Zehong Wua,
Dong Liua,
Jian Huanga,
Peter Prokschb,
Kui Zhu
c and
Wenhan Lin
*a
aState Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing 100191, P. R. China. E-mail: whlin@bjmu.edu.cn; Fax: +86-10-82802724; Tel: +86-10-82806188
bInstitute für Pharmazeutische Biologie und Biotechnologie, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, 40225, Germany
cCollege of Veterinary Medicine, China Agricultural University, Beijing 100193, P. R. China
First published on 28th November 2018
Bioassay-guided fractionation and chromatographic separation of a sponge-derived fungus Hansfordia sinuosae, resulted in the isolation of thirteen new polyesters namely hansforesters A–M (1–13), along with five known analogues involving ascotrichalactone A, ascotrichester B, 15G256π, 6R-hydroxymellein, and (−)orthosporin. The structures of the new compounds were determined through extensive spectroscopic analysis, in addition to the chemical conversion for the configurational assignment. The polyesters incorporating the motifs of orsellinic acid, 2,4-dihydroxy-6-acetonylbenzoic acid, and orcinotriol were found from nature for the first time. Hansforester A (1) and ascotrichalactone A exhibited potent inhibition against a panel of bacterial strains, including the agricultural pathogenic bacteria, Pseudomonas lachrymans, Agrobacterium tumefaciens, Xanthomonas vesicatoria, and Ralstonia solanacearum, with the MIC values of 15.6 μM, and the human infected bacterium Staphylococcus aureus with the MIC values of 3.9 μM. These findings suggested that hansforester A and ascotrichalactone A are the potential leads to be developed as the antibacterial agents for the treatment of agriculture bacterial pathogens.
Fraction | MIC (μg mL−1) | |||
---|---|---|---|---|
P. lachrymans | A. tumefaciens | X. vesicatoria | R. solanacearum | |
EA extract | 96 | 128 | 64 | 128 |
F1 | >256 | >256 | >256 | >256 |
F2 | >256 | >256 | >256 | >256 |
F3 | >256 | >256 | >256 | >256 |
F4 | 64 | 128 | 64 | 96 |
F5 | 64 | 96 | 32 | 96 |
F6 | 64 | 96 | 32 | 96 |
Streptomycin sulfate | 75 | 50 | 100 | 50 |
No. | 1 | 2 | 3 | 4 | 5 | 6 | 7 |
---|---|---|---|---|---|---|---|
3 | 6.13, d (1.5) | 6.23, d (2.2) | 6.22, d (2.2) | 6.22, d (2.3) | 6.22, d (2.0) | 6.23, d (1.5) | 6.22, d (1.5) |
5 | 6.14, d (1.5) | 6.14, d (2.2) | 6.14, d (2.2) | 6.14, d (2.3) | 6.13, d (2.0) | 6.13, d (1.5) | 6.13, d (1.5) |
8 | 2.19, s | 3.81, d (17.5) | 3.81, d (17.5) | 3.87, d (17.5) | 3.90, d (17.4) | 3.87, d (17.4) | 3.86, d (17.6) |
4.07, d (17.5) | 4.06, d (17.5) | 3.72, d (17.5) | 3.75, d (17.4) | 3.77, d (17.4) | 3.76, d (17.6) | ||
10 | 6.12, dd (1.2, 15.6) | 6.12, dd (1.6, 15.6) | 2.05, s | 2.52, dd (7.4, 15.7) | 2.61, dd (7.0, 14.0) | 2.64, dd (7.0, 14.0) | |
2.39, dd (5.3, 15.7) | 2.43, dd (6.0, 14.0) | 2.43, dd (6.0, 14.0) | |||||
11 | 6.86, dq (6.8, 15.6) | 6.86, dq (6.7, 15.6) | 4.02, ddq (5.3, 6.3, 7.4) | 3.67, ddq (6.0, 6.1, 7.0) | 3.67, tq (6.0, 7.0) | ||
12 | 1.88, dd (1.2, 6.8) | 1.87, dd (1.6, 6.7) | 1.07, d (6.3) | 1.07, d (6.1) | 1.06, d (6.0) | ||
2′ | 2.66, d (7.0) | 2.50, d (7.0) | 2.49, d (7.0) | 2.57, d (6.5) | 2.58, d (7.0) | 2.58, d (7.0) | 2.56, d (7.0) |
