CN114573568B - 2-quinolinone-citrinin hybrid dimer compound and preparation method and application thereof - Google Patents
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Abstract
The invention belongs to the technical field of natural polymers, and particularly relates to a 2-quinolinone-citrinin hybrid dimer compound and a preparation method and application thereof. The 4-hydroxy-1-methylquinolin-2-one alkaloid, a monomer in the 2-quinolinone-citrinin heterodimer compounds of the present invention, is currently mainly obtained from the plant rutaceae, euphorbiaceae and soil-derived actinomycetes dactylosporium sp. Has wide biological activity, such as platelet aggregation resistance, chymotrypsin inhibition, anti-tumor, antibacterial, antituberculosis and other activities. Can be applied to the field of preparing antibacterial drugs.
Description
Technical Field
The invention belongs to the technical field of natural small molecular medicines, and particularly relates to a 2-quinolinone-citrinin hybrid dimer compound and a preparation method and application thereof.
Background
2-quinolinones are a class of biologically active compounds with a broad range of actions. Beginning in the seventh eighties of the last century, scholars at home and abroad have conducted extensive synthesis research on quinolinone compounds, and the quinolinone compounds are mainly used for developing inhibitory active drugs for central systems such as anticonvulsant, analgesic and hypnotic effects. Natural 2-quinolinone alkaloids mainly originate from plants and animals, and are reported as traditional Chinese medicines such as dung beetles (1-5), cortex Dictamni (6-14), and fructus Zanthoxyli (15-21), and are shown in the following formula (I). The alkaloid from dung beetles is basically consistent with natural coumarin, and only 2, 3-double bonds are frequently disappeared. The natural alkaloid of pricklyash peel is characterized in that substitution is carried out on one side of a lactam ring, 3-hydroxyl group is poly-isopentenyl group and forms a pyran ring, methyl substitution is carried out on 1-N, and a natural 2-quinolinone alkaloid zanthobisquinone is obtained and is connected through methylene. The alkaloid is mainly used for treating tumors, skin diseases and the like.
Quinolinone alkaloids are widely distributed in plants, but rarely in filamentous fungi. The structures reported in the literature are of a significantly more complex origin than in plants, and are characterized by the fact that they contain a 4-phenyl-3, 4-dihydroxyquinolinone-2-one structural unit, while at the same time there is often one or two isopentenyl substitutions at the 7-position on the benzene ring side, while isopentenyl groups can form furan, pyran or six-membered carbocycles, while the corresponding isomers of such structures are also produced from the cyclic structural positions, and to date no chiral enantiomer on the lactam side has been reported. 2-quinolinone alkaloids derived from marine filamentous fungi are rich in backbone and often exist in enantiomeric forms. The compounds have almost no activity in the prior antitumor activity report; the plant has nematicidal and antiviral activities at all times, and has no report on antibacterial activity. The activity of such compounds is therefore to be further exploited.
Disclosure of Invention
To overcome the above-mentioned drawbacks and disadvantages of the prior art, a primary object of the present invention is to provide a class of 2-quinolinone-citrinin heterodimer compounds.
It is another object of the present invention to provide a process for preparing the above 2-quinolinone-citrinin heterodimer compound.
It is still another object of the present invention to provide the use of the above 2-quinolinone-citrinin heterodimer compound in the preparation of antibacterial drugs. The 2-quinolinone-citrinin hybrid dimer compound of the present invention has excellent antibacterial activity.
