Baohuoside I carbamate prodrug as well as salt, preparation and application thereof

Abstract

The invention provides baohuoside I carbamate prodrugs and salts thereof as well as preparation and application thereof, belongs to the technical field of medicines, and discloses carbamate prodrugs formed between natural isoprene flavone baohuoside I and amino acid and salts thereof, and pharmaceutical salts thereof comprise potassium salts, sodium salts and amine salts (ammonium salts). According to the invention, amino acid methyl ester is used as a modification group and is covalently bound with a phenolic hydroxyl group on the 7th site in a baohuoside I structure to form the baohuoside I mono-substituted carbamate prodrug. The prodrug can significantly improve the water solubility and fat solubility of baohuoside I, compared with a baohuoside I bulk drug, the oral bioavailability of the monosubstituted prodrug is significantly improved, clinical medication can effectively reduce the administration frequency and improve patient compliance, and an effective strategy is expected to be provided for improvement of the oral bioavailability of phenolic drugs.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and specifically relates to a prodrug of baohuoside I carbamate and a salt thereof, and a preparation and application thereof. Background Art

[0002] Baohuoside Ⅰ (BI), also known as icariside Ⅱ (ICS-Ⅱ), is an active flavonoid compound derived from Epimedium, which has multiple biological and pharmacological effects such as anti-inflammatory, anti-osteoporosis, antioxidant, anti-aging and anti-cancer. Baohuoside Ⅰ contains two phenolic hydroxyl groups in its structure, of which the 5-hydroxyl group forms an intramolecular hydrogen bond to stabilize, and the 7-hydroxyl group can form an intermolecular hydrogen bond, so Baohuoside Ⅰ has poor water solubility and fat solubility. Phenolic hydroxyl groups are prone to II-binding reactions in the intestine and liver, and there is an intestinal first-pass effect, so the low oral bioavailability (absolute bioavailability is about 4.1%) limits its drug research and development. The structure of Baohuoside Ⅰ is as follows:

[0003]

[0004] Most of the existing technologies use preparation methods to promote the absorption of Baohuoside I, but a large amount of carriers need to be introduced, and the drug loading is low; there are also methods to synthesize Baohuoside I derivatives by structural modification, but the newly synthesized derivatives need to undergo long preclinical and clinical studies before they can be launched as new drugs, which is time-consuming and laborious. If we start from the molecular structure characteristics of Baohuoside I itself, chemically modify it with amino acid esters to form carbamate prodrugs, thereby improving its water solubility, membrane permeability and metabolic stability, it will be an economically feasible and effective strategy to improve the bioavailability of Baohuoside I, but there is no report on Baohuoside I prodrugs modified with amino acid methyl esters. Summary of the invention

[0005] In view of the problems existing in the prior art, the present invention proposes a prodrug of Baohuoside I carbamate and its salt, as well as preparation and application. The present invention provides a prodrug of Baohuoside I carbamate and its salt, and also provides application of the prodrug of Baohuoside I carbamate in improving the water solubility, lipid solubility and bioavailability of Baohuoside I.

[0006] The Baohuoglycoside I carbamate prodrug is obtained by modifying the 7-position phenolic hydroxyl group of Baohuoglycoside I with amino acid methyl ester;

[0007] The amino acid methyl ester is one of glycine methyl ester, alanine methyl ester, valine methyl ester, leucine methyl ester, isoleucine methyl ester or phenylalanine methyl ester;

[0008] The amino acid methyl ester is preferably an L-configuration amino acid methyl ester, and the L-configuration amino acid methyl ester includes one of L-valine methyl ester, L-leucine methyl ester or L-isoleucine methyl ester, and the structural formula is shown below:

[0009]

[0010] The general structural formula of the Baohuoglycoside I carbamate prodrug is:

[0011]

[0012] Wherein, R is the side chain of amino acid methyl ester;

[0013] The synthetic route of the Baohuoglycoside I carbamate prodrug is as follows:

[0014]

[0015] The preparation of the prodrug of amino acid methyl ester of Baohuoside I and its salt of the present invention comprises the following steps:

[0016] Step 1: dissolving compound I and a carbonylation agent in a reaction solvent, and then adding a basic catalyst dropwise to react to obtain compound II;

[0017] Step 2: dissolving Baohuo glycoside I in an aprotic solvent, and then dropping the compound II in step 1 to react to obtain a reaction solution; diluting the reaction solution with distilled water, extracting it three times with dichloromethane, washing it with distilled water and a saturated sodium chloride solution in turn, and filtering it with suction; the filtered solution is concentrated by vacuum decompression on a rotary evaporator to dryness, and then redissolved with methanol, and purified by liquid phase separation to obtain compound III;

