Total Synthesis, structure revision and antifungal activity of palmarumycin B8 and B7.

Palmarymycins B8 (1), its regioisomer (2) and B7 (3) were synthesized via 10-, 9-, and 11-steps in 6.5 %, 2.3 % and 0.54 % overall yields from chroman-4-one (4), 4-hydroxyindanone (12), and 2,5-dimethoxybenzaldehyde (20) as the starting materials, using benzyl protection, enol trimethylsilyl ether by TMSOTf, Rubottom oxidation and deprotection with hydrogenation under Pd/C catalyst as the key steps, respectively. Their structures were characterized by 1H, 13C NMR, COSY, HSQC, HMBC and HR-ESI-MS spectral data. The structure of palmarumycin B8 was revised from 1 to 2 based on the total synthesis, 2D NMR analysis and DFT calculation. The antifungal assay results indicated that palmarumycin B8 (1) showed moderate inhibitory activity against Phytophthora capsica. Compounds 15 and 16 exhibited excellent in vitro antifungal activities against P. capsica with EC50 values of 2.17 and 8.50 µg/mL, respectively.

Elucidation of Palmarumycin Spirobisnaphthalene Biosynthesis Reveals a Set of Previously Unrecognized Oxidases and Reductases.

Spirobisnaphthalenes (SBNs) are a class of highly oxygenated, fungal bisnaphthalenes containing a unique spiroketal bridge, that displayed diverse bioactivities. Among the reported SBNs, palmarumycins are the major type, which are precursors for the other type of SBNs structurally. However, the biosynthesis of SBNs is unclear. In this study, we elucidated the biosynthesis of palmarumycins, using gene disruption, heterologous expression, and substrate feeding experiments. The biosynthetic gene cluster for palmarumycins was identified to be distant from the polyketide synthase gene cluster, and included two cytochrome P450s (PalA and PalB), and one short chain dehydrogenase/reductase (PalC) encoding genes as key structural genes. PalA is an unusual, multifunctional P450 that catalyzes the oxidative dimerization of 1,8-dihydroxynaphthalene to generate the spiroketal linkage and 2,3-epoxy group. Chemical synthesis of key intermediate and in vitro biochemical assays proved that the oxidative dimerization proceeded via a binaphthyl ether. PalB installs the C-5 hydroxy group, widely found in SBNs. PalC catalyzes 1-keto reduction, the reverse 1-dehydrogenation, and 2,3-epoxide reduction. Moreover, an FAD-dependent oxidoreductase, encoded by palD, which locates outside the cluster, functions as a 1-dehydrogenase. These results provided the first genetic and biochemical evidence for the biosynthesis of palmarumycin SBNs.

Design, synthesis, antifungal activity and molecular docking of novel pyrazole-4-carboxamides containing tertiary alcohol and difluoromethyl moiety as potential succinate dehydrogenase inhibitors.

BACKGROUND: Resistance problems with the long-term and frequent use of existing fungicides, and the lack of structure diversity of traditional pyrazole-4-carboxamide succinate dehydrogenase inhibitors, it is highly required to design and develop new fungicides to address the resistance issue. RESULTS: Different from previous pyrazole-4-carboxamide succinate dehydrogenase inhibitors by breaking the norm of difluoromethyl at the C-3 position of pyrazole and introducing a tertiary alcohol group at the C-3 position, 27 novel pyrazole-4-carboxamide derivatives were designed, synthesized and characterized by proton (1 H) nuclear magnetic resonance (NMR), carbon-13 (13 C) NMR, fluorine-19 (19 F) NMR and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). The crystal structures of compounds A14 and C5 were analyzed by single crystal X-ray diffraction. Their in vitro antifungal activities were evaluated against phytopathogen Fusarium graminearum, Botrytis cinerea, Phytophthora capsica, Sclerotinia sclerotiorum, Thanatephorus cucumeris. The results displayed that most of them exhibited significant antifungal activities against S. sclerotiorum at 50 mg/L, the half maximal effective concentration (EC50 ) data of A8 and A14 were 3.96 and 2.52 mg/L, respectively. Their in vivo antifungal activities were evaluated against Pseudoperonospora cubensis, Puccinia sorghi Schw, Colletotrichum gloeosporioides, F. graminearum, Erysiphe graminis, Thanatephorus cucumeris, the control efficacies of A6, B3, C3, and C6 against E. graminis reached 100% at a concentration of 400 mg/L. The molecular docking results showed that the binding mode of the target compounds containing tertiary alcohols were similar to that of fluxapyroxad in succinate dehydrogenase. In addition, tertiary alcohols were involved in the formation of hydrogen bonds. CONCLUSION: The excellent in vitro and in vivo inhibitory activities of novel pyrazole-4-carboxamide derivatives against succinate dehydrogenase were reported for the first time, and they could be used as the potential lead compounds. © 2023 Society of Chemical Industry.