The aim of the invention is achieved by the following scheme:
the 2-quinolinone-citrinin hybrid dimer compound is one of the following structural formulas:
the 2-quinolinone-citrinin heterozygous dimer compound can be prepared by the following steps: fermenting and culturing marine fungus Penicillium sp.GGF16-1-2 (GDMCC No. 61080), extracting the obtained fermentation broth with ethyl acetate, concentrating to obtain fermentation broth EtOAc phase extract; separating by silica gel column chromatography, gradient eluting with petroleum ether/ethyl acetate at volume ratio of 100:0-0:100 as eluent, and distinguishing and combining according to thin layer chromatography result to obtain 6 crude components Fr1-Fr6; components vpe:vetoac=85:15-75:25 were selectedCarrying out Sephdex LH-20 column chromatography, methanol elution and thin layer chromatography tracking on Fr2 to obtain five subflows Fr2-1 to Fr 2-5; HPLC separation of Fr2-3 subfraction, V Acetonitrile :V Water and its preparation method The mixture is eluted with the concentration of 80:20 and the concentration of 1.0mL/min, 4 components are obtained by separation, wherein 3 compounds with the residence time of 40-60min are the 2-quinolinone-citrinin heterozygous dimer compound.
In one example, the residence time of the 2-quinolinone-citrinin heterodimer compound in the HPLC separation is 45.410min (Compound I), 51.927min (Compound II), 47.327min (Compound III), respectively.
The Fr2-3 subfraction is a bright yellow solution, and is expressed as R in TCL f =0.3-0.8, 254nm dark spots, 365nm pale yellow spots.
The Fr2-1 subfraction was a pale yellow solution with no apparent spots in TCL.
The Fr2-2 subfraction is a dark red solution, and is expressed as R in TCL f =0.1-0.45, 254nm dark spots, 365nm yellow spots.
The Fr2-4 subfraction is a yellow solution, and is expressed as R in TCL f =0.3-0.9, 254nm dark spots, 365nm beige→orange-red spots (ethanol sulfate can develop).
The Fr2-5 subfraction is a light yellow solution, and is expressed as R in TCL f =0.3-0.9, 254nm dark spots, 365nm yellow spots.
The HPLC column was a Kromasil semi-preparative column, 10mm X250 mm,5 μm, akzo Nobel, sweden.
In one embodiment, the Fr2-3 subfraction is applied at 143mg and the resulting compound I is 30.8mg; 6.0mg of compound II; compound III is 2.0mg.
The parameters of the silica gel column chromatography are 10cm in diameter, 110cm in length, silica gel H and 3.85L in column volume. 165g (7 cm in height) of sample-mixing silica gel and 1.65kg (57 cm in height) of column-packed silica gel.
The Sephadex LH-20 column chromatography parameters are phi=3.0 cm, l=200 cm, LH-20 (40-60 μm) 100g, height 160cm and column volume 300mL.
The fermentation culture can be obtained by conventional culture methods. If the strain is activated in PDA culture medium, inoculating in liquid culture medium for culturing to obtain fermentation liquid.
The formula of the PDA culture medium can be as follows: 200g of potato, 20g of glucose and 35g of sea salt in each 1L of water.
The formula of the liquid culture medium can be as follows: in each 1L of liquid culture medium, maltose 40g, tryptophan 5g, sorbitol 50g, monosodium glutamate 10g and KH 2 PO 4 5g、MgSO 4 ·7H 2 O3 g, yeast extract 13g, and the balance of aged seawater, and the pH is 6.5.
More specifically, the activation can be performed by picking the strain into a flask containing PDA medium, and culturing in a constant temperature shaker (165 r/min) at 28℃for 2 days to obtain a seed solution. In a 250mL flask, the amount of PDA medium is preferably 100mL.
More specifically, the inoculation culture may be performed by adding the activated seed solution to a liquid medium and performing stationary culture at 28℃for 60 days. The amount of inoculation is preferably 1.5mL seed solution per 400mL liquid medium. 150L of medium was co-cultured. A2L blank liquid medium was used as a control.
The resulting fermentation broth may be inactivated by the addition of methanol, more preferably by the addition of 10-20mL methanol to 400mL liquid medium. And (5) inactivating and filtering to obtain fermentation liquor and mycelium.
The fermentation broth is preferably concentrated from 150L to 10L and then subjected to an extraction operation.
The ethyl acetate used is preferably equal in volume to the fermentation broth. Preferably 3 times.
The 2-quinolinone-citrinin hybrid dimer compound has extremely strong antibacterial activity and can be applied to the field of preparation of antibacterial medicines.
