Article pubs.acs.org/JAFC
Cite This: J. Agric. Food Chem. 2018, 66, 5368−5372
New Antimicrobial Cyclopentenones from Nigrospora sphaerica ZMT05, a Fungus Derived from Oxya chinensis Thunber Zhihui Wu,† Zihui Xie,† Manlin Wu,† Xiaoqi Li,† Weilin Li,† Weijia Ding,† Zhigang She,‡ and Chunyuan Li*,† †
College of Materials and Energy, South China Agricultural University, Guangzhou 510642, China School of Chemistry and Chemical Engineering, Sun Yat-sen University, Guangzhou 510275, China
‡
S Supporting Information *
ABSTRACT: Six new cyclopentenone derivatives (+)-nigrosporione A (+)-1, (−)-nigrosporione A (−)-1, nigrosporione B (2), nigrosporione C (3), (+)-nigrosporione D (+)-4, and (−)-nigrosporione D (−)-4 were isolated from an endophytic fungus Nigrospora sphaerica ZMT05, collected from the rice grasshopper (Oxya chinensis Thunberg), which is an insect pest in rice and which is also used as a food for people in some countries. Their planar and spatial structures were determined by spectroscopic analyses and eletronic circular dichroism (ECD) calculations. Compounds (+)-1, (−)-1, and 2 inhibited the plant pathogens Fusarium oxysporum, Colletotrichum musae, Penicillium italicum, and Fusarium graminearum, compounds 3 and (−)-4 inhibited F. oxysporum, C. musae, and P. italicum, and compound (+)-4 inhibited F. oxysporum, C. musae, and F. graminearum, showing antifungal activities stronger than triadimefon. Additionally, compounds (+)-1, (−)-1, 2, and 3 displayed moderate antibacterial activities against Staphyloccocus aureus and Escherichia coli. KEYWORDS: insect-derived fungi, Nigrospora sphaerica, cyclopentenone, antifungal activity, Oxya chinensis Thunberg
■
Ultraviolet (UV), infrared radiation (IR), circular dichroism (CD), and high resolution electrospray ionization mass spectroscopy (HRESIMS) spectra were obtained using UV-2550 spectrophotometer (Shimadzu Corporation, Tokyo, Japan), Nicolet iS10 Fourier transform infrared spectrophotometer (Thermo Electron Corporation, Madison, WI), Chirascan CD spectrometer (Applied Photophysics Ltd., London, U.K.), and quadrupole-time of flight (Q-TOF) mass spectrometer (Thermo Fisher Scientific Inc., Frankfurt, Germany), respectively. Column and thin layer chromatography (TLC) were performed using 200−300 mesh silica gel and G60, F-254 silica gel plates (Qingdao Haiyang Chemicals Co., Ltd., Qingdao, China), respectively. Chiral purification was carried out using a 1260 Infinity Series highperformance liquid chromatography (HPLC) system (Agilent Corporation, Santa Clara, CA). The HPLC column used was a 250 mm × 4.6 mm i.d., 5 μm, Lux Cellulose-2, without a guard column. The HPLC solvents and other chemicals adopted were of chromatographic and analytical pure grades, respectively. Fungal Material. Nigrospora sphaerica ZMT05 was isolated from Oxya chinensis Thunberg and was stored in the College of Materials and Energy, South China Agricultural University. This fungus was identified through molecular analyses. The use of BLAST disclosed that its internal transcribed spacers (ITS) sequence (No. MG171196 in GenBank) was identical to those of two Nigrospora sphaerica strains KM893076.1 and KM111472.1. Fermentation, Extraction, and Isolation. A scraping of the agar with mycelium of the fungus growing on potato dextrose agar medium at 28 °C for 3−4 days was put into the liquid medium (2% glucose, 2% peptone, 1% sea salt) and was cultured at 28 °C, 180 rpm for about 4 days as seed culture. Then, 6 mL of the culture was transferred to an Erlenmeyer flask (1 L) with the autoclaved rice medium (80 mL H2O,
INTRODUCTION In recent years, fungi inhabiting insect organs have been recognized as abundant sources of biologically active natural products with novel structures.1−3 Nigrospora sphaerica, a filamentous fungus belonging to the phylum Ascomycota,4 has been proven to produce different types of secondary metabolites with specific agricultural and pharmaceutical values.5−8 Oxya chinensis Thunberg (Orthoptera: Acrididae) is a main insect pest that threatens Oryza sativa L. It is distributed in rice-growing zones all over China.9 A recent report shows that the pest feeds on various plants including Oryza sativa L., Saccharum off icinarum L., Zea mays L., Sorghum vulgare Pers., and others.10 Since this insect contains rich nutrients, such as proteins, fatty acids, and vitamins, it is also widely used for food among people in some areas in China, Japan, and Thailand.11 As part of our program to search for leads of new fungicides used in agriculture from microorganisms,12,13 a fungus Nigrospora sphaerica (collection No. ZMT05) isolated from Oxya chinensis Thunberg collected from Guangzhou, China, attracted our interest because of its antifungal activities against several plant pathogens in vitro. Our investigation on the metabolites of this fungus led to the separation of six new cyclopentenone derivatives, whose structures were identified by spectroscopic and spectrometric method and whose antifungal and antibacterial activities were determined.