3′ | 5.36, tq (6.0, 7.0) | 5.27, tq (6.2, 7.0) | 5.26, tq (6.2, 7.0) | 5.30, tq (6.4, 6.5) | 5.27, tq (6.2, 7.0) | 5.26, tq (6.0, 7.0) | 5.27, tq (6.0, 7.0) |
4′ | 1.26, d (6.0) | 1.07, d (6.2) | 1.07, d (6.2) | 1.14, d (6.4) | 1.14, d (6.2) | 1.15, d (6.0) | 1.13, d (6.0) |
3′′ | 6.18, d (1.5) | 6.19, d (2.0) | 6.18, d (2.2) | 6.18, d (2.3) | 6.18, d (2.0) | 6.18, d (1.5) | 6.18, d (1.5) |
5′′ | 6.15, d (1.5) | 6.16, d (2.0) | 6.15, d (2.2) | 6.15, d (2.3) | 6.15, d (2.0) | 6.16, d (1.5) | 6.17, d (1.5) |
8′′ | 2.75, dd (5.0, 13.2) | 2.75, dd (8.5, 13.5) | 2.71, dd (8.5, 13.5) | 2.75, dd (8.5, 13.6) | 2.87, dd (5.0, 13.6) | 2.88, dd (4.8, 13.6) | 2.86, dd (5.0, 14.0) |
2.87, dd (8.5, 13.2) | 2.87, dd (4.7, 13.5) | 2.86, dd (4.6, 13.5) | 2.88, dd (5.0, 13.6) | 2.75, dd (8.2, 13.6) | 2.76, dd (8.0, 13.6) | 2.71, dd (7.0, 14.0) | |
9′′ | 4.98, ddq (5.0, 6.1, 8.5) | 5.00, ddq (4.7, 6.2, 8.5) | 4.98, ddq (4.6, 6.2, 8.5) | 5.00, ddq (5.0, 6.2, 8.5) | 5.00, ddq (5.0, 6.2, 8.2) | 5.01, ddq (4.8, 6.5, 8.0) | 4.98, ddq (5.0, 6.0, 7.0) |
10′′ | 1.11, d (6.1) | 1.10, d (6.2) | 1.10, d (6.2) | 1.11, d (6.2) | 1.10, d (6.2) | 1.11, d (6.5) | 1.09, d (6.0) |
2′′′ | 2.62, d (6.5) | 2.66, d (6.5) | 2.60, d (6.5) | 2.66, d (6.5) | 2.66, d (6.5) | 2.66, d (6.5) | 2.61, d (6.0) |
3′′′ | 5.31, tq (6.0, 6.5) | 5.36, tq (6.2, 6.5) | 5.35, tq (6.2, 6.5) | 5.36, tq (6.3, 6.5) | 5.37, tq (6.3, 6.5) | 5.37, tq (5.8, 6.5) | 5.33, tq (5.7, 6.0) |
4′′′ | 1.24, d (6.0) | 1.25, d (6.2) | 1.21, d (6.2) | 1.26, d (6.3) | 1.26, d (6.3) | 1.26, d (5.8) | 1.21, d (5.7) |
1′′′′ | 6.05, s | 6.05, s | 6.04, s | 6.06, s | 6.05, s | ||
3′′′′ | 6.05, s | 6.05, s | 6.15, d (3.3) | 6.04, s | 6.06, s | 6.05, s | 6.17, d (2.0) |
5′′′′ | 6.05, s | 6.05, s | 6.17, d (3.3) | 6.04, s | 6.06, s | 6.05, s | 6.14, d (2.0) |
7′′′′ | 2.50, m; 2.67, m | 2.50, dd (7.0, 14.2) | 2.88, dd (4.5, 14.4) | 2.50, dd (5.0, 14.0) | 2.66, dd (5.0, 13.5) | 2.67, dd (5.0, 14.0) | 3.14, dd (5.0, 14.0) |
2.67, dd (6.5, 14.2) | 3.14, dd (7.0, 14.4) | 2.66, dd (7.0, 14.0) | 2.50, dd (6.5, 13.5) | 2.50, dd (6.5, 14.0) | 2.86, dd (6.0, 14.0) | ||
8′′′′ | 4.93, m | 4.92, ddq (6.2, 6.5, 7.0) | 5.04, ddq (4.5, 6.2, 7.0) | 4.93, ddq (5.0, 6.3, 7.0) | 4.93, ddq (5.0, 6.3, 6.5) | 4.93, ddq (5.0, 6.3, 6.5) | 5.05, ddq (5.0, 6.0, 6.3) |
9′′′′ | 1.08, d (6.1) | 1.09, d (6.2) | 1.11, d (6.2) | 1.09, d (6.3) | 1.09 (d, 6.3) | 1.09, d (6.3) | 1.11, d (6.3) |
OH-2 | 10.93, s | 11.22, s | 11.20, s | 11.01, s | 10.98, s | 10.99, s | 11.01, s |
OH-4 | 10.07, s | 10.28, s | 10.27, s | 10.23, s | 10.23, s | 10.22, s | 10.25, brs |
OH-2′′ | 10.56, s | 10.56, s | 10.59, s | 10.55, s | 10.58, s | 10.56, s | 10.62, s |
OH-4′′ | 9.98, brs | 10.00, s | 10.01, s | 9.96, s | 9.99, s | 9.98, s | 10.03, s |
OH-3′′′′ | 9.10, brs | 9.11, s | 11.86, s | 9.08, s | 9.11, s | 9.09, s | 11.89, brs |
OH-5′′′′ | 9.10, brs | 9.11, s | 10.14, s | 9.08, s | 9.11, s | 9.09, s | 10.17, brs |
COOH | 13.52, brs | 13.63, brs | |||||
OMe | 3.17, s | 3.17, s |
No. | 1 | 2 | 3 | 4 | 5 | 6 | 7 |
---|---|---|---|---|---|---|---|
a 13C NMR data were measured in 125 MHz. | |||||||
1 | 106.8, C | 105.9, C | 105.9, C | 106.4, C | 106.7, C | 106.8, C | 106.8, C |
2 | 162.9, C | 163.7, C | 163.7, C | 163.2, C | 163.0, C | 163.0, C | 163.0, C |
