CN110981888B - N-aryl dithiopyrryl ketonuria and amino ester derivatives, preparation and application thereof - Google Patents

N-aryl dithiopyrryl ketonuria and amino ester derivatives, preparation and application thereof Download PDF

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CN110981888B
CN110981888B CN201911186804.4A CN201911186804A CN110981888B CN 110981888 B CN110981888 B CN 110981888B CN 201911186804 A CN201911186804 A CN 201911186804A CN 110981888 B CN110981888 B CN 110981888B
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谭相端
吕良
蒙杰雲
周异欢
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Shenzhen Hepalink Pharmaceutical Group Co Ltd
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Guilin Medical University
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Abstract

The invention discloses a series of N-aryl dithiopyrryl ketonuria and amino ester derivatives, and preparation and application thereof, in particular to a derivative shown as a formula (I), a preparation method and application thereof,
Figure DEST_PATH_IMAGE001
wherein: r is selected from the following unsubstituted or optionally substituted groups: C1-C10 alkyl, C2-C10 alkenyl, C3-C10 cycloalkyl, C1-C10 alkyl with C5-C10 aryl, C5-C10 aryl, or 3-10 membered heterocyclyl containing 1-3 heteroatoms independently selected from N, O or S; x is selected from NH and O. The preparation method comprises the steps of preparing the compound by using 6-amino-4- (2, 4-dimethoxyphenyl) - [1,2]Disulfide [4,3-b]Pyrrole-5 (4H) -ketone hydrochloride is used as a raw material, and N-aryl dithiopyrryl ketonuria and amino ester derivatives are prepared through multi-step chemical reactions. In addition, the derivative shown in the formula (I) or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof is used as a bacterial RNA polymerase inhibitor and is used for developing antibacterial drugs.

Description

N-aryl dithiopyrryl ketonuria and amino ester derivatives, preparation and application thereof
Technical Field
The invention belongs to the field of new drug design and synthesis, and particularly relates to N-aryl dithiopyrryl ketonuria and amino ester derivatives, a preparation method thereof and application of the derivatives as a bacterial RNA polymerase inhibitor.
Background
Since the 60 s of the 20 th century, the problem of drug resistance caused by pathogenic bacteria has become a worldwide problem, and particularly, resistant Acinetobacter baumannii (Acinetobacter baumannii), Escherichia coli (ESB L-producing Escherichia coli) producing extended spectrum β -lactamase, Pseudomonas aeruginosa (Pseudomonas aeruginosa) and Enterococcus faecalis (Enterococcus faecalis), methicillin-resistant Staphylococcus aureus (MRSA), etc. have caused an increasingly serious threat to human health
Figure BDA0002292580890000011
Oxazolidinone and lipopeptide drugs are on the market, but these drugs still do not meet the current rapidly expanding demand for resistance to drug-resistant bacterial infections. Therefore, development of novel antibacterial agents has become urgent.
Bacterial RNA polymerase (RNAP) is a protein complex composed of a plurality of subunits, participates in the synthesis of almost all mRNA, rRNA and tRNA in a bacterium body, and is one of important targets for screening broad-spectrum antibacterial drugs. Based on the structure and transcriptional process of RNAP, the following 3 mechanisms for inhibiting bacterial RNAP can be attributed: (1) the active substance binds near the RNAP active center, preventing the formation of RNA primary products; (2) affecting the RNAP active center, overlapping two subunits of the active center, and inhibiting the conformation transformation of the RNAP active center; (3) binding at sites distant from the RNAP active center interferes with the opening of the RNAP active space to prevent DNA entry and unwinding, or with the binding of RNAP to the DNA template strand.
Myxopyrronins, a polyketide containing α -pyranone structure, isolated from fermentation broth of Myxopyrronins (Myxococcus fulvus) Mx f50 by Irschik et al in 1983, contains two structural analogues, Myxopyrronin A and Myxopyrronin B, which have a narrow inhibitory spectrum but are more active, inhibiting part of gram-positive and gram-negative bacteria, and no inhibitory activity against yeast and fungi.
Dithiopyrrolone (DTP) antibiotics are a generic term for a class of compounds having a unique pyrrolone dithiole (4H- [1,2] dithio [4,3-b ] pyrrol-5-one) backbone. Based on the difference of the side chain of the C-6 amido group and whether the N-4 position contains methyl, the N-methyl-N-acetylthionine, N-acetylthionine and thiomarinols can be classified. To date, nearly 30 DTP compounds have been reported, including holomycin (holomycin), thiolutin (thiolutin), aureomycin (aureothricin), and thiomarinols, among others. The marketed anti-tuberculosis drug rifamycin inhibits the activity of RNAP by binding near the RNAP active center, preventing the formation of RNA initial products, while there is a literature (Mar. drugs,2013,11,3970-3997) that natural DTP derivatives may inhibit the activity of RNAP by preventing the elongation of bacterial RNAP chain. In view of this, DTP-based compounds are considered to be one of the hot-gate framework structures for the design of next-generation RNAP inhibitors.
There are many documents reporting that N-aryl dithiopyrrolidone derivatives have antitumor, leukocyte increasing, etc. activities, for example: CN102219724A pyrrolidone compounds and a preparation method and application thereof, CN101522688B dithiolopyrrolone compounds and therapeutic application thereof, CN1642959 novel dithiolopyrrolone compounds with therapeutic effect, and CN1276723 dithiolopyrrolone used as antitumor agents and corresponding monoxide and dioxide thereof, but no literature and patent report that N-aryl dithiolopyrrolone derivatives have bacterial RNA polymerase inhibition activity. In the previous research, the N-aryl dithiopyrrone-pyrone hybrid derivative is found to have good antibacterial activity and can inhibit the action of bacterial RNAP, and CN109535174A is applied for the N-aryl dithiopyrrone-pyrone hybrid derivative and the preparation method and the application thereof. The invention discloses a series of N-aryl dithiopyrryl ketonuria and amino ester derivatives, and in-vitro antibacterial experiments, in-vitro bacterial RNAP inhibitory activity researches and in-vivo antibacterial experiments are carried out to prove that the N-aryl dithiopyrryl ketonuria and the amino ester derivatives have the function of inhibiting bacterial RNAP.
