Av 600 mhz spectrometer

Optical rotations were performed on a P-1020 polarimeter. IR spectra were measured on a Bruker FT-IR Tensor 27 spectrometer with KBr pellets. UV spectra were obtained on Shimadzu UV-2401A spectrometer. 1D and 2D-NMR spectra were recorded on Bruker AV-600 MHz spectrometer. Coupling constants were expressed in Hertz and chemical shifts were given on a ppm scale with tetramethylsilane as an internal standard. HRESIMS were recorded on an API QSTAR Pulsar 1 spectrometer. Column chromatography (CC) was performed on silica gel (200–300 mesh, Qingdao Marine Chemical Ltd., Qingdao, People’s Republic of China), Sephadex LH-20 (Pharmacia Fine Chemical Co., Ltd., Sweden), and MCI-gel CHP 20P (75–100 μm, Mitsubishi Chemical Co., Ltd). Thin-layer chromatography (TLC) was carried out on pre-coated silica gel plates (Qingdao Marine Chemical Co., Ltd.) with CHCl₃/MeOH (15:1, 4:1, v/v) as developing solvents and spots were visualized by Dragendorff’s reagent. High performance liquid chromatography (HPLC) was performed using Waters 600 pump with semi-preparative C₁₈ columns (150 × 9.4).

For isolation of compounds, the ethyl acetate extract of Actinomadura sp. ATCC 31491 grown on MSF agar was fractionated on a Sephadex LH-20 column with MeOH elution. After analysis with HPLC, the fractions containing metabolites of interest were combined, and the target compounds were purified from these fractions by reversed-phase semipreparative HPLC (YMC-Triart C18, 5 μm, 10 mm ID × 250 mm) (UV detection at 280 nm). Compounds 1 (40 mg, t_R = 33.0 min) and 2 (30.1 mg, t_R = 45.0 min) were purified with 52% (acetonitrile+ 0.05% FA)/(H₂O + 0.05% FA) at 4 mL/min. The ¹H- and ¹³C and 2D NMR spectra were recorded on a Bruker AV-600 MHz spectrometer using CDCl₃ as the solvent.

¹H-¹⁵N-HSQC spectra were recorded as SOFAST versions48 (link) at 35 °C on a Bruker AV 600 MHz spectrometer (Bruker, Karlsruhe, Germany) equipped with a cryo-probehead. TopSpin (version 3.1, Bruker) was used for data processing, including zero filling and linear prediction. The transfer of previous assignment13 (link) and general data evaluation were done using the CCPN software package (version 2.1.5)49 (link).
CSP was calculated using the following formula50 (link):

where ¹H and ¹⁵N refer to the mathematical difference of individual hydrogen and nitrogen chemical shifts of two distinct peak maxima. Gyromagnetic ratio (γ_i) of nuclei i, where i is ¹H or ¹⁵N, is used for normalization.

IR spectra were measured on a Bruker Tensor 27 FTIR spectrometer using KBr disks. NMR spectra were performed with a Bruker AV600 MHz spectrometer with TMS as an internal standard. HRMS–ESI data were obtained through an Agilent 1290 UPLC/6540 Q-TOF mass instrument. Column chromatography (CC) was performed using silica gel (30–400 mesh, Qingdao Marine Chemical Inc., China), Sephadex LH-20 (25–100 µm, Pharmacia Biotech Ltd., Sweden), and MCI gel (75–150 µm, Mitsubishi Chemical Corporation, Tokyo, Japan). Semipreparative HPLC was conducted on a HITACHI Chromaster system equipped with a DAD detector, a YMC-Triart C18 column (250 × 10 mm, i.d. 5 μm), and a flow rate of 3.0 mL/min. LC–MS was performed using an Agilent 1200 series HPLC system coupled to an Agilent q TOF 6540 mass spectrometer with a YMC-Triart C18 column (250 × 4.6 mm, i.d. 5 μm) and a flow rate of 1.0 mL/min.

All reactions were performed in atmosphere unless noted. All reagents were commercially available and use as supplied without further purification. NMR spectra were collected on either an Agilent DD2400 MHz spectrometer or a Bruker AV-600 MHz spectrometer with internal standard tetramethylsilane (TMS) and signals as internal references, and the chemical shifts (δ) were expressed in ppm. High-resolution Mass (ESI) spectra were obtained with Bruker Micro-TOF spectrometer. The Fourier transform infrared (FTIR) samples were prepared as thin films on KBr plates, and spectra were recorded on a Bruker Tensor 27 spectrometer and are reported in terms of frequency of absorption (cm⁻¹). X-ray data were collected on a Bruker Smart APEX-2 CCD diffractometer.

Melting points were measured on a YRT-3 melting point apparatus and were uncorrected. NMR spectra were recorded at 22 °C on a Bruker AV 600 MHz spectrometer, and chemical shifts (δ) are given in parts per million (ppm). The ¹H and ¹³C chemical shift were referenced to the CDCl₃ solvent peaks at δ_H 7.26 and δ_C 77.0 ppm, respectively. Electron ionization mass spectra (EI-MS) were determined by Waters micromass ZQ-2000 mass spectrometer, and HRMS were acquired on an Angilent 1290–6540B UPLC-Q-TOF mass spectrometer, respectively. Flash column chromatography was performed with silica gel (200–300 mesh) purchased from Qingdao Haiyang Chemical Co. Ltd. All chemical reagents were obtained from Aldrich Chemical Co. and treated with standard methods before use.