The 4-hydroxy-1-methylquinolin-2-one alkaloid, a monomer in the 2-quinolinone-citrinin heterodimer compounds of the present invention, is currently mainly obtained from the plant rutaceae, euphorbiaceae and soil-derived actinomycetes dactylosporium sp. Such substances have a wide range of biological activities, such as anti-platelet aggregation, chymotrypsin inhibition, anti-tumor, antibacterial and anti-tuberculosis activities.
Drawings
FIG. 1 is an HPLC chart of HPLC separation of Fr 2-3.
FIG. 2 shows the compounds I and II 1 H- 1 H COSY and HMBC correlation diagrams.
FIG. 3 is a diagram of a compound III 1 H- 1 H COSY and HMBC correlation diagrams.
FIG. 4 is a CD graph of compound I (A) and compound II (B).
Figure 5 is an XRD diffractogram of compound iii.
FIG. 6 is a CD plot of Compound III.
Detailed Description
The present invention will be described in further detail with reference to examples and drawings, but embodiments of the present invention are not limited thereto. The materials referred to in the examples below are available commercially unless otherwise specified. The method is conventional unless otherwise specified.
The marine fungus Penicillium sp.GGF16-1-2 is obtained by separating a No. 16 starfish sample from a sea area near the Gong island of Sandong in south China, and is preserved in the microorganism strain preservation center, GDMCC for short, address: guangzhou early China No. 100 university 59 building 5 Guangdong province microbiological institute, deposit No. GDMCC No. 61080, date of deposit: 7 months and 7 days 2020.
Example 1: isolation of 2-quinolinone-citrinin heterodimer compounds
(1) Strains: penicillium sp.GGF16-1-2GDMCC No. 61080.
PDA medium: 200g of potato, 20g of glucose, 35g of sea salt, 1L of pure water and natural pH;
liquid medium: 1L, maltose 40g, tryptophan 5g, sorbitol 50g, monosodium glutamate 10g, KH 2 PO 4 5g、MgSO 4 ·7H 2 O3 g, yeast extract 13g, and the balance of aged seawater, and the pH is 6.5.
(2) Activating strains: the seed solution was obtained by picking up the seed from the seed tube with an inoculating loop into a flask (100 mL/250 mL) containing PDA medium, and culturing it in a thermostatic shaker (165 r/min) at 28℃for 2 days.
(3) Inoculating: sucking 1.5mL of the activated seed culture solution into a 1000mL culture flask filled with 400mL of liquid culture medium, standing and culturing at 28 ℃ for 60 days, and co-culturing for 150L.
(4) Extracting: adding 10-20mL of methanol into each culture flask to inactivate strains, filtering with gauze, and separating fermentation broth and mycelium; concentrating the obtained fermentation liquor to 10L, extracting with an equal volume of EtOAc three times, concentrating the obtained extract, and finally obtaining a fermentation liquor EtOAc phase extract;
(5) Separating: crude extracts of EtOAc portions (111.9 g of EtOAc phase extract of fermentation broth) were obtained in the 150L liquid medium above and subjected to column chromatography on silica gel (phi=10 cm, l=110 cm, H, 165g of sample-mixed silica gel (height 7 cm), column-packed silica gel 1.65kg (height 57 cm), column volume 3.85L) and petroleum ether-ethyl acetate system elution (VPE: VEtOAc=100:0.fwdarw.0:100, 1L/bottle), and 7 crude fractions Fr.1 to Z1-Fr.7 were obtained by combining according to the thin layer TLC. Fr2 (VPE: VEtOAc=85:15-75:25 fractions) was selected for Sephdex LH-20 column chromatography (phi=3.0 cm, l=200 cm, LH-20 (40-60 μm), 100g, height 160cm, column volume 300mL, pure methanol elution), 1.5g samples were loaded each time, 25mL test tubes were subjected to liquid, and TLC plate tracking yielded five subflows Fr2-1 to Fr2-5 in total. Wherein Fr2-1 (pale yellow solution): TLC showed no apparent spots; fr2-2 (dark red solution): r is R f =0.1-0.45, 254nm dark spots, 365nm yellow spots; fr2-3 (bright yellow solution): r is R f =0.3-0.8, 254nm dark spots, 365nm pale yellow spots; fr2-4 (yellow solution): r is R f =0.3-0.9, 254nm dark spots, 365nm beige→orange-red spots (ethanol sulfate can develop); fr2-5 (pale yellow solution): r is R f =0.3-0.9, 254nm dark spots, 365nm yellow spots.