3 | 100.9, CH | 101.9, CH | 101.9, CH | 102.0, CH | 102.0, CH | 102.1, CH | 102.1, CH |
4 | 161.2, C | 162.4, C | 162.4, C | 162.3, C | 162.1, C | 162.1, C | 162.1, C |
5 | 111.0, CH | 112.9, CH | 112.9, C | 112.6, CH | 112.6, CH | 112.5, CH | 112.5, CH |
6 | 141.9, C | 139.7, C | 139.7, C | 139.3, C | 139.1, C | 138.9, C | 138.9, C |
7 | 169.7, C | 169.7, C | 169.5, C | 169.2, C | 169.2, C | 169.3, C | 169.2, C |
8 | 22.9, CH3 | 47.8, CH2 | 47.8, CH2 | 50.7, CH2 | 50.7, CH2 | 50.4, CH2 | 50.4, CH2 |
9 | 196.6, C | 196.6, C | 205.4, C | 206.4, C | 205.6, C | 205.6, C | |
10 | 131.4, CH | 131.4, CH | 29.5, CH3 | 51.4, CH2 | 48.7, CH2 | 48.7, CH2 | |
11 | 143.6, CH | 143.4, CH | 63.2, CH | 72.6, CH | 72.6, CH | ||
12 | 18.3, CH3 | 18.3, CH3 | 24.1, CH3 | 19.3, CH3 | 19.3, CH3 | ||
1′ | 169.8, C | 169.4, C | 169.4, C | 169.5, C | 169.5, C | 169.5, C | 169.5, C |
2′ | 40.7, CH2 | 40.4, CH2 | 40.4, CH2 | 40.5, CH2 | 40.5, CH2 | 40.4, CH2 | 40.4, CH2 |
3′ | 68.6, CH | 68.6, CH | 68.6, CH | 68.6, CH | 68.6, CH | 68.7, CH | 68.7, CH |
4′ | 19.7, CH3 | 19.3, CH3 | 19.3, CH3 | 19.4, CH3 | 19.4, CH3 | 19.4, CH3 | 19.4, CH3 |
1′′ | 108.6, C | 108.8, C | 108.5, C | 108.6, C | 108.6, C | 108.6, C | 108.3, C |
2′′ | 162.0, C | 160.9, C | 161.2, C | 161.1, C | 161.2, C | 161.1, C | 161.3, C |
3′′ | 101.7, CH | 101.7, CH | 101.7, CH | 101.7, CH | 101.7, CH | 101.7, CH | 101.7, CH |
4′′ | 161.2, C | 161.1, C | 161.2, C | 161.1, C | 161.0, C | 161.1, C | 161.2, C |
5′′ | 111.0, CH | 110.8, CH | 110.9, CH | 111.0, CH | 111.0, CH | 111.0, CH | 111.1, CH |
6′′ | 140.3, C | 140.2, C | 140.4, C | 140.3, C | 140.3, C | 140.3, C | 140.5, C |
7′′ | 168.9, C | 168.8, C | 168.9, C | 168.9, C | 168.9, C | 168.9, C | 168.9, C |
8′′ | 40.6, CH2 | 40.6, CH2 | 40.7, CH2 | 40.9, CH2 | 40.7, CH2 | 40.7, CH2 | 40.8, CH2 |
9′′ | 71.5, CH | 71.4, CH | 71.4, CH | 71.5, CH | 71.4, CH | 71.4, CH | 71.4, CH |
10′′ | 20.0, CH3 | 20.0, CH3 | 20.0, CH3 | 20.0, CH3 | 20.0, CH3 | 20.0, CH3 | 20.0, CH3 |
1′′′ | 169.6, C | 169.6, C | 169.6, C | 169.7, C | 169.7, C | 169.6, C | 169.4, C |
2′′′ | 40.7, CH2 | 40.7, CH2 | 40.6, CH2 | 40.7, CH2 | 40.7, CH2 | 40.7, CH2 | 40.7, CH2 |
3′′′ | 68.6, CH | 68.5, CH | 68.4, CH | 68.6, CH | 68.5, CH | 68.5, CH | 68.5, CH |
4′′′ | 19.7, CH3 | 19.7, CH3 | 19.6, CH3 | 19.7, CH3 | 19.7, CH3 | 19.7, CH3 | 19.6, CH3 |
1′′′′ | 107.6, CH | 107.6, CH | 105.7, C | 107.6, CH | 107.6, CH | 107.6, CH | 105.7, C |
2′′′′ | 158.7, C | 158.7, C | 164.2, C | 158.7, C | 158.7, C | 158.7, C | 164.2, C |
3′′′′ | 101.2, CH | 101.1, CH | 101.8, CH | 101.1, CH | 101.1, CH | 101.1, CH | 101.8, CH |
4′′′′ | 158.7, C | 158.7, C | 162.0, C | 158.7, C | 158.7, C | 158.7, C | 162.0, C |
5′′′′ | 107.6, CH | 107.6, CH | 112.0, CH | 107.6, CH | 107.6, CH | 107.6, CH | 112.0, CH |
6′′′′ | 139.7, C | 139.7, C | 142.4, C | 139.7, C | 139.7, C | 139.7, C | 142.4, C |
7′′′′ | 41.9, CH2 | 41.8, CH2 | 41.7, CH2 | 41.8, CH2 | 41.8, CH2 | 41.8, CH2 | 41.7, CH2 |
8′′′′ | 71.8, CH | 71.9, CH | 71.9, CH | 71.9, CH | 71.9, CH | 71.9, CH | 71.9, CH |
9′′′′ | 20.0, CH3 | 19.5, CH3 | 20.1, CH3 | 19.5, CH3 | 19.5, CH3 | 19.5, CH3 | 20.2, CH3 |
10′′′′ | 172.9, C | 172.9, C | |||||
MeO | 55.8, CH3 | 55.8, CH3 |