■
MATERIALS AND METHODS
General Experimental Procedures. Nuclear magnetic resonance (NMR) data were recorded using an AVIII 600 MHz NMR spectrometer (Bruker BioSpin GmbH company, Rheinstetten, Germany), adopting the residual solvent signals of (CD3)2SO or CDCl3 as references. Optical rotation was measured on a P-1020 digital polarimeter (Jasco International Co., Ltd., Tokyo, Japan). © 2018 American Chemical Society
Received: Revised: Accepted: Published: 5368
March 15, 2018 May 4, 2018 May 10, 2018 May 10, 2018 DOI: 10.1021/acs.jafc.8b01376 J. Agric. Food Chem. 2018, 66, 5368−5372
Article
Journal of Agricultural and Food Chemistry
(−)-Nigrosporione D, (−)-4. Yellow oil; UV (CH3CN) λmax (log ε) 265 (1.94) nm; IR (KBr) νmax 3414, 2920, 1708, 1572, 1357, 1230, 1038, 823 cm−1; [α]25 D −13.50 (c 0.17, MeOH); HRESIMS m/z 169.0860 ([M + H]+, calcd. for C9H13O3 169.0859); 13C NMR and 1H NMR (Tables 1 and 2). ECD Calculations. The theoretical eletronic circular dichroism (ECD) spectra of the isolated compounds were calculated on the basis of the relative configurations determined by their nuclear overhauser enhancement spectroscopy (NOESY). Conformational analyses were accomplished by the MMFF94 force field calculation through the software Spartan′10 (Wave function Inc., Irvine, CA). Density functional theory (DFT) calculations were used to generate and optimize the conformers with energy ≤10 kcal/mol at the 6-31G(d,p) level. The ECD calculations were performed for the stable conformers by the method of time-dependent density functional theory (TDDFT) using Gaussian 09 (Gaussian Inc., Wallingford, CT) software at the B3LYP/6-311+G(d,p) level. The rotary strengths of 30 excited states were calculated. MeOH was used as the integral equation formalism polarizable continuum model (IEF-PCM) solvent. A halfbandwidth of 0.3 eV was applied to Gaussian according to dipolelength rotational strengths. Then, the softwares SpecDis 1.64 (University of Wurzburg, Wurzburg, Germany) and OriginPro 8.5 (OriginLab, Ltd., Northampton, MA) were used to generate the ECD curves. Antimicrobial Activity Assay. Four fungi including Fusarium oxysporum (F. oxysporum), Colletotrichum musae (C. musae), Fusarium graminearum (F. graminearum), and Penicillium italicum (P. italicum) along with two bacteria Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) used for bioassay were acquired from College of Agriculture, South China Agricultural University. The antimicrobial effects were examined as the minimum inhibitory concentration (MIC) values.16 In brief, a stock solution of each test sample was prepared in 5% aqueous dimethyl sulfoxide (DMSO) (v/v), following which 0.5 mL of the solution was serially diluted with 0.5 mL of potato dextrose broth (PDB) to final concentrations of 200, 100, 50, 25, 12.5, 6.25, and 3.13 μg/mL in a set of capped test tubes. Ten microliters of an inoculum suspension of the test microorganism (105 colonyforming units/mL in PDB) was added to each test tube. Then, the fungi and bacteria were cultured at 28 °C for 48 h and at 37 °C for 24 h, respectively. The MIC value for each sample was determined to be the lowest concentration with invisible microbial growth. The solvent (0.5 mL of 5% DMSO/H2O + 0.5 mL PDB) was used as negative control, whereas triadimefon (Aladdin Bio-Chem Technology Co., Ltd., Shanghai, China) and kanamycin (Yuanye Bio-Technology Co., Ltd., Shanghai, China) were adopted as positive controls with respect to fungi and bacteria, respectively.