Fr2-3 (yellow gum, 143 mg) was purified by HPLC (Kromasil semi-preparative chromatography column, 10 mm. Times.250 mm,5 μm, akzo Nobel, sweden), acetonitrile water eluted (V Acetonitrile :V Water and its preparation method =80:20, 1.0 ml/min) and the result is shown in fig. 1, giving 4 component compounds in sequence, peak1 (4.5 mg, τ R = 15.027 min), peak2 (compound i, 30.8mg, τ R = 45.410 min), peak3 (compound iii, pale yellow crystalsBody, 2.0mg, τ R = 47.327 min) and peak4 (compound ii, 6.0mg, τ R ＝51.927min)。
Example 2: identification of 2-quinolinone-citrinin heterodimer compounds
(1) Compound i, orange amorphous powder, positive ion hresis gives excimer ion peak m/z:394.1647[ M+H ]] + (calculated as m/z 394.1649[ M+H ]] + ) Determining the molecular formula of C 23 H 23 NO 5 The unsaturation was 13. The maximum absorption wavelength in the ultraviolet spectrum is shown as: 220nm, 286nm and 320nm. Carbonyl group (1722 cm) is shown in the infrared spectrum -1 、1635cm -1 ) Benzene ring (1608 cm) -1 、1519cm -1 、1494cm -1 And 1457cm -1 ) A group.
1 H and 13 the results of the C NMR spectrum show that most of the signals occur in pairs with a 1:1 ratio, possibly due to the compound in CDCl 3 The presence of intramolecular hydrogen bonds in the solvent results in its rotation being hindered and two sets of spectra appear. 1 The H NMR spectrum shows 4 aromatic proton signals delta H 7.27 (1H, t,7.6,2.0Hz, H-7), 7.35 (1H, d,8.0Hz, H-9), 7.55 (1H, t,7.2,1.2Hz, H-8), 8.16 (1H, d,8.0Hz, H-6) and methyl signals 3.74/3.75 (3H, s, 1-CH) 3 ) The display structure contains quinolinone structural fragments; 3 methyl signals delta H 1.19/1.12 (3H, d,7.2Hz, H-11 '), 1.32/1.21 (3H, d,6.8Hz, H-12 '), 1.99/1.98 (3H, s, H-13 '), 2 methine signals δH2.86/2.86 (1H, m,7.2Hz, H-3 ') and δH2.59/4.58 (1H, q,2.4Hz, H-2 ') and 1 double bond proton signal δH H 7.91/7.91 (1H, s, H-10') shows that the structure contains a citrinin fragment A (dashed box portion in FIG. 2); 1 methylene signal delta H 3.82/3.85 (2H, d,8.0Hz, H-1 "); 2 active proton signals delta H 12.08/11.99(1H,s,8'-OH)、12.78/12.71(1H,s,4-OH)。 13 There are 40 total carbon signals in the C NMR spectrum, most of which occur in pairs, showing 11 quaternary carbons (including 1 ketocarbonyl group), 7 methines, 1 methylene and 4 methyl signals in combination with DEPT 135.