[0018] Step 3: dissolving compound III in a base to obtain a salt prodrug;

[0019] The carbonylation reagent in step 1 includes one of dimethyl carbonate, 4-nitrophenyl carbonate or phosgene;

[0020] The reaction solvent in step 1 includes one of dichloromethane, N,N-dimethylformamide, tetrahydrofuran, dioxane, and N,N-dimethylhexanamide;

[0021] The basic catalyst in step 1 includes one of triethylamine, potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide or pyridine;

[0022] In step 1, the molar ratio of compound I, carbonylation reagent and reaction solvent is: 1: (0.1-10): (0.1-10);

[0023] The ratio of the compound I and the alkaline catalyst in step 1 is: 1: (0.1-10) mol / ml;

[0024] The alkaline catalyst is added dropwise in step 1 under a nitrogen atmosphere at a temperature of -20°C to 0°C, and the reaction is carried out at room temperature for 1 to 10 hours after the addition is completed;

[0025] The aprotic solvent in step 2 includes one of dimethyl sulfoxide, N,N-dimethylformamide or N-methylpyrrolidone;

[0026] The molar ratio of the baohuoside I and the aprotic solvent in step 2 is 1:(0.1-10);

[0027] In step 2, the ratio of Baohuo glycoside I to compound II is 1: (0.1-10) mol / ml;

[0028] In step 2, the compound II is added dropwise under a nitrogen atmosphere at room temperature; the reaction is carried out at room temperature for 1 to 10 hours after the addition is completed;

[0029] In step 2, the reaction solution is diluted with distilled water, and the volume ratio of the reaction solution to distilled water is 1: (1-100);

[0030] The washing in step 2 is specifically performed by first washing with distilled water for 1 to 5 times, and then washing with a saturated sodium chloride solution for 1 to 5 times;

[0031] The pressure of the vacuum decompression in step 2 is 10 to 30 mmHg;

[0032] The solid material described in step 2 is redissolved in methanol, and the ratio of the solid material to methanol is 1: (1-100) mg / ml;

[0033] The base in step 3 comprises one of potassium hydroxide, potassium bicarbonate, potassium carbonate, sodium hydroxide, sodium bicarbonate, sodium carbonate, aqueous ammonia, diethylamine or triethylamine;

[0034] The molar ratio of compound III to the base in step 3 is 1:(0.1-10);

[0035] The carbamate prodrug of baohuoside I and its salt can obviously improve the solubility and fat solubility of baohuoside I, and can improve the bioavailability of baohuoside I in vivo.

[0036] The beneficial effects of the present invention are:

[0037] The amino acid methyl ester-modified monosubstituted carbamate prodrug of baohuoside I synthesized by the present invention can significantly improve the water solubility and fat solubility of baohuoside I. Taking baohuoside I as a reference, when 7-isoleucine baohuoside I is orally administered, the relative bioavailability of baohuoside I in rats is 197.53%. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 The blood concentration-time curves of Baohuoside I carbamate prodrug and Baohuoside I (n=3).

[0039] Reference numerals: Oral administration of Baohuo glycoside I (30 mg / kg), It is an oral prodrug 7-isoleucyl baohuoside I (30 mg / kg based on baohuoside I). It is an oral prodrug 7-isoleucyl baohuoside I and an inhibitor Gly-Sar (30 mg / kg based on baohuoside I). DETAILED DESCRIPTION

[0040] The following specific examples are used to further illustrate the present invention, but are by no means intended to limit the scope thereof.

[0041] According to the general synthetic route of Baohuo glycoside I carbamate prodrug, the following compound was prepared, and the synthetic route is as follows:

[0042]

[0043] Example 1

[0044] The first step: dissolve L-isoleucine methyl ester hydrochloride and phosgene in dichloromethane, fill with nitrogen, control the dropping temperature to -20°C-0°C, and add triethylamine dropwise to obtain compound II.

[0045] Step 2: Dissolve Baohuoside I in N,N-dimethylformamide, add dropwise the reaction solution containing compound II, fill with nitrogen, react at room temperature for 2 hours, dilute the reaction solution with water, extract three times with dichloromethane, wash with aqueous solution and saturated brine in turn, filter, concentrate the filtrate under vacuum, dissolve in methanol, and separate and purify by preparative liquid phase to obtain the final product, L-isoleucine methyl ester-modified Baohuoside I carbamate prodrug.

[0046] Step 3: dissolve the final product, L-isoleucine methyl ester-modified baohuoside I carbamate prodrug, in sodium bicarbonate to obtain its sodium salt.

[0047] Example 2

[0048] The first step: dissolve L-valine methyl ester hydrochloride and phosgene in dichloromethane, fill with nitrogen, control the dropping temperature to -20°C-0°C, and add triethylamine dropwise to obtain compound II.

