Analysis of the Thiol Adduction of Linoleate 9,10-Dihydroxy-11<i>E</i>-13-ketones of the Mammalian Skin Barrier and Their Cyclization to Hemiketals

Enzymic transformations of esterified linoleate (C18:2ω6) in acylceramides help to seal the mammalian skin permeability barrier by promoting Michael addition to protein thiols via the synthesis of 9,10-<i>trans</i>-epoxy-11<i>E</i>-13-oxo and 9,10-<i>erythro</i>-dihydroxy-11<i>E</i>-13-oxo oxidation products. In modeling the conjugation of these oxidized linoleates with cysteine and glutathione, we observed anomalous chromatographic and proton NMR results for the thiol conjugates of the dihydroxy-ketones. Adduction eliminates the 11,12 double bond and allows spontaneous C9-C13 cyclization to hemiketal derivatives. This produces a polar diastereomer that chromatographs as interconverting hemiketal species and a less polar diastereomer favored as a single hemiketal and stereochemically pure as indicated by proton NMR. Structural assignments were aided by precedents from carbohydrate research. For the stable diastereomer, COSY analysis revealed a highly unusual 6-bond <i>J</i>-coupling between the ring H10 proton and the two H14 protons on the omega-side chain, its detection facilitated by the rigidity of the structure and well-dispersed chemical shifts. Hemiketal formation on protein thiol-adducted oxidized acylceramides in the skin barrier should alter the shape and structure and influence access to enzymes or other proteins. Given the common occurrence of hemiketals and hemiacetals in natural products and synthetic chemistry, the current findings should contribute to their secure structural analysis and characterization.