Open ring resorcinol glycoside derivatives, methods of making and uses thereof

Abstract

The application belongs to the technical field of medicine, and discloses a kind of open loop resin glycoside derivatives and its preparation method and application.The application takes three color spinflower seed as separation target, and is obtained by comprehensively using a variety of separation technology and means open loop resin glycoside derivatives with novel chemical structure.The in-vitro activity experiment proves that the open loop resin glycoside derivatives provided by the application have significant anti-neuroinflammatory activity, can be used as active ingredient for treating neuroinflammatory related diseases, and have wide application.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to an open-ring resin glycoside derivative, its preparation method, and its application. Background Technology

[0002] The incidence of neurodegenerative diseases, traumatic brain injury, and stroke is rising year by year, placing a heavy burden on society and families. Studies have shown that neuroinflammation plays a crucial role in the pathogenesis of various neurological diseases. Microglial hyperactivation is considered a typical marker of neuroinflammation; excessive microglial activation can produce and secrete large amounts of neurotoxic factors, including chemokines and pro-inflammatory factors, leading to damage to the nervous system. Therefore, inhibiting neuroinflammation mediated by abnormal microglial activation is an important strategy for developing drugs to treat neurological diseases.

[0003] Resin glycosides are characteristic components of plants in the Convolvulaceae family, and are classified into macrocyclic resin glycosides and open-ring resin glycosides. Open-ring resin glycosides consist of three parts: an oligosaccharide chain, a long-chain fatty acid aglycone, and a modified fatty acyl group. The terminal carboxyl group of the aglycone usually exists in a free state or as a methyl ester. In recent years, many open-ring resin glycoside derivatives have been reported to possess significant biological activities, such as anti-epileptic, sedative, and vasodilatory activities, providing diverse template molecules for drug development. Therefore, these novel structures are increasingly attracting the attention of drug researchers.

[0004] Therefore, it is necessary to study the active components of some plants and develop novel active resin glycoside compounds. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide an open-ring resin glycoside derivative, its preparation method, and its application.

[0006] To address the above problems, the present invention provides the following technical solution:

[0007] In a first aspect, the present invention provides an open-ring resin glycoside derivative having a chemical structure shown in any of the following structural formulas:

[0008]

[0009] In a second aspect, the present invention provides a method for preparing the above-mentioned open-ring resin glycoside derivative, specifically: the resin glycoside derivative is isolated from the seeds of Convolvulus tricolor L.

[0010] In some embodiments of the present invention, the above preparation method specifically involves selecting seeds of Tricolor Convolvulella tricolor, extracting them by reflux at 95%, and then obtaining the above-mentioned resin glycoside derivatives of formulas I to III through various separation techniques and methods. These derivatives have significant anti-neuroinflammatory activity and can be used as active ingredients for treating nervous system diseases, thus having a wide range of applications.

[0011] In some embodiments of the present invention, the above preparation method includes the following steps:

[0012] This invention utilizes the seeds of *Convolvulus tricolor* (tricolor bindweed) and extracts them by reflux with a 95% (v / v) ethanol aqueous solution. The compound of this invention is prepared through various separation techniques and methods. The *Convolvulus tricolor* seeds were purchased in Beijing.

[0013] 1) Extraction

[0014] The seeds of Tricolor Convolvulia were extracted several times by hot reflux with an ethanol aqueous solution. The extracts were combined and concentrated under reduced pressure to obtain an extract.

[0015] 2) Separation by silica gel chromatography

[0016] The extract was subjected to silica gel column chromatography with gradient elution using a dichloromethane / methanol mixed solvent (1:0, 10:1, 5:1, 2:1, 1:1, 0:1, v / v) to obtain fractions 1 to 8 sequentially.

[0017] 3) Reverse-phase ODS column chromatography

[0018] Fraction 2 was subjected to reversed-phase ODS column chromatography with methanol-water solution (50:50, 70:30, 80:20, 85:15, 90:10, 95:5, 100:0, v / v) as eluent to obtain fractions 2.1 to 2.3.

