CN113995745B - Application of diterpenoid compound - Google Patents
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- CN113995745B CN113995745B CN202010730033.7A CN202010730033A CN113995745B CN 113995745 B CN113995745 B CN 113995745B CN 202010730033 A CN202010730033 A CN 202010730033A CN 113995745 B CN113995745 B CN 113995745B
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- -1 diterpenoid compound Chemical class 0.000 title claims abstract description 18
- 150000001875 compounds Chemical class 0.000 claims abstract description 14
- 239000003814 drug Substances 0.000 claims abstract description 12
- 240000004670 Glycyrrhiza echinata Species 0.000 claims abstract description 5
- 235000001453 Glycyrrhiza echinata Nutrition 0.000 claims abstract description 5
- 235000017382 Glycyrrhiza lepidota Nutrition 0.000 claims abstract description 5
- 235000004294 Ononis spinosa Nutrition 0.000 claims abstract description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 38
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 36
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 32
- 239000003208 petroleum Substances 0.000 claims description 20
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 16
- 239000011259 mixed solution Substances 0.000 claims description 12
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 10
- 239000000741 silica gel Substances 0.000 claims description 10
- 229910002027 silica gel Inorganic materials 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 8
- 238000010828 elution Methods 0.000 claims description 8
- 239000002027 dichloromethane extract Substances 0.000 claims description 7
- 239000000243 solution Substances 0.000 claims description 7
- 208000001072 type 2 diabetes mellitus Diseases 0.000 claims description 7
- 239000003480 eluent Substances 0.000 claims description 6
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- 238000000605 extraction Methods 0.000 claims description 5
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- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
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- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- XELZGAJCZANUQH-UHFFFAOYSA-N methyl 1-acetylthieno[3,2-c]pyrazole-5-carboxylate Chemical compound CC(=O)N1N=CC2=C1C=C(C(=O)OC)S2 XELZGAJCZANUQH-UHFFFAOYSA-N 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
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- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 1
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000202807 Glycyrrhiza Species 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 206010022489 Insulin Resistance Diseases 0.000 description 1
- 241000013557 Plantaginaceae Species 0.000 description 1
- 244000265563 Scoparia dulcis Species 0.000 description 1
- 235000004500 Scoparia dulcis Nutrition 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
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- 239000003472 antidiabetic agent Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 244000309466 calf Species 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 238000001516 cell proliferation assay Methods 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 235000021551 crystal sugar Nutrition 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- HWJHWSBFPPPIPD-UHFFFAOYSA-N ethoxyethane;propan-2-one Chemical compound CC(C)=O.CCOCC HWJHWSBFPPPIPD-UHFFFAOYSA-N 0.000 description 1
- 230000005713 exacerbation Effects 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000005802 health problem Effects 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- LTINPJMVDKPJJI-UHFFFAOYSA-N iodinated glycerol Chemical compound CC(I)C1OCC(CO)O1 LTINPJMVDKPJJI-UHFFFAOYSA-N 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
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- 238000010992 reflux Methods 0.000 description 1
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- 210000002966 serum Anatomy 0.000 description 1
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- 235000000346 sugar Nutrition 0.000 description 1
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/21—Esters, e.g. nitroglycerine, selenocyanates
- A61K31/215—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids
- A61K31/235—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids having an aromatic ring attached to a carboxyl group
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C67/00—Preparation of carboxylic acid esters
- C07C67/48—Separation; Purification; Stabilisation; Use of additives
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C67/00—Preparation of carboxylic acid esters
- C07C67/48—Separation; Purification; Stabilisation; Use of additives
- C07C67/56—Separation; Purification; Stabilisation; Use of additives by solid-liquid treatment; by chemisorption
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C2602/00—Systems containing two condensed rings
- C07C2602/02—Systems containing two condensed rings the rings having only two atoms in common
- C07C2602/14—All rings being cycloaliphatic
- C07C2602/26—All rings being cycloaliphatic the ring system containing ten carbon atoms
- C07C2602/28—Hydrogenated naphthalenes
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Diabetes (AREA)
- Emergency Medicine (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Endocrinology (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Hematology (AREA)
- Obesity (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines Containing Plant Substances (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention belongs to the field of biochemical medicines, and discloses application of diterpenoid compounds, wherein the diterpenoid compounds have the following structures:
the compound is extracted and separated from the wild licorice, and cell experiments prove that the compound can obviously resist islet cell apoptosis and has important application prospect in the treatment of T2 DM.