Disclosure of Invention
In order to develop resources of clinical medicines, the invention provides N-aryl dithiopyrrone urea and amino ester derivatives, and a preparation method and application thereof, 6-amino-4- (2, 4-dimethoxyphenyl) - [1,2] dithio [4,3-b ] pyrrole-5 (4H) -ketone hydrochloride containing 2, 4-dimethoxyphenyl at N-4 position is selected as a raw material, a series of N-aryl dithiopyrrone urea and amino ester derivatives are obtained by extending a side chain on C-6 position amino and carrying out amidation reaction with corresponding acyl chloride, and thus, the N-aryl dithiopyrrone urea and amino ester bacterial RNA polymerase inhibitor is provided and is used for developing antibacterial medicines.
The first aspect of the present invention provides a derivative represented by the following formula (I),
Figure BDA0002292580890000021
wherein R is selected from the following unsubstituted or optionally substituted groups: C1-C10 alkyl, C2-C10 alkenyl, C3-C10 cycloalkyl, C1-C10 alkyl with C5-C10 aryl, C5-C10 aryl, or 3-10 membered heterocyclyl containing 1-3 heteroatoms independently selected from N, O or S; x is selected from NH and O. In addition, the derivative shown in the formula (I) or the pharmaceutically acceptable salt thereof has antibacterial pharmacological action and is a bacterial RNA polymerase inhibitor.
In a preferred embodiment, the derivative of formula (I) wherein X is NH or O and R is selected from methyl, ethyl, cyclopropyl, furyl, phenyl, pyridyl, 4-fluorophenyl and 4-cyanophenyl.
The derivative shown in the formula (I) is easy to separate and can be purified by a conventional separation method; for example, the derivatives of formula (I) may be purified by column chromatography.
In a first aspect of the invention, more preferred inventive compounds are:
Figure BDA0002292580890000031
Figure BDA0002292580890000041
in a second aspect of the invention, a preparation method and a preparation route of the derivative shown in the formula (I) are provided, wherein the preparation route is as follows:
Figure BDA0002292580890000042
the preparation method comprises the following steps:
reacting a compound 6-amino-4- (2, 4-dimethoxyphenyl) - [1,2] dithio [4,3-b ] pyrrole-5 (4H) -one hydrochloride shown in a formula VI, an organic base and phenyl chloroformate in an organic solvent to obtain an intermediate V, and reacting the intermediate V with ethanolamine to obtain a compound IV;
in an organic solvent, reacting the compound VI with triphosgene to obtain an intermediate III, and reacting the intermediate III with ethylene glycol to obtain a compound II;
and respectively carrying out esterification reaction on the obtained compound IV and the compound II and different acyl chloride derivatives RCOCl to obtain the derivatives shown in the formula (I).
The organic solvent in the preparation method is one of tetrahydrofuran, dichloromethane or trichloromethane.
The organic base in the preparation method is one of triethylamine, pyridine, diisopropylethylamine or 4-dimethylaminopyridine.
In a third aspect of the present invention, there is provided an antibacterial pharmaceutical composition comprising the derivative of formula (I) as defined in the first aspect and a pharmaceutically acceptable carrier.
The pharmaceutical composition may be formulated into various types of administration unit dosage forms such as tablets, pills, powders, liquids, suspensions, emulsions, granules, capsules, suppositories, injections (solutions and suspensions) and the like, depending on the purpose of treatment.
In a fourth aspect of the invention, there is provided the use of a derivative of formula (I) or a pharmaceutical composition thereof as an inhibitor of bacterial RNA polymerase.
In addition, the invention provides application of the derivative shown as the formula (I) or the pharmaceutical composition thereof in antibiosis.
The content of the derivative represented by the formula (I) in the pharmaceutical composition of the present invention is not particularly limited, and may be selected from a wide range, and may be generally a mass percentage.
In the present invention, the method of administration of the pharmaceutical composition is not particularly limited. The formulation of various dosage forms can be selected for administration according to the age, sex and other conditions and symptoms of the patient. For example, tablets, pills, solutions, suspensions, emulsions, granules and capsules are administered orally; the injection can be administered alone, or mixed with injectable delivery solution (such as glucose solution and amino acid solution) for intravenous injection, or simply injected into muscle, skin or abdomen if necessary; the suppository is administered to the rectum.
In the present invention, the administration dose can be appropriately selected depending on the administration method, the age, sex and other conditions of the patient and the symptoms. Typical dosages administered may be: about 0.1 to 300mg of the pharmaceutically active ingredient per kg body weight per day. Generally, each unit dosage form for administration may contain 1 to 200mg of the pharmaceutically active ingredient.
The invention discloses a unique application of N-aryl dithiopyrryl ketonuria and amino ester derivatives as a bacterial RNA polymerase inhibitor, and particularly performs in-vitro antibacterial activity research, in-vitro bacterial RNA polymerase activity inhibition research and in-vivo antibacterial activity research of mice, thereby confirming that the derivatives have the in-vivo and in-vitro antibacterial activity and the in-vitro and in-vitro antibacterial activity aiming at bacterial RNAP.