[0019] 4) Amino silica gel column chromatography

[0020] Fraction 2.2 was subjected to amino silica gel column chromatography with dichloromethane-methanol (5:1, 2:1, 1:1, 0:1, v / v) as the eluent to obtain fractions 2.2.1 to 2.2.4.

[0021] 5) Reversed-phase ODS column chromatography

[0022] Component 2.2.2 was subjected to reversed-phase ODS column chromatography with methanol-water solution (60:40, 70:30, 80:20, 85:15, 90:10, 95:5, 100:0, v / v) as eluent to obtain components 2.2.2.1 to 2.2.2.8.

[0023] 6) Reversed-phase high-performance liquid chromatography purification

[0024] Component 2.2.2.5 was purified by reversed-phase high-performance liquid chromatography. The preparation conditions were as follows: YMC RP-C18 column (10×200 mm), methanol-water (79:21, V / V), flow rate 2.0 mL / min, and Shimadzu RID 10A detector, to obtain compounds of formulas I to III.

[0025] In a third aspect, the present invention provides the use of the above-mentioned resin glycoside derivatives or pharmaceutically acceptable salts thereof as active ingredients in the preparation of anti-neuroinflammatory drugs.

[0026] In a fourth aspect, the present invention provides a pharmaceutical composition comprising the above-described open-ring resin glycoside derivatives or pharmaceutically acceptable salts thereof.

[0027] Compared with existing technologies, this invention selects the seeds of *Convolvulus tricolor* as the target, extracts them using ethanol under reflux, and then obtains the aforementioned resin glycoside derivatives of formulas I to III through various separation techniques and methods. All compounds of formulas I to III are open-ring resin glycoside derivatives, among which formulas I and II are rare resin glycoside derivatives containing eight sugar residues. These novel resin glycoside compounds exhibit significant anti-neuroinflammatory activity and can be used as active ingredients in anti-neuroinflammatory processes, possessing a wide range of applications. Attached Figure Description

[0028] Figure 1 This is a hydrogen spectrum of compound I obtained in Example 1 of the present invention;

[0029] Figure 2 This is a carbon spectrum of compound I obtained in Example 1 of the present invention;

[0030] Figure 3 This is the proton NMR spectrum of compound II obtained in Example 1 of the present invention;

[0031] Figure 4 This is a carbon spectrum of compound II obtained in Example 1 of the present invention;

[0032] Figure 5 This is the proton NMR spectrum of compound III obtained in Example 1 of the present invention;

[0033] Figure 6 This is a carbon spectrum of compound III obtained in Example 1 of the present invention. Detailed Implementation

[0034] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Operating methods not specifically specified in the following embodiments are generally performed under conventional conditions or as recommended by the manufacturer.

[0035] Example 1: Preparation of the compound

[0036] This invention utilizes the seeds of *Convolvulus tricolor* (tricolor bindweed) and extracts them by reflux with a 95% (v / v) ethanol aqueous solution. The compound of this invention is prepared through various separation techniques and methods. The *Convolvulus tricolor* seeds were purchased in Beijing.

[0037] 1) Extraction

[0038] Five kilograms of Tricolor Convolvulia seeds were crushed and then refluxed with five times the volume of 95% ethanol aqueous solution for three times, each time for three hours. The extracts were combined and concentrated under reduced pressure to obtain an extract (500g).

[0039] 2) Separation by silica gel chromatography

[0040] The extract was subjected to silica gel column chromatography with gradient elution using a dichloromethane / methanol mixed solvent (1:0, 10:1, 5:1, 2:1, 1:1, 0:1, v / v) to obtain fractions 1 to 8 sequentially.

[0041] 3) Reverse-phase ODS column chromatography

[0042] Fraction 2 was subjected to reversed-phase ODS column chromatography with methanol-water solution (50:50, 70:30, 80:20, 85:15, 90:10, 95:5, 100:0, v / v) as eluent to obtain fractions 2.1 to 2.3.

