Emodin Mannich base as well as preparation method and application thereof

Abstract

The invention relates to the field of medicinal chemistry, and particularly discloses an emodin Mannich base as well as a preparation method and application thereof. The emodin Mannich base has a structure as shown in a formula I. The emodin Mannich base is simple in structure and has relatively strong in-vitro antimicrobial activity; the compound has very high inhibitory activity on bacteria such as staphylococcus aureus, bacillus subtilis, micrococcus luteus, escherichia coli, proteusbacillus vulgaris, pseudomonas aeruginosa, shigella dysenteriae and salmonella typhimurium, and fungi such as candida utilis, candida albicans, saccharomyces cerevisiae and monascus purpureus. The compound can be used for preparing antibacterial and / or antifungal drugs. # imgabs0 #

Description

Technical Field

[0001] The invention belongs to the field of pharmaceutical chemistry, and specifically relates to a rhubarb Mannich base and a preparation method and application thereof. Background Art

[0002] Rhein is an effective component of the Chinese herbal medicine rhubarb. Studies have shown that rhein has multiple biological activities, such as vasodilation, immune suppression, anti-tumor, anti-inflammatory, antioxidant, liver protection, catharsis, diuresis, etc., and thus has good clinical medical value. At present, there are few studies on the antimicrobial activity of rhein and its derivatives, so the present invention aims to provide a rhein Mannich base compound with good antimicrobial activity. Summary of the invention

[0003] In view of this, the object of the present invention is to provide a rhamnosin Mannich base with good antimicrobial activity and a preparation method and application thereof.

[0004] In order to achieve the above object, the present invention provides the following technical solutions:

[0005] According to an embodiment of the present invention, in a first aspect, a rhamnosine Mannich base is provided, having a structure as shown in Formula I:

[0006]

[0007] Wherein, Im is a nitrogen-containing five-membered heteroaryl group.

[0008] It should be noted that nitrogen-containing five-membered heteroaryl groups, such as thiazolyl, oxazolyl, imidazolyl, 1,2,4-triazolyl, tetrazolyl, etc., have a unique aromatic structure containing at least one heteroatom and electron-rich properties, making it easier to interact with enzymes, ion channels, carrier proteins, DNA, RNA, etc. in the body through a variety of non-covalent bonds, so they show low drug resistance. The present invention introduces a nitrogen-containing five-membered heteroaryl group into the parent structure of rheumatoid arthritis to obtain a new rheumatoid arthritis Mannich base with a different mechanism of action from traditional drugs. The present invention has found that the rheumatoid arthritis Mannich base with the structure shown in Formula I exhibits excellent antibacterial and antifungal activity and is expected to be developed into a highly effective antimicrobial drug.

[0009] In an optional embodiment, Im is any one of 1,2,3-triazolyl, 1,2,4-triazolyl, oxazolyl, imidazolyl, pyrazolyl, thiazolyl, tetrazolyl, benzotriazolyl, benzimidazolyl, and carbazolyl.

[0010] According to an embodiment of the present invention, in a second aspect, there is provided a method for preparing the rhamnosin Mannich base described in the first aspect, comprising the following steps:

[0011] In a polar organic solvent, under the action of an acidic catalyst, Im-NH2 reacts with formaldehyde and rhamnosine to obtain a rhamnosine Mannich base having a structure as shown in formula I.

[0012] The preparation method provided by the invention has a simple synthesis route and is easy to operate. The target product can be obtained through a one-step reaction, and the product yield is high and the pollution is small, so the method is suitable for industrial production.

[0013] In an optional embodiment, in a polar organic solvent, under reflux conditions, Im-NH2 is reacted with formaldehyde and an acidic catalyst for 0.5-1 h, and then rheum is added and the reflux reaction is continued for 3-6 h. In this way, rheum Mannich base azole compounds can be quickly prepared.

