Development of 2-hydroxy-1,4-naphthoquinone derivatives and use against s. aureus

Abstract

The present application relates to the field of chemical synthesis and application technology, and particularly relates to 3-mercapto addition derivatives of 2-hydroxy-1,4-naphthoquinone with a general formula (A), discloses a preparation method of the compounds, and inhibitory and bactericidal effects of the compounds on bacteria. The bacteriostatic and bactericidal properties of the compounds make them have potential uses in the research and application of antibacterial agents and antibacterial drugs.

Description

Technical Field

[0001] This invention relates to the field of chemical synthesis and application technology, specifically to the development of a 2-hydroxy-1,4-naphthoquinone derivative and its use against Staphylococcus aureus. Background Technology

[0002] In today's world, various pathogenic microorganisms, such as bacteria, fungi, and molds, have caused harm to crops, livestock, and human clinical treatment. Currently, bacterial infection has become the biggest threat to human health, with Staphylococcus aureus infection being the most common. At the same time, with the rapid development of antibiotics and antimicrobial drugs, bacterial resistance is inevitable. This has a serious impact on human health and life. Therefore, research on antimicrobial drugs is crucial. 2-Hydroxy-1,4-naphthoquinone, as a widely existing active substance, has received increasing attention for its biological activity, especially its antimicrobial effect. Yang (J Food Sci Technol, 2015, 52(4): 2446-2451) et al. experimentally confirmed that 2-hydroxy-1,4-naphthoquinone purified and isolated from Lophatherum gracile leaves has inhibitory activity against Bacillus cereus, Listeria monocytogenes, Salmonella enterica, and Shigella sonnei. Summary of the Invention

[0003] This invention relates to an antibacterial agent with 2-hydroxy-1,4-naphthoquinone as the core structure, which is synthesized by adding thiols with different side chains at the 3-position. The Gram-positive bacterium Staphylococcus aureus was used as the test strain, and 2-hydroxy-1,4-naphthoquinone was used as the control. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the newly synthesized compound against the strain were detected.

[0004] The compound A of this invention has the following general formula:

[0005]

[0006] In A:

[0007]

[0008] The compounds of the above general formula can be:

[0009] 2-Hydroxy-3-methmero-1,4-naphthoquinone (A1),

[0010] 2-Hydroxy-3-ethmercapto-1,4-naphthoquinone (A2),

[0011] 2-Hydroxy-3-propyl-1,4-naphthoquinone (A3),

[0012] 2-Hydroxy-3-butyroxy-1,4-naphthoquinone (A4),

[0013] 2-Hydroxy-3-pentylmercapto-1,4-naphthoquinone (A5),

[0014] 2-Hydroxy-3-hexylmercapto-1,4-naphthoquinone (A6),

[0015] 2-Hydroxy-3-heptamer-1,4-naphthoquinone (A7),

[0016] 2-Hydroxy-3-octylmercapto-1,4-naphthoquinone (A8),

[0017] 2-Hydroxy-3-nonylmercapto-1,4-naphthoquinone (A9),

[0018] 2-Hydroxy-3-decylmercapto-1,4-naphthoquinone (A10),

[0019] 2-Hydroxy-3-undecylmercapto-1,4-naphthoquinone (A11)

[0020] 2-Hydroxy-3-dodecylmercapto-1,4-naphthoquinone (A12)

[0021] 2-Hydroxy-3-n-tetadexamer-1,4-naphthoquinone (A13)

[0022] 2-Hydroxy-3-n-tetradecyl-1,4-naphthoquinone (A14)

[0023] 2-Hydroxy-3-n-pentadecanthomer-1,4-naphthoquinone (A15)

[0024] 2-Hydroxy-3-n-hexadecyl-1,4-naphthoquinone (A16)

[0025] 2-Hydroxy-3-n-heptadecyl-1,4-naphthoquinone (A17)

[0026] 2-Hydroxy-3-n-octadecyl-1,4-naphthoquinone (A18)

[0027] 2-Hydroxy-3-n-N-Nicotinyl-1,4-Naphthoquinone (A19)

[0028] 2-Hydroxy-3-eicosemercapto-1,4-naphthoquinone (A20)

[0029] 2-Hydroxy-3-benzylmercapto-1,4-naphthoquinone (A21)

[0030] 2-Hydroxy-3-p-tert-butylbenzylmercapto-1,4-naphthoquinone (A22)

[0031] The preparation methods for compounds A1-A22 are described in Scheme 1.

[0032] Scheme 1

[0033]

[0034] The antibacterial activity results of the 2-hydroxy-1,4-naphthoquinone derivatives showed that compounds A1-A22 had antibacterial activity against Staphylococcus aureus. The MIC and MBC results are shown in Table 1.

