Spectrophotometric versus spectrofluorometric assessment in the study of the relationships between lipid peroxidation and metabolic dysregulation

Reactive oxygen species are crucial to normal cell function, but are also part of the pathogenesis of multiple modern maladies. As such, sensitive, fast, and reliable methods of appreciating redox status are needed. We aimed to optimize the Amplex Red (AR) and ferric–xylenol orange (FOX) methods usi...

Full description

Saved in:
Bibliographic Details
Published in:Chemical biology & drug design 2019-06, Vol.93 (6), p.1026-1035
Main Authors: Ungurianu, Anca, Șeremet, Oana, Grădinaru, Daniela, Ionescu‐Tîrgoviște, Constantin, Margină, Denisa, Dănciulescu Miulescu, Rucsandra
Format: Article
Language:English
Subjects:
Aged
Animals
Biomarkers - metabolism
carbohydrate
chemical biology
Diabetes Mellitus, Type 2 - metabolism
Female
fluorescence spectroscopy
Humans
lipid
Lipid Peroxidation
Male
Middle Aged
Mitochondria - metabolism
nucleic acid
Oxidative Stress
Phenols - chemistry
Pilot Projects
Rats
Rats, Wistar
Reactive Oxygen Species - metabolism
Spectrometry, Fluorescence - methods
Spectrophotometry, Ultraviolet - methods
Sulfoxides - chemistry
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Reactive oxygen species are crucial to normal cell function, but are also part of the pathogenesis of multiple modern maladies. As such, sensitive, fast, and reliable methods of appreciating redox status are needed. We aimed to optimize the Amplex Red (AR) and ferric–xylenol orange (FOX) methods using human serum samples, rat tissue homogenates, and mitochondrial preparations. For AR, we intended to reduce probe concentration, maintaining method sensitivity, as well as extending its use from isolated lipoproteins samples, and readjust it for a high‐throughput application. Also, we evaluated the usefulness of a modified xylenol orange‐based spectrophotometric protocol, comparing and contrasting these methods in terms of clinical relevance and suitability for their further use in assessing redox status of various biological samples in different pathological conditions. Our results show that these optimized protocols are suitable for complex in vivo studies, as they require low quantities of sample and reagents, and are sensitive, rapid, and economical, with the option of adapting them for high‐throughput analysis. For a better assessment of oxidative status of serum‐derived samples, the two methods can be used concurrently, while for tissue‐derived ones, either can be employed for the measurement of a global redox status. This paper was focused on the optimization of two methods (based on Amplex Red and, respectively, xylenol orange) for the detection of redox imbalances in serum samples, rat tissue homogenates, and mitochondrial preparations. The developed protocols are suitable for studies involving various biological samples in different pathological conditions, as they require low quantities of sample and reagents, and are sensitive, rapid, and economical, with the possibility of adjusting them for a high‐throughput analysis.
ISSN:1747-0277
1747-0285
DOI:10.1111/cbdd.13474