No. | 8 | 9 | 10 | 11 | 12 | 13 |
---|---|---|---|---|---|---|
3 | 6.23, d (1.5) | 6.22, d (1.5) | 6.18, d (1.5) | 6.23, d (1.7) | 6.23, d (1.5) | 6.05, s |
1/5 | 6.12, d (1.5) | 6.13, d (1.5) | 6.15, d (1.5) | 6.14, d (1.7) | 6.10, d (1.5) | 6.05, s |
7 | 2.53, dd (8.0, 13.5) | |||||
2.66, dd (6.5, 13.5) | ||||||
8 | 3.77, d (17.5) | 3.75, d (17.5) | 2.74, dd (8.4, 13.5) | 3.77, d (17.5) | 3.91, d (18.8) | 4.91, ddq (6.2, 6.5, 8.0) |
4.02, d (17.5) | 3.90, d (17.5) | 2.88, dd (4.0, 13.5) | 3.91, d (17.5) | 4.37, d (18.8) | ||
9 | 4.99, ddq (4.0, 6.0, 8.4) | 1.13, d (6.2) | ||||
10 | 2.75, dd (7.0, 17.0) | 2.38, dd (5.2, 15.5) | 1.15, d (6.0) | 2.40, dd (5.2, 15.5) | 2.52, dd (2.0, 13.0) | |
2.84, dd (6.7, 17.0) | 2.54, dd (7.3, 15.5) | 2.54, dd (7.3, 15.5) | 2.83, dd (11.5, 13.0) | |||
11 | 5.19, ddq (6.2, 6.7, 7.0) | 4.01, ddq (5.2, 6.0, 7.3) | 4.01, ddq (5.2, 6.0, 7.3) | 5.29, ddq (2.0, 6.1, 11.5) | ||
12 | 1.18, d (6.2) | 1.07, d (6.0) | 1.07, d (6.0) | 1.29, d (6.1) | ||
2′ | 2.72, dd (5.7, 16.5) | 2.63, d (7.0) | 2.67, d (6.4) | 2.64, d (6.7) | 2.63, dd (9.6, 12.6) | 2.25, dd (6.4, 14.3) |
2.78, dd (7.0, 16.5) | 2.70, dd (2.0, 12.6) | 2.34, dd (7.0, 14.3) | ||||
3′ | 5.34, ddq (5.7, 6.2, 7.0) | 5.34, tq (6.2, 7.0) | 5.37, tq (6.1, 6.4) | 5.32, tq (6.2, 6.7) | 5.19, ddq (2.0, 6.1, 9.6) | 3.94, ddq (6.2, 6.4, 7.0) |
4′ | 1.33, d (6.2) | 1.31, d (6.2) | 1.28, d (6.1) | 1.20, d (6.2) | 1.34, d (6.1) | 1.02, d (6.2) |
1′′ | 6.05, s | 6.05, s | ||||
3′′ | 6.15, d (1.5) | 6.18, d (1.5) | 6.05, s | 6.05, s | ||
5′′ | 6.18, d (1.5) | 6.16, d (1.5) | 6.05, s | 6.05, s | ||
7′′ | 2.53, m; 2.67, m | 2.52, dd (7.2, 13.5) | ||||
2.66, dd (6.6, 13.5) | ||||||
8′′ | 2.73, dd (5.0, 13.5) | 2.77, dd (8.7, 13.2) | 4.93, m | 4.93, ddq (6.0, 6.6, 7.2) | ||
3.00, dd (8.8, 13.5) | 2.91, dd (4.6, 13.2) | |||||
9′′ | 4.85, ddq (5.0, 6.1, 8.8) | 5.00, dd1 (4.6, 6.0, 8.7) | 1.10, d (6.0) | 1.07, d (6.0) | ||
10′′ | 1.15, d (6.1) | 1.11, d (6.0) | ||||
2′′′ | 2.51, dd (7.0, 15.5) | 2.58 d (6.0) | 2.18, dd (6.7, 14.2) | |||
2.55, dd (6.5, 15.5) | 2.30, dd (6.5, 14.2) | |||||
3′′′ | 5.22, ddq (6.2, 6.5, 7.0) | 5.29, tq (6.0, 6.2) | 3.90, m | |||
4′′′ | 1.24, d (6.2) | 1.15, d (6.2) | 0.97, d (6.0) | |||
OH-2 | 10.77, s | 10.96, s | 10.62, s | 10.94, s | 11.08, s | 9.11, s |
OH-4 | 10.17, s | 10.22, s | 10.01, s | 10.21, s | 10.23, s | 9.11, s |
OH-2′′ | 10.14, s | 10.54, s | 9.11, s | 9.10, s | ||
OH-4′′ | 9.83, s | 9.98, s | 9.11, s | 9.10, s | ||
OH-11 | 4.59, brs | 4.60, brs | ||||
COOH | 12.44, brs |
No. | 8 | 9 | 10 | 11 | 12 | 13 |
---|---|---|---|---|---|---|
1 | 107.4, C | 106.7, C | 108.6, C | 106.9, C | 106.8, C | 107.6, CH |
2 | 162.4, C | 162.9, C | 161.2, C | 162.9, C | 163.2, C | 158.6, C |
3 | 102.1, CH | 102.0, CH | 101.6, CH | 102.0, CH | 102.0, CH | 101.0, CH |
4 | 161.9, C | 162.1, C | 161.1, C | 162.1, C | 162.2, C | 158.6, C |
5 | 112.3, CH | 112.7, CH | 111.1, CH | 112.5, CH | 112.5, CH | 107.6, CH |
6 | 138.4, C | 139.1, C | 140.5, C | 139.0, C | 138.0, C | 139.8, C |
7 | 169.1, C | 169.2, C | 169.0, C | 169.2, C | 170.1, C | 41.9, CH2 |
8 | 49.4, CH2 | 50.7, CH2 | 40.8, CH2 | 50.7, CH2 | 51.9, CH2 | 71.1, CH |
9 | 204.3, C | 206.4, C | 70.8, CH | 206.5, C | 205.8, C | 19.7, CH3 |