Detailed Description
Example 1:
preparation of Compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1,2] dithio [4,3-b ] pyrrol-6-yl) -3- (2-hydroxyethyl) urea)
Figure BDA0002292580890000061
Mixing 6-amino-4- (2, 4-dimethoxyphenyl) - [1,2]Dithio [4,3-b ]]Pyrrole-5 (4H) -one hydrochloride (compound VI) (3g, 8.70mmol) was dissolved in tetrahydrofuran (80m L), triethylamine (2.64g, 26.10mmol) and phenyl chloroformate (1.77g, 11.31mmol) were added, respectively, the reaction was allowed to react at room temperature for 8H, and T L C monitored (chloroform/methanol ═ 20:1, v/v, Rf0.61.) after completion of the reaction, concentration was performed, distilled water (60m L) was added to the residue, extraction was performed with dichloromethane (3 × 30m L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30m L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain a yellow solid (compound V);
the yellow solid (Compound V) from the above step was dissolved in tetrahydrofuran (30m L), and addedEthanolamine (0.75g, 12.26mmol) was added, the reaction was allowed to proceed at room temperature for 7h, and monitored at T L C (chloroform/methanol 10:1, v/v, R)f(0.51.) after completion of the reaction, concentration was carried out, distilled water (60M L) was added to the residue, extraction was carried out with methylene chloride (3 × 30M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30M L), dried over anhydrous magnesium sulfate, filtered, the solvent was distilled off under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was carried out to obtain a yellow solid, i.e., compound IV (2.89g), with a total yield of 84% in two steps, mp 144.3-146.5 ℃, ESI-MS M/z:396.09[ M + H/z ℃. (M + H)]+；1H NMR(400MHz,DMSO-d6):8.52(s,1H),7.27(d,J＝8.6Hz,1H),6.88(t,J＝5.6Hz,1H),6.80(d,J＝2.5Hz,1H),6.70(s,1H),6.68(dd,J＝8.7,2.6Hz,1H),3.88(s,3H),3.79(s,3H),3.48(t,J＝5.6Hz,2H),3.21(q,J＝5.5Hz,2H)；13C NMR(125MHz,DMSO-d6)166.30,161.21,156.69,155.52,136.67,130.71,127.99,116.65,109.60,105.64,100.03,60.69,56.23,56.02,42.45。
Example 2:
preparation of Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1,2] dithio [4,3-b ] pyrrol-6-yl) formate))
Figure BDA0002292580890000062
Triphosgene (0.43g, 1.45mmol) is dissolved in ultra-dry tetrahydrofuran (10m L), cooled to 0 ℃ under ice bath, and a mixed solution of 6-amino-4- (2, 4-dimethoxyphenyl) - [1,2] dithio [4,3-b ] pyrrol-5 (4H) -one hydrochloride (compound VI) (0.5g,1.45mmol), triethylamine (0.45g, 4.35mmol) and ultra-dry tetrahydrofuran (20m L) is slowly dropped, reacted for 2-3H under ice bath, and concentrated to dryness under reduced pressure to obtain an intermediate product III;
intermediate III was dissolved in ultra dry tetrahydrofuran (10m L), cooled to 0 ℃, then a solution of ethylene glycol (0.27g, 4.35mmol) in ultra dry tetrahydrofuran (10m L) was added dropwise rapidly, after completion of the addition, the reaction was allowed to react at room temperature for 12h, and monitored at T L C (chloroform/methanol 10:1, v/v, R L C)f0.86. The reaction was complete, concentrated, water (50m L) was added to the residue, extraction was carried out with dichloromethane (3 × 30m L), the organic phases were combined and saturated with waterWashing with NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtering, distilling off solvent under reduced pressure, eluting with chloroform/methanol, and performing silica gel column chromatography to obtain yellow solid, i.e. compound II (0.39g), with the two-step total yield of 68%, mp 198 ℃; ESI-MS M/z of 398.00[ M + H/z%]+；1H NMR(400MHz,DMSO-d6)9.56(s,1H),7.27(d,J＝8.6Hz,1H),6.85(s,1H),6.81(d,J＝2.5Hz,1H),6.69(dd,J＝8.6,2.6Hz,1H),4.91(dd,J＝12.2,6.8Hz,1H),4.18(t,J＝4.8Hz,2H),3.89(s,3H),3.79(s,3H),3.66(d,J＝4.6Hz,2H)；13C NMR(150MHz,DMSO-d6)165.93,161.23,156.66,154.51,136.44,133.40,130.74,116.56,115.37,110.76,105.63,99.99,67.27,59.68,56.20,56.03。
Example 3:
preparation of Compound 1
Figure BDA0002292580890000071
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ and acetyl chloride (48.0mg,0.60mmol) was added followed by reaction at 35 ℃ and monitoring (CHCl) at 8 h.T L C3/MeOH＝10:1，v/v，RfNo. 0.43), after completion of the reaction, concentration was performed, distilled water (20M L) was added to the residue, extraction was performed with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 1(153mg) at a yield of 70.3% ESI-MS M/z 438.13[ M + H438.13 ]]+；1H NMR(400MHz,CDCl3)8.66(s,1H),7.33–7.03(m,1H),6.66–6.41(m,2H),6.31(s,2H),3.78(s,3H),3.75–3.62(m,5H),3.07(d,J＝3.6Hz,2H),1.94(s,3H)；13C NMR(100MHz,CDCl3)169.77,166.24,160.40,155.40,154.55,135.73,128.86,128.36,115.29,114.98,109.64,103.61,98.73,62.02,54.86,54.62,37.60,19.84。
Example 4:
preparation of Compound 2
Figure BDA0002292580890000081
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was carried out, cyclopropylcarbonyl chloride (63.0mg,0.60mmol) was added, reaction was carried out at 35 ℃ for 8h, and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.63. after completion of the reaction, concentration was performed, distilled water (20M L) was added to the residue, extraction was performed with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 2(180mg) in 77.5% yield ESI-MS M/z 464.12[ M + H ],]+；1H NMR(400MHz,CDCl3)8.65(s,1H),7.23–7.09(m,1H),6.58–6.43(m,2H),6.35–6.23(m,2H),3.78(s,3H),3.74–3.64(m,5H),3.08(q,J＝5.7Hz,2H),1.54–1.44(m,1H),0.95–0.86(m,2H),0.84–0.73(m,2H)；13C NMR(100MHz,CDCl3)173.57,166.26,160.40,155.40,154.53,135.76,128.88,128.35,115.31,114.97,109.55,103.63,98.76,61.94,54.87,54.64,37.69,11.82,7.49。
example 5:
preparation of Compound 3
Figure BDA0002292580890000082
