Radiation-Chemical Perspective of the Radiobiology of Pulsed (High Dose-Rate) Radiation (FLASH): A Postscript.

An earlier commentary (Wardman P, Radiat Res. 2020; 194:607-617) discussed possible chemical reaction pathways that might be involved in the differential responses of tissues to high- vs. low-dose-rate irradiation, focusing on reactions between radicals, and radiolytic depletion of a chemical influencing radiosensitivity.

Factors Important in the Use of Fluorescent or Luminescent Probes and Other Chemical Reagents to Measure Oxidative and Radical Stress.

Numerous chemical probes have been used to measure or image oxidative, nitrosative and related stress induced by free radicals in biology and biochemistry. In many instances, the chemical pathways involved are reasonably well understood. However, the rate constants for key reactions involved are often not yet characterized, and thus it is difficult to ensure the measurements reflect the flux of oxidant/radical species and are not influenced by competing factors.

Radiolysis Studies of Oxidation and Nitration of Tyrosine and Some Other Biological Targets by Peroxynitrite-Derived Radicals.

The widespread interest in free radicals in biology extends far beyond the effects of ionizing radiation, with recent attention largely focusing on reactions of free radicals derived from peroxynitrite (i.e., hydroxyl, nitrogen dioxide, and carbonate radicals).

Approaches to modeling chemical reaction pathways in radiobiology.

Purpose: Recent interest in understanding effects of high dose-rate ('FLASH') radiobiology has prompted a number of groups to model the chemical reactions that might be involved, either to estimate radiolytic oxygen consumption in tissues, or the yields and persistence of specific reactive intermediates or products. However, most models have been either not biomimetic and/or inadequately supported by kinetic data. This review summarizes issues which should be addressed in developing models for chemical reactions in radiobiology.

The effects of nitric oxide or oxygen on the stable products formed from the tyrosine phenoxyl radical.

Tyrosine is a critical component of many proteins and can be the subject of oxidative posttranslational modifications. Furthermore, the oxidation of tyrosine residues to phenoxyl radicals, sometimes quite stable, is essential for some enzymatic functions. The lifetime and fate of tyrosine phenoxyl radicals in biological systems are largely driven by the availability and proximity of oxidants and reductants.

Radiotherapy Using High-Intensity Pulsed Radiation Beams (FLASH): A Radiation-Chemical Perspective.

Radiation chemists have been routinely using high-dose microsecond-pulsed irradiation for almost 60 years, involving many thousands of studies, in the technique of "pulse radiolysis". This involves dose rates broadly similar to the FLASH regimen now attracting interest in radiotherapy and radiobiology. Using the experience gained from radiation chemistry, two scenarios are examined here that may provide a mechanistic basis for any differential response in normal tissues versus tumors in FLASH radiotherapy.

Nitroimidazoles as hypoxic cell radiosensitizers and hypoxia probes: misonidazole, myths and mistakes.

Nitroimidazoles have been extensively explored as hypoxic cell radiosensitizers but have had limited clinical success, with efficacy restricted by toxicity. However, they have proven clinically useful as probes for tumour hypoxia. Both applications, and probably much of the dose-limiting toxicities, reflect the dominant chemical property of electron affinity or ease of reduction, associated with the nitro substituent in an aromatic structure.

Calculation of Standard Reduction Potentials of Amino Acid Radicals and the Effects of Water and Incorporation into Peptides.

Guanine (Guo) is generally accepted as the most easily oxidized DNA base when cells are subjected to ionizing radiation; calculations of the standard reduction potential of the guanyl radical, E(Guo/Guo) are within ∼0.1 V of experimental values in aqueous solution extrapolated to standard conditions. While a number of experimental studies have shown some amino acid radicals have redox properties at pH 7 which suggest or confirm a capacity for radical "repair" by electron transfer from the amino acid to Guo (or its deprotonated conjugate), the redox properties of the radicals of other amino acids, including methionine, lysine and cystine, are less well characterized.

Calculations of the Energetics of Oxidation of Aqueous Nucleosides and the Effects of Prototropic Equilibria.

Recently the calculated standard reduction potentials of the radical-cations of N-methyl substituted DNA bases have been reported that agree fairly well with the experimental results. However, there are issues reflecting the fact that the experimental results usually relate to the couple E(o)(Nuc(•),H(+)/NucH(+)), whereas the calculated results are for the E(o)(Nuc(•+)/Nuc) couple. To calculate the midpoint reduction potential at pH 7 (Em7), it is important to have accurate acid dissociation constants (pKs) for both ground-state bases and their radicals, and the effects of uncertainty in some of these values (e.

Kinetics of oxidation of tyrosine by a model alkoxyl radical.

Oxidation of tyrosine moieties by radicals involved in lipid peroxidation is of current interest; while a rate constant has been reported for reaction of lipid peroxyl radicals with a tyrosine model, little is known about the reaction between tyrosine and alkoxyl radicals (also intermediates in the lipid peroxidation chain reaction). In this study, the reaction between a model alkoxyl radical, the tert-butoxyl radical and tyrosine was followed using steady-state and pulse radiolysis. Acetone, a product of the β-fragmentation of the tert-butoxyl radical, was measured; the yield was reduced by the presence of tyrosine in a concentration- and pH-dependent manner.

