Antioxidant assays
Roger L. McMullen in Antioxidants and the Skin, 2018
The TRAP value is calculated using the following equation, originally derived from Wayner et al.:The stoichiometric factor of 2 in Equation 6.4 represents the number of peroxyl radicals trapped per added molecule of antioxidant. The induction period for plasma and Trolox is given by τ and τ, respectively. Finally, the concentration of Trolox must be factored into the calculation of TRAP as well as dilution factor, f, in the event that the sample is prepared in a buffer solution. The original TRAP assay has undergone many revisions since it was originally reported by the Ingold group in 1985. For example, it was found that using an oxygen electrode in the original assay was disadvantageous, because the electrode was not stable over the course of the entire experiment. As a result, this led to the introduction of the fluorescent protein, beta-phycoerythrin (ex. 495 nm; em. 575 nm). Thus, as before, the initiator initially reacts with residence antioxidants, and once these have been depleted continues by damaging the fluorescent probe. The decrease in fluorescence of the probe is analogous to original experiments in which the oxygen concentration was monitored. Another frequently used modification to the TRAP assay is to use luminol-enhanced chemiluminescence. Luminol, a molecule frequently used in crime scene investigations to detect blood, exhibits chemiluminescence when it undergoes oxidation. It is believed that peroxyl radicals react with luminol and its chemiluminescence may be monitored using a luminometer. The experimental design is slightly different from the original TRAP assay; however, one may still calculate a TRAP value. The TRAP assay is not only limited to measuring the antioxidant capacity of biological fluids, such as blood plasma. It has also been applied to the study of olive oil, wine, solid food extracts, and plant extracts. To the best of this author’s knowledge, there have been no reports in the literature where the TRAP assay has been used to study the effects of antioxidants in skin. However, one possible application of this assay might be to investigate the effect of topical antioxidant treatment in skin on the antioxidant status of blood plasma.
Principles of forensic science and crime scene investigation
Jason Payne-James, Richard Jones in Simpson's Forensic Medicine, 2019
The presence of blood is normally suggested by its colour and the chemical reaction it gives when a presumptive test is applied. Blood, however, does not always appear as red/brown in colour and may have been diluted. This can make it very challenging to locate stains, particularly on a darker surface. Stains that are to be tested are scraped with the edge of a piece of folded sterile filter paper. The presumptive tests used are generally leuco-malachite green (LMG) or Kastle–Meyer (K-M). Both involve the addition of the reduced form (colourless) of each reagent to the filter paper followed a few seconds later by hydrogen peroxide. If a rapid colour change occurs after the addition of both chemicals, and the colour of the scraping is typical (green for LMG, pink for K-M) of a bloodstain, then the presence of blood is indicated. The colour change occurs as blood has a peroxidase-like activity due to haemoglobin, which catalyses the oxidation of each chemical to its coloured form. When bloodstains cannot be seen, different methods of detection can be used. For example, luminol, in solution, provides a blue chemiluminescent signal in the presence of iron (present in haemoglobin) and provides a very sensitive technique for latent bloodstains.
Why Does Moderate Exercise Enhance, But Intense Training Depress, Immunity ?
Husband Alan J. in Behaviour and Immunity, 2019
Sterile Hank’s balanced salt solution containing 5mM D-glucose (HBSS) and phosphate buffered saline (PBS) were prepared by conventional methods. Luminol was prepared as a chemiluminogenic indicator by suspending 10 mg of luminol (5-amino-2,3-dihydro-l,4-phthalazinedione) (Boehringer, Mannheim, FRG) in 5ml of PBS containing 8mM triethylamine. The mixture was sonicated for 1 minute and shaken to give a clear solution. A single batch of opsonized zymosan (used for all experiments) was prepared by incubating 10 mg of Zymosan-A particles (highly glycosylated fragments of yeast cell walls) (Sigma. St Louis, MO:) with 1 ml of fresh human plasma for 30 min at 37°C. The suspension was centrifuged at 400 χ g for 2 minutes and the pellet washed twice in PBS before resuspension at a final concentration of 10 mg/ml. Aliquots (1ml) were stored frozen and used only once after thawing. Counting by haemocytometer revealed that this preparation contained approx. 106 zymosan particles/microlitre.
