The use of Raman microscopy to determine and localize vitamin E in biological samples

Abstract

Alpha-tocopherol (aT), the predominant form of vitamin E in mammals, is thought to prevent oxidation of polyunsaturated fatty acids. In the lung, aT is perceived to be accumulated in alveolar type II cells and secreted together with surfactant into the epithelial lining fluid. Conventionally, determination of aT and related compounds requires extraction with organic solvents. This study describes a new method to determine and image the distribution of aT and related compounds within cells and tissue sections using the light-scattering technique of Raman microscopy to enable high spatial as well as spectral resolution. This study compared the nondestructive analysis by Raman microscopy of vitamin E, in particular aT, in biological samples with data obtained using conventional HPLC analysis. Raman spectra were acquired at spatial resolutions of 2-0.8 microm. Multivariate analysis techniques were used for analyses and construction of corresponding maps showing the distribution of aT, alpha-tocopherol quinone (aTQ), and other constituents (hemes, proteins, DNA, and surfactant lipids). A combination of images enabled identification of colocalized constituents (heme/aTQ and aT/surfactant lipids). Our data demonstrate the ability of Raman microscopy to discriminate between different tocopherols and oxidation products in biological specimens without sample destruction. By enabling the visualization of lipid-protein interactions, Raman microscopy offers a novel method of investigating biological characterization of lipid-soluble compounds, including those that may be embedded in biological membranes such as aT.

The chemical and biological activities of quinones: overview and implications in analytical detection

“…Gas chromatography (GC) (Raspotnig et al 2010;Zuo et al 2008), Raman microscopy (Beattie et al 2007), high-performance liquid chromatography (HPLC) (Sakunphueak and Panichayupakaranant 2010;Xue et al 2008), and mass spectrometry (MS) (Zhao et al 2010) Scheme 2 Illustration, using benzoquinone as an example, of the nucleophilic addition with formation of mono-,di-,tri, and tetra-susbtitution identification and quantification of quinones. An extensive literature search showed that among the methods, HPLC or HPLC/MS are the most often used methods.…”

“…This non-destructive analysis may allow the discrimination between different tocopherols and oxidation specimens as well as the visualization of lipid-protein interactions. As an imaging technique, Raman microscopy may help to identify biological functions of alpha tocopherol especially, with regards to intracellular distributions and metabolic fate (Beattie et al 2007).…”

Exploring Raman spectroscopy for the evaluation of glaucomatous retinal changes

“…The diverse applications have included characterization of different cancers by obtaining biochemical information from an in situ sample such as lung cancer, [7][8][9] vitamin distribution in tissues 10,11 and the investigation of bone properties. 12 Once the Raman spectra of a tissue sample are acquired, mathematical classification techniques are utilized to differentiate the spectral signatures of diseased and normal tissues.…”

Raman spectroscopy of advanced glycation end products (AGEs), possible markers for progressive retinal dysfunction

“…The diverse applications have included the study of a range of cancers such as lung cancer, 1 -3 vitamin distribution in tissue, 4,5 and the investigation of bone, 6,7 among others. 8,9 The defining characteristic of Raman scattering, of course, is structural specificity, coupled with spatial resolution in the case of Raman microscopy.…”