60 g rice, and 0.1 g sea salt) and was incubated for 30 days at room temperature. One hundred Erlenmeyer flasks containing fermented solid rice media and mycelia were continuously extracted three times using 95% ethanol. The solvent was evaporated in vacuo to 2 L and was extracted three times with ethyl acetate to afford 28.0 g of a yellow extract. The extract was divided into seven fractions (Fr. 1−Fr. 7) by column (40 × 6 cm) chromatography, using gradient of petroleum ether/ethyl acetate (92:8, 1000 mL; 82:18, 510 mL; 76:24, 750 mL; 50:50, 510 mL; 24:76, 510 mL; 20:80, 500 mL; 0:100, 1000 mL; v/v) as eluents. The Fr. 3 was purified on column chromatography (40 × 1.5 cm) and was eluted with petroleum ether/ethyl acetate (92:8, 200 mL; 83:17, 200 mL; 80:20, 200 mL; 76:24, 200 mL; v/v), affording 80 subfractions (Fr. 3.1− Fr. 3.80, 10 mL per subfraction). The Fr. 3.31 and 36 were slowly recrystallized in the solvent acetone at room temperature to give compounds (+)-4 (2.2 mg) and (−)-4 (1.5 mg), respectively. The Fr. 4 was fractioned by column chromatography (40 × 1.5 cm) using petroleum ether/ethyl acetate (92:8, 200 mL; 82:18, 200 mL; 76:24, 250 mL; 50:50, 300 mL; v/v) as eluents to afford Fr. 4.1−Fr. 4.4. The Fr. 4.2 was separated by preparative TLC (petroleum ether/ethyl acetate, 3:1, v/v) to give compound 2 (5 mg, Rf = 0.62). The Fr. 4.3 was separated in the same way to afford compounds (±)-1 (3.8 mg, Rf = 0.36) and 3 (4.3 mg, Rf = 0.51). Racemic (±)-1 was separated into a pair of enantiomers (+)-1 (1.2 mg, tR = 7.6 min) and (−)-1 (1.1 mg, tR = 9.0 min) through chiral HPLC (CH3CN/H2O, 70:30; 25 °C; 1.0 mL/min). (+)-Nigrosporione A, (+)-1. Yellow oil; UV (CH3CN) λmax (log ε) 239 (1.94) nm; IR (KBr) νmax 3374, 2944, 2872, 1684, 1588, 1357, 1264, 1142, 990 cm−1; [α]25 D 25.29 (c 0.28, MeOH); HRESIMS m/z 185.0803 ([M + H]+, calcd. for C9H13O4 185.0808); 13C NMR and 1H NMR (Tables 1 and 2).
Table 1. 13C NMR Data of the Isolated Compounds δc/ppm a
position
(± )- 1
2
3b
(+)-4b
(−)-4b
1 2 3 4 5 6 7 8 9 10
207.4 104.1 188.2 61.0 54.8 72.7
208.5 102.6 189.2 49.8 52.7 67.6 59.4 58.9 15.9
209.0 103.9 189.7 54.4 52.8 64.5 59.1 59.0 22.8
206.8 104.4 180.8 145.9 51.3 109.5 67.3 58.5 18.9
206.3 104.4 180.5 146.2 51.2 109.2 67.4 58.4 18.9
98.2 59.8 18.8
b
■
a Measured in (CD3)2SO at 150 MHz. bMeasured in CDCl3 at 150 MHz.