[0014] It is understandable that the chemical structure of the reaction raw material Im-NH2 can be: 2-aminothiazole, 2-aminooxazole, 2-aminoimidazole, 3-aminotriazole, 5-aminotetrazolyl.

[0015] In an optional embodiment, the molar ratio of the Im-NH2 to the formaldehyde and the emodin is 1-3:1-5:1-1.2, thereby allowing an electrophilic reaction to occur at a relatively high conversion rate to obtain the emodin Mannich base.

[0016] In an optional embodiment, the polar organic solvent is at least one of n-butanol, propanol, ethanol and methanol, thereby allowing the electrophilic iminium ions to have a certain concentration.

[0017] In an optional embodiment, the acidic catalyst is at least one of formic acid, glacial acetic acid, and hydrochloric acid, thereby forming an imine carbonium ion.

[0018] According to an embodiment of the present invention, in a third aspect, there is provided use of the rhamnosin Mannich base described in the first aspect in the preparation of an antimicrobial drug.

[0019] In an alternative embodiment, the microorganism is a bacterium and / or a fungus.

[0020] In an optional embodiment, the bacteria are one or more of Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Escherichia coli, Proteus, Pseudomonas aeruginosa, Shigella dysenteriae or Salmonella typhimurium.

[0021] In an optional embodiment, the fungus is one or more of Candida utilis, Candida albicans, Saccharomyces cerevisiae or Monascus.

[0022] In an optional embodiment, the drug is any one of a tablet, a capsule, an aerosol, and an ointment.

[0023] The beneficial effects of the present invention are as follows: the rhamnosin Mannich base provided by the present invention has a simple chemical structure and strong in vitro antimicrobial activity, especially showing high inhibitory activity against bacteria such as Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Escherichia coli, Proteus, Pseudomonas aeruginosa, Shigella dysenteriae, Salmonella typhimurium, and fungi such as Candida utilis, Candida albicans, Saccharomyces cerevisiae, and Monascus, and can be used to prepare antibacterial and / or antifungal drugs, thereby providing more efficient and safe candidate drugs for clinical antimicrobial treatment, and helping to solve clinical treatment problems such as increasingly serious drug resistance, stubborn pathogenic microorganisms, and newly emerging harmful microorganisms. DETAILED DESCRIPTION

[0024] The technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. In addition, the technical features involved in the different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0025] If no specific experimental steps or conditions are specified in the examples and comparative examples, the conventional experimental steps or conditions described in the literature in the art can be used. If no manufacturer is specified for the reagents or instruments used, they are all conventional reagent products that can be purchased commercially.

[0026] Example 1

[0027] The preparation of rhubarb Mannich base I-1, the reaction formula is as follows:

[0028]

[0029] The specific steps are as follows:

[0030] 0.602 g of 2-aminothiazole, 0.180 g of formaldehyde, 0.045 g of formic acid and 70 mL of n-butanol were added to a 100 mL round-bottom flask. After stirring under reflux for half an hour, 1.350 g of rheum was added and the reaction was continued under reflux for 4 hours. After the reaction was completed, the n-butanol was removed by distillation under reduced pressure, and the residue was extracted with dichloromethane (3×30 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and then dried by spin drying. The mixture was purified by silica gel column chromatography using dichloromethane as an eluent and dried to obtain 0.786 g of rheum Mannich base I-1 as a yellow solid.

[0031] The yield of this example is 41%; melting point 200-202°C; HRMS (TOF) calcd. for C 19 H 14N2O5S[M+H] + ,383.0696;found,383.0706.

[0032] Example 2

[0033] Rheum officinale Mannich base I-2, the reaction formula is as follows:

[0034]

[0035] The specific steps are as follows:

[0036] 0.251 g of 2-aminooxazole, 0.092 g of formaldehyde, 0.023 g of formic acid and 30 mL of n-butanol were added to a 50 mL round-bottom flask. After stirring under reflux for half an hour, 0.676 g of rhamnolide was added and the reaction was continued under reflux for 4 hours. After the reaction was completed, the n-butanol was removed by distillation under reduced pressure, and the residue was extracted with dichloromethane (3×20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and then dried by spin drying. The mixture was purified by silica gel column chromatography using dichloromethane as an eluent and dried to obtain 0.415 g of rhamnolide Mannich base I-2 as a yellow solid.

