A quinic acid compound, a preparation method and application thereof

Abstract

The application relates to a quinic acid compound and a preparation method and application thereof, the quinic acid compound has an alpha-glucosidase semi-inhibition concentration IC 50 The value is 0.000019 mol / L, indicating that the quinic acid compound has good alpha-glucosidase inhibition activity, has good blood glucose lowering activity, and has important significance for the research and development of a drug for treating diabetes.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to a quinic acid compound, its preparation method, and its application. Background Technology

[0002] Diabetes mellitus is a metabolic and chronic disease caused by factors such as insulin secretion defects. Its main characteristic is hyperglycemia. Long-term hyperglycemia can lead to a series of complications such as cardiovascular disease and kidney disease, seriously threatening human health. Currently, drugs used to treat diabetes include insulin (sulfonylureas and meglitinides), biguanides, and thiazolidinediones, which are very effective in controlling blood sugar. However, long-term use usually produces varying degrees of toxic side effects. Alpha-glucosidase inhibitors lower blood sugar by altering the absorption of carbohydrates in the intestine, and have a good preventive effect on diabetic complications. They are widely used clinically in the treatment of type 2 diabetes. Currently, commonly used alpha-glucosidase inhibitors such as acarbose and voglibose can also cause adverse reactions such as bloating and diarrhea. Therefore, finding new alpha-glucosidase inhibitors with better activity and mild or no adverse reactions is of great significance for the treatment of diabetes. Summary of the Invention

[0003] The technical problem to be solved by this invention is to provide a drug for treating diabetes. The quinic acid compounds of this invention have good α-glucosidase inhibitory activity and can be used to treat diabetes.

[0004] The technical solution of this invention to solve the above-mentioned technical problems is as follows: a quinic acid compound, wherein the structural formula of the quinic acid compound is:

[0005]

[0006] The beneficial effects of this invention are: the quinic acid compound of this invention (named methyl-3-O-caffeoyl-4-O-galloylquinic acid) has an IC50 half-inhibitory concentration (IC50) against α-glucosidase. 50 The value was 0.000019 mol / L (IC50 value of the positive control acarbose). 50 The value was 0.0019 mol / L, indicating that this quinic acid compound has good α-glucosidase inhibitory activity and good hypoglycemic activity, which is of great significance for the development of drugs for the treatment of diabetes.

[0007] To achieve the second objective mentioned above, this invention provides a method for preparing quinic acid compounds, comprising:

[0008] After crushing and extracting the leaves of *Cephalotaxus fortunei*, the filtrate was concentrated, floated in water, defatted, and concentrated again. The concentrated water fraction was then separated sequentially by Sephadex LH-20 gel column chromatography, MCI gel CHP 20P column chromatography, Sephadex LH-20 gel column chromatography, Chromatorex C18 column chromatography, and Diaion HP20SS column chromatography, with the eluent being a mixture of methanol and water.

[0009] The beneficial effects are: the potent α-glucosidase inhibitor obtained by extracting and separating the leaves of *Cinnamomum yuanjiangense* from the leaves of this invention is simple to prepare and easy to operate. Moreover, the extraction is carried out using leaves as raw materials, which are abundant and have great potential economic benefits.

[0010] Furthermore, the eluent for the first Sephadex LH-20 gel column chromatography separation is a mixture of methanol and water with methanol volume contents of 0%, 20%, 40%, 60%, 80%, and 100% respectively.

[0011] The eluent for the MCI gel CHP 20P column chromatography separation is a mixture of methanol and water with methanol volume contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% respectively.

[0012] The eluent for the second Sephadex LH-20 gel column chromatography separation was a mixture of methanol and water with methanol volume contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% respectively.

[0013] The eluent for the Chromatorex C18 column chromatography separation is a mixture of methanol and water with a methanol volume content of 0% to 50%.

[0014] The eluent for the Diaion HP20SS column chromatography separation is a mixture of methanol and water with a methanol volume content of 0% to 40%.

[0015] Furthermore, the extraction includes: extracting with methanol at a volume concentration of 65-75% 2-3 times, each extraction lasting 6-8 days, and combining the filtrates;

[0016] The reagent used for degreasing is petroleum ether.

[0017] To achieve the third objective mentioned above, this invention provides a pharmaceutical salt of quinic acid compounds, wherein the pharmaceutical salt is a salt synthesized from the aforementioned quinic acid compounds with organic acids and / or inorganic acids.

[0018] To achieve the fourth objective mentioned above, this invention provides a pharmaceutically acceptable esterified derivative of a quinic acid compound, wherein the pharmaceutically acceptable esterified derivative is prepared by an esterification reaction of the aforementioned quinic acid compound.

[0019] To achieve the fifth objective mentioned above, this invention provides the application of quinic acid compounds in the preparation of hypoglycemic drugs.