Further analyzing the two-dimensional spectrogram (HSQC), 1 H- 1 H COSY, HMBC) (see Table1) Analysis of CH 3 -11' and H-2', H-2' and H-3', H-3' and CH 3 Between-12 1 H- 1 H COSY-related signals, H-2 'and C-3', C-4', C-9', H-3 'and C-4', C-5', C-9', C-12', H-10' and C-2', C-8', C-9',11' -CH in HMBC spectra 3 With C-2', C-3', CH 3 -12' and C-2', C-3', C-4',13' -CH 3 The signals associated with C-4', C-5', C-6', C-7' and 8' -OH and C-8', C-9' illustrate that the structure contains a 7-substituted decarboxylated citrinin fragment A; between H-6, H-7, H-8 and H-9 1 H- 1 H COSY-related signals and signals related to H-6 and C-4, C-5, C-10, H-8 and C-7, C-9, C-10, H-9 and C-5, C-10, N-CH3 and C-10, C-2 in HMBC spectra indicate that the structures contain 4-hydroxy-1-methylquinolin-2-one fragments. The two relevant fragments of H-1 'and C-2, C-3, C-4, C-6', C-7', C-8' are directly linked via a methylene group, thereby determining the planar structure of compound I (see FIG. 2).
To determine the steric configuration of compound I, NOESY tests were performed on it to determine the relative configuration of C-2', C-3'. NOESY results showed H-2 'and 12' -CH 3 H-3 'and 11' -CH 3 There are associated signals, respectively, indicating that the methyl groups on C-2', C-3' are trans, i.e., 2'R,3' S. The absolute configuration of citrinin compounds C-2', C-3' is determined to be 2'R,3' S from the biological genetic point of view by ECD and VCD calculation according to the prior literature. Optical rotation test and CD of Compound I with specific optical rotation value
-79.1 (c 0.055, meOH). The CD test results showed a (+) cotton effect at 230, 256, and 283nm and a (-) cotton effect at 332nm (see A in FIG. 4), which was found to be a new compound, designated as penicilloneine A.
TABLE 1 NMR data for Compound I (CDCl 3 )
Annotation: 1 H NMR(400MHz), 13 c NMR (100 MHz), a represents weightAnd (5) stacking peaks.
(2) Compound ii, orange amorphous powder, positive ion hresis gives excimer ion peak m/z:408.1812[ M+H ]] + (calculated as m/z:408.1811[ M+H)] + ) Determining the molecular formula of C 24 H 25 NO 5 The unsaturation was 13. The maximum absorption wavelength in the ultraviolet spectrum is shown as: 220nm and 320nm. Carbonyl group (1726 cm) is shown in the infrared spectrum -1 、1631cm -1 ) Benzene ring (1494 cm) -1 And 1448cm -1 ) A group.
The 1D and 2D NMR data (see Table 2) for compound II are highly similar to the nuclear magnetic data for compound I, indicating that compound II is of the same type of structure as compound I. The difference is that the compound II 1 The H NMR spectrum has no methylene signal and one more methyl signal delta H 1.79/1.78 (3H, d,7.2Hz, H-2'), delta H 4.86/4.78 (1H, q,6.4Hz, H-1 "). Bonding of 13 The disappearance of the methylene signal was found in C NMR and DEPT135 patterns and 1 methyl signal delta was present C 15.5,15.4/15.3,15.3 (q, C-1 "). Comparison of HMBC 1 H- 1 The H COSY pattern shows that the planar structure of the compound II is similar to that of the compound I, and the compound II consists of a 4-hydroxy-1-methylquinolin-2-one fragment and a citrinin fragment A which are connected through a carbon bridge, but one proton on C-1' is replaced by a methyl group (see figure 2). 1 H NMR spectrum results show that most of the signal occurs in pairs and in a 1:1 ratio, possibly due to the compound in CDCl 3 The presence of intramolecular hydrogen bonds in the solvent results in two sets of signals due to blocked rotation, and furthermore, since C-1' is chiral carbon, epimers may be present resulting in four sets of signals (1:1:1:1) of carbon spectrum.
TABLE 2 NMR data for Compound II (CDCl) 3 )
Annotation: 1 H NMR(400MHz), 13 c NMR (100 MHz), a represents an overlapping peak.
