroform. 30.5g of chloroform extract is obtained, and is subjected to silica gel column chromatography, gradient elution by a petroleum ether-acetone system and a chloroform-methanol system (the volume ratio is 100: 1-1: 1), and further separation and purification by Sephadex LH-20 column chromatography, semi-preparative high performance liquid chromatography and other methods to obtain the following 4 compounds.

Compound 4: white powder (CH3OH), easily soluble in methanol, purple in vanillin sulfate solution, and no dark spot under UV 254 nm. In the 1H-NMR (400MHz, CD3OD) spectrum, δ 4.38(1H, d, J ═ 7.8Hz, H-1 ″), 4.33(1H, d, J ═ 7.6Hz, H-1'), 1.03(3H, s, 19-CH3), 0.95(3H, d, J ═ 6.5Hz, 27-CH3), 0.87(3H, t, J ═ 8.2Hz, 29-CH3), 0.87(3H, s, 18-CH3), 0.86(3H, d, J ═ 8.0Hz, 21-CH3), 0.84(3H, d, J ═ 8.0Hz, 26-CH3) proton signals are present. Where δ 4.33(1H, D, J ═ 7.6Hz, H-1'), 4.38(1H, D, J ═ 7.8Hz, H-1 ″) are sugar end group proton signals and suggest that the sugar is in the β -D configuration. In the 13C-NMR (100MHz, CD3OD) spectrum, δ 142.0(C-5), 122.9(C-6), 105.4(C-1 '), 102.7(C-1'), 80.2(C-3), 69.7(C-6'), 62.6 (C-6'), 58.3(C-14), 57.6(C-17), 51.8(C-9), 47.4(C-24), 43.6(C-13), 41.3(C-12), 39.9(C-4), 38.6(C-1), 37.9(C-10), 37.5(C-20), 35.2(C-22), 33.3(C-8), 33.2(C-7), 30.9(C-2), 30.5(C-25), 29.5(C-16), 27.2(C-23), 25.4 (C-24), 15.2 (C-22), 11.11 (C-4), 28.11 (C-4 20.0(C-27), 19.9(C-19), 19.5(C-26), 19.4(C-21), 12.4(C-29), 12.4(C-18) carbon signals. δ 78.0, 77.0, 76.9, 75.2, 75.1, 72.6, 71.6, 70.4 are the other carbon signals on 2 glucose, respectively; δ 105.4, 102.7 are sugar end group carbon signals; delta.69.7, 62.6 are carbon signals at sugar 6, and indicate 2 glucose 1-6 linked. From the above data, it was concluded that compound 4 was β -sitosterol-3-O-gentiobioside (β -sitosterol-3-O-gentiobioside). Compound 1 comprises compound 4 and its chiral isomer.

Compound 5: white solid (CH3OH), vanillin sulfate solution appeared bluish purple. In the 1H-NMR (400MHz, CD3OD) spectrum, δ 3.69(1H, d, J ═ 7.0Hz, H-7a), 3.45(1H, d, J ═ 7.0Hz, H-7b), 3.07(1H, m, H-2'), 2.22(3H, s, 3' -CH3), 2.08(1H, dd, J ═ 14.0, 6.1Hz, H-4a), 1.97(1H, dd, J ═ 13.7, 2.4Hz, H-2a), 1.85(2H, m, H-1'a, 1' b), 1.77(1H, dd, J ═ 13.9, 10.9Hz, H-2b), 1.53(1H, dd, J ═ 13.8, 1.7, 1H-4 b), 1.3H-3H (3H-3 s-3H, 3H-3H, 2H, 1H, 2H, 1H, 2H, b, 2H, 1H, 2H, 1H, 2H, 1H, 2H, 1H, 2H, 1H, 2H, b, 1H, 2H. In the 13C-NMR (100MHz, CD3OD) spectrum, δ 213.1(s, C-3'), 85.6(s, C-8), 80.1(t, C-7), 77.9(t, C-5), 69.4(s, C-3), 56.6(d, C-2'), 54.6(t, C-2), 47.0(t, C-4), 45.4(s, C-1), 33.4(q, 3-CH3), 26.7(t, C-1'), 20.2(q, 5-CH3), 17.8(q, 1-CH3) carbon signals are present. From the above data, compound 5 was concluded to be curvone (drummondoneA). Compound 2 comprises compound 5 and its chiral isomer.

Compound 6: white powder (CH3OH), vanillin sulfate solution appeared orange-yellow. In the 1H-NMR (400MHz, CD3OD) spectrum, δ 8.07(1H, dd, J ═ 7.7, 1.3Hz, H-4), 7.95(1H, s, H-2), 7.43(1H, dd, J ═ 7.7, 1.3Hz, H-7), 7.20(1H, ddd, J ═ 7.6, 7.5, 1.5Hz, H-6), 7.17(1H, ddd, J ═ 7.6, 7.5, 1.4Hz, H-5) proton signals were present. In the 13C-NMR (100MHz, CD3OD) spectrum, there were delta 169.3(COOH), 138.2(C-9), 133.4(C-2), 127.5(C-8), 123.6(C-4), 122.4(C-6), 122.0(C-5), 112.9(C-7), 108.8(C-3) carbon signals. It is concluded from the above data that compound 6 and compound 3 are both 3-indolecarboxylic acid (indole-3-carboxylic acid).

Compound 7: white powder, vanillin sulfate solution appears grayish purple. In the 1H-NMR (400MHz, CDCl3) spectrum, δ 0.86(3H, t, J ═ 6.12Hz, CH3), 1.25(brs, 58H), 1.58(2H, m, H-2), 3.62(2H, t, CH2OH) proton signals were present. In the 13C-NMR spectrum (100MHz, CDCl3), delta 14.07(CH3), 32.84-25.75 (CH2) and 63.10(CH2OH) carbon signals exist. EI-MSm/z (rel. int.) 448([ M ] + -18, 36), 421(8), 365(5), 153(18), 71(80), 57 (100). From the above data, it was concluded that compound 7 was n-dotriacontanol (n-dotriacontanol).