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Simultaneous photoreduction and Raman spectroscopy of red blood cells to investigate the effects of organophosphate exposure.

Journal of Biophotonics 2019 January 22
Simultaneous photoreduction and Raman spectroscopy with 532 nm laser has been used to study the effects of organophosphate (chlorpyrifos) exposure on human red blood cells. Since in red blood cells, auto-oxidation causes oxidative stress, which, in turn, is balanced by the cellular detoxicants, any possible negative effect of chlorpyrifos on this balance should results in an increased level of damaged (permanently oxygenated) hemoglobin. Therefore, when 532 nm laser, at a suitable power, was applied to photoreduce the cells, only common oxygenated form of hemoglobin got photoreduced leaving the permanently oxygenated hemoglobin detectable in the Raman spectra simultaneously excited by the same laser. Using the technique effects of chlorpyrifos to build up oxidative stress on red blood cells could be detected at concentrations as low as 10 ppb from a comparison of relative strengths of different Raman bands. Experiments performed using simultaneously exposing the cells, along with chlorpyrifos, to H2 O2 (oxidative agent) and/or 3-Aminotriazole (inhibitor of anti-oxidant catalase), suggested role of chlorpyrifos to suppress the cellular anti-oxidant mechanism. Since the high level of damaged hemoglobin produced by the action of chlorpyrifos (at concentrations > 100 ppm) is expected to cause membrane damage, atomic force microscopy was used to identify such damages. Upper panel: Raman spectra of normal, photoreduced chlorpyrifos exposed and unexposed red blood cells. Lower panel: The weak Fe-O2 Raman band for chlorpyrifos exposed cells shown on the left. The atomic force microscopy images of unexposed and exposed cells are shown on the right. Scale bar, 2.5 μm. This article is protected by copyright. All rights reserved.

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