[0049] Step 2: Dissolve Baohuoside I in N,N-dimethylformamide, add dropwise the reaction solution containing compound II, fill with nitrogen, react at room temperature for 2 hours, dilute the reaction solution with water, extract three times with dichloromethane, wash with aqueous solution and saturated brine in turn, filter, concentrate the filtrate under vacuum, dissolve in methanol, and separate and purify by preparative liquid phase to obtain the final product, L-valine methyl ester-modified Baohuoside I carbamate prodrug.

[0050] Step 3: dissolve the final product, L-valine methyl ester-modified baohuoside I carbamate prodrug, in sodium bicarbonate to obtain its sodium salt.

[0051] Example 3

[0052] The first step: dissolve L-leucine methyl ester hydrochloride and phosgene in dichloromethane, fill with nitrogen, control the dropping temperature to -20°C-0°C, and add triethylamine dropwise to obtain compound II.

[0053] Step 2: Dissolve Baohuoside I in N,N-dimethylformamide, add the reaction solution containing compound II dropwise, fill with nitrogen, react at room temperature for 2 hours, dilute the reaction solution with water, extract three times with dichloromethane, wash with aqueous solution and saturated brine in turn, filter, concentrate the filtrate under vacuum, dissolve in methanol, and separate and purify by preparative liquid phase to obtain the final product, L-leucine methyl ester-modified Baohuoside I carbamate prodrug.

[0054] Step 3: dissolve the final product, L-leucine methyl ester-modified baohuoside I carbamate prodrug, in sodium bicarbonate to obtain its sodium salt.

[0055] The identification information of Baohuoside I and L-isoleucine methyl ester, L-valine methyl ester, and L-leucine methyl ester modified Baohuoside I carbamate prodrug compounds is shown in Table 1:

[0056] Table 1. Relevant information of Baohuoglycoside I and its prodrugs

[0057]

[0058] Example 4

[0059] Determination of the Oil-Water Partition Coefficient of the Carbamate Prodrug of Baohuoside Ⅰ

[0060] The poor membrane permeability of Baohuoside I is also an important reason that limits its bioavailability, and an appropriate oil-water partition coefficient is conducive to the drug's penetration through the lipid biological membrane. Moreover, determining the oil-water partition coefficient of a drug helps predict the dissolution, absorption, distribution, and transport of the drug in the body. The oil-water partition coefficient is generally expressed as Log P, where P represents the concentration ratio of the drug in the oil-water phase. The solubility parameter of n-octanol is similar to that of the cell lipid membrane, so the present invention selects the n-octanol-water system to determine the oil-water partition coefficient of the drug. First, prepare a water-saturated n-octanol solution and an n-octanol-saturated aqueous solution, and seal them for standby use. Baohuoside I and its prodrug are completely dissolved in a water-saturated n-octanol solution, respectively, and shaken at a constant temperature to equilibrium. Take a certain volume of the saturated solution and mix it with an equal volume of water saturated with n-octanol, shake it at a constant temperature to equilibrium, and obtain the concentration ratio of Baohuoside I and its prodrug in the oil-water phase at equilibrium, and calculate the oil-water partition coefficient.

[0061] The results show that compared with Baohuoside I, the solubility of 7-isoleucyl Baohuoside I, 7-valyl Baohuoside I and 7-leucyl Baohuoside I in water increased by 2-6 times, and the solubility in n-octanol increased by 30-50 times, among which 7-isoleucyl Baohuoside I had the greatest improvement in solubility in water and n-octanol. At the same time, compared with Baohuoside I, the oil-water partition coefficient of the prodrug was also improved.

[0062] Table 3. Water solubility of baohuoside I and its prodrugs

[0063]

[0064] Example 5

[0065] Pharmacokinetic study in rats

[0066] After Baohuoside I was modified with isoleucine methyl ester to form carbamate, its water solubility and membrane permeability were enhanced, and after the phenolic hydroxyl group was protected, the phase II metabolites should also be reduced. Next, we conducted in vivo pharmacokinetics studies on the isoleucine carbamate prodrug of Baohuoside I.

[0067] Sprague-Dawley rats in the experimental and control groups were gavaged with sodium carboxymethylcellulose (CMC) suspension of carbamate prodrug (7-I) of baohuoside I, CMC suspension of prodrug (7-I) with added inhibitor of Gly-Sar, and CMC suspension of baohuoside I (30 mg / kg for each baohuoside I), and the concentration of baohuoside I in plasma was measured. The results are shown in Tables 4 and Figure 1 .