10 | 47.5, CH2 | 51.4, CH2 | 20.2, CH3 | 51.5, CH2 | 48.3, CH2 | |
11 | 67.1, CH | 63.2, CH | 63.2, CH | 69.8, CH | ||
12 | 20.2, CH3 | 24.2, CH3 | 24.2, CH3 | 20.6, CH3 | ||
1′ | 169.9, C | 170.2, C | 170.7, C | 169.7, C | 170.1, C | 170.9, C |
2′ | 40.1, CH2 | 40.4, CH2 | 44.7, CH2 | 40.5, CH2 | 40.8, CH2 | 44.8, CH2 |
3′ | 68.3, CH | 68.8, CH | 63.7, CH | 68.7, CH | 71.1, CH | 63.8, CH |
4′ | 19.9, CH3 | 19.9, CH3 | 23.4, CH3 | 19.6, CH3 | 20.3, CH3 | 23.6, CH3 |
1′′ | 110.7, C | 108.8, C | 107.6, CH | 107.6, CH | ||
2′′ | 159.4, C | 162.0, C | 158.6, CH | 158.7, C | ||
3′′ | 101.6, CH | 101.7, CH | 101.1, CH | 101.1, CH | ||
4′′ | 160.4, C | 161.1, C | 158.6, C | 158.7, C | ||
5′′ | 109.9, CH | 110.9, CH | 107.6, CH | 107.6, CH | ||
6′′ | 139.6, C | 140.2, C | 139.7, C | 139.7, CH | ||
7′′ | 168.5, C | 168.9, C | 41.8, CH2 | 41.8, CH2 | ||
8′′ | 39.9, CH2 | 40.6, CH2 | 71.9, CH | 71.9, CH | ||
9′′ | 72.8, CH | 71.7, CH | 19.5, CH3 | 19.5, CH3 | ||
10′′ | 20.3, CH3 | 20.0, CH3 | ||||
1′′′ | 169.4, C | 169.5, C | 169.7, C | |||
2′′′ | 40.3, CH2 | 40.5, CH2 | 40.7, CH2 | |||
3′′′ | 68.6, CH | 68.6, CH | 68.5, CH | |||
4′′′ | 19.5, CH3 | 19.5, CH3 | 19.7, CH3 |
Alkaline hydrolysis of compound 1 in NaOH derived four components (Fig. 3), which were purified and identified as orsellinic acid (1a),14 3-hydroxybutyric acid (1b), 6-hydroxymellein (1c), and orcinotriol (1d). The specific rotation of 1b ([α]25D −43.0°, MeOH) was comparable to that for 3R-hydroxybutyric acid ([α]25D −48.5°, MeOH),4 which was co-isolated from the same fraction. The specific rotation of 1c ([α]25D −45.6°, MeOH) was in agreement with 6R-hydroxymellein ([α]25D −51°, MeOH).15 It is noted that the cyclization of the sodium 2,4-dihydroxy-6-(2-hydroxy-n-propyl)benzoate to the corresponding lactone 1c took place spontaneously during the acidic workup. The derivative 1d was identified to R configuration due to the similar specific rotation between 1d ([α]25D −16.8, MeOH) and the authentic sample for (R)-orcinotriol ([α]25D −22.4, MeOH), which was in contrast to that of (S)-orcinotriol ([α]25D +6.0, MeOH).16 This assignment was also supported by the data of the chemical shift differences for the (R)-MPA ester and (S)-MPA ester of 1d (Fig. 4).
Hansforester B (2) has a molecular formula of C39H44O15 as determined by the HRESIMS data, requiring 18 degrees of unsaturation. Its NMR data (Tables 2 and 3) resembled those of 1, while the 2D NMR data (Fig. 2) established the substructure from unit B to unit E to be identical to those of 1. The significant difference was attributed to the substituent in unit A, in which a (E)-pent-3-en-2-one moiety to replace a methyl group of 1 was recognized by the COSY relationships from the olefinic proton H-11 (δH 6.86, dq, J = 6.8, 15.6 Hz) to H-10 (δH 6.12, dd, J = 1.2, 15.6 Hz) and H3-12 (δH 1.88, dd, J = 1.2, 6.8 Hz), in association with the HMBC correlation from a ketone carbon C-9 (δC 196.6) to H-10, H-11 and H2-8 (δH 3.81, 4.07, d, J = 17.5 Hz). The substitution of this moiety at C-6 (δC 139.7) was evident from the HMBC correlations from H2-8 to C-5 (δC 112.9), C-6, and C-1 (δC 105.9) and from H-5 (δH 6.14, d, J = 2.2 Hz) to C-8 (δC 47.8) (Fig. 2).
