Article pubs.acs.org/jnp
Cite This: J. Nat. Prod. 2018, 81, 1860−1868
Spiro-Phthalides and Isocoumarins Isolated from the MarineSponge-Derived Fungus Setosphaeria sp. SCSIO41009 Xiaoyan Pang,†,‡ Xiuping Lin,† Jie Yang,§ Xuefeng Zhou,† Bin Yang,† Junfeng Wang,*,† and Yonghong Liu*,†,‡
J. Nat. Prod. 2018.81:1860-1868. Downloaded from pubs.acs.org by UNIV OF SOUTH DAKOTA on 08/24/18. For personal use only.
†
CAS Key Laboratory of Tropical Marine Bio-resources and Ecology/Guangdong Key Laboratory of Marine Materia Medica/RNAM Center for Marine Microbiology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510220, China ‡ University of Chinese Academy of Sciences, Beijing 100049, China § Guangdong Provincial Key Laboratory of New Drug Screening, Guangzhou Key Laboratory of Drug Research for Emerging Virus Prevention and Treatment, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515, China S Supporting Information *
ABSTRACT: Fourteen new polyketides classified as four phthalides, setosphalides A and B, 5-O-desmethylcolletotrialide, and (S)-colletotrialide (1−4), three isocoumarin derivatives, exserolides I−K (5−7), four pyrones, setosphapyrones A−D (8−11), one furanone (12), and two depsidones (13 and 14), along with 17 known polyketides were isolated from cultures of the sponge-derived fungus Setosphaeria sp. SCSIO41009. The structures and absolute configurations of these new compounds (1−14) were determined by spectroscopic analyses, X-ray diffraction, chiral-phase HPLC analysis, modified Mosher’s method, and comparison of ECD spectra to calculations. Setosphalides A (1) and B (2) are the first examples possessing a 5,5 spiroketal skeleton in phthalide derivatives. Botryorhodines I (13) and J (14) showed moderate antifungal activities against the phytopathogenic fungi Colletotrichum asianum and Colletotrichum acutatum. Compound 18 (7-O-demethylmonocerin) exhibited potent radical scavenging activity against DPPH.
R
compounds consist of four phthalides, setosphalides A and B, 5-O-desmethylcolletotrialide and (S)-colletotrialide (1−4), three isocoumarin derivatives, exserolides I−K (5−7), four pyrones, setosphapyrones A−D (8−11), one furanone (12), and two depsidones (13 and 14). Previously, researchers isolated phthalide derivatives and dimeric octaketide spiroketals from the jellyfish-derived fungus Paecilomyces variotii.22,23 Setosphalides A and B are the first examples possessing a 5,5 spiroketal skeleton in phthalide derivatives. All new compounds were evaluated for their antifungal, antibacterial, antioxidant, antiviral, and Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) inhibitory activities. Herein, we report the isolation, structure elucidation, absolute configurations, and bioactivities of the new compounds 1−14.
ecently, because of the structurally unique as well as interesting biological and pharmacological properties, secondary metabolites obtained from marine-derived fungi have attracted considerable attention.1 The fungal genus Setosphaeria has been mainly isolated from terrestrial plants, with fewer strains isolated from marine sources.2,3 Among this genus, S. rostrata and S. turcica are well known as plant pathogenic fungi and widely researched for their pathogenicity.4−7 Nearly 30 bioactive secondary metabolites have been isolated from the fungal genus Setosphaeria together with the genus Exserohilum, which is an anamorph of Setosphaeria.8−16 Those secondary metabolites showed phytotoxic,9 anticancer,10,14 antiplasmodial,12 antifungal,13 antibacterial,13 αglucosidase inhibitory, and porcine pancreatic alpha amylase inhibitory activities.16 Among marine sources, fungi derived from sponges are one of the richest sources, which can produce structurally unique and biologically active secondary metabolites.17 As part of our continuing search for structures with antiviral and antifungal activities from sponge-derived fungi,18−21 the fungus Setosphaeria sp. SCSIO41009 isolated from a Callyspongia sp. sponge was studied. From the extracts of the culture of Setosphaeria sp. SCSIO41009, 14 new (1−14) and 17 known (including 15−22) metabolites were isolated. The new © 2018 American Chemical Society and American Society of Pharmacognosy
■
RESULTS AND DISCUSSION Setosphalide A (1) was obtained as colorless crystals. Its molecular formula was established as C16H20O7 by 13C NMR data and a protonated molecule at m/z 325.1290 [M + H]+ in the HRESIMS spectrum. A detailed analysis of its 1H NMR Received: May 2, 2018 Published: August 9, 2018 1860
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
Table 1. 1H NMR and 13C NMR Data for Compounds 1−4 1a no.
δC, type
1 3
168.7, C 114.9, C
3a 4 5 6 7 7a 8
142.9, C 102.1, CH 160.3, C 139.1, C 150.7, C 108.2, C 78.7, CH
9
40.0, CH2
δH (J in Hz)
1b δC, type
δH (J in Hz)
165.2, C 112.4, C
6.85, s
4.19, d (4.0)
10 11
83.4, CH 40.3, CH2
12
20.3, CH2
13 5OCH3 6OCH3 7-OH 8-OH
14.4, CH3 56.9, CH3
2.26, dd (13.5, 6.0) 2.09, ddd (14.0, 9.5, 4.5) 4.65, m 1.73, m 1.60, m 1.50, m 1.45, m 0.97, t (7.5) 3.95, s
61.2, CH3
3.80, s
141.5, C 100.9, CH 158.1, C 137.5, C 149.0, C 106.9, C 76.6, CH 38.9, CH2
6.74, s
4.10, t (4.2)
13.9, CH3 56.3, CH3 60.4, CH3
3.69, s
18.8, CH2
δC, type
δH (J in Hz)
168.9, C 112.2, C
2.16, dd (13.3, 5.6) 1.94, ddd (14.0, 9.8, 4.2) 4.55, m 1.61, m 1.51, m 1.35, m 1.35, m 0.90, t (7.0) 3.88, s
81.1, CH 38.7, CH2
2a
3a δC, type 171.8, C 81.1, CH
143.9, C 98.5, CH 160.9, C 139.1, C 150.8, C 108.8, C 76.1, CH
4.50, t (9.0)
148.5, C 101.8, CH 159.4, C 136.9, C 151.7, C 105.2, C 29.7, CH2
36.8, CH2
2.27, m
38.1, CH2
6.67, s
14.3, CH3 57.0, CH3
2.19, ddd (12.0, 8.5, 3.5) 4.44, m 1.72, m 1.63, m 1.48, m 1.43, m 0.98, t (7.5) 3.96, s
61.2, CH3
3.80, s
80.2, CH 39.8, CH2 19.9, CH2
212.4, C 45.4, CH2
δH (J in Hz) 5.35, dd (8.5, 3.5) 6.44, s
2.32, m 1.87, m 2.60, dt (18.0, 7.5) 2.50, ddd (17.5, 8.0, 5.5) 2.44, td (7.0, 4.5)
4a δC, type 171.5, C 81.3, CH 148.8, C 97.6, CH 160.9, C 137.8, C 151.6, C 106.7, C 29.6, CH2 38.1, CH2
212.3, C 45.4, CH2
δH (J in Hz) 5.38, brdd (8.0, 3.0) 6.65, s
2.30, m 1.85, m 2.60, dt (18.0, 7.5) 2.50, ddd (17.5, 8.0, 5.5) 2.41, dq (14.0, 6.5)
18.2, CH2
1.58, dq (14.5, 7.5)
18.2, CH2
1.54, dq (15.0, 7.5)
14.0, CH3
0.91, t (7.5)
14.0, CH3 57.0, CH3
0.88, t (7.0) 3.93, s
61.0, CH3
3.80, s
61.2, CH3
3.76, s
10.11, brs 5.67, d (4.9)
a
Measured in methanol-d4 and at 500, 125 MHz NMR. bMeasured in DMSO-d6 and at 700, 175 MHz NMR.
Figure 1. COSY and key HMBC correlations of compound 1.
