Letter: Free radical cyclization of unsaturated hydroperoxides.
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Cyclization↗
Biomedical subjects
N A Porter
Publications and source records attributed to N A Porter.
A phospholipase C protocol for phospholipid peroxidation analysis.
A new method has been developed to analyze the primary products of phospholipid peroxidation. The procedure utilizes the ability of phospholipase C to hydrolyze phospholipid hydroperoxides to their corresponding diacylglycerol derivatives. 1-Palmitoyl-2-linoleoylphosphatidylcholine (1P,2L-GPC), 1-stearoyl-2-linoleoylphosphatidylcholine (1S,2L-GPC), and 1-stearoyl-2-arachidonylphosphatidylcholine (1S,2A-GPC) were autoxidized. The diacylglycerol hydroxides derived from the phosphatidylcholine hydroperoxides were separated by reverse-phase high-pressure liquid chromatography (RP-HPLC) and normal-phase high-pressure liquid chromatography (NP-HPLC). 1P,2L-diglyceride (1P,2L-DG) and 1P,2A-DG products were easily separated from 1S,2L-DG and 1S,2A-DG products by RP-HPLC. The linoleate diglyceride oxidation mixture was separated into the 13-trans/cis, 13-trans/trans, 9-trans/cis, and 9-trans/trans isomers by NP-HPLC. Likewise, 1P,2A-DG and 1S,2A-DG oxidation products were resolved into the 15-trans/cis, 15-trans/trans, 12-trans/cis, 11-trans/cis, 9-trans/cis, 8-trans/cis, and 5-trans/cis isomers. In both of the above cases, the 1,2-diacylglycerol isomers could be separated from the 1,3 isomers. Moreover, the diastereomers of the 9-, 8-, and 5-hydroxides could be separated. Each of the diacylglycerol oxidation products was characterized by (1) proton nuclear magnetic resonance (proton NMR), (2) electron ionization-mass spectrometry (EI-MS), and (3) NP-HPLC of the corresponding fatty acids. The diacylglycerol analysis provided the same results for the autoxidation of 1P,2L-GPC as the fatty acid methyl ester analysis. In addition, when 1S,2A-GPC was autoxidized in the presence of 5% alpha-tocopherol, both diastereomers of the 5-hydroxide were observed in the same proportions as the other hydroxides.
Diglycerides↗
The resolution of racemic hydroperoxides: a chromatography-based separation of perketals derived from arachidonic, linoleic, and oleic acid hydroperoxides.
In spite of the importance of optically pure unsaturated hydroperoxides in biology, chemistry, and medicine, no general method for synthesizing these labile compounds has been reported. We have developed a chiral vinyl ether that reacts with racemic hydroperoxides to give diastereomeric perketals in high yield. These perketals can be separated by liquid chromatography and the chiral group removed to provide highly enriched hydroperoxide enantiomers (greater than 99% enantiomeric excess). The chiral reagent that has been most successful in our hands is the 2-propenyl ether derived from trans-2-phenylcyclohexanol. By use of this vinyl ether, perketals are readily formed from hydroperoxides, they are stable to normal- and reverse-phase chromatography, and the hydroperoxide is regenerated from the perketal without racemization in high yield with mild acid. Several chiral hydroperoxides have been resolved by this procedure: alpha-phenethyl hydroperoxide, 2-octyl hydroperoxide, and a number of hydroperoxides derived from oleic, linoleic, and arachidonic acids.
Chromatography, Liquid↗
Analytical study of comprehensive and targeted multidimensional gas chromatography incorporating modulated cryogenic trapping.
This paper reports the results of an analytical study comparing capillary gas chromatography (GC) operated in the normal mode with 2 new GC techniques, comprehensive GC (GC x GC) and targeted (or selective) multidimensional GC, which use a longitudinally modulated cryogenic system (LMCS), recently developed in our laboratory. A high-temperature application of derivatized sterols, of interest in fecal pollution monitoring, was chosen for this work. A directly connected coupled-column ensemble was used, comprising a nonpolar column and a moderately polar column. With LMCS, effluent from the first column is zone-compressed in a cryogenic trap and then pulsed to a short second column, producing narrower peaks with sharp, tall peak responses at the detector. The modulator is operated at a constant frequency, e.g., 0.25 s(-1), to produce the GC x GC result, or is moved in a predefined manner so that whole peaks are selectively trapped and subsequently pulsed through to the second column in the targeted mode. Standard solutions containing a mixture of 7 sterols and 5-alpha-cholestane internal standard were used. Detection sensitivity is increased by a factor of >25 with the use of LMCS. The estimated limit of detection was about 0.1 microg/mL when normal GC with flame ionization detection (GC/FID) and a 1.0 microL splitless injection volume were used, compared with 0.02 and 0.004 microg/mL for the LMCS operated in GC x GC and selective modes, respectively. Calibration curves for GC/FID were linear over the 0.1-2.0 microg/mL range tested. Reproducibilities for the GC x GC and normal GC modes were comparable; generally, relative standard deviations (RSD) were on the order of 3-4%, based on raw peak responses. Improved reproducibility was found for selective LMCS operation, at an RSD of around 2%; with internal standardization, better results were achieved. The coupled-column arrangement allowed complete separation of sterol peaks from overlapping impurity peaks in a number of instances with LMCS modes, and its use should improve data quality over that of normal GC operation, in which the overlapping peaks interfere with measurement of peak response in the normal mode.
Calibration↗