NOESY spectrum results show that the relative spatial configuration of C-2', C-3' of the compound II is the sameWhile the steric configuration was determined to be 2'R,3' S according to compound I. In addition, C-1 ' cannot determine its relative configuration by NOESY, NOE, etc. due to its spatial position far from C-2', C-3', and thus cannot determine the spatial configuration of that position. The compound is subjected to optical rotation test and CD, and has specific optical rotation value of
-38.2 (c 0.21, meoh). The CD test results showed a (+) chip effect at 215, 233, and 283nm and a (-) chip effect at 334nm (see B in FIG. 4). The new compound is identified by searching and named as penicilloneine B.
(3) Compound iii, pale yellow bulk crystals, positive ions hresis gives excimer ion peaks m/z:410.1598[ M+H ]] + (calculated as m/z:410.1598[ M+H ]] + ) Excimer ion peak m/z given by negative ion hresis: 408.1447[ M-H ]] - (calculated as m/z:408.1452[ M-H ]] - ) Determining the molecular formula of C 23 H 23 NO 6 The unsaturation was 13. The maximum absorption wavelength in the ultraviolet spectrum is shown as: 220nm, 280nm and 320nm. The infrared spectrum shows hydroxyl groups (3288 cm -1 ) Carbonyl (1644 cm) -1 ) Benzene ring (1544 cm) -1 ) A group.
The 1D NMR data (see Table 3) of compound III was similar to the nuclear magnetic data of compound I, and the structure also contained a 4-hydroxy-1-methylquinolin-2-one fragment, except that the citrinin structural fragment of compound III was changed. 1 Two methine signals delta containing citrinin fragments in H NMR spectra H 4.68 (1H, q,7.2Hz, H-2 '), 2.98 (1H, q,7.2Hz, H-3'), and three methyl signals delta H 1.25(3H,m,H-11′)、δ H 1.25 (3H, m, H-12 '), 2.14' (3H, s, H-13 ') all move to the low field. 13 There are 23 carbon signals in total in the C NMR spectrum, showing 12 quaternary carbons (including 2 ketocarbonyl signals), 6 methines, 1 methylene and 4 methyl signals in combination with DEPT 135.
Further analyzing the two-dimensional spectrogram (HSQC), 1 H- 1 H COSY, HMBC) (see Table 3), analysis of 11' -CH 3 And H-2', H-2' and H-3', H-3' and 12' -CH 3 Between (a) and (b) 1 H- 1 H COSY-related signals, H-2' and C-3', C-4', C-10', C-12', H-3' and C-4', C-5', C-9', C-12',11' -CH in HMBC spectra 3 With C-2', C-3',12' -CH 3 With C-2', C-3',13' -CH 3 The signals associated with C-4', C-5', C-6', C-7' and 8' -OH and C-7', C-8', C-9' indicate that C-10' in the citrinin fragment is a ketocarbonyl group, thus confirming that the planar structure thereof is composed of a 4-hydroxy-1-methylquinolin-2-one fragment and citrinin fragment B linked by a carbon bridge (see FIG. 3).
The stereochemistry of compound III was assumed to be 2'R,3' S from the C-2', C-3' stereochemistry of compound I and compound II, and was confirmed by single crystal diffraction (XRD) experiments for the first time (see Table 4, XRD transmission pattern 5), with an absolute configuration constant (Flack parameter) of 0.08 (7). The compound is subjected to optical rotation test and CD, and has specific optical rotation value of
66.5 (c 0.04, meOH). The CD test results showed a (+) cotton effect at 230, 270 and 320nm (see FIG. 6). The compound was not reported by searching, and was identified as a novel compound, designated as penicilloneine C.
TABLE 3 NMR data for Compound III (CDCl 3 )
Annotation: 1 H NMR(400MHz), 13 c NMR (100 MHz), a represents an overlapping peak.
TABLE 4 Crystal data for Compound III
Example 3: antibacterial property test of 2-quinolinone-citrinin hybrid dimer compound:
because only antibacterial performance tests are performed, the invention applies for the protection compound and the application thereof, and the protection compound is fully disclosed and characterized for various antibacterial performances, so that the application range of the protection compound is enlarged, and various strains including positive bacteria and negative bacteria can be considered for the test.