δH 4.65 (1H, m, H-10) and 4.19 (1H, d, J = 4.0 Hz, H-8); and an aromatic proton at δH 6.85 (1H, s, H-4). The 1H NMR data of 1 in DMSO-d6 showed two hydroxy protons at δH 10.31
data in methanol-d4 (Table 1) exhibited the presence of three methyl signals at δH 3.95 (s, 5-OCH3), 3.80 (s, 6-OCH3), and 0.97 (t, J = 7.5 Hz, H3-13); two oxygenated methine protons at 1861
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
(1H, brs, 7-OH) and 5.67 (1H, d, J = 4.9 Hz, 8-OH). The 13C NMR and DEPT data (Table 1) displayed 16 carbon signals including three methyls, three sp3 methylenes, two oxygenated sp3 methines, an sp2 methine, an oxygenated sp3 nonprotonated carbon, five sp2 nonprotonated carbons, and an ester carbonyl. Besides the presence of a benzene ring and an ester carbonyl, two degrees of unsaturation were left, which indicated that there were another two rings in the structure of 1. Part of the NMR data were similar to those of (S)-3-ethyl-7hydroxy-5,6-dimethoxyphthalide,24 which suggested 1 was a phthalide derivative. This speculation was confirmed by HMBC correlations from H-4 to C-3 (δC 114.9), C-3a (δC 142.9), C-5 (δC 160.3), C-6 (δC 139.1), and C-7a (δC 108.2), from 5-OCH3 to C-5, and from 6-OCH3 to C-6 (Figure 1). However, the spiroketal fragment connected to C-3 in 1 was a very unique feature. The relative configuration was ascertained by NOESY experiment. The cross-peaks of H-8 to H-9b, of H9a to H-10 and 8-OH, and of 8-OH to H-4 in the NOESY spectrum (Figure 2) suggested that if the 8-OH were α-
Figure 2. Key NOESY correlations of compounds 1 and 2 in DMSOd6.
oriented, then H-10 should be α-oriented and the configuration at C-3 should be R. These orientations were further confirmed by single-crystal X-ray diffraction using the anomalous dispersion of Cu Kα radiation (ORTEP drawing in Figure 3). The value of the Flack parameter, 0.01(8), allowed assignment of the absolute configuration of 1 as 3R, 8S, and 10S. Setosphalide B (2), a colorless oil, had the same molecular formula as 1. The 1H and 13C NMR data of 2 closely resembled those of 1 (Table 1), except some of the chemical shifts changed about 2−3 ppm, such as C-4 (δC/H 98.5/6.67), C-8 (δC/H 76.1/4.50), C-9 (δC/H 36.8/2.27 and 2.19), and C10 (δC/H 80.2/4.44) in 2. The 2D NMR data of 2 (Figure S6) established the same planar structure as 1, which was confirmed by HMBC correlations from H-4 to C-3 (δC 112.2), C-3a (δC 143.9), C-5 (δC 160.9), C-6 (δC 139.1), and C-7a (δC 108.8), from H-8 to C-3 and C-3a, from H2-9 to C-3, and from H-10 to C-8 and COSY cross-peaks of H-8/H29/H-10/H2-11 (δH 1.72 and 1.63)/H2-12 (δH 1.48 and 1.43) and H3-13. The NOESY correlations (Figure 2) of 2 from H-4 to H-8 and from H-8 to H2-11 indicated the configuration of C-3 in 2 was opposite that in 1. Interestingly, 2 could convert to 1 when kept at room temperature in MeOH, while under the same conditions only a small amount of compound 1 converted to 2. This interconversion might be due to reversible ketal formation.23 The absolute configuration of 2 was established based on comparison of its experimental electronic circular dichroism (ECD) curve with the calculated ECD curve of the 3S-8S-10S-2 model and the 3R-8R-10R-2 model at the B3LYP/6-31G(d,p) level in Gaussian 03, and the former was
Figure 3. ORTEP drawings of compounds 1, 3, and 10.
consistent with the experimental one (Figure 4). Thereby, the absolute structure of 2 was defined as 3S, 8S, and 10S. The molecular formulas of 5-O-desmethyl-colletotrialide (3) and (S)-colletotrialide (4) were assigned as C15H18O6 and C16H20O6, respectively, through analysis of their HRESIMS data together with the 13C NMR data (Table 1). Their NMR data (Table 1) were almost identical to those of colletotrialide isolated from Colletotrichum sp.,25 indicating that 3 and 4 were structure analogues of colletotrialide, except for the absence of a methoxy group at C-5 in 3. This was deduced by the chemical shift of the single methoxy group (δC/H 61.0/3.80) in 3, which was nearly the same as the methoxy group at C-6 in 1, 2, and colletotrialide. Also, the HMBC correlation (Figure S6) from OCH3-6 to C-6 (δC 136.9) confirmed the location of the lone methoxy group at C-6 in 3. A single-crystal X-ray 1862
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
Figure 4. Comparison between calculated and experimental ECD spectra of 2 and 5 as well as experimental ECD spectra of 1 and 6.
Table 2. 1H NMR and 13C NMR Data for Compounds 5−7 in Methanol-d4 5a no.
δC, type
1 3 4 4a 5 6 7 8 8a 9
169.4, C 83.1, CH 75.6, CH 133.3, C 106.2, CH 160.1, C 138.0, C 156.8, C 102.9, C 40.0, CH2
10 11
79.7, CH 33.6, CH2
12
30.0, CH2
13 6-OCH3 7-OCH3
62.7, CH3 56.9, CH3 60.1, CH3
6b δH (J in Hz)
5.11, d (3.5) 4.62, brs 6.75, s
2.65, 2.05, 4.13, 1.65, 1.65, 1.61, 1.53, 3.53, 3.96, 3.83,
ddd (14.0, 8.4, 6.3) dd (14.7, 5.6) dq (11.9, 6.3) m m td (14.0, 7.0) m t (6.3) s s
δC, type
7b δH (J in Hz)
169.4, C 83.1, CH 75.8, CH 133.3, C 106.2, CH 160.1, C 138.1, C 156.9, C 102.9, C 39.8, CH2
5.12, m 4.64, brs 6.76, s
2.67, 2.07, 4.17, 1.92, 1.84, 2.37, 2.33,
78.9, CH 32.3, CH2 31.4, CH2 177.1, C 56.9, CH3 61.0, CH3
ddd (13.5, 7.5, 5.5) dd (14.5, 5.0) m dt (13.5, 7.0) dt (14.5, 7.5) td (16.0, 7.0) td (16.0, 7.5)
3.94, s 3.81, s
δC, type 171.2, C 81.3, CH 66.1, CH 138.9, C 108.6, CH 157.4, C 136.3, C 158.4, C 101.5, C 38.7, CH2
δH (J in Hz) 4.67, t (6.5) 4.55, brs 6.50, s
14.4, CH3
2.12, 1.97, 3.85, 1.51, 1.51, 1.51, 1.41, 0.96,
60.9, CH3
3.85, s
68.4, CH 40.8, CH2 19.8, CH2
ddd (13.5, 9.5, 3.5) m m m m m m t (6.5)
a
Measured at 700, 175 MHz NMR. bMeasured at 500, 125 MHz NMR.