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was performed at 0 deg.C, furoyl chloride (78mg,0.60mmol) was added, reaction was carried out at 35 deg.C for 8h, and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.37. after completion of the reaction, concentration was carried out, distilled water (20m L) was added to the residue, extraction was carried out with methylene chloride (3 × 10m L), and the organic phases were combined and organicWashing the phases with saturated NaCl solution (10M L), drying over anhydrous magnesium sulfate, filtering, removing the solvent by evaporation under reduced pressure, eluting with chloroform/methanol, and performing silica gel column chromatography to obtain compound 3(170mg) with yield 69.3%. ESI-MS M/z 490.13[ M + H ] M]+；1H NMR(400MHz,DMSO-d6)8.52(s,1H),8.05(d,J＝0.8Hz,1H),7.44–7.37(m,1H),7.26(d,J＝8.6Hz,1H),6.96(t,J＝5.8Hz,1H),6.84–6.75(m,2H),6.74–6.62(m,2H),4.34(t,J＝5.2Hz,2H),3.88(s,3H),3.79(s,3H),3.51(d,J＝5.3Hz,2H)；13C NMR(100MHz,DMSO-d6)165.76,160.72,157.78,156.17,154.87,147.70,143.67,136.10,130.23,127.78,118.66,116.03,115.90,112.31,109.49,105.11,99.50,63.73,55.71,55.52,38.36。
Example 6:
preparation of Compound 4
Figure BDA0002292580890000091
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ followed by addition of succinic acid monoethyl ester acid chloride (99mg,0.60mmol), reaction was carried out at 35 ℃ for 8h and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.69.) after completion of the reaction, concentration was performed, distilled water (20M L) was added to the residue, extraction was performed with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 4(170mg) with a yield of 66.8% ESI-MS M/z 524.24[ M + H ],]+；1H NMR(600MHz,DMSO-d6)8.52(s,1H),7.26(d,J＝8.5Hz,1H),6.89(t,J＝5.8Hz,1H),6.80(d,J＝2.6Hz,1H),6.72–6.66(m,2H),4.14–4.07(m,4H),3.88(s,3H),3.79(s,3H),3.50–3.38(m,2H),2.68–2.58(m,4H),1.23(t,J＝7.1Hz,3H)；13C NMR(100MHz,DMSO-d6)172.36,166.28,161.23,156.68,155.39,136.62,130.73,128.23,116.55,116.44,109.95,105.62,100.01,63.93,60.53,56.22,56.02,38.73,29.11,29.08,14.52。
example 7:
preparation of Compound 5
Figure BDA0002292580890000092
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was performed at 0 deg.C, benzoyl chloride (84mg,0.60mmol) was added, reaction was carried out at 35 deg.C for 8h, and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.78. The reaction was completed, concentration was carried out, distilled water (20M L) was added to the residue, extraction was carried out with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtered, the solvent was distilled off under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was carried out to obtain compound 5(190mg) in 74.5% yield ESI-MS M/z 500.07[ M + H + E]+；1H NMR(400MHz,CDCl3)8.75(s,1H),8.75(s,1H),8.13–7.90(m,2H),7.54(t,J＝7.4Hz,1H),7.41(t,J＝7.7Hz,2H),7.07(d,J＝8.6Hz,1H),6.54(d,J＝2.5Hz,1H),6.51–6.45(m,2H),6.35(s,1H),4.04(t,J＝6.0Hz,2H),3.77(s,3H),3.69(s,3H),3.27(q,J＝6.0Hz,2H)；13C NMR(100MHz,CDCl3)166.28,165.28,160.38,155.37,154.65,135.78,131.92,129.07,128.84,128.71,128.35,127.33,115.30,114.87,109.57,103.52,98.70,62.38,54.81,54.58,37.83。
Example 8:
preparation of Compound 6
Figure BDA0002292580890000101
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was performed at 0 deg.C, nicotinoyl chloride (85mg,0.60mmol) was added, reaction was allowed to proceed for 8h at 35 deg.C, and the reaction was monitored by T L CYing (CHCl)3/MeOH＝10:1，v/v，Rf0.70.) reaction, concentration, addition of distilled water (20M L) to the residue, extraction with dichloromethane (3 × 10M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (10M L), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent under reduced pressure, chloroform/methanol as eluent, column chromatography on silica gel to give compound 6(150mg) in 62.1% yield ESI-MS M/z 501.12[ M + H3 []+；1H NMR(400MHz,CDCl3)9.17(s,1H),8.75(s,2H),8.26(d,J＝8.0Hz,1H),7.35(dd,J＝7.9,4.8Hz,1H),7.10(d,J＝8.5Hz,1H),6.63–6.41(m,3H),6.36(s,1H),4.05(t,J＝5.9Hz,2H),3.78(s,3H),3.70(s,3H),3.27(q,J＝5.9Hz,2H)；13CNMR(100MHz,CDCl3)166.26,163.96,160.40,155.39,154.64,152.36,149.98,136.25,135.71,128.85,128.49,124.98,122.29,115.20,114.91,109.80,103.51,98.67,62.91,54.84,54.60,37.66。
Example 9:
preparation of Compound 7
Figure BDA0002292580890000111
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ followed by addition of 6-chloronicotinyl chloride (107mg,0.60mmol) and reaction was carried out at 35 ℃ for 8h, monitoring the reaction by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.74.) reaction, concentration, addition of distilled water (20M L) to the residue, extraction with dichloromethane (3 × 10M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (10M L), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent under reduced pressure, chloroform/methanol as eluent, column chromatography on silica gel to give compound 7(210mg) in 77.9% yield ESI-MS M/z 535.12[ M + H3 []+；1H NMR(400MHz,DMSO-d6)9.03(d,J＝2.2Hz,1H),8.55–8.33(m,2H),7.77(d,J＝8.3Hz,1H),7.26(d,J＝8.6Hz,1H),6.99(t,J＝5.8Hz,1H),6.79(d,J＝2.5Hz,1H),6.76–6.61(m,2H),5.81(s,1H),4.40(s,2H),3.87(s,3H),3.78(s,3H),3.57(d,J＝5.1Hz,2H)；13C NMR(100MHz,DMSO-d6)165.75,163.81,160.71,156.16,154.95,154.46,150.77,140.32,136.08,130.21,127.80,125.18,124.62,116.02,115.87,109.55,105.11,99.50,64.86,55.71,55.51,54.87。
Example 10:
preparation of Compound 8
Figure BDA0002292580890000112
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea) (0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was performed at 0 deg.C, p-fluorobenzoyl chloride (95mg,0.60mmol) was added, reaction was performed at 35 deg.C for 8h, and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.77. after completion of the reaction, concentration was carried out, distilled water (20M L) was added to the residue, extraction was carried out with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtered, the solvent was distilled off under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was carried out to obtain compound 8(140mg) in 54.7% yield ESI-MS M/z 518.18[ M + H ],]+；1H NMR(400MHz,CDCl3)8.68(s,1H),8.11–7.94(m,2H),7.17–6.94(m,3H),6.55–6.46(m,2H),6.41(t,J＝5.8Hz,1H),6.37(s,1H),4.01(t,J＝5.9Hz,2H),3.78(s,3H),3.70(s,3H),3.28(d,J＝4.5Hz,2H)；13C NMR(100MHz,CDCl3)166.28,164.36,160.40,155.38,154.63,135.72,131.34,131.25,128.85,128.47,125.29,115.22,114.88,114.58,114.36,109.78,103.53,98.70,62.63,54.82,54.60,37.79。