(1) Hypha growth rate method: compound liquid medicine with different concentrations (3 times of repetition are set for each concentration) is prepared and poured into a culture dish for standby, and a PDA culture medium plate with sterile water is used as a control. Taking cultured pathogenic bacteria cake (Staphylococcus epidermidis, staphylococcus aureus, methicillin-resistant Staphylococcus aureus, escherichia coli and Pseudomonas aeruginosa supplied by Guangdong microorganism) with sterile puncher, inoculating to the center of PDA culture medium plate, and culturing in 28 deg.C constant temperature incubator for 3 days. The positive control was carbendazim. The colony diameter was measured by the crisscross method, and the hypha growth inhibition rate was calculated. Solving a virulence regression equation, and further calculating LC 50 。
Hypha growth inhibition ratio = [ (control colony diameter-treated colony diameter)/(control colony diameter-cake diameter) ] ×100%.
(2) Antibacterial activity against anthrax bacteria:
compound liquid medicine with different concentrations (3 times of repetition are set for each concentration) is prepared and poured into a culture dish for standby, and a PDA culture medium plate with sterile water is used as a control. Taking cultured pathogenic bacteria cake (anthracnose strain and powdery mildew of fruit tree are provided by fruit tree institute of agricultural sciences, guangdong province) with a sterile puncher, inoculating to the center of PDA culture medium plate, and culturing in a constant temperature incubator at 28deg.C for 3 days. The positive control was carbendazim. The colony diameter was measured by the crisscross method, and the hypha growth inhibition rate was calculated. Solving a virulence regression equation, and further calculating LC 50 。
Hypha growth inhibition ratio = [ (control colony diameter-treated colony diameter)/(control colony diameter-cake diameter) ] ×100%.
TABLE 5 antibacterial Activity of Compounds I, II, III (LC 50 、μg/mL)
Note that: staphylococcus epidermidis A, staphylococcus aureus B, methicillin-resistant staphylococcus aureus C, escherichia coli D, pseudomonas aeruginosa E, papaya anthracis, powdery mildew G, H carbendazim (positive control) and kanamycin I (positive control). "-" represents no test.
The 2-quinolinone-citrinin hetero-dimer compound has extremely strong antibacterial activity (see table 5).
The above examples are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above examples, and any other changes, modifications, substitutions, combinations, and simplifications that do not depart from the spirit and principle of the present invention should be made in the equivalent manner, and the embodiments are included in the protection scope of the present invention.
Claims (10)
1. A2-quinolinone-citrinin hybrid dimer compound is characterized by being one of the following structural formulas:
2. a process for the preparation of a 2-quinolinone-citrinin heterodimeric compound as defined in claim 1, comprising the steps of:
fermenting and culturing marine fungus Penicillium sp.GGF16-1-2, extracting the obtained fermentation broth with ethyl acetate, and concentrating to obtain fermentation broth EtOAc phase extract; separating by silica gel column chromatography, gradient eluting with petroleum ether/ethyl acetate at volume ratio of 100:0-0:100 as eluent, and distinguishing and combining according to thin layer chromatography result to obtain 6 crude components Fr1-Fr6; selecting V PE :V EtOAc Component Fr 2=85:15-75:25 was subjected to Sephdex LH-20 column chromatography, methanol elution, and thin layer chromatography tracking to obtain five subfractions Fr2-1 to Fr 2-5; HPLC separation of Fr2-3 subfraction, V Acetonitrile :V Water and its preparation method Eluting with the solution of (E) =80:20, eluting with the solution of 1.0mL/min, and separating to obtain 4 components, wherein 3 compounds with residence time of 40-60min are 2-quinolineKetone-citrinin hybrid dimer compounds;
the marine fungus Penicillium sp.GGF16-1-2 has a deposit number of GDMCC No. 61080.
3. The preparation method according to claim 2, characterized in that:
the residence time of the 2-quinolinone-citrinin hetero-dimer compound in the HPLC separation is 45.410min compound I, 51.927min compound II and 47.327min compound III respectively.