Exserolide I (5) was obtained as a colorless oil. The 13C NMR data and a sodium adduct ion at m/z 347.1113 in the HRESIMS spectrum established its molecular formula as C16H20O7. The NMR data (Table 2) of 5 closely resembled those of (12R)-12-hydroxymonocerin (15),28 except for two sp3 methylenes (δC 62.7, C-13; 30.0, C-12) in 5 instead of the corresponding signals, an oxygenated methine (C-12) and a methyl (C-13) in 15, which was confirmed by the COSY correlations of H-10 (δH 4.13)/H2-11 (δH 1.65)/H2-12 (δH 1.61 and 1.53)/H2-13 (δH 3.53) (Figure S6). The NOESY spectrum of 5 (Figure S7) displayed cross-peaks from H-4 to H-3 and H-10, which indicated H-3, H-4, and H-10 were on the same side. The absolute configuration of 5 was determined to be 3R, 4R, and 10S, which was supported by comparing the calculated ECD spectrum of the 3R-4R-10S-model at the B3LYP/6-31G(d,p) level in Gaussian 03 and the experimental ECD spectrum of 5 (Figure 4). As well, the experimental ECD spectrum of 5 has a similar Cotton effect to those of known compounds 15−18 (Figure S9). Accordingly, the absolute configuration of 5 was determined as 3R, 4R, and 10S. Exserolide J (6) possessed the elemental composition C16H18O8 as established by the 13C NMR data and a deprotonated molecule at m/z 337.0929 in the HRESIMS
diffraction analysis of a suitable crystal of 3 (Figure 3) confirmed the structure. The HMBC correlations (Figure S6) of 4 from H-3 (δH 5.38) to C-1 (δC 171.5), C-3a (δC 148.8), and C-9 (δC 38.1), from H-4 (δH 6.65) to C-3 (δC 81.3), C-5 (δC 160.9), C-6 (δC 137.8), and C-7a (δC 106.7), and from H28 (δH 2.30 and 1.85), H2-9 (δH 2.60 and 2.50), H2-11 (δH 2.41), and H2-12 (δH 1.54) to C-10 (δC 212.3) and COSY correlations of H-3/H2-8/H2-9 and of H2-11/H2-12/H3-13 (δH 0.88) revealed that 4 shared the same planar structure as colletotrialide. In general, the 3R-phthalides display positive specific rotations, while 3S-phthalides have negative ones.24−27 In our study, the positive specific rotation of 3 ([α]25D +9.6) also suggested C-3 had the R configuration, which was further confirmed by its single-crystal X-ray diffraction experiment with Cu Kα radiation (Figure 3). Compound 4 was proposed as S, based on its negative specific rotation ([α]25D −22), which was opposite that of colletotrialide ([α]25D +17) and indicated 4 was the enantiomer of colletotrialide. Besides, the experimental ECD spectra of 3 and 4 showed opposite Cotton effects, further confirming their different configurations (Figure S8). Thus, the absolute configurations of 3 and 4 were determined as 3R and 3S, respectively. 1863
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products Table 3. 1H and
13
Article
C NMR Data for Compounds 8−12 (500, 125 MHz) 8a
no.
δC, type
δH (J in Hz)
2 3 4 5 6 7
167.4, C 88.3, CH 173.4, C 110.9, C 161.4, C 33.1, CH2
8
73.4, CH
9
31.0, CH2
2.58, dd (14.0, 6.0) 2.41, dd (14.0, 8.5) 3.55, tt (8.0, 4.5) 1.52, m;1.47, m
10 11 4-OCH3 COOCH3
10.7, CH3 17.9, CH3 57.1, CH3
0.98, t (7.5) 2.28, s 3.88, s
5.57, s
9a δC, type 167.4, C 88.4, CH 173.3, C 113.3, C 159.9, C 21.6, CH2
10a
δH (J in Hz)
δC, type
5.56, s
39.1, CH2
2.53, ddd (14.0, 10.0, 5.5) 2.42, tdd (14.0, 10.0, 6.5) 1.52, m
68.0, CH 23.6, CH3 17.1, CH3 57.1, CH3
167.3, C 88.4, CH 173.2, C 112.8, C 160.3, C 24.4, CH2
11a δH (J in Hz)
5.46, s
2.34, t (7.5)
δC, type 167.2, C 88.4, CH 173.1, C 112.6, C 160.3, C 24.3, CH2
12b δH (J in Hz)
5.56, s
2.43, t (7.5)
δC, type 167.3, C 141.5, C 142.6, C 104.5, C 24.1, CH2 29.6, CH2
δH (J in Hz)
2.31, t (8.0) 1.59, dt (15.5, 7.5)
25.3, CH2
1.64, m
25.2, CH2
1.76, m
23.0, CH2
3.71, dq (12.5, 6.0)
34.1, CH2
2.21, t (7.0)
33.9, CH2
2.35, t (7.5)
13.8, CH3
1.39, dq (15.0, 7.5) 0.94, t (7.5)
1.18, d (6.0) 2.27, s 3.88, s
177.1, C 17.2, CH3 57.2, CH3
2.16, s 3.78, s
175.4, C 17.2, CH3 57.1, CH3 52.0, CH3
24.5, CH3 58.7, CH3
1.65, s 3.92, s
2.26, s 3.88, s 3.65, s
a
Measured in methanol-d4. bMeasured in CDCl3.
2.41)/H-8 (δH 3.55)/H2-9 (δH 1.52 and 1.47)/H3-10 (δH 0.98) (Figure S6). The small value of [α]25D (−0.6) suggested that 8 might be a racemic mixture. Subsequently, setosphapyrone A was resolved by HPLC using a chiral-phase column to afford a 45/55 ratio of (+)/(−)-enantiomers (Figure S2). Mosher’s ester analysis (Figure S4) established that the (−)-enantiomer of 8 had the 8R configuration;30 thus the (+)-enantiomer of 8 had the 8S configuration. Setosphapyrone B (9) had the same molecular formula as 8, which was established by their HRESIMS and 13C NMR data. Comparison of the 13C NMR data (Table 3) of setosphapyrone B with those of setosphapyrone A revealed a different oxygenated methine (δC 68.0, C-9) was shielded by 5.4 ppm, while the methyl (δC 23.6, C-10) was deshielded by 12.9 ppm compared to those of setosphapyrone A. Furthermore, in the 1 H NMR spectra an obvious distinction was the peak of H3-10 [1.18, d, J = 6.0 Hz] in setosphapyrone B was a doublet, while in setosphapyrone A it was a triplet. These changes demonstrated setosphapyrone B was a C-9 hydroxy derivative of 21, which was established by its COSY cross-peaks of H2-7 (δH 2.53 and 2.42)/H2-8 (δH 1.52)/H-9 (δH 3.71)/H3-10 (Figure S6). Setosphapyrone B was also a racemic mixture, which was suggested by the small value of [α]25D (+0.5). Chiral-phase separation of setosphapyrone B afforded a 42/58 ratio of (+)/(−)-enantiomers (Figure S3). The absolute configuration of C-9 in the (−)-enantiomer was determined as R by the modified Mosher’s method (Figure S4).30 Therefore, the absolute configuration of the (+)-enantiomer was 9S. The molecular formulas of setosphapyrones C (10) and D (11) were assigned as C11H14O5 and C12H16O5, respectively, based on their HRESIMS and 13C NMR data (Table 3). Analysis of their NMR data showed that compounds 10 and 11 differed by only one methoxy, while both of them possessed the same skeleton as 21. The presence of a carboxyl carbon (δC 177.1, C-10) in 10 and the disappearance of a methyl signal in 21 suggested that the carboxyl carbon was connected to C-9 in 10. The deduction was supported by the HMBC correlations from H2-8 (δH 1.64) and H2-9 (δH 2.21) to C-10 (Figure S6). The structure of 10 was further determined by a single-crystal
