Shim pack velox c18 column

The PEAME was analyzed by UPLC-PDA-MS/MS using a previously reported methodology [19 (link),20 (link)]. Liquid chromatography system of A Nexera-i LC-2040 (Shimadzu, Kyoto, Japan) connected to a UPLC Shim-pack Velox C18 Column, 2.1 × 50 mm; 2.7 μm particle, was used. The solvent gradient and sample preparation were as reported [19 (link),20 (link)]. An LC-2030/2040 PDA detector and an LC-MS 8045 triple quadrupole mass spectrometer equipped with electrospray ionization (ESI) source in negative mode (Shimadzu, Kyoto, Japan) were used for detection of compounds using the reported settings [19 (link),20 (link)].

Infected (race 1 and 3) and non-infected common bean leaf extracts were subjected to analysis on a liquid chromatography-quadrupole time-of flights tandem mass spectrometry instrument (LCMS-9030 qTOF, Shimadzu Corporation, Kyoto, Japan) for quantification of metabolites at different time intervals. A Shim-pack Velox C18 column (100 mm × 2.1 mm with a 2.7 µm particle size) was used for chromatographic separation at 55°C (Shimadzu Corporation, Kyoto, Japan). An injection volume of 3 µL was used for all samples and were run on a binary mobile phase including solvent A: 0.1% formic acid in Milli-Q HPLC grade water (Merck, Darmstadt, Germany) and solvent B: UHPLC grade methanol with 0.1% formic acid (Romil Ltd., Cambridge, United Kingdom). Chromatographic analysis was done using qTOF high-definition mass spectrometer that was set to negative electrospray ionisation for data acquisition. Parameters set included nebulization, interface voltage (4.0 kV), interface temperature (300°C), dry gas flow (3 L/min), detector voltage (1.8 kV), heat block (400°C), DL (280°C) and flight tube (42°C) temperatures. Ion fragmentation was achieved using argon gas for collision with an energy of 30 eV and 5 eV spread (Ramabulana et al., 2021 (link)).

An ultra-high-performance liquid chromatography-mass spectrometry (UPLC-MS) analysis of metabolites was performed on an AB SCIEX LC-30A-Triple TOF 5600+ ESI MS system (AB SCIEX, Boston, MA, USA) equipped with a Shim-Pack Velox C18 column (2.7 μm, 150 mm × 2.1 mm, SHIMADZU, Japan), and a Thermo QE Focus instrument (Thermo Fisher Scientific, Waltham, MA, USA) with an Accucore AQ C18 column (2.6 μm, 150 mm × 2.1 mm, Thermo Fisher Scientific, Waltham, MA, USA). High-resolution electrospray ionization mass spectrometry (HRESIMS) data of novel metabolites were acquired by a Thermo Scientific Q Exactive mass spectrometer. Analytical high-performance liquid chromatography (HPLC) and semipreparative HPLC were carried out on an Agilent 1100 series system (Agilent Technologies Inc., Santa Clara, CA, USA) with a YMC-Pack ODS-A column (5 μm, 250 mm × 4.6 mm; 5 μm, 250 mm × 10 mm, YMC CO., LTD., Japan). Medium-pressure liquid chromatography (MPLC) was performed on a Buchi Medium pressure preparative chromatography (BUCHI Labor Technik AG, Switzerland) equipped with a medium-pressure column containing YMC reversed-phase (RP)-C₁₈ silica gel. Gel chromatography was carried out on a gel column with Sephadex LH-20 (40–70 μm; Amersham Pharmacia Biotech AB, Sweden). ¹H, ¹³C, and 2D NMR data were acquired on a Bruker Avance Neo 400 MHz apparatus (Bruker Corporation, Faellanden, Switzerland).