LC-ESI mass spectra of anabaenopeptin (panel a) and anabaenopeptin B (panel b)

The externally calibrated spectra (no internal lock mass correction) were acquired with a sampling cone setting at 30 V. The data were acquired by using a Synapt G2 quadrupole/time-of-flight (Q-TOF) tandem mass spectrometer (Waters Co., Manchester, UK) equipped with an Acquity UPLC system (Waters) and an ESI source. Samples were injected into a reversed-phase column (ACQUITY UPLC BEH C18 1.7 µm, 2.1×50 mm, at 25°C) and eluted with an isocratic solvent system (solvent A: 0.1% formic acid in water; Solvent B: acetonitrile) at a flow rate of 0.2 mL/min for short gradient (30% B to 70% B in 1 to 2 min) LC-MS runs. The ions generated at positive ESI capillary (2.5 kV) under atmospheric pressure went into the vacuum system through the sampling cone (30 V). The ions went through the transfer optics were mass analyzed at the orthogonal TOF analyzer for the recording of a high-resolution (approx. 10,000 FWHM) externally calibrated product ion spectrum in each second. In post-run data processing, a few spectra at the retention time of each analyte were combined and converted to a centroid spectrum. </p

Product ion spectra from the [M+2H]²⁺ ions of anabaenopeptin A (3) (panel a), B (4) (panel b), citrullinated-B (5) (panel c), and C (6) (panel d), acquired at a 20 eV laboratory-frame collision energy

Doubly charged ions are marked with #. The numbers in parentheses beside each dashed line are nominal masses corresponding to the side of each formal cleavage site. The italicized numbers are nominal masses of actual fragment ions that were generated with hydrogen rearrangement(s). The data were acquired by using a Synapt G2 quadrupole/time-of-flight (Q-TOF) tandem mass spectrometer (Waters Co., Manchester, UK) equipped with an Acquity UPLC system (Waters) and an ESI source. Samples were injected into a reversed-phase column (ACQUITY UPLC BEH C18 1.7 µm, 2.1×50 mm, at 25°C) and eluted with an isocratic solvent system (solvent A: 0.1% formic acid in water; Solvent B: acetonitrile; 30% B) at a flow rate of 0.2 mL/min for short LC-MS/MS runs. The ions generated at positive ESI capillary (3 kV) under atmospheric pressure went into the vacuum system through the sampling cone (15 V). The ions were transferred to the quadrupole mass analyzer for isolation of precursor ions of interests (i.e., the protonated molecules of analytes). The isolated precursor ions were subjected to low-energy CID (20 eV laboratory-frame collision energy) with Ar (99.9999%, 0.4 mL/min) at the trap collision cell (2.5×10−2 mbar). Ions went through the collision cell and the following ion optics were mass analyzed at the orthogonal TOF analyzer for the recording of a high-resolution (approx. 20,000 FWHM) externally calibrated product ion spectrum in each second. In post-run data processing, a few spectra at the retention time of each analyte were combined and converted to a centroid spectrum. The precursor ion peak in each combined spectrum was used as an internal lock mass for recalibration to ensure ppm-level mass accuracy. </div

Product ion spectra from the protonated molecules of citrullinated-anabaenopeptin B (5) (panel a) and anabaenopeptin C (6) (panel b) acquired at a 40 eV laboratory-frame collision energy

The numbers in parentheses beside each dashed line are nominal masses corresponding to the side of each formal cleavage site on the chemical structures. The italicized numbers are nominal masses of actual fragment ions that were generated with hydrogen rearrangement(s). The data were acquired by using a Synapt G2 quadrupole/time-of-flight (Q-TOF) tandem mass spectrometer (Waters Co., Manchester, UK) equipped with an Acquity UPLC system (Waters) and an ESI source. Samples were injected into a reversed-phase column (ACQUITY UPLC BEH C18 1.7 µm, 2.1×50 mm, at 25°C) and eluted with an isocratic solvent system (solvent A: 0.1% formic acid in water; Solvent B: acetonitrile; 30% B) at a flow rate of 0.2 mL/min for short LC-MS/MS runs. The ions generated at positive ESI capillary (3 kV) under atmospheric pressure went into the vacuum system through the sampling cone (15 V). The ions were transferred to the quadrupole mass analyzer for isolation of precursor ions of interests (i.e., the protonated molecules of analytes). The isolated precursor ions were subjected to low-energy CID (40 eV laboratory-frame collision energy) with Ar (99.9999%, 0.4 mL/min) at the trap collision cell (2.5×10−2 mbar). Ions went through the collision cell and the following ion optics were mass analyzed at the orthogonal TOF analyzer for the recording of a high-resolution (approx. 20,000 FWHM) externally calibrated product ion spectrum in each second. In post-run data processing, a few spectra at the retention time of each analyte were combined and converted to a centroid spectrum. The precursor ion peak in each combined spectrum was used as an internal lock mass for recalibration to ensure ppm-level mass accuracy. </div