spectrum. The NMR data (Table 2) of 6 were very similar to those of 5, and the obvious difference was that the oxygenated methylene (δC 62.7, C-13) in 5 was replaced by a carboxyl (δC 177.1, C-13) in 6. The planar structure of 6 was established by the HMBC correlations (Figure S6) from H2-11 (δH 1.92 and 1.84) and H2-12 (δH 2.37 and 2.33) to C-13 (δC 177.1). Its NOESY cross-peaks (Figure S7) and the ECD spectrum (Figure 4) closely matched those of 5. Thereby, the absolute configuration of 6 was also determined as 3R, 4R, and 10S. Exserolide K (7) was obtained as a white powder with a molecular formula of C15H18O6, as confirmed by the HRESIMS analysis together with the 13C NMR data. The NMR data of 7 (Table 2) were closely related to those of exserolide C (20),13 except for the disappearance of a methoxy signal at C-6 in 20, which was replaced by a hydroxy group in 7. This was deduced by the chemical shift of the only methoxy group (δC/H 60.9/3.85) in 7, which was nearly the same as for the methoxy group at C-7 in 5 and 6. Also, the HMBC correlation (Figure S6) from OCH3 (δH 3.85) to C-7 (δC 136.3) further confirmed the conjecture. Compound 7 shared a planar structure with exserolide D.13 The NOESY spectrum of 7 (Figure S7) showed cross-peaks from H-3 to H2-11 and from H-4 to H-10, which indicated H-3 and H-4 possessed the trans-configuration as opposed to the cis-configuration in exserolide D. The absolute configuration of C-3 in 7 was deduced via the ECD exciton chirality method.13 The ECD spectrum of 7 exhibited a negative Cotton effect at 271 nm (Δε = −2.78) in accordance with that of exserolide C (270 nm, Δε = −4.66),13 which allowed assignment of the 3R absolute configuration. Therefore, the absolute configuration of 7 was determined as 3R, 4S, and 10R. Setosphapyrone A (8) was obtained as a white powder, and the molecular formula was established as C11H16O4 based on the HRESIMS and 13C NMR data. The NMR data for setosphapyrone A (Table 3) were quite similar to those of 5butyl-4-methoxy-6-methyl-2H-pyran-2-one (21),29 except for the presence of an oxygenated methine (δC 73.4, C-8), and the chemical shifts of C-7 (δC 33.1) and C-9 (δC 31.0) were deshielded by 9 ppm compared to those of 21. This deduction was confirmed by the COSY correlations of H2-7 (δH 2.58 and 1864
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
Table 4. 1H NMR and 13C NMR Data for Compounds 13 and 14
X-ray diffraction experiment with Cu Kα radiation (Figure 3). The 13C NMR data of 11 showed one more methoxy (δC 52.0) and C-10 (δC 175.4) was shielded by 1.7 ppm compared to C10 in 10, which demonstrated that 11 was the esterification product of 10 at C-10. This was confirmed by the key HMBC correlation from OCH3-11 (δH 3.65) to C-10 (Figure S6). Compound 12 was isolated as a yellowish oil. The molecular formula was established as C10H16O4 by the 13C NMR data and a sodium adduct ion at m/z 223.0946 [M + Na]+ in the HRESIMS spectrum. The 1H NMR spectrum displayed three methyls at δH 3.92 (s, H3-11), 1.65 (s, H3-10), and 0.94 (t, J = 7.5 Hz, H3-9) and three methylenes. The 13C NMR spectrum showed three methyls (δC 58.7, C-11; 24.5, C-10; 13.8, C-9), three sp3 methylenes (δC 29.6, C-7; 24.1, C-6; 23.0, C-8), an sp3 oxygenated nonprotonated carbon (δC 104.5, C-5), two sp2 nonprotonated carbons (δC 142.6, C-4; 141.5, C-3), and a conjugated carbonyl carbon (δC 167.3, C-2). Besides one carbonyl carbon and one double bond, there is one degree of unsaturation left, which indicated a ring in the structure of 12. Comparing the NMR data with 3-methoxy-4-methyl-5propylfuran-2(5H)-one31 suggested that 12 was a furan-2one derivative. The differences were the location of substituent groups on the furan ring. The COSY cross-peaks (Figure S6) of H2-6 (δH 2.31)/H2-7 (δH 1.59)/H2-8 (δH 1.39)/H3-9 indicated the presence of an n-butyl group. The HMBC correlations (Figure S6) from H3-11 to C-3, from H2-6 to C-3, C-4, and C-5, and from H3-10 to C-5 and C-4 demonstrated the methoxy group, n-butyl group, and methyl were connected at C-3, C-4, and C-5, respectively. The deshielded C-5 (δC 104.5) suggested a hydroxy connected at C-5. Thus, the planar structure of 12 was established as shown. Due to the existence of the cyclic hemiketal, compound 12 spontaneously racemized under protic solvent conditions. The racemic 12 was named 5-hydroxy-3-methoxy-5-methyl-4-butylfuran-2(5H)one. Botryorhodines I (13) and J (14) were isolated as white powders, and their molecular formulas were assigned as C18H18O6 and C21H24O6, respectively, based on their HRESIMS and 13C NMR data. The NMR data of 13 and 14 (Table 4) exhibited resonances for a methoxy group (δC/H 58.5/3.42, CH3-1″) and a butoxy group (δC/H 71.4/3.63, CH21″; 31.5/1.65, CH2-2″; 19.3/1.41, CH2-3″; 13.8/0.94, CH34″), respectively. The NMR data of 13 and 14 displayed similarity to those of botryorhodine C,32,33 except for the presence of methoxy and n-butoxy groups in 13 and 14, respectively, replacing the hydroxy group in botryorhodine C. These deductions were further supported by the key HMBC correlations from H-8 to C-1″ in 13 and 14 (Figure S6), key COSY cross-peaks of H2-1″/H2-2″/H2-3″/H3-4″ in 14 (Figure S6), and key NOESY cross-peaks of H-5 to H3-9 and of H-8′ to H-8 and H-1′ in 13 and 14. Thus, the structures of 13 and 14 were determined as new depsidones. By comparing NMR and specific rotation data with reported ones, the structures of known compounds 15−22 were identified as (12R)-12-hydroxymonocerin (15),28 11-hydroxymonocerin (16),11,12 exserolide E (17),13 7-O-demethylmonocerin (18),34 exserolide B (19),13 exserolide C (20),13 5butyl-4-methoxy-6-methyl-2H-pyran-2-one (21),29 and 5butyl-6-(hydroxymethyl)-4-methoxy-2H-pyran-2-one (22),29 respectively. The nine known compounds were identified and are listed in the Supporting Information (p S6). All of the new compounds were evaluated for their antifungal, antibacterial, antioxidant, antiviral, and MptpB inhibitory activities.
13a no.
δC, type
1 2 3 4 5 6 7 8 9 1′ 2′ 3′ 4′ 5′ 6′ 7′ 8′ 1″ 2″ 3″ 4″
113.6, C 162.7, C 114.3, C 163.7, C 116.1, CH 146.6, C 165.9, C 64.2, CH2 21.6, CH3 114.0, CH 154.0, C 115.3, C 144.7, C 144.0, C 128.6, C 9.3, CH3 16.5, CH3 58.5, CH3
14b
δH (J in Hz)
6.62, s
4.72, s 2.39, s 6.44, s
2.13, s 2.37, s 3.42, s
δC, type 113.1, C 160.8, C 110.9, C 160.1, C 116.5, CH 145.2, C 163.6, C 66.5, CH2 21.3, CH3 113.2, CH 151.1, C 114.5, C 143.6, C 143.6, C 126.9, C 9.2, CH3 16.9, CH3 71.4, CH2 31.5, CH2 19.3, CH2 13.8, CH3
δH (J in Hz)
6.60, s
5.05, s 2.45, s 6.41, s
2.22, 2.31, 3.63, 1.65, 1.41, 0.94,
s s t (7.0) m dq (14.7, 7.0) t (7.0)
a
Measured in methanol-d4 and at 500, 125 MHz NMR. bMeasured in CDCl3 and at 700, 175 MHz NMR.
Compounds 13 and 14 showed moderate antifungal activities against phytopathogenic Colletotrichum asianum with MIC values of 0.16 and 0.63 mg/mL and Colletotrichum acutatum with MIC values of 0.31 and 0.63 mg/mL, while cycloheximide was used as the positive control with MIC values of 0.08 and 0.04 mg/mL, respectively. Compound 18 exhibited radical scavenging activity against DPPH with an IC50 value of 38 μM, which was comparable to ascorbic acid with an IC50 value of 39 μM as the positive control. Unfortunately, none of the new compounds displayed antibacterial, antiviral, or MptpB inhibition activities.
