Toward methods for analysis of primary autoxidation in neural lipids: detection and characterization of autoxidation in neural lipids and their mitigation by bis-allylic deuterated HUFA

Abstract: Chemical oxidation that destroys highly unsaturated fatty acids (HUFA) is a normal byproduct of metabolism in HUFA-rich neural tissue. Oxidative chain reactions are initiated by abstraction of a bis-allylic H to yield a radical that adds O and propagates via further H abstraction; a single initial H abstraction can destroy scores of HUFA before quenching. Runaway oxidation is likely to be an underlying event in the pathology of neurodegenerative diseases. Methods for kg-scale preparation of D-HUFA have recently been developed, enabling animal and human feeding studies. We developed methods for intramolecular characterization of D-HUFA, and for initial characterization of chemical oxidation products in retinal and neural extracts. We use docosahexaenoic acid (DHA) as an example.

For oxidation analysis, we use high mass resolution orbitrap MS to establish by exact mass measurements that an envelope of isotopologues result from catalytic preparation of D-DHA. In separate experiments, we perform ex vivo oxidation on lipid extracts of bovine retina over several days at room temperature. Using HPLC we demonstrate appearance of peroxides of native DHA-containing phospholipids. Signal was lower from addition of single O to native phospholipids than O-O in these ex vivo studies.

We fed D-DHA and D-ARA to mice, and performed ex vivo oxidation of brain extracts. Our data show the presence of peroxidized phospholipids containing native (H) DHA phospholipids. Preliminary data indicate that D-DHA-containing PL were more robust to oxidation than native PL. These data show that peroxidation of HUFA-containing PL can be detected in complex, real neural extracts.