Highly sensitive fluorescent method for the detection of cholesterol aldehydes formed by ozone and singlet molecular oxygen

Fernando V. Mansano, Rafaella M.A. Kazaoka, Graziella E. Ronsein, Fernanda M. Prado, Thiago C. Genaro-Mattos, Miriam Uemi, Paolo Di Mascio, Sayuri Miyamoto

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Abstract

Cholesterol oxidation gives rise to a mixture of oxidized products. Different types of products are generated according to the reactive species being involved. Recently, attention has been focused on two cholesterol aldehydes, 3β-hydroxy-5β-hydroxy-B-norcholestane-6β- carboxyaldehyde (1a) and 3β-hydroxy-5-oxo-5,6-secocholestan-6-al (1b). These aldehydes can be generated by ozone-, as well as by singlet molecular oxygen-mediated cholesterol oxidation. It has been suggested that 1b is preferentially formed by ozone and 1a is preferentially formed by singlet molecular oxygen. In this study we describe the use of 1-pyrenebutyric hydrazine (PBH) as a fluorescent probe for the detection of cholesterol aldehydes. The formation of the fluorescent adduct between 1a with PBH was confirmed by HPLC-MS/MS. The fluorescence spectra of PBH did not change upon binding to the aldehyde. Moreover, the derivatization was also effective in the absence of an acidified medium, which is critical to avoid the formation of cholesterol aldehydes through Hock cleavage of 5α-hydroperoxycholesterol. In conclusion, PBH can be used as an efficient fluorescent probe for the detection/quantification of cholesterol aldehydes in biological samples. Its analysis by HPLC coupled to a fluorescent detector provides a sensitive and specific way to quantify cholesterol aldehydes in the low femtomol range.

Original languageEnglish (US)
Pages (from-to)6775-6781
Number of pages7
JournalAnalytical chemistry
Volume82
Issue number16
DOIs
StatePublished - Aug 15 2010

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ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Mansano, F. V., Kazaoka, R. M. A., Ronsein, G. E., Prado, F. M., Genaro-Mattos, T. C., Uemi, M., Mascio, P. D., & Miyamoto, S. (2010). Highly sensitive fluorescent method for the detection of cholesterol aldehydes formed by ozone and singlet molecular oxygen. Analytical chemistry, 82(16), 6775-6781. https://doi.org/10.1021/ac1006427