阅读说明：本技术 一类2-甲基-4-(1-丙三醇)-呋喃类化合物及其制备方法和应用 (2-methyl-4- (1-glycerol) -furan compounds and preparation method and application thereof ) 是由 马忠俊 蒋永俊 黄昀 于 2019-10-31 设计创作，主要内容包括：本发明公开了一类2-甲基-4-(1-丙三醇)-呋喃类化合物,结构式分别如下式(I)～下式(Ⅲ)所示,该三种化合物均为由放线菌通过发酵培养及提纯后得到的天然产物,结构新颖,大大增加了2-甲基-4-(1-丙三醇)-呋喃类似物的种类。经活性试验发现,该三种2-甲基-4-(1-丙三醇)-呋喃类化合物均具有可显著抑制Hep-G2细胞内脂质堆积,可用于制备降血脂的药物或保健食品。<Image he="1000" wi="493" file="DDA0002255178840000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The invention discloses 2-methyl-4- (1-glycerol) -furan compounds, the structural formulas of which are respectively shown as the following formulas (I) to (III), the three compounds are natural products obtained by fermenting, culturing and purifying actinomycetes, the structure is novel, and the types of 2-methyl-4- (1-glycerol) -furan analogues are greatly increased. Activity tests show that the three 2-methyl-4- (1-glycerol) -furan compounds can remarkably inhibit lipid accumulation in Hep-G2 cells, and can be used for preparing blood fat reducing medicines or health foods.)

1. The 2-methyl-4- (1-glycerol) -furan compounds are characterized in that the structural formulas are respectively shown as the following formulas (I) to (III):

2. a method for preparing 2-methyl-4- (1-propanetriol) -furans according to claim 1, comprising the steps of:

1) activating actinomycetes, inoculating the activated actinomycetes into a Gao's first liquid culture medium, and performing shake culture on a shaking table at constant temperature to obtain a fermentation liquid;

the actinomycetes are streptomyces sp.CICC 21933 sold by China Industrial microorganism strain preservation and management center;

2) extracting the fermentation liquor by using an organic solvent to obtain an organic extraction liquid;

3) and concentrating the organic extract, and separating and purifying to respectively obtain the 2-methyl-4- (1-glycerol) -furan compounds with the structures as shown in formulas (I) to (III).

3. The method for producing 2-methyl-4- (1-propanetriol) -furans according to claim 2, wherein the step 1) of activating actinomycetes comprises:

and inoculating the actinomycetes onto a Gao's first agar solid culture medium, standing in a constant-temperature incubator at 28-30 ℃, and performing activation culture for 3-5 days.

4. The method for producing 2-methyl-4- (1-propanetriol) -furans according to claim 2, characterized in that in step 1):

the preparation of the Gao's No. one liquid culture medium comprises the following steps:

uniformly mixing soluble starch, potassium nitrate, dipotassium hydrogen phosphate, magnesium sulfate, ferrous sulfate, sea salt and water, and adjusting the pH value to 7.0-7.2;

the conditions of shake culture are as follows: culturing for 6-10 days at 28-32 ℃ in a shaking table at 180-200 rpm.

5. The method for producing 2-methyl-4- (1-propanetriol) -furans according to claim 2, wherein in the step 2), the organic solvent is ethyl acetate.

6. The method for producing 2-methyl-4- (1-propanetriol) -furans according to claim 2, characterized in that in step 3):

the concentration specifically comprises the following steps:

drying the ethyl acetate extract in vacuum to remove the solvent;

the separation and purification specifically comprises the following steps:

(a) separating the concentrated product by reverse phase column chromatography, wherein the volume ratio is 1:9 to 1: performing gradient elution on the methanol-water system of 0, collecting fractions containing the target compound and merging the fractions;

(b) separating the fractions obtained in step (a) by gel column chromatography, using pure methanol as eluent, collecting fractions containing target compounds, and combining;

(c) and (c) separating and purifying the fraction obtained in the step (b) by adopting a reverse-phase high performance liquid chromatography to respectively obtain the 2-methyl-4- (1-glycerol) -furan compounds shown as formulas (I) to (III).