■
EXPERIMENTAL SECTION
General Experimental Procedures. Melting points were measured using an SGWX-4 digital display micromelting point instrument. Optical rotations were measured using a MCP-500 polarimeter (Anton). UV spectra were recorded on a UV-2600 UV− vis spectrophotometer (Shimadzu). ECD spectra were measured with a Chirascan circular dichroism spectrometer (Applied Photophysics). IR spectra were recorded on an IR Affinity-1 FT-IR spectrometer (Shimadzu). 1D and 2D NMR spectra were measured on a Bruker AV 500 MHz or AVANCE HD 700 MHz NMR spectrometer with tetramethylsilane as an internal standard. HRESIMS spectra were recorded on a Bruker maXis Q-TOF mass spectrometer in positive/ negative ion mode. HPLC was carried out on a Hitachi Primaide with a YMC ODS Series column (YMC-Pack ODS-A, YMC Co. Ltd., 250 × 10 mm i.d., S-5 μm, 12 nm) and chiral-phase analysis (Phenomenex Lux cellulose-2 column, 4.6 mm × 25 mm). The TLC plates with silica gel GF254 (0.4−0.5 mm, Qingdao Marine Chemical Factory) were used for analysis and preparative TLC. Column chromatography was carried out on silica gel (200−300 mesh, Jiangyou Silica Gel Development Co.), Sephadex LH-20 (40−70 μm, Amersham Pharmacia Biotech AB), and YMC Gel ODS-A (12 nm, S-50 μm YMC). Spots were detected on TLC under UV light or by heating after spraying with the mixed solvent of saturated vanillin and 5% H2SO4 in H2O. X-ray diffraction intensity data were collected on an 1865
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
mg, tR = 26.0 min) and 6 (37.3 mg, tR = 30.0 min). Fr-4 (5.9 g) was subjected to CC eluting with a PE and acetone mixed solvent system in a step gradient (10:1−0:1, v/v) to gain seven fractions (Fr-4-1−Fr4-5). Fr-4-1 (308.7 mg) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by semipreparative HPLC (70% CH3CN/H2O, 1.5 mL/min) to afford 14 (3.9 mg, tR = 18.0 min). Fr-4-2 (771.9 mg) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by semipreparative HPLC (45% CH3CN/H2O, 2 mL/min) to afford 13 (7.0 mg, tR = 19.3 min). Fr-4-4 (1.3 g) was subjected to reversed-phase MPLC with MeOH/H2O (1:9−10:0, v/ v), then purified by semipreparative HPLC to afford 11 (32.3 mg, 15% CH3CN/H2O, 2 mL/min, tR = 41.0 min), 10 (25.9 mg, 15% CH3CN/H2O, 2 mL/min, tR = 42.6 min), and 7 (26.4 mg, 25% CH3CN/H2O, 2 mL/min, tR = 23.5 min). Setosphalide A (1): colorless needles; mp 151.6−152.4 °C; [α]25D +64 (c 0.10, MeOH); UV (MeOH) λmax (log ε) 221 (3.53), 262 (3.02), 298 (2.53) nm; ECD (0.53 mM, MeOH) λmax (Δε) 218 (−4.81), 228 (+1.95), 243 (−4.67), 266 (13.49) nm; IR (film) νmax 3406, 2957, 2936, 2872, 1748, 1738, 1614, 1479, 1474, 1368, 1269, 1123, 1105, 1020; 1H and 13C NMR data, Table 1; HRESIMS m/z 325.1290 [M + H]+ (calcd for C16H21O7, 325.1282). Setosphalide B (2): colorless oil; [α]25D +13 (c 0.10, MeOH); UV (MeOH) λmax (log ε) 221 (3.37), 262 (2.85), 298 (2.37) nm; ECD (0.49 mM, MeOH) λmax (Δε) 216 (−0.57), 261 (3.37) nm; IR (film) νmax 3420, 2957, 2936, 2870, 1734, 1717, 1614, 1474, 1456, 1373, 1265, 1123, 1022; 1H and 13C NMR data, Table 1; HRESIMS m/z 347.1108 [M + Na]+ (calcd for C16H20NaO7, 347.1101). 5-O-Desmethylcolletotrialide (3): colorless plate; [α]25D +9.6 (c 0.14, MeOH); UV (MeOH) λmax (log ε) 219 (3.46), 260 (3.03) 293 (2.49) nm; ECD (0.68 mM, MeOH) λmax (Δε) 218 (+0.54), 246 (−0.06), 262 (+0.10), 291 (−0.22) nm; IR (film) νmax 3442, 2965, 2938, 2873, 1732, 1717, 1705, 1616, 1609, 1508, 1456, 1373, 1339, 1069, 1009; 1H and 13C NMR data, Table 1; HRESIMS m/z 317.1017 [M + Na]+ (calcd for C15H18NaO6, 317.0996). (S)-Colletotrialide (4): yellowish oil; [α]25D −22 (c 0.10, MeOH); UV (MeOH) λmax (log ε) 219 (3.54), 259 (3.07), 297 (2.47) nm; ECD (0.36 mM, MeOH) λmax (Δε) 225 (−1.70), 247 (+0.13), 261 (−0.45), 298 (+0.36) nm; IR (film) νmax 3420, 2961, 2936, 1732, 1713, 1607, 1489, 1476, 1433, 1368, 1259, 1194, 1136, 1098, 1009; 1 H and 13C NMR data, Table 1; HRESIMS m/z 331.1161 [M + Na]+ (calcd for C16H20NaO6, 331.1152). Exserolide I (5): colorless oil; [α]25D +43 (c 0.10, MeOH); UV (MeOH) λmax (log ε) 219 (4.29), 273 (3.98), 306 (3.52) nm; ECD (0.60 mM, MeOH) λmax (Δε) 216 (+8.17), 232 (+4.17), 274 (−5.87) nm; IR (film) νmax 3509, 2943, 2870, 1668, 1662, 1653, 1522, 1456, 1394, 1379, 1339, 1275, 1121, 1034, 1013; 1H and 13C NMR data, Table 2; HRESIMS m/z 347.1113 [M + Na]+ (calcd for C16H20NaO7, 347.1101). Exserolide J (6): yellowish oil; [α]25D +21 (c 0.10, MeOH); UV (MeOH) λmax (log ε) 220 (4.27), 274 (3.94), 306 (3.49) nm; ECD (0.60 mM, MeOH) λmax (Δε) 215 (+6.64), 233 (+5.56), 274 (−9.44) nm; IR (film) νmax 3576, 2943, 2874, 1732, 1717, 1668, 1662, 1616, 1520, 1508, 1456, 1375, 1273, 1028; 1H and 13C NMR data, Table 2; HRESIMS m/z 337.0929 [M − H]− (calcd for C16H17O8, 337.0929). Exserolide K (7): white powder; [α]25D −35 (c 0.01, MeOH); UV (MeOH) λmax (log ε) 219 (4.21), 273 (3.93) nm; ECD (0.68 mM, MeOH) λmax (Δε) 219 (+3.37), 273 (−2.78), 305 (+0.38) nm; IR (film) νmax 3300, 2959, 2936, 2874, 1651, 1522, 1514, 1454, 1362, 1269, 1097; 1H and 13C NMR data, Table 2; HRESIMS m/z 295.1191 [M + H]+ (calcd for C15H19O6, 295.1176). Setosphapyrone A (8): white powder; [α]25D −0.6 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 205 (3.51), 285 (2.83) nm; IR (film) νmax 3443, 2965, 2938, 2874, 1697, 1645, 1560, 1456, 1408, 1250; 1H and 13 C NMR data, Table 3; HRESIMS m/z 213.1130 [M + H]+ (calcd for C11H17O4, 213.1121). 8S-(+)-Setosphapyrone A (3.8 mg): [α]25D +20 (c 0.1, MeOH); ECD (0.32 mM, MeOH) λmax (Δε) 219 (+0.95), 291 (−0.64) nm. 8R-(−)-Setosphapyrone A (5.3 mg): [α]25D −14 (c 0.1, MeOH); ECD (0.32 mM, MeOH) λmax (Δε) 216 (−0.73), 273 (+0.39).
