Gf254

All of the reagents were obtained from commercial sources and used without further purification unless stated otherwise. Thin-layer chromatography (TLC) on the silica gel plates GF254 (200–300 mm; Qingdao Haiyang Chemical Co., Ltd., Qingdao, China) with UV light illumination was used to monitor chemical reactions. ¹H NMR (500 MHz) and ¹³C NMR (100 MHz) spectra were measured by Avance spectrometer (Bruker, Varian Mercury, Billerica, MA). Chemical shifts were reported in δ values (ppm) with tetramethylsilane as the internal standard. High-resolution mass spectrometry (HRMS) was performed using the Thermo Scientific^™ Exactive^™ Plus mass spectrometer (Thermo, Waltham, MA). The melting points were recorded with a Mettler Toledo melting point apparatus. The purity of all the target compounds was determined by high-performance liquid chromatography (HPLC) on a Waters Acquity machine with a BEH C18 column (1.7 µm, 50 × 2.1 mm); mobile phase A = water (containing 0.1% formic acid) and mobile phase B = acetonitrile; the flow rate was 0.25 mL/min.

The ¹H NMR and ¹³C NMR spectra were acquired at 25 °C in a nuclear magnetic resonance spectrophotometer (DRX—400, Bruker, Germany) at 400 MHz for ¹H NMR in DMSO-d₆ and 100 MHz for ¹³C NMR in DMSO-d₆. Thin-layer chromatography (TLC) was performed on silica gel (GF254, Qingdao Haiyang Chemical Co., Ltd., Qingdao, China) with a visualization of the spots under UV light at 254 nm. Fluorescence spectra used a fluorescence spectrophotometer (F-2710, Hitachi, Hiroshima-shi, Japan). Absorption measurements were conducted using a UV–Vis spectrophotometer (UH5300, Hitachi, Japan) within a 190–1100 nm wavelength range.

Optical rotations were measured using an Autopol VI polarimeter. Ultraviolet absorption spectra were recorded on a UV-2401 PC spectrophotometer. ECD spectra were recorded on a Chirascan-plus spectrometer (Applied photophysics Ltd., Surrey, United Kingdom). IR spectra were obtained from a Perkin-Elmer 577 spectrometer. NMR spectra were measured on a Bruker AV-600 or AV-400 spectrometer and calibrated by the solvent peak used. Mass spectrometry was performed on a SYNAPT G2-Si HDMS (Waters Corp., Manchester, UK) with an electrospray ion source (Waters, Milford, MA) connected to a lock-mass apparatus, which performed real-time calibration correction. Column chromatography was performed with CHP20P MCI gel (75–150 μm, Mitsubishi Chemical Corporation, Japan), silica gel (100–200, or 200–300 mesh, Qingdao Haiyang Chemical Co., Ltd.), Sephadex LH-20 (GE Healthcare Bio-Sciences AB, Sweden), and reversed-phase C₁₈ silica gel (50 μm, YMC, Kyoto, Japan). Precoated TLC sheets of silica gel 60 GF254 (Qingdao Haiyang Chemical Co., Ltd.) were used. A Waters 2535 Series machine equipped with a Xbridge C₁₈ column (4.6 × 250 mm, 5 μm) was used for HPLC analysis, and a preparative Xbridge Prep C₁₈ OBD column (19 × 250 mm, 5 μm) was used for the sample preparation.

Optical rotations were measured with an MCP-200 polarimeter. UV spectra were recorded on a Shimadzu spectrophotometer. 1D and 2D NMR spectra were acquired on Bruker ARX-600, 600-MHz spectrometers. Column chromatography (CC) was performed on silica gel (200–300 and 100–200 mesh, Qing-dao Haiyang Chemical Co., Ltd., Qingdao, China), RP-18 silica gel (20 × 45 mm, Merck, Japan), and Sephadex LH-20 gel (Pharmacia, Sweden). Fractions were monitored by TLC on silica gel plates (GF254, Qingdao Haiyang Chemical Co., Ltd., Qingdao, China). HPLC was performed using Waters 1,525 pumps coupled with analytical preparative YMC-C₁₈ columns (4.6 × 250 mm, 5 μm). The HPLC system employed a Waters 996 photodiode array detector.

