Novel Photoinduced Squalene Cyclic Peroxide Identified, Detected, and Quantified in Human Skin Surface Lipids

Abstract

Skin surface lipids (SSLs) form the first barrier that protects the human organism from external stressors, disruption of the homeostasis of SSLs can result in severe skin abnormalities. One of the main causes of this disruption is oxidative stress that is primarily due to SSLs oxidation. Squalene (SQ), the most abundant lipid among SSLs, was shown to first undergo singlet molecular oxygen (1O2) oxidation to yield 6 SQ-monohydroperoxide (SQ-OOH) isomers as the primary oxidation products. However, due to the instability and lability of hydroperoxides, we found that when total SQ-OOH isomers are further photooxidized, they form a unique higher molecular weight secondary oxidation product. To generate the compound, we photooxidized total SQ-OOH isomers in the presence of ground state molecular oxygen (3O2), after its isolation and purification, we studied its structure using MS/MS, NMR, derivatization reactions, and chemical calculations. The compound was identified as 2-OOH-3-(1,2-dioxane)-SQ. Photooxidation of individual SQ-OOH isomers revealed that 6-OOH-SQ is the precursor of 2-OOH-3-(1,2-dioxane)-SQ and indicated the possibility of the formation of similar cyclic peroxides from each isomer following the same photoinduced chain reaction mechanism. An HPLC-MS/MS method was developed for the analysis of 2-OOH-3-(1,2-dioxane)-SQ and its presence on the skin was confirmed in SSLs of six healthy individuals. Its quantity on the skin correlated directly to that of SQ and was not inversely proportional to its precursor, indicating the possibility of its accumulation on the skin surface and the constant regeneration of 6-OOH-SQ from SQ’s oxidation. In general, research on lipid cyclic peroxides in the human organism is very limited, and especially on the skin. This study shows for the first time the identification and presence of a novel SQ cyclic peroxide “2-OOH-3-(1,2-dioxane)-SQ” in SSLs, shedding light on the importance of further studying its effect and role on the skin.

Analytical and Structural Tools of Lipid Hydroperoxides: Present State and Future Perspectives

“…Quantum chemical calculations of NMR chemical shifts and coupling constants have been extensively utilized to confirm proposed structures or to aid reassignment of structures [ 142 , 143 , 144 , 145 , 146 , 147 , 148 , 149 , 150 , 151 , 152 , 153 , 154 ]. Nevertheless, in the field of free fatty acids and their hydroperoxide derivatives, only a limited number of computational studies have so far been reported [ 112 , 138 , 141 , 155 , 156 ]. Recently, Venianakis et al [ 112 ] presented a density functional theory (DFT) study of the 1 H- and 13 C-NMR chemical shifts of the geometric isomers of 18:2 ω-7-conjugated linoleic acid (CLA) and nine model compounds, using five functionals and two basis sets.…”

“…Quantum chemical calculations of NMR parameters have also been extensively used for the precise determination of diastereomerism of natural products with multiple asymmetric centers [ 138 , 141 , 142 , 143 , 158 , 159 , 160 , 161 ]. On the contrary, in the field of lipid hydroperoxides, only three papers have so far been reported [ 138 , 141 , 156 ]. It has been demonstrated that the oxidation of methyl linolenate results in the formation of four peroxyl radicals which lead to conjugated dienoic 9-, 12-, 13-, and 16-hydroperoxides [ 10 , 98 , 139 , 162 ] ( Figure 48 ).…”

“…Recently, Khalifa et al [ 156 ] isolated from human-skin-surface lipids a novel photoinduced squalene cyclic endo hydroperoxide. DFT calculations of 13 C chemical shifts of several model compounds, at the ωB97X-V level of theory, using Boltzmann distribution of the various stereoisomers, showed that the most probable structure is the cis 1A ( Table 11 ).…”

“…These data combined with prediction software will greatly facilitate structural information from 1 H, 13 C, 1 H- 1 H COSY, 1 H- 1 H TOCSY, 1 H- 13 C HSQC, and 1 H- 13 C HMBC NMR data [ 194 , 195 , 196 , 197 ]. DFT calculations of 1 H-, and 13 C- chemical shifts and J couplings can contribute significantly to the unequivocal resonance assignment, identification of cis / trans geometric isomers, and diastereomeric pairs of complex hydroperoxides and solvent effects [ 142 , 143 , 144 , 145 , 146 , 147 , 148 , 149 , 150 , 151 , 152 , 153 , 154 , 155 , 156 , 198 , 199 ]. Further, research is also required for the determination of chirality in solution [ 200 ].…”

Density functional theory calculations of δ(13C) and δ(1H) chemical shifts and 3J(13COO1H) coupling constants as structural and analytical tools in hydroperoxides: Prospects and limitations of 1H13C heteronuclear multiple bond correlation experiments

“…[31] In the last few decades, a great effort has been made to compute NMR parameters to confirm proposed structures or to aid reassignment of structures [33][34][35][36][37][38][39][40][41][42][43][44] and to investigate the effects of solvents on molecular structures. [26,37,40,[44][45][46] To date, however, only a handful examples of NMR computational studies of fatty acids [47][48][49] and their hydroperoxide derivatives [31,50] have been published. We report herein detailed DFT calculations of δ( 13 C) and δ( 1 H) chemical shifts and 3 J ( 13 C O O 1 H) coupling constants as structural tools of model hydroperoxides and provide a critical evaluation of the prospects and limitations of 1 H 13 C HMBC experiments in the analysis of hydroperoxide moieties.…”