7. The process for producing 2-methyl-4- (1-propanetriol) -furans according to claim 6, wherein in the step (c):

detecting the wavelength of 210nm, performing gradient elution for 40 minutes at a rate of 10mL/min by adopting a methanol-water system with the methanol volume percentage of 10-100%, and collecting the eluent for 13.5-14 minutes to obtain the 2-methyl-4- (1-glycerol) -furan compound with the structure as shown in the formula (I);

detecting the wavelength of 210nm, performing gradient elution for 40 minutes at a rate of 10mL/min by adopting a methanol-water system with the methanol volume percentage of 10-100%, and collecting the eluent for 19.5-20 minutes to obtain the 2-methyl-4- (1-glycerol) -furan compound with the structure as shown in the formula (II);

the detection wavelength is 210nm, a methanol-water system with 10-100% of methanol volume percentage is adopted, gradient elution is carried out for 40 minutes at 10mL/min, and eluent of 7.0-7.5 minutes is collected to obtain the 2-methyl-4- (1-glycerol) -furan compound with the structure shown in the formula (III).

8. The application of the 2-methyl-4- (1-glycerol) -furan compounds according to claim 1 in preparing medicines for reducing blood fat.

9. The application of the 2-methyl-4- (1-glycerol) -furan compound in the preparation of health-care food for reducing blood fat according to claim 1.

Technical Field

The invention relates to the field of preparation of active compounds from secondary metabolites of actinomycetes, and particularly relates to 2-methyl-4- (1-glycerol) -furan compounds and a preparation method and application thereof.

Background

2-methyl-4- (1-propanetriol) -furan (MFPT, structural formula (1) below), was isolated by Japanese researchers from fermentation products of Streptomyces MC41-M1 and MC340-A1, first in 1971. The research shows that the compound has an inhibiting effect on herpes virus and mouse lymphocyte leukemia cell strains (L1210). Therefore, the compounds have potential anticancer and antiviral effects.

To date, 2-methyl-4- (1-propanetriol) -furan has been discovered for over 40 years, and the continuous development of compounds similar in structure and function to MFPT has been the main direction of researchers based on the potential pharmaceutical efficacy of this class of compounds.

Actinomycetes are widely regarded as an important class of microorganisms for producing various kinds of pharmacologically active substances. At present, actinomycetes are still the main microbial source of lead compounds, and a plurality of active compounds are widely applied to clinic, such as streptomycin, erythromycin, polyoxin, aureomycin, kanamycin, chloramphenicol, gentamicin and the like, which are used for clinically treating a plurality of diseases of human. With the improvement of the requirement for new drug development, the search for a lead compound with a novel action mechanism is necessary.

However, as of yet, no 2-methyl-4- (1-glycerol) -furan analogues of natural origin have been reported as being extracted from actinomycetes.

Disclosure of Invention

The invention extracts natural 2-methyl-4- (1-glycerol) -furan analogues from actinomycetes for the first time, and the three compounds are totally three compounds, thereby greatly increasing the types of the 2-methyl-4- (1-glycerol) -furan analogues. Activity tests show that the three 2-methyl-4- (1-glycerol) -furan compounds can remarkably inhibit lipid accumulation in Hep-G2 cells, and can be used for preparing blood fat reducing medicines or health foods.

The specific technical scheme is as follows:

the structural formulas of the 2-methyl-4- (1-glycerol) -furan compounds are respectively shown as the following formulas (I) to (III):

the two compounds shown in the formula (I) and the formula (II) are dimers formed by adding 5-position in a 2-methyl-4- (1-glycerol) -furan nucleus with another furan ring derivative to form ethanol bridge connection for the first time.

The compound shown in the formula (III) is a dimer formed by adding 2-methyl-4- (1-glycerol) -furan nucleus and another pyran ring derivative for the first time.

Obviously, the three compounds have novel structures, and compared with the mother nucleus structure of 2-methyl-4- (1-glycerol) -furan, the structure of the compound has been changed remarkably.

The invention also discloses a preparation method of the 2-methyl-4- (1-glycerol) -furan compound, which comprises the following steps:

1) activating actinomycetes, inoculating the activated actinomycetes into a Gao's first liquid culture medium, and performing shake culture on a shaking table at constant temperature to obtain a fermentation liquid;

the actinomycetes are streptomyces sp.CICC 21933 sold by China Industrial microorganism strain preservation and management center;

2) extracting the fermentation liquor by using an organic solvent to obtain an organic extraction liquid;

3) and concentrating the organic extract, and separating and purifying to respectively obtain the 2-methyl-4- (1-glycerol) -furan compounds with the structures as shown in formulas (I) to (III).

In step 1):

the process of actinomycete activation comprises the following steps:

and inoculating the actinomycetes onto a Gao's first agar solid culture medium, standing in a constant-temperature incubator at 28-30 ℃, and performing activation culture for 3-5 days.