Following the same protocol as that of 1, alkaline hydrolysis of 2 in NaOH also derived four components, which were isolated by semi-preparative HPLC chromatography and identified by the comparison of their NMR and MS data as well as the specific rotation with those of authentic samples. Apart from 2,;4-dihydroxy-6-(2-oxo-3-pentenyl)benzoic acid, the remaining derivatives were identical to 3R-hydroxybutyric acid (1b), 6R-hydroxymellein (1c), and (R)-orcinotriol (1d). Thus, the absolute configurations in the stereogenic centers of 2 were the same as those of 1.
The HRESIMS and NMR data provided the molecular formula of hansforester C (3) to be C40H44O17, bearing 19 degrees of unsaturation and with a CO2 unit more than that of 2. Analyses of 1D and 2D NMR data (Fig. 2) revealed the structure of 3 to be mostly identical to that of 2. The distinction was attributed to unit E, where a orcinotriol moiety of 2 was replaced by a 6-(2-hydroxypropyl)-2,4-dihydroxybenzoic moiety. This assignment was ascertained by the presence of two meta-coupling aromatic protons H-3′′′′ (δH 6.15, d, J = 3.3 Hz) and H-5′′′′ (δH 6.17, d, J = 3.3 Hz) and a carboxylic resonance at δC 172.9 (C-10′′′′) but the absence of H-1′′′′. The 4J long range HMBC correlations of both H-3′′′′ and H-5′′′′ to the carboxylic carbon confirmed its location at C-1′′′′ (δC 105.7). The absolute configuration of 3 was identified as R for C-3′, C-9′′, C-3′′′, and C-8′′′′, based on the alkaline hydrolyzates of 3 to be identical to those derived from 2, including 3R-hydroxybutyric acid (1b) and 6R-hydroxymellein (1c) as evidenced by the comparison of their NMR data and the specific rotation.
Hansforester D (4) possessing a molecular formula of C37H42O15 was determined by the HRESIMS data, containing 17 degrees of unsaturation. The NMR data (Tables 2 and 3) speculated that 4 is an analogue of 1 with a different substituent at C-6 in unit A. The presence of a methyl singlet at δH 2.05 (s, H3-10) and the methylene protons at δH 3.72 (d, J = 17.5 Hz, H-8a) and 3.87 (d, J = 17.5 Hz, H-8b) in the 1H NMR spectrum, in association with the HMBC correlation of a ketone carbon C-9 (δC 205.4) with H3-10 and H2-8 clarified an acetonyl group, which was located at C-6 of unit A to replace a methyl group of 1, accordingly to the HMBC correlations of H2-8 to C-1 (δC 106.4), C-5 (δC 112.6), and C-6 (δC 139.3) in addition to the correlation between H-5 (δH 6.14, d, J = 2.2 Hz) and C-8 (δC 50.7). The absolute configurations of 4 were the same as those of 2 based on the derivatives generated by the alkaline hydrolysis to be identical to those derived from 2.
The molecular formula of hansforester E (5) was assigned to C39H46O16 by the HRESIMS data. Comparison of the NMR data indicated the structure of 5 to be closely related to 2. The distinction was attributed to the side chain in unit A, where five carbon resonances including two methylenes (δC 50.7, C-8) and (δC 51.4, C-10), a ketone (δC 206.4, C-9), a hydroxymethine (δC 63.2, C-11), and a methyl carbon (δC 24.1, C-12) were observed in the DEPT spectrum. The COSY relationships from H-11 (δH 4.02, ddq, J = 5.3, 6.3, 7.4 Hz) to H3-12 (δH 1.07, d, J = 6.3 Hz) and H2-10 (δH 2.39, 2.52) along with the HMBC correlations from the ketone carbon C-9 to H2-8 (δH 3.75, 3.90, d, J = 17.4 Hz), H2-10, and H-11, confirmed the presence of a 4-hydroxypentane-2-one moiety, which was located at C-6 (δC 139.1) on the basis of the HMBC correlation of H2-8 with C-1 (δC 106.7), C-5 (δC 112.6), and C-6.
Alkaline hydrolysis of 5 afforded four components, and three of them were identical to 3R-hydroxybutyric acid (1b), 6R-hydroxymellein (1c), and (R)-orcinotriol (1d) by the comparison of the NMR data and specific rotation with the authentic samples. In addition, 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic acid was isolated, which was cyclized by pTSA (para-toluene sulfonamide) in benzene to yield orthosporin (Fig. 3). The derived orthosporin (1e) showed the specific rotation as [α]25D −11.8 (c 0.15, MeOH), that was in agreement with R configuration in comparison with the data reported in literature.1 This was also supported by the natural (S)-isomer with opposite phase of specific rotation ([α]D +22).17 Thus, the absolute configurations in 5 were determined as R for all stereogenic centers.
Hansforester F (6) was identified as methoxylated analogue of 5 based on the similar NMR data between 5 and 6 (Tables 2 and 3) with the exception of the methoxy resonances (δH 3.17, s; δC 55.8) observed in the HMQC spectrum. The methoxy location at C-11 (δC 72.6) was confirmed by the HMBC correlation. Alkaline hydrolysis of 6 yielded four derivatives. Apart from 6R-hydroxymellein, 3R-hydroxybutyric acid, and (R)-orcinotriol which were identified by the comparison of the HPLC retention times and the specific rotation with the authentic samples, 2,4-dihydroxy-6-(4′-methoxy-2′-oxopentyl)benzoic acid were isolated. Treatment of this derivative by pTSA in benzene to yield a methylated orthosporin, whose specific rotation ([α]25D −18, MeOH) was comparable to that of R-orthosporin ([α]25D −11.8, MeOH). This finding assumed C-11 to be R configuration.