example 11:
preparation of Compound 9
Figure BDA0002292580890000121
Mixing compound IV (1- (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b]Pyrrol-6-yl) -3- (2-hydroxyethyl) urea (D)0.20g,0.50mmol) was dissolved in tetrahydrofuran (10m L), triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ and p-cyanobenzoyl chloride (99mg,0.60mmol) was added and the reaction was carried out at 35 ℃ for 8h, and the reaction was monitored at T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.78. The reaction was completed, concentration was carried out, distilled water (20M L) was added to the residue, extraction was carried out with dichloromethane (3 × 10M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (10M L), dried over anhydrous magnesium sulfate, filtered, the solvent was distilled off under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was carried out to obtain compound 9(190mg) in 76.2% yield ESI-MS M/z 525.13[ M + H + E]+；1H NMR(400MHz,DMSO-d6)8.47(s,1H),8.17(d,J＝8.4Hz,2H),8.03(d,J＝8.4Hz,2H),7.21(d,J＝8.6Hz,1H),6.96(t,J＝5.8Hz,1H),6.74(d,J＝2.5Hz,1H),6.71–6.56(m,2H),4.35(t,J＝5.0Hz,2H),3.82(s,3H),3.73(s,3H),3.52(d,J＝4.8Hz,2H)；13C NMR(100MHz,DMSO-d6)165.75,164.45,160.71,156.16,154.97,136.08,133.58,132.75,130.21,129.95,127.79,118.06,116.02,115.88,115.47,109.55,105.11,99.50,64.94,55.71,55.51,38.22。
Example 12:
preparation of Compound 10
Figure BDA0002292580890000122
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, acetyl chloride (119mg,1.51mmol) was added, reaction was carried out at 35 ℃ C for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.66.) reaction, concentration, addition of water (100M L) to the residue, extraction with dichloromethane (3 × 30M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtration, removal of the solvent under reduced pressure, chloroform/methanol as eluent, silica gel column chromatography to give compound 10(390mg) with 70.5% yield ESI-MS M/z:439.09[ M + H ],]+；1H NMR(400MHz,CDCl3)7.20(s,1H),7.11(d,J＝8.4Hz,1H),6.54–6.45(m,2H),6.27(s,1H),4.38–4.30(m,2H),4.26–4.20(m,2H),3.77(s,3H),3.69(s,3H),2.04(s,3H)；13C NMR(100MHz,CDCl3)169.80,165.15,160.31,155.33,152.28,136.13,130.10,128.99,115.12,113.56,108.08,103.77,98.87,62.76,61.20,54.79,54.62,19.82。
example 13:
preparation of Compound 11
Figure BDA0002292580890000131
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate)) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, cyclopropylcarbonyl chloride (158mg,1.51mmol) was added and the reaction was carried out at 35 ℃ for 8h, monitoring the reaction by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.68.) reaction, concentration, addition of water (100M L) to the residue, extraction with dichloromethane (3 × 30M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent under reduced pressure, chloroform/methanol as eluent, column chromatography on silica gel to give compound 11(456mg) with 78.4% yield ESI-MS M/z 465.09[ M + H ],]+；1H NMR(400MHz,CDCl3)7.22–7.17(m,1H),6.58-6.54(m,2H),6.34(s,1H),4.42–4.38(m,2H),4.32(dd,J＝5.6,2.9Hz,2H),3.84(s,3H),3.77(s,3H),1.71–1.55(m,1H),1.06-1.02(m,2H),0.94–0.86(m,2H)；13C NMR(100MHz,CDCl3)174.73,166.22,161.34,156.37,153.37,137.17,131.15,130.02,116.16,114.64,109.15,104.79,99.90,63.90,62.24,55.82,55.66,12.79,8.79。
example 14:
preparation of Compound 12
Figure BDA0002292580890000132
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 deg.C, furoyl chloride (197mg,1.51mmol) was added and reaction was carried out at 35 deg.C for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，RfNo. 0.58.) after the reaction was completed, concentration was performed, water (100M L) was added to the residue, extraction was performed with dichloromethane (3 × 30M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 12(400mg) with a yield of 65.3% ESI-MS M/z:491.16[ M + H ],]+；1H NMR(600MHz,DMSO-d6)9.76(s,1H),8.04(d,J＝0.8Hz,1H),7.39(d,J＝3.4Hz,1H),7.26(d,J＝8.6Hz,1H),6.85(s,1H),6.79(d,J＝2.6Hz,1H),6.76(dd,J＝3.5,1.7Hz,1H),6.67(dd,J＝8.6,2.6Hz,1H),4.53(s,2H),4.47(s,2H),3.87(s,3H),3.78(s,3H)；13C NMR(100MHz,DMSO-d6)165.90,161.23,158.20,156.66,154.17,148.27,144.05,136.41,133.82,130.74,119.29,116.56,115.20,112.85,110.90,105.63,100.00,63.51,63.25,56.20,56.02。
example 15:
preparation of Compound 13
Figure BDA0002292580890000141
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate)) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, monoethyl succinyl chloride (248mg,1.51mmol) was added and reaction was carried out at 35 ℃ for 8h, and reaction was monitored at T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.61.) after completion of the reaction, concentration was performed, water (100M L) was added to the residue, extraction was performed with dichloromethane (3 × 30M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 13(440mg) with a yield of 66.6% ESI-MS M/z:525.13[ M + H ],]+；1H NMR(400MHz,CDCl3)7.31–7.10(m,1H),6.57-6.54(m,2H),6.35(d,J＝3.7Hz,1H),4.21–4.04(m,6H),3.84(s,3H),3.76(s,3H),2.68–2.59(m,4H),1.26(s,3H)；13C NMR(100MHz,CDCl3)172.74,172.23,171.23,168.04,166.22,166.16,161.32,156.36,153.38,137.14,130.02,109.23,109.08,104.78,99.88,60.79,60.66,55.81,55.64,29.02,27.99,14.16。
example 16:
preparation of Compound 14
Figure BDA0002292580890000151