Kinetics of reduction of tyrosine phenoxyl radicals by glutathione.

Modification of tyrosine (TyrOH) is used as a marker of oxidative and nitrosative stress. 3,3'-Dityrosine formation, in particular, reflects oxidative damage and results from the combination of two tyrosyl phenoxyl radicals (TyrO·). This reaction is in competition with reductive processes in the cell which 'repair' tyrosyl radicals: possible reductants include thiols and ascorbate.

Radiation chemistry comes before radiation biology.

Purpose: This article seeks to illustrate some contributions of radiation chemistry to radiobiology and related science, and to draw attention to examples where radiation chemistry is central to our knowledge of specific aspects. Radiation chemistry is a mature branch of radiation science which is continually evolving and finding wider applications. This is particularly apparent in the study of the roles of free radicals in biology generally, and radiation biology specifically.

Reaction of hydrated electrons with guanine derivatives: tautomerism of intermediate species.

Here, we show that two tautomers are produced by the protonation of the guanine-electron adduct. The fate of electron adducts of a variety of substituted guanosines was investigated by radiolytic methods and addressed computationally by means of time-dependent DFT (TD-B3LYP/6-311G**//B1B95/6-31+G**) calculations. The reaction of e(aq-) with guanosine and 1-methylguanosine produces two transient species, whereas the reaction with N2-ethylguanosine and N2,N2-diethylguanosine produces only one.

Kinetics of reaction of nitrogen dioxide with dihydrorhodamine and the reaction of the dihydrorhodamine radical with oxygen: implications for quantifying peroxynitrite formation in cells.

Dihydrorhodamine 123 (RhH2) has been used to detect 'reactive nitrogen species', including peroxynitrite and its radical decomposition products, peroxynitrite probably oxidizing RhH2 to rhodamine (Rh) via radical products rather than directly. In this study, the radical intermediate (RhH(.)) was generated by pulse radiolysis, and shown to react with oxygen with a rate constant k approximately 7 x 10(8) M(-1) s(-1).

Methods to measure the reactivity of peroxynitrite-derived oxidants toward reduced fluoresceins and rhodamines.

The commonest probes for "reactive oxygen and nitrogen species" are reduced fluorescein and rhodamine dyes that fluoresce when oxidized. The reduced dyes are reactive toward peroxynitrite, although probably not directly but via free radical oxidants derived from it: hydroxyl, carbonate, and nitrogen dioxide free radicals. The reaction with peroxynitrite can be monitored by rapid mixing and stopped-flow spectrophotometry, but reliable measurement of reactivity of the peroxynitrite-derived radicals requires specialized techniques such as flash photolysis or pulse radiolysis to monitor the fast reactions in real time.

Thiyl radicals react with nitric oxide to form S-nitrosothiols with rate constants near the diffusion-controlled limit.

A possible route to S-nitrosothiols in biology is the reaction between thiyl radicals and nitric oxide. D. Hofstetter et al.

The roles of thiol-derived radicals in the use of 2',7'-dichlorodihydrofluorescein as a probe for oxidative stress.

2',7'-Dichlorodihydrofluorescein (DCFH2) is one of the most widely used probes for detecting intracellular oxidative stress, but requires a catalyst to be oxidized by hydrogen peroxide or superoxide and reacts nonspecifically with oxidizing radicals. Thiyl radicals are produced when many radicals are "repaired" by thiols, but are oxidizing agents and thus potentially capable of oxidizing DCFH2. The aim of this study was to investigate the reactivity of thiol-derived radicals toward DCFH2 and its oxidized, fluorescent form 2',7'-dichlorofluorescein (DCF).

Reaction of the carbonate radical with the spin-trap 5,5-dimethyl-1-pyrroline-N-oxide in chemical and cellular systems: pulse radiolysis, electron paramagnetic resonance, and kinetic-competition studies.

Carbonate radicals (CO3-) can be formed biologically by the reaction of OH with bicarbonate, the decomposition of the peroxynitrite-carbon dioxide adduct (ONOOCO2-), and enzymatic activities, i.e., peroxidase activity of CuZnSOD and xanthine oxidase turnover in the presence of bicarbonate.

Oxidative metabolism of combretastatin A-1 produces quinone intermediates with the potential to bind to nucleophiles and to enhance oxidative stress via free radicals.

Combretastatins are stilbene-based, tubulin depolymerization agents with selective activity against the tumor vasculature; two variants (A-1 and A-4) are currently undergoing clinical trials. Combretastatin A-1 (CA1) has a greater antitumor effect than combretastatin A-4 (CA4). We hypothesized that this reflects the enhanced reactivity conferred by the second (ortho) phenolic moiety in CA1.

Fluorescent and luminescent probes for measurement of oxidative and nitrosative species in cells and tissues: progress, pitfalls, and prospects.

Chemical probes for free radicals in biology are important tools; fluorescence and chemiluminescence offer high detection sensitivity. This article reviews progress in the development of probes for "reactive oxygen and nitrogen" species, emphasizing the caution needed in their use. Reactive species include hydrogen peroxide; hydroxyl, superoxide, and thiyl radicals; carbonate radical-anion; and nitric oxide, nitrogen dioxide, and peroxynitrite.