[0037] The yield of this example is 45%; melting point is 123-125°C; HRMS (TOF) calcd.for C 19 H 14 N2O6[M+H] + ,367.0925;found,367.0930.

[0038] Example 3

[0039] Rheum officinale Mannich base I-3, the reaction formula is as follows:

[0040]

[0041] The specific steps are as follows:

[0042] 0.250 g of 3-aminotriazole, 0.091 g of formaldehyde, 0.024 g of formic acid and 30 mL of n-butanol were added to a 50 mL round-bottom flask. After stirring under reflux for half an hour, 0.675 g of rheum was added and the reaction was continued under reflux for 4 hours. After the reaction was completed, the n-butanol was removed by distillation under reduced pressure, and the residue was extracted with dichloromethane (3×20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and then dried by spin drying. The mixture was purified by silica gel column chromatography using dichloromethane as an eluent and dried to obtain 0.456 g of rheum Mannich base I-3 as a yellow solid.

[0043] The yield of this example is 50%; melting point is 201-203°C; HRMS (TOF) calcd.for C 18 H 14 N4O5[M+H]+ ,367.1037;found,367.1043.

[0044] Example 4

[0045] The preparation of rhubarb Mannich base I-4, the reaction formula is as follows:

[0046]

[0047] The specific steps are as follows:

[0048] 0.250 g of 2-aminoimidazole, 0.090 g of formaldehyde, 0.023 g of formic acid and 30 mL of n-butanol were added to a 50 mL round-bottom flask. After stirring under reflux for half an hour, 0.675 g of rheum was added and the reaction was continued under reflux for 4 hours. After the reaction was completed, the n-butanol was removed by distillation under reduced pressure, and the residue was extracted with dichloromethane (3×20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and then dried by spin drying. The mixture was purified by silica gel column chromatography using dichloromethane as an eluent and dried to obtain 0.405 g of rheum Mannich base I-4 as a yellow solid.

[0049] The yield of this embodiment is 44%; melting point 139-140°C; HRMS (TOF) calcd.for C 19 H 15 N3O5[M+H] + ,366.1085;found,366.1083.

[0050] Example 5

[0051] The preparation of rhubarb Mannich base I-5, the reaction formula is as follows:

[0052]

[0053] The specific steps are as follows:

[0054] 0.250 g of 5-aminotetrazole, 0.091 g of formaldehyde, 0.023 g of formic acid and 30 mL of n-butanol were added to a 50 mL round-bottom flask. After stirring under reflux for half an hour, 0.676 g of rhamnolide was added and the reaction was continued under reflux for 4 hours. After the reaction was completed, the n-butanol was removed by distillation under reduced pressure, and the residue was extracted with dichloromethane (3×20 mL). The organic phases were combined, dried over anhydrous sodium sulfate, and then dried by spin drying. The mixture was purified by silica gel column chromatography using dichloromethane as an eluent and dried to obtain 0.421 g of rhamnolide Mannich base I-5 as a yellow solid.

[0055] The yield of this embodiment is 46%; melting point 212-213°C; HRMS (TOF) calcd. for C 17 H 13 N5O5[M+Na]+ ,390.0809;found,390.0808.