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, benzoyl chloride (212mg,1.51mmol) was added, reaction was carried out at 35 ℃ C for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.63.) after the reaction, concentration was performed, water (100M L) was added to the residue, extraction was performed with dichloromethane (3 × 30M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30M L), dried over anhydrous magnesium sulfate, filtration was performed, the solvent was evaporated under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was performed to obtain compound 14(420mg) with a yield of 67.3%. ESI-MS M/z:501.13[ M + H ],]+；1H NMR(400MHz,CDCl3)8.10–8.05(m,2H),7.57(t,J＝7.4Hz,1H),7.45(t,J＝7.7Hz,1H),7.25(d,J＝8.3Hz,1H),7.18(d,J＝8.4Hz,1H),6.58-6.53(m,2H),6.35(s,1H),4.56(s,2H),4.54(s,2H),3.84(s,3H),3.76(s,3H)；13C NMR(100MHz,CDCl3)166.37,166.23,161.34,156.37,153.44,137.17,133.20,131.24,130.06,130.02,129.81,129.72,128.44,128.34,116.16,114.65,109.22,104.80,99.91,63.88,62.77,55.82,55.65。
example 17:
preparation of Compound 15
Figure BDA0002292580890000152
Reacting compound II (2-hydroxy)Ethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2-]Dithio [4,3-b ]]Pyrrol-6-yl) formate) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 deg.C, nicotinoyl chloride (213mg,1.51mmol) was added, reaction was carried out at 35 deg.C for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.64. reaction, concentration, addition of water (100M L), extraction with dichloromethane (3 × 30M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent under reduced pressure, chloroform/methanol as eluent, silica gel column chromatography to give compound 15(370mg), yield 59.3%. ESI-MS M/z:502.09[ M + H ],]+；1H NMR(400MHz,CDCl3)9.26(s,1H),8.80(d,J＝3.8Hz,1H),8.33(d,J＝8.0Hz,1H),7.41(dd,J＝7.9,4.9Hz,1H),7.20–7.04(m,2H),6.56(dd,J＝8.2,5.4Hz,2H),6.34(s,1H),4.64–4.47(m,4H),3.84(s,3H),3.76(s,3H)；13C NMR(100MHz,CDCl3)166.15,165.04,161.34,156.35,153.65,153.30,151.06,137.29,131.22,130.01,125.67,123.36,116.13,114.51,109.13,104.78,99.90,63.61,63.20,55.82,55.65。
example 18:
preparation of Compound 16
Figure BDA0002292580890000161
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, 6-chloronicotinyl chloride (266mg,1.51mmol) was added, reaction was carried out at 35 ℃ C for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.67.) reaction, concentration, addition of water (100M L) to the residue, extraction with dichloromethane (3 × 30M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtration, removal of the solvent under reduced pressure, chloroform/methanol as eluent, silica gel column chromatography to give compound 16(380mg) with 56.5% yield ESI-MS M/z 535.03[ M + H ],]+；1H NMR(400MHz,CDCl3)7.39(s,1H),7.27(s,1H),7.19(d,J＝8.4Hz,1H),7.11(t,J＝8.4Hz,3H),6.56(dd,J＝4.0,2.7Hz,2H),6.37(s,1H),4.54(d,J＝2.9Hz,4H),3.84(s,3H),3.76(s,3H)；13C NMR(100MHz,CDCl3)169.44,167.18,166.31,165.39,161.36,156.35,153.49,137.12,132.78,132.69,132.45,132.35,130.01,115.70,115.48,109.55,104.79,99.89,63.76,62.91,55.81,55.64。
example 19:
preparation of Compound 17
Figure BDA0002292580890000162
Compound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate)) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ 4-fluorobenzoyl chloride (240mg,1.51mmol) was added, reaction was carried out at 35 ℃ for 8h, and reaction was monitored by T L C (CHCl)3/MeOH＝10:1，v/v，Rf0.62.) reaction, concentration, addition of water (100M L) to the residue, extraction with dichloromethane (3 × 30M L), combination of organic phases, washing of the organic phase with saturated NaCl solution (30M L), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent under reduced pressure, chloroform/methanol as eluent, column chromatography on silica gel to give compound 17(410mg) in 62.8% yield ESI-MS M/z 518.94[ M + H ],]+；1H NMR(400MHz,CDCl3)9.03(d,J＝2.1Hz,1H),8.28(dd,J＝8.3,2.4Hz,1H),7.43(d,J＝8.3Hz,1H),7.24–7.03(m,2H),6.56(dd,J＝8.2,5.5Hz,2H),6.35(s,1H),4.57(m,4H),3.84(s,3H),3.76(s,3H)；13C NMR(100MHz,CDCl3)166.12,164.20,161.34,156.33,155.93,153.26,151.37,139.77,137.11,129.99,124.65,124.24,116.09,114.44,109.26,104.78,99.88,63.45,55.81,55.64。
example 20:
preparation of Compound 18
Figure BDA0002292580890000171
Will combine withCompound II (2-hydroxyethyl (4- (2, 4-dimethoxyphenyl) -5-oxo-4, 5-dihydro- [1, 2)]Dithio [4,3-b ]]Pyrrol-6-yl) formate)) (0.5g, 1.26mmol) was dissolved in tetrahydrofuran, triethylamine (0.15g,1.5mmol) was added, stirring was carried out at 0 ℃ C, p-cyanobenzoyl chloride (250mg,1.51mmol) was added and the reaction was carried out at 35 ℃ for 8h, and T L C monitored (CHCl)3/MeOH＝10:1，v/v，Rf0.67. after completion of the reaction, concentration was carried out, water (100M L) was added to the residue, extraction was carried out with methylene chloride (3 × 30M L), the organic phases were combined, the organic phase was washed with a saturated NaCl solution (30M L), dried over anhydrous magnesium sulfate, filtered, the solvent was distilled off under reduced pressure, chloroform/methanol was used as an eluent, and silica gel column chromatography was carried out to obtain compound 18(463mg) in 69.9% yield ESI-MS M/z:526.07[ M + H ]. the reaction was carried out]+；1H NMR(400MHz,CDCl3)8.18(d,J＝8.4Hz,1H),7.75(d,J＝8.4Hz,1H),7.20–7.14(m,1H),6.61–6.52(m,1H),6.36(s,1H),4.59(d,J＝3.9Hz,1H),4.54(d,J＝4.6Hz,1H),3.84(s,1H),3.76(s,1H)；13C NMR(100MHz,CDCl3)166.15,164.72,161.38,156.33,153.31,137.11,133.53,132.29,130.34,129.99,117.98,116.61,116.05,114.47,109.34,104.81,99.90,63.54,55.83,55.66。
Example 21: minimum Inhibitory Concentration (MIC) test in vitro against standard strains
(1) Strain preparation
Staphylococcus aureus (Staphylococcus aureus), Bacillus subtilis (Bacillus subtilis), Enterococcus faecalis (Enterococcus faecalis) and Streptococcus pneumoniae (Streptococcus pneumoniae) were selected. The strain frozen with glycerol at-80 ℃ is inoculated to a corresponding solid agar culture medium or a liquid culture medium. Placing in an incubator at 37 ℃ for culturing for 18-24 h.