Product ion spectra from the protonated molecules of anabaenopeptin A (3) and B (4) acquired at a 40 eV laboratory-frame collision energy

The numbers in parentheses beside each dashed line are nominal masses corresponding to the side of each formal cleavage site on the chemical structure for anabaenopeptin A. The italicized numbers are nominal masses of actual fragment ions that were generated with hydrogen rearrangement(s). The data were acquired by using a Synapt G2 quadrupole/time-of-flight (Q-TOF) tandem mass spectrometer (Waters Co., Manchester, UK) equipped with an Acquity UPLC system (Waters) and an ESI source. Samples were injected into a reversed-phase column (ACQUITY UPLC BEH C18 1.7 µm, 2.1×50 mm, at 25°C) and eluted with an isocratic solvent system (solvent A: 0.1% formic acid in water; Solvent B: acetonitrile; 30% B) at a flow rate of 0.2 mL/min for short LC-MS/MS runs. The ions generated at positive ESI capillary (3 kV) under atmospheric pressure went into the vacuum system through the sampling cone (15 V). The ions were transferred to the quadrupole mass analyzer for isolation of precursor ions of interests (i.e., the protonated molecules of analytes). The isolated precursor ions were subjected to low-energy CID (40 eV laboratory-frame collision energy) with Ar (99.9999%, 0.4 mL/min) at the trap collision cell (2.5×10−2 mbar). Ions went through the collision cell and the following ion optics were mass analyzed at the orthogonal TOF analyzer for the recording of a high-resolution (approx. 20,000 FWHM) externally calibrated product ion spectrum in each second. In post-run data processing, a few spectra at the retention time of each analyte were combined and converted to a centroid spectrum. The precursor ion peak in each combined spectrum was used as an internal lock mass for recalibration to ensure ppm-level mass accuracy. </div

Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation

We have recently shown that 2-aminoimidazole is a superior nucleotide activating group for nonenzymatic RNA copying. Here we describe a prebiotic synthesis of 2-aminoimidazole that shares a common mechanistic pathway with that of 2-aminooxazole, a previously described key intermediate in prebiotic nucleotide synthesis. In the presence of glycolaldehyde, cyanamide, phosphate and ammonium ion, both 2-aminoimidazole and 2-aminooxazole are produced, with higher concentrations of ammonium ion and acidic pH favoring the former. Given a 1:1 mixture of 2-aminoimidazole and 2-aminooxazole, glyceraldehyde preferentially reacts and cyclizes with the latter, forming a mixture of pentose aminooxazolines, and leaving free 2-aminoimidazole available for nucleotide activation. The common synthetic origin of 2-aminoimidazole and 2-aminooxazole and their distinct reactivities are suggestive of a reaction network that could lead to both the synthesis of RNA monomers and to their subsequent chemical activation

ORAC assay.

<p>Antioxidant capacities of the F7 lipid fraction and a lipid mixture containing Lyso-PAF C18:0 (44%), Lyso-PAF (9<i>Z</i>)-C18:1 (44%), and LPC C18:0 (12%). Lyso-PAF C18:0, Lyso-PAF (9<i>Z</i>)-C18:1, and LPC are expressed in μmol Trolox equivalents (TE)/g, calculated from a Trolox standard curve.</p

Antiapoptotic activity of the F7 fraction in H₂O₂-treated cells.

<p>J774A.1 cells were preincubated with the F7 lipid fraction for 12 h. H₂O₂ was then added to the cells (112.5 μM). After 12 h, the TUNEL-positive cells were quantified and the results expressed as the mean ± standard deviation of triplicate experiments. (A) TUNEL staining of J774A.1 cells treated with the F7 lipid fraction. (B) Frequency of apoptosis. (*) According to Student’s <i>t</i> test, the difference between the lipid-treated cells and untreated cells (control) was significant (<i>P</i> < 0.05).</p

LC/TOF-MS data for the F7 fraction and its ozonolysis products.

<p>ND; not done.</p><p>LC/TOF-MS data for the F7 fraction and its ozonolysis products.</p

Viability of J774A.1 cells treated with LPC or Lyso-PAF.

<p>J774A.1 cells were preincubated with LPC or Lyso-PAF for 12 h, and H₂O₂ was then added to the cells (25 μM). After 12 h, cell viability was measured and is expressed as the mean viability ± standard deviation of triplicate experiments. (A) F7 lipid fraction. (B) Lipid mixture containing Lyso-PAF C18:0 (44%), Lyso-PAF (9<i>Z</i>)-C18:1 (44%), and LPC C18:0 (12%). (C) Lyso-PAF C18:0. (D) Lyso-PAF (9<i>Z</i>)-C18:1. (E) LPC. (*) Significant (<i>P</i> < 0.05) according to ANOVA followed by Tukey’s multiple comparisons.</p

Structures of lysophospholipids.

<p>Compounds <b>1</b>–<b>6</b> were identified as Lyso-PAF and LPC in the F7 lipid fraction. Compounds <b>7</b>–<b>9</b> were the products of ozonolysis.</p