RESULTS AND DISCUSSION (±) Nigrosporione A, (±)-1, was a yellow oil, and its molecular formula was elucidated as C9H12O4 on the basis of highresolution ESIMS data, suggesting that (±)-1 had four degrees of unsaturation. The IR spectrum displayed the functional groups of hydroxyl (3374 cm−1), carbonyl (1684 cm−1), and double bond (1588 cm−1). The 1H NMR spectrum (Table 2) exhibited one hydroxyl at δH 6.52 (8-OH, exchangeable), one methyl at δH 1.17 (H-10), two methines at δH 2.82 (H-4) and 5.26 (H-8), one oxymethylene at δH 3.76 (H-6a) and 3.59 (H6b), one oxymethyl at δH 3.84 (H-9), and one olefinic methine at δH 5.37 (H-2). The 13C NMR and HSQC (heteronuclear singular quantum correlation) spectra (Table 1) exhibited nine signals consisting of one carbonyl group at δC 207.4 (C-1), one double bond at δC 104.1 (C-2) and 188.2 (C-3), one methyl at δC 18.8(C-10), one oxymethyl at δC 59.8(C-9), one oxymethylene at δC 72.7 (C-6), two methines at δC 98.2 (C-8) and 61.0 (C-4), and one quaternary carbon at δC 54.8(C-5), accounting for two of the four degrees of unsaturation in (±)-1. This result also implied that (±)-1 contained two rings. HMBC
(−)-Nigrosporione A, (−)-1. Yellow oil; UV (CH3CN) λmax (log ε) 239 (1.94) nm; IR (KBr) νmax 3374, 2944, 2872, 1684, 1588, 1357, 1264, 1142, 990 cm−1; [α]25 D −25.18 (c 0.28, MeOH); HRESIMS m/z 185.0803 ([M + H]+, calcd. for C9H13O4 185.0808); 13C NMR and 1H NMR (Tables 1 and 2). Nigrosporione B (2). Yellow oil; UV (CH3CN) λmax (log ε) 238 (1.98) nm; IR (KBr) νmax 3364, 2916, 1679, 1596, 1355, 1038, 829 cm−1; [α]25 D −36.63 (c 0.17, MeOH); HRESIMS m/z 209.0789 ([M + Na]+, calcd. for C9H15O4 209.0784); 13C NMR and 1H NMR (Tables 1 and 2). Nigrosporione C (3). Yellow oil; UV (CH3CN) λmax (log ε) = 239 (1.88) nm; IR (KBr) νmax 3342, 2931, 1679, 1594, 1447, 1347, 1367, 1239, 1038, 829, 551 cm−1; [α]25 D 20.56 (c 0.16, MeOH); HRESIMS m/z 187.0967 ([M + H]+, calcd. for C9H15O4 107.0964); 13C NMR and 1H NMR (Tables 1 and 2). (+)-Nigrosporione D, (+)-4. Yellow oil; UV (CH3CN) λmax (log ε) 265 (1.94) nm; IR (KBr) νmax 3414, 2920, 1708, 1572, 1357, 1230, 1038, 823 cm−1; [α]25 D 12.35 (c 0.17, MeOH); HRESIMS m/z 169.0860 ([M + H]+, calcd. for C9H13O3 169.0859); 13C NMR and 1H NMR (Tables 1 and 2). 5369
DOI: 10.1021/acs.jafc.8b01376 J. Agric. Food Chem. 2018, 66, 5368−5372
Article
Journal of Agricultural and Food Chemistry Table 2. 1H NMR Data of the Isolated Compounds δH/ppm, multi (J/Hz) position 2 4 6
( ± )1a
2b
5.37,s 2.82,s 3.76,d(9.0) 3.59,d(9.0)
5.24,d(1.2) 3.04,ddd(1.8,4.8,4.8) 3.57,d(10.2) 3.63,d(10.2) 3.75,dd(4.8,11.4) 3.95,dd(4.8,11.4) 3.84,s 1.17,s
5.34,s(3.0) 2.70,d(4.8) 3.67,d(12.0) 3.84,dd(3.0,12.0) 4.01,dd(2.4,12.0) 3.92,dd(6.0,12.0) 3.86,s 1.10,s
unobservable unobservable
4.51−4.66c 4.51−4.66c
7 8 9 10 6-OH 7-OH 8-OH
5.26,d(3.6) 3.84,s 1.17,s
3b
(+)-4b
(−)-4b
5.44,d(1.2)
5.43,d(1.2)
5.22,d(1.2) 5.69,d(1.2) 3.60,d(10.8) 3.71,d(10.8) 3.92,s 1.20,s
5.22,d(1.2) 5.69,d(1.2) 3.60,d(10.8) 3.70,d(10.8) 3.92,s 1.22,s
6.52,s
a
Measured in (CD3)2SO at 600 MHz. bMeasured in CDCl3 at 600 MHz. cThe chemical shifts of 6-OH and 7-OH of compound 3 were not differentiated because no HMBC correlations for them were observed.