(2) Preparation of Compound mother plate
Compounds were dissolved using 100% DMSO and configured as a stock solution at a concentration of 12.8mg/m L, and standard antibiotics and compounds were configured in V-bottom 96-well plates with 100% DMSO as 11 two-fold serial dilutions of 100 × high-concentration working solutions, respectively.
100% DMSO was used as a negative control.
(3) Preparation of inoculum
Collection of appropriate amount of solid plate culture/liquid culture heavy suspension in rawSaline solution was treated, mixed well, and the turbidity of the bacterial suspension was adjusted to an OD600 of 0.2 with a turbidimeter to about 1 × 108cfu/m L bacteria the turbidity adjusted suspension was then diluted with test medium to a bacteria concentration of 1 × 106cfu/m L this is the inoculum.
(4) Detection of minimum inhibitory concentration
The compound master plate was transferred to 100 × high concentration working solution serially diluted 2 μ L2 times to a round bottom 96 well plate, and then 198 μ L of bacterial inoculum was added to each well to obtain test plates all of which were prepared and incubated at 35 ℃ for 16-20 h.
(5) The experimental results are as follows: TABLE 1 Minimum Inhibitory Concentration (MIC) of Compounds 1-18 against Standard strains in vitro
Figure BDA0002292580890000181
In the table, Cip: ciprofloxacin, Rif: rifampin, Van: vancomycin.
The results show that most compounds show stronger inhibitory activity to streptococcus pneumoniae, and the inhibitory activity is superior to that of positive control compounds such as ciprofloxacin, rifampicin and vancomycin; compounds 1,2, 11, 13 and 15 showed strong activity against Staphylococcus aureus with inhibitory activity comparable to or better than vancomycin.
Example 22: in vitro Minimal Inhibitory Concentration (MIC) test for clinical drug-resistant strains
In the experiment, four compounds (1, 11, 13 and 15) with strong activity on inhibiting Staphylococcus aureus (ATCC 27660) are selected as compounds to be tested, and the MIC value of the compounds to clinical drug-resistant strains is further tested. The strain can be Methicillin-resistant Staphylococcus aureus (MRSA), Vancomycin-resistant Staphylococcus aureus (VRSA), rifamycin-resistant Staphylococcus aureus (RRSA), and intermediate penicillin-resistant Streptococcus pneumoniae (MPRP). The experimental procedure was as in example 21. The results of the experiment are shown in table 2.
TABLE 2 Minimal Inhibitory Concentration (MIC) of preferred compounds against drug-resistant strains in vitro
Figure BDA0002292580890000191
Cip: ciprofloxacin, Rif: rifampin, Van: vancomycin;amethicillin-resistant staphylococcus aureus (MRSA);bvancomycin-resistant staphylococcus aureus (VRSA);crifampicin Resistant Staphylococcus Aureus (RRSA);dmedium penicillin-resistant streptococcus pneumoniae (MPRSP); ND: and (5) not testing.
Test results show that the selected compound has a good inhibition effect on staphylococcus aureus resistant to different drugs and also has a good inhibition effect on moderate penicillin-resistant streptococcus pneumoniae (MPRPSP). Wherein, the MIC values of the compounds 11 and 13 to methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Staphylococcus aureus (VRSA) are both 2, and the MIC value to moderate penicillin-resistant Streptococcus pneumoniae (MPRPSP) is less than 0.125. The selected compounds have small difference of MIC values of rifampicin-resistant staphylococcus aureus (RRSA) and standard staphylococcus aureus, and the compounds have potential effect of avoiding cross resistance with rifampicin.
Example 23: bacterial RNA polymerase inhibitory Activity assay
Bacterial RNA polymerase inhibitory activity tests were performed on preferred compounds 1,2, 11, 13 and 15, with the bacterial RNA polymerase inhibitors myxopyronin B, which is a bacterial RNA polymerase switch region inhibitor, acting primarily on the bacterial RNA polymerase switch region, and rifampicin, which acts primarily on the β -subunit of bacterial RNA polymerase, as positive control compounds.