[0035] Table 1. MIC and MBC results for compounds A8, A10, A12, A14, and A22

[0036] Attached Figure Description

[0037] Figure 1 This is the mass spectrum of compound A8 from Example 1 of the present invention;

[0038] Figure 2 This is the carbon NMR spectrum of compound A8 in Example 1 of this invention;

[0039] Figure 3 The above is the 1H NMR spectrum of compound A8 from Example 1 of this invention;

[0040] Figure 4 The mass spectrum of compound A10 in Example 2 of this invention is as follows:

[0041] Figure 5 This is the mass spectrum of compound A12 from Example 3 of the present invention;

[0042] Figure 6 This is the mass spectrum of compound A14 from Example 4 of the present invention;

[0043] Figure 7 This is the mass spectrum of compound A22 from Example 5 of the present invention; Detailed Implementation

[0044] The synthesis, structural characterization, and antibacterial experiments of the 2-hydroxy-1,4-naphthoquinone derivative antibacterial agent of the present invention are described below through specific embodiments.

[0045] Example 1

[0046] 2-Hydroxy-3-octylmercapto-1,4-naphthoquinone (A8),

[0047] Under argon protection, 2-hydroxy-1,4-naphthoquinone (0.2 g, 1 eq) was dissolved in 15 mL of DMF solution, and octylthiol (0.299 mL, 1.5 eq) was added. The mixture was stirred, and then triethylamine (0.16 mL, 1 eq) was added. The reaction was carried out at 70 °C for 6 h. The solvent was evaporated under reduced pressure to obtain the crude product, which was separated by silica gel column chromatography using a mixture of petroleum ether and ethyl acetate as elution, yielding 2-hydroxy-3-octylthio-1,4-naphthoquinone. Product A3 (0.186 g, 0.58 mmol, yield 50.8%) was obtained as a red solid; Rf = 0.27 (methanol-dichloromethane 1:20). HR-MS (ESI) m / z: Calcd for C 18 H 22 O3S[M+H + ]319.1362, found: 319.1360. 13 C NMR (151MHz, DMSO-d6) δ182.47, 179.12, 158.37, 134.87, 133.85, 132.93, 130.52, 1 26.54, 126.44, 120.51, 32.13, 31.64, 30.06, 29.03, 28.91, 28.36, 22.52, 14.36. 1H NMR (600MHz, DMSO-d6) δ8.05-7.69 (m, 4H), 3.01 (t, J=7.3Hz, 2H), 1.49 (p, J=7.4Hz, 2H), 1.32 (p, J=7.1Hz, 2H), 1.20 (dt, J=13.1, 6.2H z， z, 8H), 0.81 (t, J = 6.9 Hz, 3H). Mass spectrum attached. Figure 1 The attached carbon NMR spectrum is shown in the figure. Figure 2 The hydrogen NMR spectrum is shown in the attached image. Figure 3 As shown.

[0048] Example 2

[0049] 2-Hydroxy-3-decylmercapto-1,4-naphthoquinone (A10),

[0050] Under argon protection, 2-hydroxy-1,4-naphthoquinone (0.2 g, 1 eq) was dissolved in 15 mL of DMF solution, followed by the addition of decyl mercaptan (0.355 mL, 1.5 eq), stirring, and then the addition of triethylamine (0.16 mL, 1 eq). The reaction was carried out at 70 °C for 6 h, and the solvent was evaporated under reduced pressure to obtain the crude product. Separation was performed by silica gel column chromatography using a mixture of petroleum ether and ethyl acetate as elution, yielding the product 2-hydroxy-3-decylmercapto-1,4-naphthoquinone. Product A4 (0.205 g, 0.59 mmol, yield 51.5%) was obtained as a red solid; Rf = 0.28 (methanol-dichloromethane 1:20). HR-MS (ESI) m / z: Calcd for C 20 H 26 O3S[M+H + 347.1675, found: 347.1672. Mass spectrum attached. Figure 4 As shown.

[0051] Example 3

[0052] 2-Hydroxy-3-dodecylmercapto-1,4-naphthoquinone (A12),

[0053] Under argon protection, 2-hydroxy-1,4-naphthoquinone (0.2 g, 1 eq) was dissolved in 15 mL of DMF solution, and n-dodecyl mercaptan (0.413 mL, 1.5 eq) was added. The mixture was stirred, and then triethylamine (0.16 mL, 1 eq) was added. The reaction was carried out at 70 °C for 6 h. The solvent was evaporated under reduced pressure to obtain the crude product, which was separated by silica gel column chromatography using a petroleum ether and ethyl acetate mixture as elution to give the product 2-hydroxy-3-n-dodecylmercapto-1,4-naphthoquinone. Product A5 (0.218 g, 0.58 mmol, yield 50.6%) was obtained as a red solid; Rf = 0.26 (methanol-dichloromethane 1:20). HR-MS (ESI) m / z: Calcd for C 22 H 30 O3S[M+H + 375.1988, found: 375.2033. Mass spectrum attached. Figure 5 As shown.

