Lipid peroxidation is believed to play a key role in the onset and progression of degenerative disease. Interestingly, although cholesterol is the most abundant lipid in the human body, our understanding of its autoxidation and subsequent decomposition is relatively limited. In fact, until recently, cholesterol-7-hydroperoxide was accepted as the only primary product of cholesterol autoxidation in organic solution, however, our group exhibited that the 4-, 5-, and 6-hydroperoxides are also formed. Although this work facilitated thorough investigation of the complexities of both H-atom abstraction and addition in cholesterol autoxidation in organic solution, it did not account for the dynamic environment of a cell membrane. Herein, we report on the product distribution of these primary autoxidation products in lipid bilayers and how antioxidant supplementation, H-bonding interactions, and concentration of polyunsaturated fatty acid (PUFA) substrate influence both the product distribution and efficiency of autoxidation. Indeed, not only does H-bonding of the 3β-OH of cholesterol appear to shut-down C4 H-atom abstraction, the absence of kinetic chol-5α-OOH product is likely due to the poor potency of α- tocopherol (α-TOH), also as a result of H-bonding with phosphate head group of lipid membrane phospholipids. Therefore, within a lipid membrane the 7-hydroperoxide products predominate, consistent with literature precedent, however the factors involved are more complex than previously understood. Moreover, with the authentic cholesterol hydroperoxides in hand, we sought to determine if the different regioisomers exhibit different cytotoxicity. Glutathione peroxidases (GPXs) are cytoprotective enzymes that reduce harmful hydroperoxides to benign alcohols in vivo. Using RSL3, a small-molecule inhibitor for GPX4, we were able to sensitize mammalian cells to ferroptotic cell death via administration of our exogenously prepared chol-OOHs. Surprisingly, we found that the toxicities of each of 7α-OOH, 6β-OOH and 5α-OOH were only marginally augmented by RSL3 treatment, suggesting that they do not substantially sensitize cells to ferroptosis, perhaps because their decomposition to lipid peroxidation chain-initiating species (i.e. alkoxyl radicals) is not particularly efficient. Instead their cytotoxicities may derive from other mechanisms, such as the induction of apoptosis. This inspired our investigation of the fate of lipid hydroperoxides in vivo, namely the secondary products of the predominant 7-hydroperoxide species. Acid-catalyzed Hock fragmentation, known for the industrial synthesis of phenol and acetone from cumene or implication in the generation of 4-hydroxynonenal (4-HNE), of 5α- and 6β-OOH has been shown by our group to produce highly electrophilic secosterol species; we sought to investigate the same decomposition mechanism for 7α-OOH in light of our investigations in the lipid membrane. Interestingly, we found that Hock fragmentation of 7α-OOH does not exhibit products resulting from the anticipated O-vinyl oxocarbenium intermediate, rather, the mechanism appears to funnel through an α-epoxy carbenium to produce unprecedented A-ring cleavage and epoxide products. Herein, we describe our thorough analysis of this chol-7α-OOH Hock fragmentation and attempts to investigate the presence of these products in biological samples, similar to previous analyses of similar products in atherosclerotic plaque extracts. The products isolated and characterized through this work have provided new mechanistic insight with regards to the primary and secondary oxidation products of cholesterol in vivo; through further development of these findings, we hope to provide a better understanding of the implications of cholesterol oxidation in the pathogenesis of atherosclerosis.