[0056] Comparative Example 1

[0057]

[0058] Reference (Thidarat Chalothorn, Vatcharin Rukachaisirikul, Souwalak Phongpaichit, Sakawrat Pannara, Chittreeya Tansakul, Synthesis and antibacterial activity of emodin and its derivatives against methicillin-resistant Staphylococcus aureus, Tetrahedron Letters, 2019, 60, 151004), the specific synthesis method is as follows:

[0059] At room temperature, dimethylamine aqueous solution (40%, 48 μL, 0.38 mmol) and benzaldehyde (38 μL, 0.38 mmol) were added to a solution of rheumatoid artemisia II (50 mg, 0.19 mmol) in dioxane (2 mL). The reaction system was heated to 65°C for 24 hours. After the reaction was completed, dichloromethane (3×30 mL) was extracted, the organic phases were combined, dried over anhydrous sodium sulfate, and then spin-dried. 60% ethyl acetate / dichloromethane was used as the eluent for silica gel column chromatography purification and drying to obtain 0.057 g of compound III as an orange-yellow solid.

[0060] Comparative Example 2

[0061]

[0062] Reference (Thidarat Chalothorn, Vatcharin Rukachaisirikul, Souwalak Phongpaichit, Sakawrat Pannara, Chittreeya Tansakul, Synthesis of amino derivatives of 1,6,8-trihydroxy-3-methyl-9,10-anthraquinone, Chemistry of Natural Compounds, 1999, 35 (6), 619-620), the specific synthesis method is as follows:

[0063] After cooling morpholine (0.6 mL, 0.006 mol) to 0°C, add formaldehyde aqueous solution (40%, 0.23 g, 0.008 mol) dropwise, and finally add rhein II (0.45 g, 0.002 mol), and stir at room temperature for one hour. After adding 10 mL of water to the reaction system, filter, collect the filtrate, and add NaOH (10%) dropwise under stirring to adjust the pH to 9. Filter, collect the filter cake, wash with water, and dry to obtain 0.35 g of compound IV.

[0064] Experimental Example 6

[0065] The in vitro antimicrobial activity of the emodin Mannich bases prepared in the examples and comparative examples was tested using the 96-well microdilution method recommended by the Clinical Laboratory Standards Institute (CLSI) of the United States, and the minimum inhibitory concentration (MIC) of these compounds against bacteria such as Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Escherichia coli, Proteus, Pseudomonas aeruginosa, Shigella dysenteriae, and Salmonella typhimurium, as well as Candida utilis, Candida albicans, Saccharomyces cerevisiae, and Monascus was detected.

[0066] The specific test method is: the test compound and the reference drug are dissolved in dimethyl sulfoxide (DMSO) to prepare 6.4 mg mL -1 Solution. Take a sterile 96-well plate and add 100 μL of bacterial solution to each well. Take 100 μL of the test solution diluted with sterile broth to well 1, mix well and take 100 μL to well 2, and dilute to well 11 in multiples, so that the DMSO content in each well is less than 1%, and the effect of solvent DMSO on the in vitro antibacterial and antifungal activity test is eliminated. Well 12 does not contain drugs and serves as a positive control. Each drug sensitivity plate is cultured on a constant temperature shaker at 37°C for 24 hours. The results are shown in Tables 1 and 2.

[0067] Table 1 Antibacterial activity (MIC, μmol / mL)

[0068]

[0069]

[0070] Table 2 Antifungal activity (MIC, μmol / mL)

[0071] Compound Candida utilis Aspergillus flavus Brewer's Yeast Candida albicans Candida I-1 0.021 0.042 0.042 0.010 0.010 I-2 0.022 0.022 0.022 0.011 0.044 I-3 0.011 0.044 0.011 0.022 0.011 I-4 0.022 0.044 0.011 0.011 0.022 I-5 0.022 0.011 0.022 0.022 0.022 II 0.948 0.474 0.474 0.474 0.474 III 0.159 0.317 0.635 0.635 0.635 IV 0.693 0.347 0.173 0.347 0.173 Fluconazole 0.026 0.837 0.013 0.013 0.013

[0072] It can be seen from Table 1 that rhamnosin Mannich bases I-1 to I-5 all showed good inhibitory effects on the tested bacteria, and Table 2 shows that rhamnosin Mannich bases I-1 to I-5 all showed certain inhibitory effects on the tested fungi, which shows that the rhamnosin Mannich base azoles provided by the present invention have strong antimicrobial activity and can be used to prepare antibacterial and / or antifungal drugs.