(heteronuclear multiple bond correlation) correlations from H10 to C-1, C-5, and C-6; H-6 to C-1, C-5, and C-10; and H-4 to C-1, C-5, and C-10 unequivocally established the direct connections of C-5 with C-1, C-4, C-6, and C-10. Similarly, those from H-2 to C-1, C-3, C-4, and C-5; and H-9 to C-3 established the five-membered enone ring and placed the oxymethyl group at C-3. Furthermore, the correlations from H8 to C-4, C-5, and C-6; and 8-OH to C-4, C-8, indicated a tetrahydrofuran ring and placed 8-OH at C-8. Accordingly, the planar structure of (±)-1 was determined (Figure 1). Nuclear
Figure 2. Key HMBC and NOESY of the isolated compounds.
Nigrosporione B (2) was obtained as a yellow oil and showed IR bands for hydroxyl (3364 cm−1), carbonyl (1679 cm−1), and double bond groups. The molecular formula was determined as C9H14O4 according to HRESIMS (three degrees of unsaturation). The 13C NMR (Table 1) and HSQC spectroscopic data for 2 indicated nine carbon resonances, including one carbonyl group at δC 208.5 (C-1), one double bond at δC 102.6 (C-2) and 189.2 (C-3), one methyl at δC 15.9(C-9), one oxymethyl at δC 58.9(C-9), two oxymethylenes at δC 67.6 (C-6) and δC 59.4 (C-7), one methine at δC 49.8 (C4), and one quaternary carbon at δC 52.7(C-5). The carbonyl and double bond groups accounted for two of the three elements of unsaturation, indicating that the molecule only possessed one cyclic ring. These data together with the molecular formula revealed by HRESIMS indicated that compound 2 was a cyclopentenone with the substitutes of one methyl, one methoxyl, and two hydroxymethyls. Furthermore, the HMBC spectrum showed correlations from H-9 at δH 1.17 to C-1, C-4, C-5, and C-6; H-6 at δH 3.57 and 3.63 to C-1, C-4, C-5, and C-9; H-4 at δH 3.04 to C-3, C-5, and C-9; H-2 at δH 5.24 to C-1, C-3, C-4, and C-5; H-8 at δH 3.84 to C-3; and H-7 at δH 3.75 and 3.95 to C-3, C-4, and C-5, which established the planar structure of 2 (Figure 1). NOE correlations (Figure 2) between H-7 and H-9, and the absence of correlations between H-7 and H-6, revealed that H-7 was cis to H-9 and trans to H-6. Additionally, the lack of correlations between H-4 and H-9 revealed that H-4 and H-9 faced the
Figure 1. Chemical structures of the isolated compounds.
Overhauser effect (NOE) correlations (Figure 2) between H-4 and H-10 indicated that H-4 was on the same side relative to H-10. No detectable coupling constant between H-4 and H-8 in the 1H NMR spectrum of (±)-1 placed the two protons on the same side, which was supported by no visible NOE cross peak signals between H-4 and 8-OH. Notably, the optical rotation and CD maximum of (±)-1 were both close to zero (Figure 3), suggesting that (±)-1 was a raceme. A subsequent chiral HPLC analysis of (±)-1 displayed two different chromatographic peaks in the proportion of 1:1, which further confirmed this deduction. Then, the racemate was separated into a pair of enantiomers, (+) 1 and (−) 1. The theoretical ECD spectra of (+)-1 and (−)-1 were further calculated and compared with the experimental ones to determine the absolute configurations. They (Figure 3) were very similar (Figure 3), so that (+)-1 and (−)-1 were 4S, 5S, 8S and 4R, 5R, 8R, respectively. 5370
DOI: 10.1021/acs.jafc.8b01376 J. Agric. Food Chem. 2018, 66, 5368−5372
Article
Journal of Agricultural and Food Chemistry
Figure 3. Calculated and experimental ECD spectra of the isolated compounds.
