cytochrome-c-t and dihydrorhodamine-123

Polymorphonuclear neutrophils (PMNs) generate reactive oxygen species (ROS) during phagocytosis and in response to soluble agonists. This functional response, termed oxidative burst, contributes to host defense, but it can also result in collateral damage of host tissues. To study this important PMN response, different methods have been developed that are based on the assessment of oxidative burst by measuring intracellular ROS production or formation of ROS in the extracellular space. Among the different methods that were developed, the following two are particularly widely used because of their convenience and accuracy. The first method depends on the reduction of cytochrome c, which can be assessed by photometry, while the second method relies on changes in the fluorescence properties of dihydrorhodamine 123, which can be assessed by flow cytometry.

Topics: Animals; Cell Culture Techniques; Cell Separation; Cytochromes c; Flow Cytometry; Humans; Mice; Neutrophils; Reactive Oxygen Species; Respiratory Burst; Rhodamines

Three hydroquinone glucosides and four caffeoylquinic esters were examined for their effect on tyrosine nitration, as well as on the oxidation of dihydrorhodamine (DHR) 123 and cytochrome c(2+) induced by peroxynitrite. All these phenolics, which had previously been characterized as the active principles of the plant Phagnalon rupestre, were fairly active in preventing the oxidation of DHR 123, though inefficient in the cytochrome c test. While their antioxidant potency is associated with the presence of a caffeoyl moiety, not so an obvious chemical character was correlated to a greater activity against nitration of tyrosine. Here, the highest potency corresponded to 2-isoprenylhydroquinone-1-glucoside. On the basis of the fact that the susceptibility to nitration of given aromatic compound confers to it inhibitory activity of tyrosine nitration, the analysis of ultraviolet and nuclear magnetic resonance spectral shifts provides valuable information for explaining the ability of natural phenolics to interfere with that reaction.

Topics: Asteraceae; Cytochromes c; Glucosides; Hydroquinones; Molecular Structure; Nitrates; Oxidation-Reduction; Peroxynitrous Acid; Rhodamines; Tyrosine