Description
Technical Field
The invention belongs to the field of biological medicine, and in particular relates to application of diterpenoid compounds.
Background
Type 2 diabetes mellitus (T2 DM) has become a major health problem seriously jeopardizing human health and socioeconomic development. Diabetes not only seriously jeopardizes public health, but also causes great economic burden to the family and society of patients. The global economic burden of diabetes was about $1.31 trillion in 2015, and by the year 2030 it was predicted that it would increase to $2.5 trillion per year. Insulin resistance is the source of early development of T2DM, and as the course of the disease increases, the number of islet β cells decreases gradually, and the function thereof decreases progressively. Islet beta cell Apoptosis (PI beta CA) is the final form of reduction in Islet beta cell numbers and is responsible for insulin hyposecretion and thus increasing blood glucose. Although commercially available drugs can lower blood glucose levels in patients with T2DM, the decline in function is caused by apoptosis of islet beta cells, which results in progressive exacerbation of the patient's condition and concomitant failure of the drug. Therefore, besides controlling blood sugar, the use of drugs capable of reducing beta cell apoptosis is a new core of modern T2DM treatment, and clinically used antidiabetic drugs have no drugs directly aiming at islet beta cell apoptosis. Therefore, the search for safe and effective anti-islet beta cell apoptosis drugs can effectively solve the treatment problem of T2 DM.
The plant of the genus Glycyrrhiza (Scoparia dulcis L.) of the family Scrophulariaceae (Scorphulaceae) is sweet in taste and is known as "Crystal sugar grass". The wild licorice has the efficacy of clearing heat and detoxicating in areas such as Hainan and Guangdong in China, and is used as a Li medicine in areas of Hainan Li nationality. The research of the subject group shows that the wild licorice is distributed in the whole island of Hainan, has rich resources and is used for preventing and treating diabetes by local Li medical application.
Disclosure of Invention
The invention provides an application of diterpenoid compounds in preparing medicines for resisting islet cell apoptosis.
The technical scheme of the invention is realized as follows:
the application of diterpenoid compounds in preparing medicines for treating type 2 diabetes is provided, wherein the structure of the diterpenoid compounds is shown as a formula I:
the diterpenoid compound is applied to the preparation of anti-islet cell apoptosis medicines, and the structure of the diterpenoid compound is shown as a formula I:
further, the diterpenoid compound is extracted from the glycyrrhiza uralensis.
Further, the islet cells are MIN-6 cells.
Further, in the application, the concentration of the diterpenoid compound is 25-50 mu M.
In the above application, the extraction method of diterpenoid compounds comprises the following steps:
(1) Drying the whole plant of the wild licorice; reflux-extracting with ethanol under heating; filtering to obtain total extractive solution of radix Glycyrrhizae, concentrating under reduced pressure until no alcohol smell is present to obtain total extractive solution; dissolving the obtained total extract with proper amount of water, adding petroleum ether for extraction, recovering the upper organic phase, extracting the water layer with dichloromethane, recovering the lower organic phase, and concentrating under reduced pressure to obtain dichloromethane extract;
(2) Performing gradient elution on the obtained dichloromethane extract by using silica gel column chromatography, and collecting fractions according to the used eluents with different gradients by using petroleum ether and acetone mixed solution;
(3) The volume ratio of petroleum ether to acetone used in the step (2) is 100: the obtained fraction is loaded onto a silica gel chromatographic column, the mixed solution of petroleum ether, dichloromethane and acetone is used as an eluent, the mixed solution of V petroleum ether/V dichloromethane/V acetone=50:1:1-2:1:1 is subjected to gradient elution, the eluted fraction of petroleum ether, dichloromethane and acetone with the volume ratio of 10:1:1 is further subjected to Sephadex LH-20 (MeOH) column chromatography, and then semi-prepared high performance liquid chromatography separation is carried out, and the elution is carried out by using the mixed solution of methanol and water with the volume ratio of 60:40, so that the compound I is obtained.