Table 3. Antifungal and Antibacterial Activity of the Isolated Compounds Measured as MIC values MIC, μg/mL
a
compds
F. oxysporum
C. musae
P. italicum
F. graminearum
E. coli
S. aureus
(+)-1 (−)-1 2 3 (+)-4 (−)-4 Triadimefona Kanamycinb
12.5 12.5 12.5 25 50 3.13 100 NT
6.25 6.25 25 12.5 25 3.13 80 NT
12.5 12.5 12.5 12.5 200 6.25 50 NT
25 25 3.13 >200 100 >200 150 NT
3.13 3.13 3.13 12.5 >200 >200 NT 1.0
3.13 3.13 6.25 12.5 50 >200 NT 1.0
Positive control toward fungi. bPositive control toward bacteria.
HRESIMS, indicating one fewer degree of unsaturation than 3. There were great resemblances between the NMR (Tables 1 and 2) spectra of (+)-4 and those of 3. However, compared with 3, the proton signals of H-4 at δH 2.70 and of H-7 at δH 4.01 and 3.92 were missing and were changing to exocyclic olefinic methylene signals at δH 5.22 and 5.69 in (+)-4. Together, the signals at δC 54.4 (CH, C-4) for the methine group and at δC 59.1 (CH2, C-7) for the oxymethylene group were replaced by two olefinic signals at δC 145.9 (C, C-4) and δC 109.5 (CH2, C-7), respectively, in (+)-4. It could be concluded that the hydroxymethyl group (CH2, C-7) at C-4 in 3 was changed to the exocyclic olefin Δ4(7) substituent at C-4 in (+)-4. The clear HMBC correlations from H-7 to C-3, -4, and -5 supported the deduction (Figure 2). Finally, all NMR data for (+)-4 were readily assigned by HMBC analysis. Compound (+)-4 only has one chiral carbon, and its experimental ECD was similar to the calculative one of (S)-4 (Figure 3), with the absolute configuration of (+)-4 being 5S. (−)-Nigrosporione D, (−)-4, shared almost the same 1D and 2D NMR spectra with compound (+)-4, suggesting that their planar structures were identical. However, the specific rotation ([α]25 D −13.50) and the Cotton effects in the CD spectrum of
opposite direction. The trend of the experimental ECD curve was almost the same as that of the theoretical one for (4R, 5R)2 (Figure 3), establishing the chiral carbons of 2 to be 4R, 5R. Nigrosporione C (3) had the same molecular formula C9H14O4 as 2 deduced from its HRESIMS spectrum. The carbon types of 3 (Table 1) were the same as those of 2. Moreover, they shared similar chemical shifts at most carbons. However, the carbon chemical shifts of compound 3 at C-4 and C-9 showed obvious differences with the deviations of 4.6 and 6.3 ppm with those of 2, respectively. Subsequently, detailed analysis of the HMBC correlations of 3 revealed that 3 had the same planar structure as 2. These results implied that 3 might be a stereoisomer of 2 (Figure 1). NOE correlations between H-4 and H-9, and the lack of correlations between H-4 and H6, indicated that H-4 was cis to H-9 and trans to H-6 (Figure 2). Moreover, NOE correlations of H-7a and H-7b to H-6a and H-6b indicated that H-6 was cis to H-7. The experimental ECD spectra of 3 were quite similar to the calculated one of compound 4S, 5R-3 (Figure 3), establishing the absolute configuration of 3 as 4S, 5R. Nigrosporione D, (+)-4, was purified as yellow oil, and its molecular formula C9H12O3 was determined on the basis of the 5371
DOI: 10.1021/acs.jafc.8b01376 J. Agric. Food Chem. 2018, 66, 5368−5372
Article
Journal of Agricultural and Food Chemistry Funding