[0020] It should be noted that the quinic acid compounds provided in this invention

[0021] methyl-3-O-caffeoyl-4-O-galloylquinic acid, or its derivatives, or pharmaceutical salts thereof, can be combined with excipients or carriers permitted in formulations or pharmaceuticals to prepare drugs or pharmaceutical compositions with α-glucosidase inhibitory activity for the treatment of diabetes. These drugs or pharmaceutical compositions can be in dosage forms such as tablets, granules, or capsules; they can also be formulated using methods known in the modern pharmaceutical industry, such as nanoparticles, controlled-release formulations, and sustained-release formulations.

[0022] Furthermore, the hypoglycemic drug includes a pharmaceutically acceptable carrier and the quinic acid compound and / or a derivative of the quinic acid compound.

[0023] It should be noted that derivatives of quinic acid compounds include pharmaceutically acceptable salts of quinic acid compounds. Attached Figure Description

[0024] Figure 1 The structural formula of the quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention is shown below.

[0025] Figure 2 The quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention 1 H-NMR spectrum;

[0026] Figure 3 The quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention 13 C-NMR spectrum;

[0027] Figure 4 The HSQC spectrum of the quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention is shown below.

[0028] Figure 5The HMBC spectrum of the quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention is shown below.

[0029] Figure 6 The quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention 1 H- 1 H COSY spectrum;

[0030] Figure 7 The image shows the HREIMS spectrum of the quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinicacid) of this invention. Detailed Implementation

[0031] The principles and features of this invention are described below. The examples given are for illustrative purposes only and are not intended to limit the scope of the invention. Where specific techniques or conditions are not specified in the embodiments, they should be performed according to the techniques or conditions described in the literature in this field, or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels.

[0032] Example

[0033] A quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinic acid) and its preparation method, comprising:

[0034] (1) After crushing 5.5 kg of dried Yuanjiang Conifer leaves, it was extracted three times by soaking in 40 L of 70% methanol for 7 days each time. After filtration, the filtrates were combined and then concentrated under reduced pressure to obtain 2.4 L of aqueous solution. The aqueous solution was defatted four times with 2.5 L of petroleum ether. The defatted aqueous solution was then concentrated under reduced pressure to remove the petroleum ether.

[0035] (2) The aqueous solution after removing petroleum ether was separated by Sephadex LH-20 gel (10cm×50cm) column chromatography. It was eluted sequentially with 2L of a mixture of methanol and water at different concentrations (the volume fractions of methanol were 0%, 20%, 40%, 60%, 80%, and 100%). The same components were combined by TLC monitoring to obtain 7 fractions (Fr.1~7).

[0036] (3) Fr.4 (227.0 g) was separated by MCI gel CHP 20P (7.5 cm × 29.5 cm) column chromatography. The fractions were eluted sequentially with 1.5 L of a mixed solution of methanol and water of varying concentrations (the volume fractions of methanol were 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100%). The same components were combined by TLC monitoring to obtain 11 fractions (Fr.4.1 to 4.11).

[0037] (4) Fr.4.4 (20.5 g) was separated by Sephadex LH-20 gel chromatography (3.5 cm × 31.5 cm). The fractions were eluted sequentially with 0.5 L of a mixture of methanol and water at different concentrations (the volume fractions of methanol were 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100%). The same components were combined by TLC monitoring to obtain 7 fractions (Fr.4.4.1 to 4.4.7).

[0038] (5) Fr.4.4.3 (6.3 g) was separated by Chromatorex C18 (2.5 cm × 23.5 cm) column chromatography, and eluted sequentially with a mixed solution of 300 mL of methanol and water (in which the volume fraction of methanol was 0%, 10%, 20%, 30%, 40%, and 50%). The same components were combined by TLC monitoring to obtain 5 fractions (Fr.4.4.3.1 to 4.4.3.5);

[0039] (6) Fr.4.4.3.1 (0.6 g) was separated by Diaion HP20SS (1.7 cm × 24.5 cm) column chromatography and eluted sequentially with a mixed solution of 100 mL of methanol and water (in which the volume fraction of methanol was 0%, 10%, 20%, 30%, and 40%) to obtain 10.0 mg of quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinic acid).

[0040] Identification of the quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinic acid) in the examples:

[0041] Figure 2 It is a quinic acid compound, methyl-3-O-caffeoyl-4-O-galloylquinic acid. 1 H-NMR spectrum; Figure 3 It is a quinic acid compound, methyl-3-O-caffeoyl-4-O-galloylquinic acid.13 C-NMR spectrum; Figure 4 This is the HSQC spectrum of the quinic acid compound methyl-3-O-caffeoyl-4-O-galloylquinic acid; Figure 5 This is the HMBC spectrum of the quinic acid compound methyl-3-O-caffeoyl-4-O-galloylquinic acid; Figure 6 It is a quinic acid compound, methyl-3-O-caffeoyl-4-O-galloylquinic acid. 1 H- 1 H COSY spectrum; Figure 7 This is the HREIMS spectrum of the quinic acid compound methyl-3-O-caffeoyl-4-O-galloylquinic acid. The quinic acid compound (methyl-3-O-caffeoyl-4-O-galloylquinic acid) is a yellow amorphous powder with a bright spot under 254 nm UV light. It reacts with 1% FeCl3-EtOH colorimetric reagent to form blue-black spots. The HREIMS spectrum shows its quasi-molecular ion peak at m / z: 519.1144 [MH]. - (Calculated value 519.1175, C) 24 H 23 O 13 The molecular formula is C. 24 H 24 O 13 The degree of unsaturation is 13. 1 H-NMR (500MHz, acetone-d6), 13 C-NMR (125MHz, acetone-d6) data are shown in Table 1 below.

[0042] Table 1. NMR data (in acetone-d6) of the quinic acid compound methyl-3-O-caffeoyl-4-O-galloylquinic acid.

[0043]

[0044]

[0045] Based on the comprehensive analysis of the above mass spectrometry, one-dimensional and two-dimensional NMR data, the structural formula of the compound was deduced as follows: Figure 1 As shown, it is named methyl-3-O-caffeoyl-4-O-galloylquinic acid.

[0046] Activity assay of quinic acid compound methyl-3-O-caffeoyl-4-O-galloylquinic acid:

[0047] Prepare a 50 mM, pH 6.8 solution of 20×PBS phosphate buffer. Use 1 mmol / L p-nitrophenyl-α-D-glucopyranoside (PNPG) as the substrate. Prepare an enzyme solution with an activity of 0.5 U / mL using 10 U / mg α-glucosidase. Use acarbose as a positive control. Add 50 μL of PBS phosphate buffer to a 96-well plate. Add 40 μL of sample (methyl-3-O-caffeoyl-4-O-galloylquinicacid) and PBS to the enzyme reaction group and blank group, respectively. Add 10 μL of α-glucosidase, mix well, and incubate at 37°C for 5 min. Then add 20 μL of 1 mmol / L PNPG solution and incubate at 37°C for 30 min. Measure the absorbance at 405 nm using a microplate reader.

[0048] According to the formula: α-glucosidase inhibition rate = [1-(A 反应组 -A 反应对照组 ) / (A 空白组 -A 空白对照组 Calculate the enzyme activity inhibition rate by multiplying the result by 100%. Repeat the above experiment three times in parallel, take the average value, and calculate the IC50. 50 value.

[0049] Activity results:

[0050] IC50 of the positive drug acarbose 50 The IC50 of methyl-3-O-caffeoyl-4-O-galloylquinic acid, a quinic acid compound, is 0.0019 mol / L. 50 The concentration was 0.000019 mol / L, showing strong α-glucosidase inhibitory activity, which can be effectively used to prepare hypoglycemic drugs, opening up new pathways for hypoglycemic drugs and new uses for *Castanopsis fargesii*.

[0051] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of different embodiments or examples.

[0052] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present invention.

Claims

1. A method for preparing quinic acid compounds, characterized in that, include: After crushing and extracting the leaves of *Cephalotaxus fortunei*, the filtrate was concentrated, floated in water, defatted, and concentrated again. The concentrated water fraction was then separated sequentially by Sephadex LH-20 gel column chromatography, MCI gel CHP 20P column chromatography, Sephadex LH-20 gel column chromatography, Chromatorex C18 column chromatography, and Diaion HP20SS column chromatography. The eluent for all separations was a mixture of methanol and water. The structural formula of the quinic acid compounds is: 。 2. The method for preparing quinic acid compounds according to claim 1, wherein the eluent for the first Sephadex LH-20 gel column chromatography is a mixture of methanol and water with methanol volume contents of 0%, 20%, 40%, 60%, 80%, and 100% in sequence; The eluent for separation by the MCI gel CHP 20P column chromatography is a mixture of methanol and water with methanol volume contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% respectively. The eluent for the second Sephadex LH-20 gel column chromatography separation was a mixture of methanol and water with methanol volume contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% respectively. The eluent for the Chromatorex C18 column chromatography separation is a mixture of methanol and water with a methanol volume content of 0% to 50%. The eluent for Diaion HP20SS column chromatography is a mixture of methanol and water with a methanol volume content of 0% to 40%.

3. The method for preparing quinic acid compounds according to claim 1, wherein the extraction comprises: Extract with methanol at a volume concentration of 65-75% 2-3 times, each extraction lasting 6-8 days, and combine the filtrates; The reagent used for degreasing is petroleum ether.

4. The use of quinic acid compounds prepared by the preparation method according to any one of claims 1 to 3 in the preparation of hypoglycemic drugs.

5. The application of the quinic acid compounds according to claim 4 in the preparation of hypoglycemic drugs, characterized in that, The hypoglycemic drug includes a pharmaceutically acceptable carrier and the quinic acid compound.

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