Agilent Xcalibur Nova single-crystal diffractometer using Cu Kα radiation. Sea salt (Guangzhou Haili Aquarium Technology Company) was a commercial product. Fungal Material. The fungal strain SCSIO41009 was obtained from a Callyspongia sp. sponge, which was collected from the sea near Xuwen County, Guangdong Province, China. The producing strain was stored on MB agar (malt extract 15 g, sea salt 10 g, agar 16 g, H2O 1 L, pH 7.4−7.8) slants at 4 °C and deposited at Key Laboratory of Tropical Marine Bioresources and Ecology, Chinese Academy of Science. The ITS1-5.8S-ITS2 sequence region (550 base pairs (bp), GenBank accession no. MF685201) of strain SCSIO41009 was amplified by PCR, and DNA sequencing showed it shared significant homology to several species of Setosphaeria. The 550 bp ITS sequence has 99% sequence identity to that of Setosphaeria rostrata Ex 1 (GenBank accession no. KC198082.1), so it was designated as a Setosphaeria sp. and was named Setosphaeria sp. SCSIO41009. Fermentation and Extraction. The strain SCSIO41009 was cultured in 100 mL flasks (×60) each containing 10 mL of seed medium (malt extract 15 g, sea salt 2.5 g, tap H2O 1 L, pH 7.4−7.8) at 27 °C on a rotary shaker (172 rpm) for 48 h. The mass fermentation of this fungus was carried out at 25 °C for 32 days using a rice medium (rice 200 g/flask, sea salt 2.5 g/flask, distilled H2O 200 mL/flask) in the 1 L flask (×60). The flasks were incubated statically at 25 °C under a normal day/night cycle. After 32 days, the cultures were soaked in acetone (500 mL/flask), mashed into small pieces, and sonicated for 20 min. Then the acetone was evaporated under reduced pressure to afford an aqueous solution, which was extracted with EtOAc three times. At the same time, the rice residue was extracted with EtOAc to give another EtOAc solution. Both of the EtOAc solutions were combined and concentrated under reduced pressure to afford a crude extract. To remove the oil, the crude extract was suspended in MeOH and then partitioned with an equal volume of petroleum ether (PE). Finally, the MeOH solution was concentrated under reduced pressure to obtain a reddish-brown extract (106.0 g). Isolation and Purification. The reddish-brown extract was subjected to silica gel column chromatography (CC) eluting with a CH2Cl2 and MeOH mixed solvent system in a step gradient (100:0− 3:1, v/v) to give eight fractions (Fr-1−Fr-8). Fr-3 (15.8 g) was subjected to CC eluting with a PE and acetone mixed solvent system in a step gradient (10:1−0:1, v/v) to gain seven fractions (Fr-3-1−Fr3-7). Fr-3-1 (777.1 mg) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by semipreparative HPLC (60% MeOH/H2O, 3 mL/min) to afford 12 (65.3 mg, tR = 13.0 min) and 21 (4.0 mg, tR = 21.0 min). Fr-3-3 (1.3 g) was subjected to a Sephadex LH-20 column eluting with MeOH, then purified by semipreparative HPLC to afford 22 (290.6 mg, 33% CH3CN/H2O, 2 mL/min, tR = 18.2 min) and 3 (32.0 mg, 33% CH3CN/H2O, 2 mL/ min, tR = 27.2 min). Fr-3-4 (1.8 g) was subjected to a Sephadex LH20 column eluting with MeOH, followed by reversed-phase MPLC with MeOH/H2O (1:9−10:0, v/v), then purified by semipreparative HPLC to afford 8 (15.3 mg, 35% MeOH/H2O, 2 mL/min, tR = 33.8 min), 4 (18.1 mg, 32% CH3CN/H2O, 1.4 mL/min, tR = 35.0 min), 20 (37.5 mg, 33% CH3CN/H2O, 2.8 mL/min, tR = 19.6 min), 18 (8.0 mg, 33% CH3CN/H2O, 2.8 mL/min, tR = 33.5 min), 16 (18.3 mg, 18% CH3CN/H2O, 2 mL/min, tR = 78.0 min), and 17 (20.2 mg, 18% CH3CN/H2O, 2 mL/min, tR = 80.0 min). Fr-3-5 (3.3 g) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by reversed-phase MPLC with MeOH/H2O (1:9−10:0, v/v), then purified by semipreparative HPLC to afford 15 (51.9 mg, 30% CH3CN/H2O, 1.5 mL/min, tR = 31.2 min), 2 (10.7 mg, 32% CH3CN/H2O, 2 mL/min, tR = 17.4 min), and 1 (11.6 mg, 32% CH3CN/H2O, 2 mL/min, tR = 17.7 min). Fr-3-6 (578.6 mg) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by reversed-phase MPLC with MeOH/H2O (1:9−10:0, v/v), then purified by semipreparative HPLC to give 9 (22.3 mg, 22% CH3CN/ H2O, 4 mL/min, tR = 12.8 min) and 19 (16.2 mg, 30% CH3CN/H2O, 1.5 mL/min, tR = 39.0 min). Fr-3-7 (347.0 mg) was subjected to a Sephadex LH-20 column eluting with MeOH, followed by semipreparative HPLC (30% CH3CN/H2O, 1.5 mL/min) to give 5 (86.2 1866
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
Setosphapyrone B (9): white powder; [α]25D +0.5 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 204 (3.49), 285 (2.79) nm; IR (film) νmax 3379, 3211, 2968, 1695, 1630, 1557, 1456, 1408, 1368, 1260, 1248, 1192, 1130, 1045; 1H and 13C NMR data, Table 3; HRESIMS m/z 235.0949 [M + Na]+ (calcd for C11H16NaO4, 235.0941). 9S(+)-Setosphapyrone B (2.8 mg): [α]25D +5.0 (c 0.1, MeOH); ECD (0.52 mM, MeOH) λmax (Δε) 207 (−0.19), 223 (+0.45), 291 (−0.64) nm. 9R-(−)-Setosphapyrone B (3.4 mg): [α]25D −9.6 (c 0.1, MeOH); ECD (0.80 mM, MeOH) λmax (Δε) 208 (+1.06), 280 (+0.24). Setosphapyrone C (10): colorless block; mp 123.2−124.1 °C; UV (MeOH) λmax (log ε) 208 (4.33), 284 (3.84) nm; IR (film) νmax 3464, 2957, 2936, 1695, 1684, 1557, 1456, 1408, 1250, 1248, 1163, 1099, 1024; 1H and 13C NMR data, Table 3; HRESIMS m/z 225.0774 [M − H]− (calcd for C11H13O5, 225.0768). Setosphapyrone D (11): colorless oil; UV (MeOH) λmax (log ε) 210 (4.30), 284 (3.97) nm; IR (film) νmax 3464, 2957, 2936, 2872, 1732, 1717, 1645, 1560, 1456, 1406, 1248, 1163, 1099, 1024; 1H and 13 C NMR data, Table 3; HRESIMS m/z 263.0895 [M + Na]+ (calcd for C12H16NaO5, 263.0890) and 241.1072 [M + H]+ (calcd for C12H17O5, 241.1071). 5-Hydroxy-3-methoxy-5-methyl-4-butylfuran-2(5H)-one (12): pale yellow oil; UV (MeOH) λmax (log ε) 200 (2.70), 231 (2.78) nm; IR (film) νmax 3402, 2959, 2874, 1749, 1684, 1456, 1373, 1227, 1153, 1084, 1061; 1H and 13C NMR data, Table 3; HRESIMS m/z 223.0946 [M + Na]+ (calcd for C10H16NaO4, 223.0941). Botryorhodine I (13): white powder; UV (MeOH) λmax (log ε) 207 (4.17), 268 (4.17) nm; IR (film) νmax 3362, 2928, 2870, 1714, 1697, 1688, 1607, 1456, 1418, 1269, 1211, 1142, 1109, 1086; 1H and 13C NMR data, Table 4; HRESIMS m/z 353.1004 [M + Na]+ (calcd for C18H18NaO6, 353.0996). Botryorhodine J (14): white powder; UV (MeOH) λmax (log ε) 203 (4.66), 268 (4.05) nm; IR (film) νmax 3325, 2959, 2916, 2849, 1715, 1697, 1608, 1576, 1456, 1423, 1267, 1142, 1107, 1086; 1H and 13 C NMR data, Table 4; HRESIMS m/z 395.1470 [M + Na]+ (calcd for C21H24NaO6, 395.1465). Crystallographic Data for Setosphalide A (1). Moiety formula: C16H20O7 (MW = 324.32), colorless plate, crystal size = 0.45 × 0.4 × 0.08 mm3, orthorhombic, space group P212121 (No. 19); unit cell dimensions: a = 7.73476(8) Å, b = 8.02415(8) Å, c = 25.5244(3) Å, V = 1584.17(3) Å3, Z = 4, ρcalcd = 1.360 mg mm−3, T = 150.00(10) K, μ(Cu Kα) = 0.904 mm−1. A total of 6385 reflections were measured with 3118 independent reflections (Rint = 0.0178, Rsigma = 0.0229). Final R indices [I > 2σ(I)]: R1 = 0.0306, wR2 = 0.0840. Final R indexes [all data]: R1 = 0.0310, wR2 = 0.0842, Flack parameter = 0.01(8). Largest diff peak and hole = 0.198 and −0.235 e