(−)-4 were completely opposite from those of (+)-4, indicating that compound (−)-4 was the enantiomer of the latter. Moreover, the experimental ECD spectra of (−)-4 were quite similar to the calculated one of the 5R-4 (Figure 3), which also confirmed the 5R configuration of (−)-4. All the compounds were examined for inhibory activity toward the plant pathogenic fungi F. oxysporum, C. musae, P. italicum, and F. graminearum and the bacteria E. coli and S. aureus (Table 3). In particular, compounds (+)-1, (−)-1, and 2 exhibited antifungal activities against all the tested plant pathogens (MIC values, 3.13−25 μg/mL), higher than triadimefon (the positive control). Compounds 3 and (−)-4 showed stronger antifungal activities toward F. oxysporum, C. musae, and P. italicum (MIC values, 3.13−25 μg/mL) than the control. Additionally, compound (+)-4 displayed antifungal activities against F. oxysporum, C. musae, and F. graminearum with MIC values of 50, 25, and 100 μg/mL, respectively, which were stronger than triadimefon. The results indicated the potential values of these new cyclopentenones as fungicides used in agriculture. As shown in Table 3, compounds (+)-1, (−)-1, 2, and 3 also displayed moderate antibacterial activities against S. aureus belonging to Gram positive bacterium and E. coli belonging to Gram negative bacterium (MIC values, 3.13− 12.5 μg/mL). It is interesting that the configuration seems to have no impact on the antimicrobial activities for compounds (+)-1 and (−)-1. In previous investigations, only one example, nosporin A, with the same bicyclic ring system as compounds (+)-1 and (−)-1 had been found as fungal metabolites.14 It was reported to have moderate antibacterial activity toward Bacillus subtilis ATCC 6633. In addition, cyclopentenones and hygrophorones A−G were reported to have inhibitory activities toward Cladosporium cucumerinum.15 (±)-(4S*,5S*)-2,4,5-trihydroxy3-methoxy-4-methoxycarbonyl-5-methyl-2-cyclopenten-1-one exhibited antifungal activity to F. graminearum.16
■
This work was supported by the National Natural Science Foundation of China (21102049), the Natural Science Foundation of Guangdong Province (2015A030313405), the Science and Technology Project of Guangdong Province (2016A020222019), and the Science and Technology Project of Guangzhou City (201707010342). Notes
The authors declare no competing financial interest.
■
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jafc.8b01376. 1 H NMR, 13C NMR, HSQC, HMBC, 1H−1HCOSY (chemical-shift correlation spectroscopy), NOESY, HRESIMS, UV, and IR spectra of the new compounds (Figures S1−S41) and the chiral-HPLC separation profile of the racemate (±) 1 (Figure S42) (PDF)
■
REFERENCES
(1) Zhang, Y. L.; Ge, H. M.; Zhao, W.; Dong, H.; Xu, Q.; Li, S. H.; Li, J.; Zhang, J.; Song, Y. C.; Tan, R. X. Unprecedented immunosuppressive polyketides from Daldinia eschscholzii, a mantisassociated fungus. Angew. Chem., Int. Ed. 2008, 47, 5823−5826. (2) Chen, C.; Wang, J.; Liu, J.; Zhu, H.; Sun, B.; Wang, J.; Zhang, J.; Luo, Z.; Yao, G.; Xue, Y.; Zhang, Y. Armochaetoglobins A−J: cytochalasan alkaloids from Chaetomium globosum TW1−1, a fungus derived from the terrestrial arthropod Armadillidium vulgare. J. Nat. Prod. 2015, 78, 1193−1201. (3) Ge, H.; Tan, R. Symbionts, an important source of new bioactive natural products. Huaxue Jinzhan 2009, 21, 30−46. (4) Metwaly, A. M.; Kadry, H. A.; El-Hela, A. A.; Mohammad, A. -E. I.; Ma, G.; Cutler, S. J.; Ross, S. A. Nigrosphaerin A a new isochromene derivative from the endophytic fungus Nigrospora sphaerica. Phytochem. Lett. 2014, 7, 1−5. (5) Zhang, Q.