Hansforester G (7) has a molecular formula of C41H48O18 as determined by the HRESIMS data, bearing a CO2 unit more than that of 6. Analyses of the 1D and 2D NMR data established the partial structure regarding units A–D to be identical to that of 6. The difference was found in unit E, where two aromatic protons with meta-coupling including H-3′′′′ (δH 6.17, d, J = 2.0 Hz) and H-5′′′′ (δH 6.14, d, J = 2.0 Hz) were observed in the COSY spectrum. Apart from the phenol protons OH-2′′′′ (δH 11.89, brs) and OH-4′′′′ (δH 10.17, brs) in unit E, the observation of four-bond HMBC correlations from H-3′′′′ and H-5′′′′ to the carboxylic carbon C-10′′′′ (δC 172.9) confirmed a carboxylic acid to be positioned at C-1′′′′ (δC 105.7). Alkaline hydrolysis of 7 yielded the derivatives, which were identical to 6R-hydroxymellein, 3R-hydroxybutyric acid, and methoxylated R-orthosporin.
Hansforester H (8) has a molecular formula of C30H34O13 according to the HRESIMS data, requiring 14 degrees of unsaturation. The 1D and 2D NMR data established four building blocks (Fig. 2), including a 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic moiety (unit A), two 3-hydroxybutanoic segments (units B and D), and a 6-(2-hydroxypropyl)-2,4- dihydroxybenzoic moiety (unit C). The segment connection was accomplished by the HMBC correlations between H-11 (δH 5.19, ddq, J = 6.2, 6.7, 7.0 Hz)/C-1′ (δC 169.9), H-3′ (δH 5.34, ddq)/C-7′′ (δC 168.5), H-9′′ (δH 4.85, ddq)/C-1′′′ (δC 169.4), and H-3′′′ (δH 5.22, ddq)/C-7 (δC 169.1). Thus, the structure of 8 was established as a cyclic polyester. Alkaline hydrolysis of 8 derived three components, two of which was identical to 6R-hydroxymellein and 3R-hydroxybutyric acid based on the comparison of their HPLC retention times and the specific rotation with those of the authentic samples. 2,4-Dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic acid was cyclized by pTSA in benzene to yield orthosporin, which showed the specific rotation as [α]25D −11.5 (c 0.05, MeOH), that was in agreement with R configuration in comparison with the data reported in literature.1 Thus, the absolute configurations in 8 were determined as R for all stereogenic centers.
The molecular formula of hansforester I (9) was determined to be C30H36O14 by the HRESIMS data, containing 13 degrees of unsaturation. The 1D and 2D NMR data established the structure to be closely related to that of 8 with the same building blocks. The distinction was attributed to units A and B, while the shielded H-11′′ (δH 4.01, m) showed the COSY correlation with a OH proton at δH 4.59 (br), indicating the presence of an alcohol group. In addition, a proton for a carboxylic acid at δH 12.44 (brs) was observed in the 1H NMR spectrum. These findings in association with the absence of the HMBC correlation between H-11 and C-1′ (δC 170.2) and one site of unsaturation less than that of 8 indicated that 9 was a hydrolyzed product of 8 by the cleavage of the ester bond between C-11 (δC 63.2) and C-1′. Alkaline hydrolysis of 9 yielded the derivatives which were identical to those of 8, supported both 8 and 9 followed the same biogenetic pathway.
Hansforester J (10) has a molecular formula of C27H34O11 as established by the HRESIMS data (m/z 557.1990, calcd for [M + Na]+, 557.1993). Analyses of the 2D NMR data enabled the assignment of four segments, including a 2,4-dihydroxy-6-(2′-hydroxypropyl)benzoic residue (unit A), two 3-hydroxybutanoic units (units B and D), and an orcinotriol moiety (unit C). The key HMBC correlation between H-9 (δH 4.99, ddq) of unit A and C-1′′′ (δC 169.7) of unit D connected units A and D by an ester bond. The linkage of unit A with unit B by esterification was evident from the HMBC correlation between H-3′ (δH 5.37, tq) and C-7 (δC 169.0). In addition, an ester bond formed between unit B and unit C (orcinotriol moiety) was established by the HMBC correlation between H-8′′ (δH 4.93, m) and C-1′ (δC 170.7). The absolute configuration of the stereogenic centers in 10 was assigned to R based on the hydrolyzed products to be identical to those of authentic samples by the comparison of the HPLC retention time and the specific rotation.
Analyses of the 1D and 2D NMR data established hansforester K (11) to possess three segments, including a 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoate (unit A), a 3-hydroxybutyrate (unit B), and an orcinotriol moiety (unit C) (Fig. 2). The connection of units A and B by an ester bond was deduced by the HMBC correlation between H-3′ (δH 5.32, tq) and C-7 (δC 169.2), while an additional ester bond formed between units B and C was deduced by the HMBC correlation of H-8′′ (δH 4.93, ddq) with C-1′ (δC 169.7). Based on the biogenetic consideration and referring to the specific rotation, the absolute configurations of 11 were assumed to be R.