Both 1D and 2D spectra were run on Bruker Avance III 600 MHz and Bruker Avance III 500 MHz spectrometers. IR spectra were obtained with a Tenor 27 spectrophotometer using KBr pellets. UV spectra were measured on a UH-5300 spectrometer. CD spectra were recorded by a Chirascan-plus circular dichromatic spectrometer. The optical rotations were measured with a Horiba Sepa-300 polarimeter. According to the solvent signal, the chemical shifts were expressed in ppm. The mass spectra were recorded on a mass spectrometer (Thermo Fisher Scientific, Bremen, Germany). Medium-pressure liquid chromatography (MPLC) was applied to the Biotage SP1 system and packed with RP-18 gel. Preparative high-performance liquid chromatography (prep-HPLC) was performed on an Agilent 1260 liquid chromatography system equipped with Zorbax SB-C18 columns (5 μm, 9.4 mm × 150 mm, or 21.2 mm × 150 mm) and a DAD detector. Column chromatography (CC) separations were carried out in silica gel (200–300 mesh, Qingdao Haiyang Chemical Co., Ltd., Qingdao) and Sephadex LH-20 (Sweden’s France Asia Fine Chemical Co., Ltd.). The components were monitored by TLC (GF₂₅₄, Qingdao Haiyang Chemical Co., Ltd., Qingdao) and the spots were observed by heating the silica gel plate and spraying with vanillin and 10% H₂SO₄ in ethanol.

Phenazine-1-carboxylic acid (PCA) and Metalaxyl were provided by the College of Agriculture, Yangtze University. All chemicals and solvents were commercially available without further purification. All target compounds were further purified by column chromatography using silica gel (200–300 mesh, Liang Chen Gui Yuan Co., Ltd., Luan, China). Thin layer chromatography (TLC) analysis was performed on silica gel GF254 (Qingdao Hai Yang Chemical Co., Ltd., Qingdao, China). ¹H NMR and ¹³C NMR spectra were recorded on an AVANCE DPX400 nuclear magnetic resonance spectrometer (Bruker Spectroscopy, France, Germany) using trimethylchlorosilane (TMS) and DMSO-d₆ or CDCl₃ as the internal standard and the solvent, respectively. High-resolution mass spectra (HRMS) data were acquired on Thermo Scientific Q Exactive (Thermo Fisher Scientific, Waltham, MA, USA). The melting points of all target compounds were confirmed by using a WRR melting point apparatus (Shanghai Jingke Industrial Co., Ltd., Shanghai, China) and are uncorrected.

Optical rotations were measured with a Rudolph Autopol I automatic polarimeter (Rudolph, Hackettstown, NJ, USA). UV spectra were obtained on a Shimadzu UV-2401PC spectrophotometer (Shimadzu, Kyoto, Japan). IR spectra were measured with a Bruker TENSOR-27 spectrophotometer (Bruker, Bremerhaven, Germany) using KBr pellets. The 1D and 2D NMR spectra were recorded on JEOL ECX 500 MHz spectrometers (JEOL Ltd, Kyoto, Japan) with TMS as an internal standard. Chemical shifts (δ) were expressed in ppm with reference to solvent signals. High-Resolution Electrospray Ionization Mass Spectrometry (HR-ESI-MS) was recorded on a Bruker Daltonics micrOTOF-Q II spectrometer (Bruker, Bremerhaven, Germany). Column chromatography (CC) was performed on Silica gel (200–300 and 300–400 mesh, Qingdao Marine Chemical Ltd., Qingdao, China). Fractions were monitored by TLC (GF 254, Qingdao Haiyang Chemical Co., Ltd., Qingdao, China), and spots were visualized by Dragendorff’s reagent. Solvents were distilled prior to use for extraction and isolation.