The experiment adopts Escherichia coli (Escherichia coli) RNA polymerase detection kit (ProFoldinCatalog No. RPA100KE) which comprises detection buffer solution, DNA template, NTP mixture, Escherichia coli RNA polymerase and fluorescent dye, a 96-well plate is prepared, and 18 mu L of H is added into each well respectively2O, 10 × buffer solution of 3. mu. L, 10 × DNA template of 3. mu. L10 × enzyme of 3 mu L and 10 × NTP of 3 mu L, then 0.5 mu L compound with different concentrations is respectively added, 0.5 mu L DMSO is added into a control group, the mixture is uniformly mixed, the reaction is carried out at room temperature for 1h, 1 × fluorescent dye of 30 mu L is added, the mixture is uniformly mixed, the reaction is carried out at room temperature for 5min, and the fluorescence value (excitation wavelength 485 nm/absorption wavelength 535nm) is measured by a microplate reader.
Results of the experiment
TABLE 3 preferred Compounds Activity of E.coli RNA polymerase inhibition in vitro
Figure BDA0002292580890000201
The results show that part of the preferred compounds have better inhibitory activity to the Escherichia coli RNA polymerase, especially the compound 11 has better inhibitory activity to the Escherichia coli RNA polymerase. Compared with the bacterial RNA polymerase switch region inhibitor myxopyronin B, the selected compounds 11 and 13 have stronger inhibitory activity on bacterial RNA polymerase and inhibit IC50Rifampicin, 22.0 + -0.6 and 28.9 + -0.6. mu.M, respectively, is also a bacterial RNA polymerase inhibitor, but it acts primarily on the β -subunit of RNA polymerase50The values are generally large, suggesting that preferred compounds may act to inhibit by acting at different sites on bacterial RNA polymerase.
In conclusion, compounds 11 and 13 have better antibacterial activity, especially the best inhibitory activity against staphylococcus aureus and streptococcus pneumoniae; the compounds 11 and 13 have strong inhibitory activity on bacterial RNA polymerase, and are bacterial RNA polymerase inhibitors.
Example 23: in vivo antimicrobial Activity test
(1) Experimental animals: kunming mouse has healthy appearance, weight of 10-20 g, and half of male and female, and is provided by animal house of Guilin medical college.
(2) Bacteria: the strain used for infecting the mice is staphylococcus aureus ATCC27660 which is clinically isolated and used after laboratory identification.
(3) Preparing experimental strains: using beef extract peptone liquid medium: egg10g of white peptone, 3g of beef extract, 15g of sodium chloride, 1000m of distilled water L, pH 7.2, culturing at 37 ℃ for 18h, and preparing 5 × 10 by using 0.5 McLee's turbidimetric solution as a control8cfu/m L bacterial suspension for use.
(4) The experimental method comprises randomly grouping experimental animals into groups of 10 animals, injecting the mouse abdominal cavity with minimum 100% lethal dose (M L D)100) 0.5m L, orally administered for 2 times at 15min and 6h after infection, respectively, observing the survival rate of animals in 7d, and calculating half Effective Dose (ED) of animals by Bliss method50) And 95% confidence limits thereof, and performing statistical processing.
(5) Results of the experiment
TABLE 4 in vivo antibacterial Effect of preferred Compounds on Staphylococcus aureus infected mice
Figure BDA0002292580890000211
The results show that the compounds 11 and 13 have protective effect on mice infected with staphylococcus aureus systemically.
Example 24: preparation of tablets
Prescription: dosage of
Compound 11 50mg
Microcrystalline cellulose 250mg
Crosslinked polyvinylpyrrolidone 50mg
Pregelatinized starch 100mg
Magnesium stearate 5mg
The preparation method comprises the following steps: according to the formula, the compound 11 after being crushed and sieved, microcrystalline cellulose, pregelatinized starch and cross-linked polyvinylpyrrolidone are uniformly mixed, then mixed with 5% ethanol solution, granulated, dried, then mixed with a lubricant, and finally tabletted. Wherein, the compound 11 is crushed and sieved to be sieved by a 60-mesh sieve; the microcrystalline cellulose, the pregelatinized starch and the cross-linked polyvinylpyrrolidone are crushed and sieved to be sieved by a sieve of 80 meshes; the particle size of the granulated particles is 20 meshes; the drying temperature is preferably controlled within 3 percent of the water mass percent at 90 ℃.
Example 25: preparation of capsules
Figure BDA0002292580890000221
The preparation method comprises the following steps: according to the formula, the medicine and the auxiliary materials are mixed evenly and filled into the capsule shell.
Example 26: preparation of injection
Figure BDA0002292580890000222
The preparation method comprises the following steps: according to the formula, the compound 11 or the salt thereof and the wetting agent are ground and mixed uniformly by using a mortar, then the mixture is mixed uniformly with the suspending agent, the preservative and the water for injection, and then the mixture is ground. Wherein the milled particle size is 0.5 μm.

Claims (4)

1. The N-aryl dithiopyrryl ketonuria and amino ester derivative is characterized in that the structural formula is shown as the formula (I),
Figure FDA0002508065490000011
a derivative of formula (I) selected from the following compounds:
Figure FDA0002508065490000012
Figure FDA0002508065490000021
2. the process for the preparation of N-aryl dithiopyrrolone urea and amino ester derivatives as claimed in claim 1, wherein the derivatives are prepared by the following route:
Figure FDA0002508065490000031
the preparation method comprises the following steps:
reacting a compound shown as a formula VI, an organic base and phenyl chloroformate in an organic solvent to obtain an intermediate V, and reacting the intermediate V with ethanolamine to obtain a compound IV;
or reacting the compound VI with triphosgene in an organic solvent to obtain an intermediate III, and reacting the intermediate III with ethylene glycol to obtain a compound II;
respectively carrying out esterification reaction on the obtained compound IV and compound II and different acyl chloride derivatives RCOCl to obtain derivatives shown in the formula (I), wherein the derivatives shown in the formula (I) are selected from the compounds in claim 1;
the organic solvent in the preparation method is one of tetrahydrofuran, dichloromethane or trichloromethane;
the organic base in the preparation method is one of triethylamine, pyridine, diisopropylethylamine or 4-dimethylaminopyridine.
3. The use of the derivative of formula (I) or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof according to claim 1 for the preparation of a medicament for the treatment of bacterial infectious diseases.
4. Use according to claim 3, characterized in that: the medicament for treating the bacterial infectious diseases is a bacterial RNA polymerase inhibitor medicament.
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