Triple tof 5600 spectrometer

Young- and mature-stage seed samples were extracted with methanol using sonication for 30 min at 60°C. After extraction, the samples were centrifuged at 12,000 rpm for 3 min at 25°C, and the supernatant was filtered with 0.2-μm Acrodisc MS Syringe Filters with a WWPTFE membrane (Pall Corporation, Port Washington, NY, United States). A quantitative analysis of the anthraquinones was performed using a Triple TOF 5600 + spectrometer with a DuoSpray ion source (AB Sciex, Framingham, MA, United States) coupled with a Nexera X2 UHPLC (Shimadzu, Kyoto, Japan) equipped with a binary solvent manager, a sample manager, a column heater, and a photodiode array detector. UHPLC was performed on an ACQUITY UPLC BEH C18 column (1.7 μm, 2.1 × 100 mm, Waters Corporation, Milford, MA, United States). The mobile phases consisted of 5 mM ammonium acetate in water (eluent A) and 100% acetonitrile (eluent B). The gradient profile was as follows: 0–1 min, 20% eluent B; 1–3.5 min, 20–30% eluent B; 3.5–8 min, 30–50% eluent B; 8–12 min, 50–100% eluent B; and 11–17 min, 100% eluent B. The flow rate was 0.5 mL/min. A total of 5 μL of each sample was injected. To detect the peaks from the test samples, the following MS parameters in ESI-negative mode were used: nebulizing gas, 50 psi; heating gas, 50 psi; curtain gas, 25 psi; desolation temperature, 500°C; and ion spray voltage floating, 4.5 kV.

Early- and late-stage of seed samples were extracted with methanol using sonication for 30 min at 60°C. After extraction, samples were centrifuged at 12,000 rpm for 3 min at 25°C and the supernatant was filtered with 0.2 μm Acrodisc^® MS Syringe Filters with WWPTFE membrane (Pall Corporation, Port Washington, NY, USA). Quantitative analysis of anthraquinones was performed by a Triple TOF 5600+ Spectrometer with a DuoSpray ion source (AB Sciex, Ontario, CA, USA) coupled with a Nexera X2 UHPLC (Shimadzu, Kyoto, Japan) equipped with binary solvent manager, sample manager, column heater, and photodiode array detector. UHPLC was performed on a ACQUITY UPLC®BEH C18 column (1.7 μm, 2.1 x 100 mm, Waters Corporation, Milford, USA) and mobile phases consisted of 5 mM ammonium acetate in water (eluent A) and 100% acetonitrile (eluent B). The gradient profile was as follows: 0–1 min, 20% B; 1–3.5 min, 10–30% B; 3.5–8 min, 30–50% B; 8–12 min, 50–100% B; 11–17 min, 100% B. The flow rate was 0.5 mL/min and five microliters of samples were injected. For detecting peaks from test samples, MS parameter in ESI-negative mode was used as follows: nebulizing gas, 50 psi; heating gas, 50 psi; curtain gas, 25 psi; desolvation temperature, 500°C; ion spray voltage floating, 4.5 kV.

Reagents and solvents employed were obtained from commercial sources and used as received (Merck, Brazil). The UV–Vis spectra were recorded using the Varian-Cary spectrophotometer (Agilent Technologies, CA, USA) with solutions produced in methanol with a concentration of 2 × 10⁻⁵ M. The Perkin Elmer/Series II 2400 (Perkin Elmer, Shelton, USA) analyzer was used to evaluate the elemental analysis of the studied substances. The infrared spectra were obtained with the aid of KBr (4000–400 cm⁻¹) using the FTIR Varian 640 equipment (Agilent Technologies, CA, USA). The ESI-MS and ESI-MS/MS spectra were obtained by the AB Sciex TripleTOF 5600⁺ spectrometer in mode positive, 5500 V and 200 °C, with solutions at a concentration of 50 µM (methanol/dimethylformamide, ratio 99/1%) and 0.1% acetic acid (SCIEX, Framingham, USA). [HPy][AuCl₄] was prepared according to the known literature [45 (link)].

Reactions were carried
out in 5 mL
vials under air. Commercial reagents were used as purchased. Reactions
were monitored by TLC analysis using Merck Silica Gel 60 F-254 thin
layer plates. Flash column chromatography was performed on Merck silica
gel 60, 0.040–0.063 mm, and visualized using both a UV lamp
(254 nm) and then a CAM solution (an aqueous solution of ceric ammonium
molybdate). Melting points were determined in capillary tubes. NMR
spectra were run at 300 MHz for ¹H and 75 MHz for ¹³C using residual nondeuterated solvent as internal standard
(CHCl₃, δ 7.26 and 77.00 ppm, respectively; MeOH,
δ 3.34 ppm and δ 49.87 ppm, respectively). Chemical shifts
are given in ppm. The carbon type was determined by DEPT experiments.
High-resolution mass spectra (ESI) were recorded on a TripleTOF 5600
spectrometer (AB Sciex, Warrington, U.K.) equipped with an electrospray
source with a capillary voltage of 4.5 kV (ESI). Specific optical
rotations were measured using sodium light (D line 589 nm). Chiral
HPLC analyses were performed in a chromatograph equipped with a UV
diode-array detector using columns with chiral stationary phases from
Daicel. 2,2,2-Trifluoroacetophenones 2 used were commercial
and alkylidenpyrazolones 1 were prepared following a
reported procedure.^{23 (link)}

HPLC was performed using an Elite P230 system (Dalian Elite Analytical Instrument Co., Ltd., Dalian, China). ESI-MS was measured on an LTQ XL linear ion trap mass spectrometer (Thermo, Wilmington, MA, USA). HR-ESI-MS was obtained using an AB SCIEX Triple TOF 5600+ spectrometer (AB SCIEX, Boston, MA, USA). Melting point was measured on a WRR melting point apparatus (Shanghai Jingke Instrument Co., Ltd., Shanghai, China) and was uncorrected. Optical rotation was performed using an Anton Paar MCP 300 polarimeter (Anton Paar Opto Tec GmbH, Seelze, Germany) and was uncorrected. 1D (¹H, ¹³C and DEPT 135°) and 2D (¹H-¹H COSY, ¹H-¹³C HSQC and ¹H-¹³C HMBC)-NMR spectra were performed using a Bruker Avance III 500 MHz instrument (Bruker, Billerica, MA) with CDCl₃ (T1, T2, and T3) or C₃D₆O (T4) as the solvent and TMS as the internal standard.