Hansforester L (12) has a molecular formula of C16H18O7 as determined by the HRESIMS data (m/z 345.0947 [M + Na]+, calcd for C16H18O7Na, 345.0944). The 2D NMR data enabled to establish two partial structures, involving a 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoate (unit A) and a 3-hydroxybutyrate (unit B). The connection of units A and B by ester bonds to form a cyclic polyester was evident from the HMBC correlations between H-11 (δH 5.29, ddq)/C-1′ (δC 170.1) and H-3′ (δH 5.19, ddq)/C-7 (δC 170.1). The R configuration for C-10 and C-3′ was assumed on the basis of the biogenetic consideration that 12 is speculated to be generated from 11.
The molecular formula of hansforester M (13) was determined as C13H18O5 on the basis of the HRESIMS data (m/z 277.1047 [M + Na]+, calcd for C13H18O5Na, 277.1046). The 1D and 2D NMR data revealed the presence of two residues, including an orcinotriol moiety (unit A) and a 3-hydroxybutyrate. The linkage between the two moieties to form an ester bond was readily assigned by key HMBC correlation from H-8 (δH 4.91, ddq) to C-1′ (δC 170.9). The absolute configuration was assumed to be 8R, 1′R based on the biogenetic consideration.
In addition, three known analogues were identical to ascotrichalactone A (14),1 ascotrichester B (15),1 LL15G256ν (16),3 6R-hydroxymellein (17)12 and (−)orthosporin (18) (Fig. 5),13 based on the comparison of their NMR and HRESIMS data in addition to the specific rotation with these reported in the literature.
Although the biosynthesis of the polyesters is rarely investigated, the biosynthetic pathway of the building blocks is well understood. A ketothiolase catalyzes acetyl-CoA to acetoacetyl-CoA, which was catalyzed by a reductase to derive R-3-hydroxybutyryl acid.18 6-Hydroxymellein is produced via the acetate-malonate pathway,19 while orthosporin is produced by polyketide biosynthetic gene cluster.20 A sequence of enzymatic reactions as induced by synthase led to the formation of polymers through ester bond.21 The synthase as a Ziegler–Natta catalyst is specific for monomers with the R configuration and will not polymerize identical compounds having the S configuration.22 In the isolated polyesters, the building blocks involved 3-hydroxybutyrate, 6-hydroxymellein, orthosporin, orthosporinin, orcinotriol, and 4,6-dihydroxy-2-methylbenzoic acid. Each motif presents as thioester with coenzyme A (CoA), while the conversion of thioester to oxyester was induced by synthase. As shown in Scheme 1, there are two manners to connect 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic acid with 3-hydroxybutyryl acid. Esterification of the carbonyl group of 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic acid or other motifs with the free hydroxyl group of hydroxybutyryl acid and followed the successive steps yielded compound 5, whereas the esterification of the free hydroxyl group in 2,4-dihydroxy-6-(4′-hydroxy-2′-oxopentyl)benzoic acid with the carbonyl group of hydroxybutyryl acid by the similar way as for 5 derived 8. Compounds 1–4 share the same sequence as that of 5 with different precursors or terminal motif. The methylation in analogues 6 and 7 were assumed after the formation of full sequence. Compound 9 was suggested to be derived from 8 by hydrolysis. The biogenetic pathway of the remaining compounds was suggested to follow the similar manner as for 5 and 8.
Compd. | B. subtilis | B. thuringiensis | S. aureus | E. coli | P. lachrymans | A. tumefaciens | X. vesicatoria | R. solanacearum |
---|---|---|---|---|---|---|---|---|
a CP: chloramphenicol, positive control. | ||||||||
1 | 15.62 | 15.62 | 3.90 | >250 | 15.62 | 15.62 | 15.62 | 15.62 |
2 | 125 | 125 | 62.5 | >250 | 125 | 125 | 125 | 125 |
3 | 250 | >250 | >250 | >250 | >250 | >250 | >250 | >250 |
4 | 125 | 62.5 | 31.25 | >250 | 62.5 | 62.5 | 62.5 | 62.5 |
5 | 125 | 125 | 62.5 | >250 | 125 | 125 | 125 | 125 |
6 | 250 | 250 | 125 | >250 | 250 | 250 | >250 | 250 |
7 | >250 | >250 | 125 | >250 | >250 | >250 | >250 | >250 |
14 | 15.62 | 15.62 | 3.90 | >250 | 15.62 | 15.62 | 15.62 | 15.62 |
CP | 3.09 | 6.19 | 3.09 | 12.38 | 12.38 | 12.38 | 12.38 | 12.38 |
Pathogenic P. lachrymans causes angular leaf spot, a common cucumber disease, resulting in significant yield reduction.23 Bacterium A. tumefaciens is a serious pathogen of many economic plants such as walnuts and grape vines, making it of great concern to the agriculture industry.24 Bacterium X. vesicatoria is a Gram-negative and it causes bacterial leaf spot on peppers and tomatoes,25 while R. solanacearum is also a Gram-negative and plant pathogenic bacterium causing bacterial wilt in a very wide range of potential host plants.26–29 Plant diseases cause major economic losses for farmers worldwide, while control of plant diseases is crucial to the reliable production of food. Natural products is a rich source for the discovery of promising leads to overcome the serious agriculture problem.
Footnote |
† Electronic supplementary information (ESI) available. See DOI: 10.1039/c8ra08606k |
This journal is © The Royal Society of Chemistry 2018 |