[0073] Example 7

[0074] Tablet preparation

[0075] Prescription: 100g of rhubarb Mannich base I-1, 40g of starch, 80g of microcrystalline cellulose, 3.0g of magnesium stearate, and an appropriate amount of hydroxypropyl methylcellulose E-30 (40% solution by mass), making a total of 4000 tablets.

[0076] Preparation method: prepare a 4% hydroxypropyl methylcellulose E-30 solution and set it aside; weigh 20g of starch, dry it at 105℃ for 5 hours to obtain dry starch and set it aside; weigh 20g of starch and the prescribed amount of rhamnosyl thiazolinone compound I-1 and microcrystalline cellulose, mix them evenly, grind them through an 80-mesh sieve, use a 4% hydroxypropyl methylcellulose E-30 solution to make a soft material, granulate it with a 20-mesh sieve, dry it at 50-60℃ to about 3% moisture, pass it through a 20-mesh sieve to size the particles, add 20g of dry starch and the prescribed amount of magnesium stearate to the dry particles, mix them evenly, and press them into tablets to obtain the product.

[0077] Usage and Dosage: According to the patient's condition and individual differences, the recommended daily dosage is 3-6 tablets, equivalent to 0.075-0.15g of large Mannich base I-1 / 60kg body weight / day, taken 3 times after meals, drink a full glass of water while taking; or follow the doctor's advice.

[0078] Implementation 8

[0079] Preparation of capsules

[0080] Prescription: Rhamnosin Mannich base I-2 100g, modified starch (120 mesh) 50g, microcrystalline cellulose (100 mesh) 30g, low-substituted hydroxypropyl cellulose (100 mesh) 10g, talcum powder (100 mesh) 10g, sweetener 5g, orange flavor 1g, appropriate amount of pigment, appropriate amount of water, make 4000 tablets.

[0081] Preparation method: After the prescribed amount of rhubarb Mannich base I-2 is micronized and crushed into extremely fine powder, it is mixed with the prescribed amount of modified starch, microcrystalline cellulose, low-substituted hydroxypropyl cellulose, talcum powder, sweetener, orange flavor and pigment, and water is used to make a soft material, 12-14 mesh sieve is used for granulation, 40-50℃ is dried, sieved and whole granules are put into empty capsules to obtain the product.

[0082] Usage and Dosage: According to the patient's condition and individual differences, the recommended daily dosage is 3 to 6 capsules, equivalent to 0.075-0.15g of rhamnosyl compound I-2 / 60kg body weight / day, taken three times after meals, and drink a full glass of water while taking; or follow the doctor's advice.

[0083] Example 9

[0084] Aerosol preparation

[0085] Prescription: Rheum officinale Mannich base I-3 2.5g, Span20 3g, talcum powder (100 mesh) 4g, trichloromonofluoromethane added to appropriate amount.

[0086] Preparation method: Place rhamnosin Mannich base I-3, Span 20 and talcum powder in a vacuum drying oven for several hours, cool them to room temperature in a dryer, grind them into fine powder using a jet mill, mix them according to the prescribed amount, pour them into a sealed container, and add trichloromonofluoromethane to the specified amount.

[0087] Usage and Dosage: Depending on the patient's condition and individual differences, it is recommended to use it 3-4 times a day.

[0088] Example 10

[0089] Preparation of ointment

[0090] Prescription: Rheum officinale Mannich base I-4 2g, stearic acid 12g, vaseline 3g, monoglyceride 3g, ethyl paraben 0.5g, distilled water 60mL, borax 1g, potassium hydroxide 0.5g, potassium sorbate 0.3g, glycerol 0.5g, liquid paraffin 0.5g, a total of 85g.