Further, in the step (2), the silica gel column chromatography is performed by dry mixing with silica gel, and the particle size of the silica gel is 200-300 mu M.
Further, in the step (2), the petroleum ether-acetone volume ratio in the gradient elution is sequentially 100:0, 100:2, 100:5, 100:10, 100:16, and 100:25.
Further, in the step (3), the semi-preparative high performance liquid chromatography column is ZORAX SB-phenyl.
The invention has the beneficial effects that:
the compound disclosed by the invention belongs to diterpenoid compounds, and cell experiments show that the compound has a remarkable effect of resisting islet cell apoptosis and has an important application prospect in the treatment of T2 DM.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions of the prior art, the drawings which are used in the description of the embodiments or the prior art will be briefly described, it being obvious that the drawings in the description below are only some embodiments of the invention, and that other drawings can be obtained according to these drawings without inventive faculty for a person skilled in the art.
FIG. 1 is a nuclear magnetic resonance carbon spectrum of compound I;
FIG. 2 shows the nuclear magnetic resonance hydrogen spectrum of compound I;
FIG. 3 is an anti-islet cell apoptosis graph of diterpenoid I according to the invention.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
EXAMPLE 1 preparation of the Compounds of the invention
10kg of Glycyrrhiza uralensis are taken, ethanol reflux extraction is adopted, the extract is dissolved in water after the concentration of the extract, petroleum ether extraction is then carried out, the upper organic phase is recovered, the remaining water layer is extracted by dichloromethane, the lower organic phase is recovered, and the reduced pressure concentration is carried out, thus obtaining dichloromethane extract (180 g).
Mixing the obtained dichloromethane extract with silica gel by dry method, adding organic solvent to dissolve the dichloromethane extract, mixing with silica gel particles, grinding after volatilizing the organic solvent, performing column chromatography with silica gel (200-300 mesh silica gel, particle size of 200-300 μm), gradient eluting with petroleum ether and acetone mixed solution, and mixing petroleum ether and acetone at volume ratio of 100: loading the obtained fraction on a silica gel chromatographic column, performing gradient elution by using a mixed solution of petroleum ether, dichloromethane and acetone as an eluent and a mixed solution of V petroleum ether/V dichloromethane/V acetone=50:1:1-2:1:1, separating the eluted fraction with the volume ratio of petroleum ether to dichloromethane to acetone of 10:1:1 by Sephadex LH-20 (MeOH) column chromatography, performing semi-preparative high performance liquid chromatography separation, and eluting by using a mixed solution with the volume ratio of methanol to water of 60:40 to obtain a compound I;
the nuclear magnetic resonance wave spectrum of the compound I is shown in figures 1 to 2. The spectrum data are shown in Table 1.