[0043] 4) Amino silica gel column chromatography

[0044] Fraction 2.2 was subjected to amino silica gel column chromatography with dichloromethane-methanol (5:1, 2:1, 1:1, 0:1, v / v) as the eluent to obtain fractions 2.2.1 to 2.2.4.

[0045] 5) Reversed-phase ODS column chromatography

[0046] Component 2.2.2 was subjected to reversed-phase ODS column chromatography with methanol-water solution (60:40, 70:30, 80:20, 85:15, 90:10, 95:5, 100:0, v / v) as eluent to obtain components 2.2.2.1 to 2.2.2.8.

[0047] 6) Reversed-phase high-performance liquid chromatography purification

[0048] Component 2.2.2.5 was purified by reversed-phase high-performance liquid chromatography. The preparation conditions were as follows: YMC RP-C18 column (10×200 mm), methanol-water (79:21, V / V), flow rate 2.0 mL / min, and Shimadzu RID 10A detector, to obtain compounds I to III.

[0049] The proton and carbon spectra of compounds I–III are as follows: Figure 1-6 As shown. By analyzing... Figures 1-6 The mass spectrometry and wavelength spectrometry data of compounds I-III are shown below.

[0050] Compound I: controlin V; major absorption peak in infrared spectrum (KBr)ν max : 3420, 2932, 1734, 1650, 1077cm -1 ;[α] 25 D -12.9(c 0.11,MeOH); UV(MeOH)λmax(logε)221(3.42)nm; HRESIMS m / z931.4070[M+2Na] 2+ (C 80 H 136 Na2O 45 (Calculated value 931.4069); the proton and carbon NMR spectra are shown in Table 1.

[0051] Compound II: controlin VI; major absorption peak in infrared spectrum (KBr)ν max : 3421, 2932, 1733, 1650, 1075 -1 ;[α] 25 D -30.2(c 0.09,MeOH); UV(MeOH)λmax(logε)220(3.56)nm; HRESIMS m / z981.4335[M+2Na] 2+ (C 85 H 144 Na2O 47 (Calculated value 981.4331); the proton and carbon NMR spectra are shown in Table 1.

[0052] Compound III: controlin VIII; major absorption peak in infrared spectrum (KBr)ν max : 3444, 2930, 1734, 1650, 1075 -1 ;[α] 25 D-21.8(c 0.12,MeOH); UV(MeOH)λmax(logε)218(3.32)nm; HRESIMS m / z850.3804[M+2Na] 2+ (C 74 H 126 Na2O 40 (Calculated value 850.3805); the proton and carbon NMR spectra are shown in Table 1.

[0053] Table 1. 1H and 1C NMR spectra of compounds I through III

[0054]

[0055]

[0056]

[0057] a Overlapped signals are reported without designating multiplicity; b Abbreviations: Glc=glucopyranosyl, Rha=rhamnopyranosyl, Fuc=fucopyranosyl, Qui=quinovopyranosyl, Ag=12-hydroxyhexadecanoyl aglycone, Nla=3-hydroxy-2-methylbutanoyl, Tga=2-methyl-2-butenoyl.

[0058] The above results indicate that the structural formulas of compounds I-III are chemical formulas I-III, respectively.

[0059] Example 2: Anti-neuroinflammatory activity of compounds I-III

[0060] 1) Experimental materials

[0061] Instruments and reagents: CO2 incubator (Thermo Fisher Scientific, USA); DMEM / RPMI 1640 medium (Gibco, Thermo Fisher Scientific, USA); DMSO and trypsin, etc. (Sigma, USA); CCK-8 and nitric oxide (NO) reagent kits (Beyotime Biotechnology Research Institute); fetal bovine serum (Hangzhou Sijiqing Biomaterials Research Institute); other commonly used inorganic salts and reagents were all of analytical grade.

[0062] The cell line used for testing was the mouse microglia BV2 cell line.