Å−3. Crystallographic Data for 5-O-Desmethylcolletotrialide (3). Moiety formula: C15H18O6 (MW = 294.29), colorless plate, crystal size = 0.6 × 0.4 × 0.02 mm3, monoclinic, space group P21 (No. 4); unit cell dimensions: a = 5.6966(3) Å, b = 5.6223(2) Å, c = 22.2355(14) Å, V = 708.40(7) Å3, Z = 2, ρcalcd = 1.380 mg mm−3, T = 150.00(10) K, μ(Cu Kα) = 0.899 mm−1. A total of 12 240 reflections were measured with 2810 independent reflections (Rint = 0.0909, Rsigma = 0.0486). Final R indices [I > 2σ(I)]: R1 = 0.0633, wR2 = 0.1729. Final R indexes [all data]: R1 = 0.0752, wR2 = 0.1907, Flack parameter = 0.0(2). Largest diff peak and hole = 0.335 and −0.333 e Å−3. Crystallographic Data for Setosphapyrone C (10). Moiety formula: C11H14O5 (MW = 226.22), colorless block, crystal size = 0.75 × 0.35 × 0.12 mm3, monoclinic, space group P21/n (No. 14); unit cell dimensions: a = 4.2311(6) Å, b = 13.8888(18) Å, c = 18.560(2) Å, V = 1090.1(2) Å3, Z = 4, ρcalcd = 1.378 mg mm−3, T = 150.00(10) K, μ(Cu Kα) = 0.924 mm−1. A total of 3531 reflections were measured with 2144 independent reflections (Rint = 0.0430, Rsigma = 0.0451). Final R indices [I > 2σ(I)]: R1 = 0.0673, wR2 = 0.1889. Final R indexes [all data]: R1 = 0.0844, wR2 = 0.2162. Largest diff peak and hole = 0.270 and −0.358 e Å−3. Compounds 1, 3, and 10 were crystallized from MeOH. A single crystal with the indicated dimensions was selected and measured on an Agilent Xcalibur Nova single-crystal diffractometer using Cu Kα
radiation and refined by full-matrix least-squares calculation. The crystallographic data for the structures 1, 3, and 10 have been deposited with the Cambridge Crystallographic Data Centre as supplementary publication number CCDC-1589160 for 1, CCDC1589350 for 3, and CCDC-1589691 for 10, respectively. ECD Calculations. The theoretical calculations of new compounds 2 and 5 were performed by using the density functional theory (DFT) as carried out in Gaussian 03.35 All ground-state geometries were optimized at the B3LYP/6-31G(d) level. Conformational analysis was initially conducted by using HyperChem 7.5 software. Solvent effects of the MeOH solution were evaluated at the same DFT level by using the SCRF/PCM method.36,37 TDDFT at the B3LYP/6-31G(d) level was employed to calculate the electronic excitation energies and rotational strengths in MeOH.38 Antimicrobial Activity Assay. Compounds 1−14 were tested for antibacterial activities against Staphylococcus aureus, and antifungal activities against C. gloeosporioides, C. asianum, C. acutatum, Fusarium oxysporum, and Pyricularia oryza, respectively, were evaluated in 96well plates using a modification of the broth microdilution method.39 Antioxidant Assay. All compounds were evaluated for their antioxidant capacity by the DPPH radical scavenging assay.40 Briefly, 20 μL of different concentrations of tested compounds including ascorbic acid as a positive control in DMSO were added to 180 μL of 0.2 mM fresh DPPH solution in EtOH with final concentrations of 50, 25, 12.5, 6.25, and 3.125 μg/mL in triplicate. After 30 min reaction in the dark, the decrease of the absorbance was measured on an Enspire Genios microplate reader at 517 nm. The blank contained EtOH instead of the DPPH solution, and the control contained DMSO in place of the sample solution. The formula DPPH scavenging activity (%) = [1 − (absorbance of sample−absorbance of blank)/absorbance of control] × 100 was used to calculate the inhibition rate. The IC50 (the concentration required to scavenge 50% of radicals) values of tested compounds were calculated using the GraphPad Prism software.
■
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jnatprod.8b00345. X-ray crystallographic file of compound 1 (CIF) X-ray crystallographic file of compound 3 (CIF) X-ray crystallographic file of compound 10 (CIF) COSY, key HMBC correlations of 2−14, key NOESY correlations of 5−7, 1D and 2D NMR spectra, HRESIMS for the new compounds 1−14 (PDF)
■
AUTHOR INFORMATION
Corresponding Authors
*Tel: +86-020-8902-3174. E-mail:
[email protected]. *Tel: +86-020-8902-3244. E-mail:
[email protected]. ORCID
Jie Yang: 0000-0003-1789-690X Xuefeng Zhou: 0000-0001-9601-4869 Junfeng Wang: 0000-0001-6702-5366 Yonghong Liu: 0000-0001-8327-3108 Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS This work was supported by the National Natural Science Foundation of China (Nos. 21172230, 21772210, 21502204, 31270402, 41476135, and 41776169), the Strategic Priority Research Program of the Chinese Academy of Sciences (XDA11030403), the Guangdong Province Public Welfare 1867
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
Journal of Natural Products
Article
(28) Zhang, W.; Krohn, K.; Draeger, S.; Schulz, B. J. Nat. Prod. 2008, 71, 1078−1081. (29) Metwaly, A. M.; Fronczek, F. R.; Ma, G. Y.; Kadry, H. A.; ElHela, A. A.; Mohammad, A. I.; Cutler, S. J.; Ross, S. A. Tetrahedron Lett. 2014, 55, 3478−3481. (30) Wang, J. F.; He, W. J.; Kong, F. D.; Tian, X. P.; Wang, P.; Zhou, X. J.; Liu, Y. H. J. Nat. Prod. 2017, 80, 1725−1733. (31) Anderson, J. R.; Edwards, R. L.; Whalley, A. J. S. J. Chem. Soc., Perkin Trans. 1 1982, 13, 215−221. (32) Abdou, R.; Scherlach, K.; Dahse, H. M.; Sattler, I.; Hertweck, C. Phytochemistry 2010, 71, 110−116. (33) Chen, S. H.; Liu, Z. M.; Liu, Y. Y.; Lu, Y. J.; He, L.; She, Z. G. Beilstein J. Org. Chem. 2015, 11, 1187−1193. (34) Fang, B.; Xie, X. G.; Zhao, C. G.; Jing, P.; Li, H. L.; Wang, Z. S.; Gu, J. X.; She, X. G. J. Org. Chem. 2013, 78, 6338−6343. (35) Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A., Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C.; Millam, J. M.; Iyengar, S. S.; Tomasi, J.; Barone, V.; Mennucci, B.; Cossi, M.; Scalmani, G.; Rega, N.; Petersson, G. A.; Nakatsuji, H.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai, H.; Klene, M.; Li, X.; Knox, J. E.; Hratchian, H. P.; Cross, J. B.; Bakken, V.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.; Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J. W.; Ayala, P. Y.; Morokuma, K.; Voth, G. A.; Salvador, P.; Dannenberg, J. J.; Zakrzewski, V. G.; Dapprich, S.; Daniels, A. D.; Strain, M. C.; Farkas, O.; Malick, D. K.; Rabuck, A. D.; Raghavachari, K.; Foresman, J. B.; Ortiz, J. V.; Cui, Q.; Baboul, A. G.; Clifford, S.; Cioslowski, J.; Stefanov, B. B.; Liu, G.; Liashenko, A.; Piskorz, P.; Komaromi, I.; Martin, R. L.; Fox, D. J.; Keith, T.; Al-Laham, M. A., Peng, C. Y.; Nanayakkara, A.; Challacombe, M.; Gill, P. M. W.; Johnson, B.; Chen, W.; Wong, M. W.; Gonzalez, C.; Pople, J. A. Gaussian 03, Revision E.01; Gaussian, Inc.: Wallingford, CT, 2004. (36) Cammi, R.; Tomasi, J. J. Comput. Chem. 1995, 16, 1449−1458. (37) Tomasi, J.; Persico, M. Chem. Rev. 1994, 94, 2027−2094. (38) Gross, E. K. U.; Dobson, J. F.; Petersilka, M. Top. Curr. Chem. 1996, 181, 81−172. (39) Wang, J. F.; Cong, Z. W.; Huang, X. L.; Hou, C. X.; Chen, W. H.; Tu, Z. C.; Huang, D. Y.; Liu, Y. H. Org. Lett. 2018, 20, 1371− 1374. (40) Huang, Z.; Nong, X.; Ren, Z.; Wang, J.; Zhang, X.; Qi, S. Bioorg. Med. Chem. Lett. 2017, 27, 787−791.