TABLE 1 spectroscopic data for Compound I of the invention
1 H-NMR test conditions were 600MHz, CD 3 OD； 13 C-NMR test conditions were 150MHz, CD 3 OD。
Example 2: anti-MIN-6 apoptosis activity experiment of the compound of the invention
The method comprises the following steps: (1) MIN6 cell reviving and culturing: taking out in a refrigerator at-80deg.C, centrifuging to remove upper frozen stock solution, adding 1640 culture medium containing 10% foetal calf serum and 1% double antibody to re-suspend cells, and culturing in incubator with relative humidity of 95% and 37 deg.C and 5% CO 2; fresh culture is replaced every 24-36 h according to the growth condition of cellsAnd (3) after the cells grow to 75% -85%, carrying out passage or freezing. (2) diterpenoid intervention model group (PA) culture: grouping of MIN6 cell proliferation: well grown MIN6 cells were plated in 96 well plates at 5X 10 5 100 μLC complete medium was added per mL. The cells were divided into the following groups: blank (CK), 300 μg/mLC model (PA), compound I25 μg/mL dosing, 50 μg/mL dosing. The original culture solution was removed, and 100. Mu.l of each well was added with the corresponding culture medium, and the culture was performed for 24 hours, respectively, to measure the proliferation of cells. (3) MTT method cell proliferation assay MIN6 cells are subjected to intervention treatment under item (2), the original culture solution is removed, 50 mu L of 1 xMTT is added to each well, the culture is incubated for 4 hours at 37 ℃, the supernatant is sucked out after 4 hours, 150UL DMSO is added to each well, the mixture is uniformly shaken on a flat shaking table for 5 minutes, and the absorbance of each well is detected at the wavelength of 490nm by an enzyme-labeled instrument after the shaking.
Experimental results: diterpenoid I obtained by the invention can obviously improve PA (300 mu M) induced MIN6 cell apoptosis at treatment concentrations of 50 mu M and 25 mu M, as shown in figure 3.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.
Claims (5)
1. The application of diterpenoid compounds in preparing a medicament for treating type 2 diabetes is characterized in that the structure of the diterpenoid compounds is shown as a formula I:
2. The use according to claim 1, wherein: the diterpenoid compounds are extracted from the glycyrrhiza uralensis.
3. The use according to claim 1 or 2, wherein the method for extracting diterpenoid compounds comprises the following steps:
(1) Drying the whole plant of the wild licorice; reflux-extracting with ethanol under heating; filtering to obtain total extractive solution of radix Glycyrrhizae, concentrating under reduced pressure until no alcohol smell is present to obtain total extractive solution; dissolving the obtained total extract with proper amount of water, adding petroleum ether for extraction, recovering the upper organic phase, extracting the water layer with dichloromethane, recovering the lower organic phase, and concentrating under reduced pressure to obtain dichloromethane extract;
(2) Performing gradient elution on the obtained dichloromethane extract by using silica gel column chromatography, and using petroleum ether and acetone mixed solution, wherein the volume ratio of petroleum ether to acetone is sequentially 100:0, 100:2, 100:5, 100:10, 100:16 and 100:25; collecting fractions according to the different gradients of eluent used;
(3) The volume ratio of petroleum ether to acetone used in the step (2) is 100: the obtained fraction is loaded onto a silica gel chromatographic column, the mixed solution of petroleum ether, dichloromethane and acetone is used as an eluent, the mixed solution of V petroleum ether/V dichloromethane/V acetone=50:1:1-2:1:1 is subjected to gradient elution, the eluted fraction of petroleum ether, dichloromethane and acetone with the volume ratio of 10:1:1 is further subjected to Sephadex LH-20MeOH column chromatography, meOH is used as the eluent, and then semi-preparative high performance liquid chromatography is carried out, and the elution is carried out by using the mixed solution of methanol and water with the volume ratio of 60:40, so that the compound I is obtained.
4. The use according to claim 3, wherein in step (2), the silica gel column chromatography is performed by dry mixing with silica gel having a particle size of 200 to 300 μm.
5. The use of claim 3, wherein the semi-preparative high performance liquid chromatography column in step (3) is ZORAX SB-phenyl.
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| Application Number | Priority Date | Filing Date | Title |
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| CN202010730033.7A CN113995745B (en) | 2020-07-27 | 2020-07-27 | Application of diterpenoid compound |
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| CN202010730033.7A CN113995745B (en) | 2020-07-27 | 2020-07-27 | Application of diterpenoid compound |
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| Publication Number | Publication Date |
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| CN113995745A CN113995745A (en) | 2022-02-01 |
| CN113995745B true CN113995745B (en) | 2023-05-19 |
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2020
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Non-Patent Citations (6)
| Title |
|---|
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