[0054] Example 4

[0055] 2-Hydroxy-3-n-tetradecyl-1,4-naphthoquinone (A14),

[0056] Under argon protection, 2-hydroxy-1,4-naphthoquinone (0.2 g, 1 eq) was dissolved in 15 mL of DMF solution, and tetradecyl mercaptan (0.472 mL, 1.5 eq) was added. The mixture was stirred, and then triethylamine (0.16 mL, 1 eq) was added. The reaction was carried out at 70 °C for 6 h. The solvent was evaporated under reduced pressure to obtain the crude product, which was separated by silica gel column chromatography using a mixture of petroleum ether and ethyl acetate as elution, yielding 2-hydroxy-3-tetradecylmercapto-1,4-naphthoquinone. Product A6 (0.237 g, 0.59 mmol, yield 51.2%) was obtained as a purple-red solid; Rf = 0.27 (methanol-dichloromethane 1:20). HR-MS (ESI) m / z: Calcd for C 24 H 34 O3S[M+H + 403.2301, found: 403.2301. Mass spectrum attached. Figure 6 As shown.

[0057] Example 5

[0058] 2-Hydroxy-3-p-tert-butylbenzylmercapto-1,4-naphthoquinone (A22),

[0059] Under argon protection, 2-hydroxy-1,4-naphthoquinone (0.2 g, 1 eq) was dissolved in 15 mL of DMF solution, and p-tert-butylbenzyl mercaptan (0.322 mL, 1.5 eq) was added. The mixture was stirred, and then triethylamine (0.16 mL, 1 eq) was added. The reaction was carried out at 70 °C for 6 h. The solvent was evaporated under reduced pressure to obtain the crude product, which was separated by silica gel column chromatography using a mixture of petroleum ether and ethyl acetate as elution, yielding the product 2-hydroxy-3-p-tert-butylbenzyl mercaptan-1,4-naphthoquinone. Product A10 (0.209 g, 0.59 mmol, yield 51.6%) was obtained as an orange-red solid; Rf = 0.28 (methanol-dichloromethane 1:20). HR-MS (ESI) m / z: Caled for C 21 H 20 O3S[M+Na + 375.1025, found: 375.1032. Mass spectrum attached. Figure 7 As shown.

[0060] Minimum inhibitory activity (MIC) test

[0061] Compounds A1-A22 and 2-hydroxy-1,4-naphthoquinone were dissolved in DMSO to prepare compound solutions. Bacteria were cultured to the logarithmic growth phase, and OD values ​​were measured. 600 =0.1, dilute the bacterial suspension to a concentration of 1×10. 7 CFU.mL -1First, add 100 μL of LB broth to each well of a 96-well plate. Then, add 1 μL of the above-mentioned bacterial culture, followed by 1 μL of the prepared A1-A22 and 2-hydroxy-1,4-naphthoquinone compound solutions. Finally, bring the volume of the mixture in each well to 200 μL with LB broth. The final concentration of the compounds in the 96-well plate is 1000 μmil·L⁻¹. 1 500 μmol·L -1 250 μmol·L -1 125 μmol·L -l 62.5 μmol·L -1 31.25 μmol·L -1 15.625 μmol·L -1 7.8125 μmol·L -1 Three parallel wells containing only culture medium and only bacterial suspension (with 1 μL of DMSO added) were set up as controls and incubated at 37°C for 16 h. The 96-well plate was then placed in a microplate reader, and the absorbance was measured at 600 nm. The inhibition rate of each compound was calculated, and the lowest concentration of the compound with an inhibition rate ≥90% was taken as the MIC of that compound.

[0062] Example 5

[0063] Minimum bactericidal activity (MBC) test

[0064] Bacteria were cultured to the logarithmic growth phase, and OD was measured. 600 =0.1, dilute the bacterial suspension to a concentration of 1×10. 7 CFU.mL -1 First, add 100 μL of LB broth to each well of a 96-well plate, then add 1 μL of the above bacterial suspension, followed by 1 μL of the prepared Al-A22 and 2-hydroxy-1,4-naphthoquinone compound solutions. Finally, bring the total volume to 200 μL with LB broth. The final compound concentration in the 96-well plate should be greater than or equal to the MIC value. Three parallel sub-wells were added, and the plates were incubated at 37°C for 24 h. Then, 50 μL of the mixture was sequentially aspirated and added to LB agar plates, incubated at 37°C for 24 h, and the colony status was observed. The concentration of the compound with fewer than 5 single colonies on the agar plate was considered the minimum bactericidal concentration.

Claims

1. Compounds of general formula (A): R is either R1 or R2 in: n is 2, 3, 4, 5, 6, 8, 10, 13, 14, 16, 18 or 19; 2. A pharmaceutical composition comprising a compound of general formula (A) according to claim 1.

3. The use of the compound of general formula (A) according to claim 1 as an antibacterial agent against Gram-positive bacteria Staphylococcus aureus.

4. According to the application described in claim 3, the bactericidal mechanism of the compound of general formula (A) is to disrupt cell membrane permeability and to bind with reactive oxygen species inside bacterial cells to achieve the purpose of sterilization.

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