[0063] Test samples: Resin glycoside derivatives of formulas I to III obtained from seeds of Convolvulia tricolor; L-arginine was selected as a positive control drug. All compounds were dissolved in DMSO and then diluted.

[0064] 2) Experimental methods

[0065] BV2 cells in the logarithmic growth phase were seeded into 96-well culture plates at a cell density of 1.5 × 10⁻⁶ cells / well. 4 / well. After culturing for 24 hours, lipopolysaccharide (LPS) was added to a final concentration of 100 ng / mL, and culturing continued for another 3 hours. Subsequently, different concentrations of samples or positive control agents were added, and culturing continued for another 24 hours. Following the kit instructions, the NO level in each group of culture media was determined using the Griess method, and the half-maximal inhibitory concentration (IC50) for each test sample inhibiting NO release was calculated. 50 The cell viability of each group was determined using the CCK-8 assay.

[0066] 3) Experimental Results

[0067] Based on the results of the Griess method and CCK-8 assay, the effects of compounds of formulas I to III of the present invention on LPS-induced NO release in BV-2 cells were calculated, and the results are shown in Table 2.

[0068] Table 2. Results of test samples inhibiting LPS-induced NO release in BV-2 cells

[0069]

[0070] a IC 50 The value is expressed as mean ± SD. b The cell viability test concentration was 50 μM.

[0071] The results showed that compounds I-III of the present invention inhibited LPS-induced NO in BV-2 microglia with an IC50. 50 With a concentration of 6.73-19.10 μM, it can be used as an active ingredient for treating neuroinflammatory diseases.

[0072] Those skilled in the art should understand that this invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to this invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the invention as claimed. The scope of protection of this invention is defined by the appended claims and their equivalents.

Claims

1. An open-ring resin glycoside derivative, characterized in that, The open-ring resin glycoside derivatives have a chemical structure shown in any of the following structural formulas:

2. A method for preparing the open-ring resin glycoside derivative as described in claim 1, characterized in that, The preparation method involves separating the open-ring resin glycoside derivative from the seeds of Convolvulia tricolor. The preparation method includes the following steps: S1. The seeds of Tricolor Convolvulia were subjected to several hot reflux extractions using an ethanol-water solution. The extracts were combined and concentrated under reduced pressure to obtain an extract. S2. The extract was subjected to silica gel column chromatography, and gradient elution was performed using a dichloromethane / methanol mixed solvent with volume ratios of 1:0, 10:1, 5:1, 2:1, 1:1, and 0:1 to obtain fractions 1 to 8 in sequence. S3. Fraction 2 was subjected to reversed-phase ODS column chromatography, with gradient elution using methanol-water solutions of volume ratios of 50:50, 70:30, 80:20, 85:15, 90:10, 95:5, and 100:0 to obtain fractions 2.1 to 2.

3. S4. Fraction 2.2 was subjected to amino silica gel column chromatography, and gradient elution was performed sequentially with dichloromethane-methanol in volume ratios of 5:1, 2:1, 1:1, and 0:1 to obtain fractions 2.2.1 to 2.2.

4. S5. Component 2.2.2 was subjected to reversed-phase ODS column chromatography, with gradient elution using methanol-water solutions of volume ratios of 60:40, 70:30, 80:20, 85:15, 90:10, 95:5, and 100:0 to obtain components 2.2.2.1 to 2.2.2.8; S6. Component 2.2.2.5 was purified by reversed-phase high-performance liquid chromatography. The preparation conditions were as follows: YMC RP-C18 column, 10×200 mm, methanol-water solution with a volume ratio of 79:21, flow rate of 2.0 mL / min, and Shimadzu RID 10A detector. Compounds with structures shown in Formulas I to III were obtained.

3. The use of the open-ring resin glycoside derivative of claim 1 as an active ingredient in the preparation of an anti-neuroinflammatory drug.

4. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the open-ring resin glycoside derivative of claim 1 or a pharmaceutically acceptable salt thereof.

Images