4. A method of preparation according to claim 3, characterized in that:
the Fr2-3 subfraction is a bright yellow solution, and is expressed as R in TCL f =0.3-0.8, 254nm dark spots, 365nm pale yellow spots;
the Fr2-1 subfraction is a light yellow solution, and no obvious spots exist in TCL;
the Fr2-2 subfraction is a dark red solution, and is expressed as R in TCL f =0.1-0.45, 254nm dark spots, 365nm yellow spots;
the Fr2-4 subfraction is a yellow solution, and is expressed as R in TCL f =0.3-0.9, 254nm dark spots, 365nm beige→orange-red spots;
the Fr2-5 subfraction is a light yellow solution, and is expressed as R in TCL f =0.3-0.9, 254nm dark spots, 365nm yellow spots.
5. The preparation method according to claim 2, characterized in that:
the HPLC column was a Kromasil semi-preparative column, 10mm X250 mm,5 μm, akzo Nobel, sweden.
6. The preparation method according to claim 2, characterized in that:
the parameters of the silica gel column chromatography are 10cm in diameter, 110cm in length, silica gel H and 3.85L in column volume; wherein, 165g of sample-mixing silica gel is mixed with the sample and the height is 7cm; column packed silica gel 1.65kg, height 57cm.
7. The preparation method according to claim 2, characterized in that:
the parameters of Sephdex LH-20 column chromatography are phi=3.0 cm, l=200 cm, LH-20 100g, height 160cm and column volume 300mL.
8. The preparation method according to claim 2, characterized in that: the fermentation culture is to obtain fermentation liquor by activating the strain in PDA culture medium and inoculating the strain in liquid culture medium for culture.
9. The method of manufacturing according to claim 8, wherein:
the formula of the PDA culture medium is as follows: 200g of potato, 20g of glucose and 35g of sea salt in each 1L of water;
the formula of the liquid culture medium is as follows: in each 1L of liquid culture medium, maltose 40g, tryptophan 5g, sorbitol 50g, monosodium glutamate 10g and KH 2 PO 4 5g、MgSO 4 ·7H 2 O3 g, yeast extract 13g, and the balance of aged seawater, wherein the pH is 6.5;
the activation is carried out by picking the strain into a culture bottle filled with PDA culture medium, and placing the culture bottle in a constant temperature shaking table at 28 ℃ for 2 days to obtain seed liquid; the inoculation culture is carried out by adding the activated seed liquid into a liquid culture medium, and standing and culturing for 60 days at 28 ℃.
10. Use of a 2-quinolinone-citrinin heterodimer compound according to claim 1 for the preparation of an antibacterial drug.
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| CN104478891A (en) * | 2014-12-18 | 2015-04-01 | 福州大学 | Penicillium citrinum sourced citrinin compound (penicitrinol O) as well as preparation method and application thereof |
| CN107325087A (en) * | 2016-05-27 | 2017-11-07 | 福州大学 | Citrinin compounds dicitrinone D and its application in terms of malignant mela noma |
| CN111139188A (en) * | 2020-01-08 | 2020-05-12 | 广州中医药大学(广州中医药研究院) | Novel skeleton heteroterpene derivative derived from marine fungi and application of novel skeleton heteroterpene derivative in preparation of anti-inflammatory drugs |
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| CN104478891A (en) * | 2014-12-18 | 2015-04-01 | 福州大学 | Penicillium citrinum sourced citrinin compound (penicitrinol O) as well as preparation method and application thereof |
| CN107325087A (en) * | 2016-05-27 | 2017-11-07 | 福州大学 | Citrinin compounds dicitrinone D and its application in terms of malignant mela noma |
| CN111139188A (en) * | 2020-01-08 | 2020-05-12 | 广州中医药大学(广州中医药研究院) | Novel skeleton heteroterpene derivative derived from marine fungi and application of novel skeleton heteroterpene derivative in preparation of anti-inflammatory drugs |
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