Aspirin can stimulate luminol-enhanced chemiluminescence of activated platelets
Published in Platelets, 2010
Zufar Gabbasov, Oksana Ivanova, Victor Kogan-Yasny, Elena Vasilieva
A preliminary investigation was conducted into the influence of aspirin on the luminol-enhanced chemiluminiscence of platelets stimulated with platelet-activating factor (PAF). Ten coronary artery disease patients and six volunteers without coronary artery disease were included in the study. All the patients received aspirin (daily dose, 100 mg) for at least 10 days before in vitro experiments. Luminol-enhanced luminescence of platelet-rich plasma samples mixed with a PAF solution was measured. After stimulation of platelets with PAF, we did not find a luminol-enhanced chemiluminescent response either in the non-coronary artery disease volunteers or in eight out of the 10 coronary artery disease patients examined. However, in samples from two patients where platelets were stimulated with PAF reactive oxygen species were formed. This ability was expressed as an intensive luminol-enhanced luminescence of activated platelets. Such a reaction was observed against the background of the administration of aspirin. The addition of aspirin to a test tube considerably enhanced the intensity of chemiluminescence. In one case, the cancellation of aspirin was accompanied by diminution of the intensity of luminol-enhanced chemiluminescence of platelets. The clinical significance of this phenomenon is unknown.
Antioxidant and Scavenging Activity of Emodin, Aloe-Emodin, and Rhein on Free-Radical and Reactive Oxygen Species
Published in Pharmaceutical Biology, 2004
Franklin Vargas, Yrene Díaz, Karla Carbonell
The objective of this study was to investigate the ability of emodin (1), aloe-emodin (2), and rhein (3) to inhibit free-radical or reactive oxygen species (.OH, 1O2, H2O2) generated in cell-free systems using isoluminol and luminol-enhanced chemiluminescence and electronic absorption spectra. In the presence of 1, 2, and 3, a dose-dependent inhibition period was observed in this system as assayed by isoluminol-enhanced chemiluminescence (ILCL) with horseradish peroxidase (HRP), as well as by luminol-enhanced chemiluminescence (LCL) with H2O2 or ferrous iron. On the other hand, these hydroxyanthraquinones showed an efficient scavenging activity of galvanoxyl radical in ethanolic solutions. In a separate experiment, we observed the trapping of singlet oxygen (1O2) generated by rose bengal in the presence of 1, 2, or 3. These results suggest that emodin, aloe-emodin, and rhein scavenge reactive oxygen and free-radical species in the following decreasing order: emodin > rhein > aloe-emodin.
Antioxidant Action of an Ethanol Extract of Ptychopetalum olacoides
Published in Pharmaceutical Biology, 2002
I.R. Siqueira, C.A.S. Cordova, T.B. Creczynski-Pasa, E. Elisabetsky, D.S. Nunes, C.A. Netto
Alcohol infusions of roots from Ptychopetalum olacoides Bentham (PO; Olacaceae) have been used for treating many diseases in which free radicals are likely to be implicated. Of particular interest are the uses amongst the elderly (to ameliorate cognitive functions), and by patients recovering from pathologies associated with damage to the central nervous system (such as stroke). The aim of this study was to evaluate the antioxidant properties of a PO ethanol extract (POEE) by using various in vitro systems. POEE acted as a scavenger of nitrogen oxides as well as superoxide generated by the xanthine-xanthine oxidase system. The extract also showed a high antioxidant capacity using a luminol chemiluminescence derived from a thermolabile diazocompound. We suggest that the therapeutic effects attributed to P. olacoides could be in part associated to its oxygen free radical scavenging capacity.
Related Knowledge Centers
- Urine
- Blood
- Hemoglobin
- Pyridazines