[0068] Table 4. Pharmacokinetic parameters of Baohuoside I in rats after oral administration of Baohuoside I and prodrug (calculated as Baohuoside I 30 mg / kg)

[0069]

[0070]

[0071] *p<0.05 and**p<0.01 versus BⅠas control。

Claims

1. A prodrug of Baohuo glycoside I carbamate, characterized in that: The Baohuoglycoside I carbamate prodrug is obtained by modifying the 7-position phenolic hydroxyl group of Baohuoglycoside I with amino acid methyl ester; The general structural formula of the Baohuoglycoside I carbamate prodrug is: Wherein, R is the side chain of amino acid methyl ester.

2. The carbamate prodrug of Baohuoside I according to claim 1, characterized in that The amino acid methyl ester includes one of glycine methyl ester, alanine methyl ester, valine methyl ester, leucine methyl ester, isoleucine methyl ester or phenylalanine methyl ester.

3. The carbamate prodrug of Baohuoside I according to claim 1, characterized in that The amino acid methyl ester is preferably an L-configuration amino acid methyl ester, and the L-configuration amino acid methyl ester includes one of L-valine methyl ester, L-leucine methyl ester or L-isoleucine methyl ester, and the structural formula is shown below:

4. The carbamate prodrug of Baohuoside I according to claim 1, characterized in that The synthetic route of the Baohuoglycoside I carbamate prodrug is as follows:

5. The preparation of the prodrug of the carbamate of baohuoside I and its salt according to any one of claims 1 to 4, characterized in that: The following steps are involved: Step 1: dissolving compound I and a carbonylation agent in a reaction solvent, and then adding a basic catalyst dropwise to react to obtain compound II; Step 2: dissolving Baohuo glycoside I in an aprotic solvent, and then dropping the compound II in step 1 to react to obtain a reaction solution; diluting the reaction solution with distilled water, extracting with dichloromethane, washing with distilled water and saturated sodium chloride solution in sequence, and filtering; the filtered solution is concentrated by vacuum decompression on a rotary evaporator to dryness, and then redissolved with methanol, and purified by liquid phase separation to obtain compound III; Step 3: Dissolve compound III in a base to obtain a salt.

6. The preparation of the prodrug of carbamate of baohuoside I and its salt according to claim 5, characterized in that: In step 1, the carbonylation reagent includes one of dimethyl carbonate, 4-nitrophenyl carbonate or phosgene; the reaction solvent includes one of dichloromethane, N,N-dimethylformamide, tetrahydrofuran, dioxane, N,N-dimethylhexanamide; the alkaline catalyst includes one of triethylamine, potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide or pyridine; The molar ratio of the compound I, the carbonylation reagent and the reaction solvent is: 1: (0.1-10): (0.1-10); the ratio of the compound I and the alkaline catalyst is: 1: (0.1-10) mol / ml; the alkaline catalyst is added under a nitrogen atmosphere at a temperature of -20°C-0°C, and the reaction is carried out at room temperature for 1-10 hours after the addition is completed.

7. The preparation of the prodrug of carbamate of Baohuoside I and its salt according to claim 5, characterized in that: The aprotic solvent in step 2 includes one of dimethyl sulfoxide, N,N-dimethylformamide or N-methylpyrrolidone; the molar ratio of the Baohuo glycoside I and the aprotic solvent in step 2 is 1: (0.1-10); the ratio of the Baohuo glycoside I to the compound II is 1: (0.1-10) mol / ml; the dropwise addition of the compound II is carried out in a nitrogen atmosphere at room temperature; the reaction is carried out at room temperature for 1-10 hours after the dropwise addition is completed; the reaction solution is diluted with distilled water, and the volume ratio of the reaction solution to distilled water is 1: (1-100); the washing is specifically, first washing with distilled water 1-5 times, and then washing with a saturated sodium chloride solution 1-5 times; the pressure of the vacuum decompression is 10-30 mmHg; the solid is redissolved with methanol, and the ratio of the solid to methanol is 1: (1-100) mg / ml.

8. The preparation of the prodrug of carbamate of baohuoside I and its salt according to claim 5, characterized in that: The base in step 3 includes one of potassium hydroxide, potassium bicarbonate, potassium carbonate, sodium hydroxide, sodium bicarbonate, sodium carbonate, aqueous ammonia, diethylamine or triethylamine; the molar ratio of compound III to the base is 1:(0.1-10).

9. The preparation of the prodrug of carbamate of Baohuoside I and its salt according to claim 5, characterized in that: The salts described in step 3 are pharmaceutically acceptable salts, including potassium salts, sodium salts or ammonium salts.

10. A pharmaceutical composition, characterized in that The invention comprises the prodrug of the carbamate of Baohuoside I as described in any one of claims 1 to 4 and a pharmaceutically acceptable salt thereof.

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