Preparing the Gao's No. one liquid culture medium: taking 1L of fermentation medium as an example, 20g of soluble starch, 1g of potassium nitrate, 0.5g of dipotassium hydrogen phosphate, 0.5g of magnesium sulfate, 0.01g of ferrous sulfate and 25g of sea salt are added with water to 1L, and the pH is adjusted to 7.0-7.2;

the conditions of shake culture are as follows: culturing for 6-10 days at 28-32 ℃ in a shaking table at 180-200 rpm; more preferably, the culture is carried out at 28 ℃ for 8 days on a shaker at 180 rpm.

In the step 2), the organic solvent is selected from ethyl acetate, and ethyl acetate extract liquor is obtained after ethyl acetate extraction.

In step 3):

the concentration specifically comprises the following steps:

and (3) drying the ethyl acetate in vacuum to remove the solvent, and continuing to perform subsequent separation and purification steps on the concentrated product.

The separation and purification specifically comprises the following steps:

(a) separating the concentrated product by reverse phase column chromatography, wherein the volume ratio is 1:9 to 1: performing gradient elution on the methanol-water system of 0, collecting fractions containing the target compound and merging the fractions;

(b) separating the fractions obtained in step (a) by gel column chromatography, using pure methanol as eluent, collecting fractions containing target compounds, and combining;

(c) and (c) separating and purifying the fraction obtained in the step (b) by adopting a reverse-phase high performance liquid chromatography to respectively obtain the 2-methyl-4- (1-glycerol) -furan compounds shown as formulas (I) to (III).

Preferably, in step (a), the packing material used for the reverse phase column chromatography is octadecyl bonded silica gel.

Preferably, in step (b), the filler used for the gel column chromatography is hydroxypropyl dextran gel.

In the step (c), three compounds shown in the formulas (I) to (III) can be respectively obtained by separating, purifying and collecting products with different elution times.

Preferably, the detection wavelength is 210nm, a methanol-water system with 10-100% methanol volume percentage is adopted, and gradient elution is carried out for 40 minutes at 10 mL/min.

Collecting the eluent for 13.5-14 minutes to obtain a 2-methyl-4- (1-glycerol) -furan compound with a structure shown in the formula (I);

collecting the eluent for 19.5-20 minutes to obtain a 2-methyl-4- (1-glycerol) -furan compound with a structure shown in a formula (II);

collecting the eluent for 7.0-7.5 minutes to obtain the 2-methyl-4- (1-glycerol) -furan compound with the structure shown in the formula (III).

The compounds shown in the formulas (I) to (III) can be respectively obtained with high yield and high purity by adopting the preparation process.

In order to further test the biological activity of the three 2-methyl-4- (1-glycerol) -furan compounds separated by the invention, an in vitro lipid accumulation activity evaluation test is carried out by using HepG2 cells.

Experiments show that the three compounds of the formulas (I) to (III) obtained by separation can obviously inhibit lipid accumulation, so that the compounds can be used for preparing medicines for reducing blood fat or health-care foods for reducing blood fat.

Compared with the prior art, the invention has the following advantages:

the invention discloses three 2-methyl-4- (1-glycerol) -furan compounds, which are obtained by actinomycetes through fermentation culture and purification for the first time, and the structures of the compounds are found in natural products for the first time, and compared with the mother nucleus structures of 2-methyl-4- (1-glycerol) -furan, the mother nucleus structures of the compounds are obviously changed and have novel structures.

Activity tests show that the three compounds can obviously inhibit lipid accumulation, so that the three compounds can be used for preparing blood fat reducing medicines or blood fat reducing health-care foods.

Drawings

FIG. 1 shows the data of the blood lipid-lowering activity of three 2-methyl-4- (1-glycerol) -furans prepared by the present invention.

Detailed Description

Bacterial source

The actinomycetes adopt Streptomyces sp.CICC 21933 sold by China industrial microorganism strain preservation and management center.

Culture medium

Gao's first agar solid medium: based on 1L of culture medium, 20g of soluble starch, 1g of potassium nitrate, 0.5g of dipotassium hydrogen phosphate, 0.5g of magnesium sulfate, 0.01g of ferrous sulfate, 20g of agar and 25g of sea salt are mixed, water is added to 1L, and the pH value is adjusted to 7.0.

Gao's No. one liquid medium: based on 1L of fermentation medium, 20g of soluble starch, 1g of potassium nitrate, 0.5g of dipotassium hydrogen sulfate, 0.5g of magnesium sulfate, 0.01g of ferrous sulfate and 25g of sea salt are mixed, water is added to 1L, and the pH value is adjusted to 7.0.