-H.; Tian, L.; Zhou, L.-D.; Zhang, Y.; Li, Z.-F.; Hua, H.M.; Pei, Y.-H. Two new compounds from the marine Nigrospora sphaerica. J. Asian Nat. Prod. Res. 2009, 11, 962−966. (6) Kim, J.-C.; Choi, G. J.; Park, J.-H.; Kim, H. T.; Cho, K. Y. Activity against plant pathogenic fungi of phomalactone isolated from Nigrospora sphaerica. Pest Manage. Sci. 2001, 57, 554−559. (7) Cutler, H. G.; Hoogsteen, K.; Littrell, R. H.; Arison, B. H. Epoxyexserohilone, a novel metabolite from Nigrospora sphaerica. Agric. Biol. Chem. 1991, 55, 2037−2042. (8) Harwooda, J. S.; Cutler, H. G.; Jacyno, J. M. Nigrosporolide, a plant growth-inhibiting macrolide from the mould Nigrospora sphaerica. Nat. Prod. Lett. 1995, 6, 181−185. (9) Wu, H.; Zhang, R.; Liu, J.; Guo, Y.; Ma, E. Effects of malathion and chlorpyrifos on acetylcholinesterase and antioxidant defense system in Oxya chinensis (Thunberg) (Orthoptera: Acrididae). Chemosphere 2011, 83, 599−604. (10) Wu, H.; Yang, M.; Guo, Y.; Xie, Z.; Ma, E. Comparisons of malathion susceptibility, target sensitivity, and detoxification enzyme activity in nine field populations of Oxya chinensis (Orthoptera: Acrididae). J. Econ. Entomol. 2007, 100, 1409−1415. (11) Zhao, Y.; Guo, X.; Li, L.; Yang, L. Analysis and evaluation of nutritional components of Oxya chinensis Thunberg in different instars. Journal of Guizhou University (Agricultural and Biological Science) 2002, 21, 115−120. (12) Li, W.; Xiong, P.; Zheng, W.; Zhu, X.; She, Z.; Ding, W.; Li, C. Identification and antifungal activity of compounds from the mangrove endophytic fungus Aspergillus clavatus R7. Mar. Drugs 2017, 15, 259. (13) Huang, S.; Chen, H.; Li, W.; Zhu, X.; Ding, W.; Li, C. Bioactive chaetoglobosins from the mangrove endophytic fungus Penicillium chrysogenum. Mar. Drugs 2016, 14, 172. (14) Kozlovsky, A. G.; Zhelifonova, V. P.; Adanin, V. M.; Ozerskaya, S. M.; Graefe, U. Nosporins A and B, new metabolites from a filamentous fungus, VKM-3750. Pharmazie 2003, 58, 76−77. (15) Lübken, T.; Schmidt, J.; Porzel, A.; Arnold, N.; Wessjohann, L. Hygrophorones A−G: fungicidal cyclopentenones from Hygrophorus species (Basidiomycetes). Phytochemistry 2004, 35, 1061−1071. (16) Wang, J.; Ding, W.; Wang, R.; Du, Y.; Liu, H.; Kong, X.; Li, C. Identification and bioactivity of compounds from the mangrove endophytic fungus Alternaria sp. Mar. Drugs 2015, 13, 4492−4504.
AUTHOR INFORMATION
Corresponding Author
*Tel: +86-20-85280319. Fax: +86-20-85282366. E-mail:
[email protected]. ORCID
Chunyuan Li: 0000-0001-8817-8994 Author Contributions
Chunyuan Li and Weijia Ding conceived and designed the experiments; Zhihui Wu, Zihui Xie, Manlin Wu, Xiaoqi Li, Weilin Li, and Chunyuan Li performed the experiments; Zhigang She, Chunyuan Li, and Weijia Ding analyzed the data; Zhihui Wu and Weijia Ding wrote the paper; Chunyuan Li and Weijia Ding revised and edited the manuscript. 5372
DOI: 10.1021/acs.jafc.8b01376 J. Agric. Food Chem. 2018, 66, 5368−5372