Research and Capacity Building Project (No. 2016A020222010), and Pearl River S&T Nova Program of Guangzhou (No. 201710010136).
■
REFERENCES
(1) Rateb, M. E.; Ebel, R. Nat. Prod. Rep. 2011, 28, 290−344. (2) De, R. P.; Pal, A. K.; Purkayastha, R. P. Indian J. Exp. Biol. 1999, 37, 706−709. (3) Guiraud, P.; Steiman, R. G.; Seigle-Murandi, F. O.; Sage, L. Antonie van Leeuwenhoek 1997, 72, 317−325. (4) Mideros, S. X.; Chung, C. L.; Wiesner-Hanks, T.; Poland, J. A.; Wu, D.; Fialko, A. A.; Turgeon, B. G.; Nelson, R. J. Phytopathology 2018, 108, 254−263. (5) Wu, D. L.; Turgeon, B. G. Fungal Genet. Biol. 2013, 61, 158− 163. (6) Gu, S. Q.; Li, P.; Wu, M.; Hao, Z. M.; Gong, X. D.; Zhang, X. Y.; Tian, L.; Zhang, P.; Wang, Y.; Cao, Z. Y.; Fan, Y. S.; Han, J. M.; Dong, J. G. Microbiol. Res. 2014, 169, 817−823. (7) Bai, Y. J.; Liu, H. Y.; Fan, Y. S.; Li, J. J. Plant Dis. Pests 2012, 3, 51−53. (8) Sugawara, K.; Sugawara, F.; Strobel, G. A. J. Org. Chem. 1985, 50, 5631−5633. (9) Bashan, B.; Levy, R. S.; Cojocaru, M.; Levy, Y. Physiol. Mol. Plant Pathol. 1995, 47, 225−235. (10) Tan, R. X.; Jensen, P. R.; Williams, P. G.; Fenical, W. J. Nat. Prod. 2004, 67, 1374−1382. (11) Sappapan, R.; Sommit, D.; Ngamrojanavanich, N.; Pengpreecha, S.; Wiyakrutta, S.; Sriubolmas, N.; Pudhom, K. J. Nat. Prod. 2008, 71, 1657−1659. (12) Sappapan, R.; Sommit, D.; Ngamrojanavanich, N.; Pengpreecha, S.; Wiyakrutta, S.; Sriubolmas, N.; Pudhom, K. J. Nat. Prod. 2008, 71, 2080. (13) Li, R. X.; Chen, S. X.; Niu, S. B.; Guo, L. D.; Yin, J.; Che, Y. S. Fitoterapia 2014, 96, 88−94. (14) Richter, J.; Sandjo, L. P.; Liermann, J. C.; Opatz, T.; Erkel, G. Bioorg. Med. Chem. 2015, 23, 556−563. (15) Pinheiro, E. A. A.; Borges, F. C.; Pina, J. R. S.; Ferreira, L. R. S.; Cordeiro, J. S.; Carvalho, J. M.; Feitosa, A. O.; Campos, F. R.; Barison, A.; Souza, A. D. L.; Marinho, P. S. B.; Marinho, A. M. R. J. Braz. Chem. Soc. 2016, 27, 1432−1436. (16) Centko, R. M.; Ratnaweera, P. B.; Tysoe, C.; Withers, S. G.; de Silva, E. D.; Andersen, R. J. Phytochem. Lett. 2017, 22, 76−80. (17) Bugni, T. S.; Ireland, C. M. Nat. Prod. Rep. 2004, 21, 143−163. (18) Wang, J. F.; Lin, X. P.; Qin, C.; Liao, S. R.; Wan, J. T.; Zhang, T. Y.; Liu, J.; Fredimoses, M.; Chen, H.; Yang, B.; Zhou, X. F.; Yang, X. W.; Tu, Z. C.; Liu, Y. H. J. Antibiot. 2014, 67, 581−583. (19) Tian, Y. Q.; Lin, X. P.; Wang, Z.; Zhou, X. F.; Qin, X. C.; Kaliyaperumal, K.; Zhang, T. Y.; Tu, Z. C.; Liu, Y. H. Molecules 2016, 21, 34. (20) Pang, X. Y.; Lin, X. P.; Tian, Y. Q.; Liang, R.; Wang, J. F.; Yang, B.; Zhou, X. F.; Kaliyaperumal, K.; Luo, X. W.; Tu, Z. C.; Liu, Y. H. Nat. Prod. Res. 2018, 32, 1−7. (21) Pang, X. Y.; Lin, X. P.; Wang, J. F.; Liang, R.; Tian, Y. Q.; Salendra, L.; Luo, X. W.; Zhou, X. F.; Yang, B.; Tu, Z. C.; Liu, Y. H. Steroids 2018, 129, 41−46. (22) Liu, J.; Li, F.; Kim, E. L.; Li, J. L.; Hong, J.; Bae, K. S.; Chung, H. Y.; Kim, H. S.; Jung, J. H. J. Nat. Prod. 2011, 74, 1826−1829. (23) Wang, H.; Hong, J.; Yin, J.; Moon, H. R.; Liu, Y.; Wei, X.; Oh, D. C.; Jung, J. H. J. Nat. Prod. 2015, 78 (11), 2832−2836. (24) Chou, T. H.; Chen, I. S.; Hwang, T. L.; Wang, T. C.; Lee, T. H.; Cheng, L. Y.; Chang, Y. C.; Cho, J. Y.; Chen, J. J. J. Nat. Prod. 2008, 71, 1692−1695. (25) Tianpanich, K.; Prachya, S.; Wiyakrutta, S.; Mahidol, C.; Ruchirawat, S.; Kittakoop, P. J. Nat. Prod. 2011, 74, 79−81. (26) Takahashi, H.; Tsubuki, T.; Higashiyama, K. Chem. Pharm. Bull. 1991, 39, 3136−3139. (27) Nakano, H.; Kumagai, N.; Matsuzaki, H.; Kabuto, C.; Hongo, H. Tetrahedron: Asymmetry 1997, 8, 1391−1401. 1868
DOI: 10.1021/acs.jnatprod.8b00345 J. Nat. Prod. 2018, 81, 1860−1868