[0091] Preparation method: Take the prescribed amount of stearic acid, vaseline, monoglyceride and ethylparaben, heat and melt, pass through a 100-mesh sieve, and keep warm at 80°C as the oil phase for later use; take the prescribed amount of distilled water, borax, potassium hydroxide, potassium sorbate, glycerin and liquid paraffin, heat and boil as the water phase; cool the water phase to 85°C, add the oil phase under constant stirring, emulsify, then add the prescribed amount of rhubarb Mannich base I-4, cool and stir to obtain the product.

[0092] Usage and Dosage: Apply to the affected area and rub repeatedly until the skin becomes warm, twice a day.

[0093] Embodiment 11

[0094] Preparation of liniment

[0095] Prescription: Rheum officinale Mannich base I-5 4g, potassium soap 7.5g, camphor 5g, distilled water added to 100mL.

[0096] Preparation method: Heat potassium soap to liquefy it and set it aside; dissolve camphor in 95% ethanol and set it aside; take the prescribed amount of rhubarb Mannich I-5, add potassium soap solution under constant stirring, then add camphor ethanol solution, gradually add water, and add distilled water to the full amount after emulsification is complete.

[0097] Usage and Dosage: Apply 1 to 2 drops to the undamaged affected area, or use a cotton swab to apply an appropriate amount of the solution to the affected area, twice a day.

[0098] Obviously, the above embodiments are merely examples for the purpose of clear explanation, and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived therefrom are still within the scope of protection of the invention.

Claims

1. Rheum officinale Mannich base, having the structure shown in Formula I: in, Im is a nitrogen-containing five-membered heteroaryl group.

2. The rhamnosine Mannich base according to claim 1, characterized in that: Im is any one of 1,2,3-triazolyl, 1,2,4-triazolyl, oxazolyl, imidazolyl, pyrazolyl, thiazolyl, tetrazolyl, benzotriazolyl, benzimidazolyl and carbazolyl.

3. The method for preparing the rhamnosin Mannich base according to claim 1 or 2, characterized in that: The steps include: In a polar organic solvent, under the action of an acidic catalyst, Im-NH2 reacts with formaldehyde and rhamnosine to obtain a rhamnosine Mannich base having a structure as shown in formula I.

4. The method for preparing the rhamnosin Mannich base according to claim 3, characterized in that: In a polar organic solvent, under reflux conditions, Im-NH2 reacts with formaldehyde and an acidic catalyst for 0.5-1h, and then rhein is added and the reflux reaction is continued for 3-6h.

5. The method for preparing the rhamnosin Mannich base according to claim 3 or 4, characterized in that: The molar ratio of the Im-NH2 to the formaldehyde and the rhamnolide is 1-3:1-5:1-1.

2.

6. The method for preparing the rhamnosin Mannich base according to claim 3 or 4, characterized in that: The polar organic solvent is at least one of n-butanol, propanol, ethanol and methanol.

7. The method for preparing the rhamnosin Mannich base according to claim 3 or 4, characterized in that: The acidic catalyst is at least one of formic acid, glacial acetic acid and hydrochloric acid.

8. Use of the rhamnosin Mannich base according to any one of claims 1 to 2 in the preparation of antimicrobial drugs.

9. The use of the rhamnosin Mannich base in the preparation of antimicrobial drugs according to claim 8, characterized in that: The microorganisms are bacteria and / or fungi; And / or, the medicine is any one of tablets, capsules, aerosols, and ointments.

10. Use of the rhamnosin Mannich base according to claim 8 or 9 in the preparation of antimicrobial drugs, characterized in that: The bacteria are one or more of Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Escherichia coli, Proteus, Pseudomonas aeruginosa, Shigella dysenteriae or Salmonella typhimurium; The fungus is one or more of Candida utilis, Candida albicans, Saccharomyces cerevisiae or Monascus.

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