crocin and 6-hydroxy-2-5-7-8-tetramethylchroman-2-carboxylic-acid

Antioxidants could improve sperm media, extending the viability of spermatozoa and protecting their DNA. The protective ability of lipoic acid, melatonin, Trolox and crocin was tested on red deer spermatozoa incubated at 37 degrees C. Cryopreserved spermatozoa were thawed and incubated with 1 mM or 0.1 mM of each antioxidant, with or without oxidative stress (100 muM Fe(2+)). Motility (CASA), viability, mitochondrial membrane potential and acrosomal status were assessed. Lipoperoxidation (malondialdehyde production), intracellular reactive oxygen species (ROS) and DNA status (TUNEL) were checked at 4 h. Incubation alone increased ROS and decreased motility. Oxidative stress intensified these effects, increasing lipoperoxidation and DNA damage. Lipoic acid had little protective effect, whereas 1 mM melatonin showed limited protection. Trolox lowered ROS and lipoperoxidation both in oxidised and non-oxidised samples. In oxidised samples, Trolox prevented DNA and acrosomal damage, and ameliorated motility. Crocin at 1 mM showed similar results to Trolox, but noticeably stimulated motility and had no effect on lipoperoxidation. In a second experiment, a broader range of crocin and melatonin concentrations were tested, confirming the effects of crocin (positive effects noticeable at 0.5-0.75 mM), but showing an increase in lipoperoxidation at 2 mM. Melatonin was increasingly effective at 2.5 and 5 mM (ROS, lipoperoxidation and DNA status). Crocin seems a promising new antioxidant, but its particular effects on sperm physiology must be further studied, especially the consequences of motility stimulation and confirming its effect on lipoperoxidation. Melatonin might be useful at relatively high concentrations, compared to Trolox.

Topics: Acrosome; Animals; Antioxidants; Carotenoids; Chromans; Cryopreservation; Deer; DNA; DNA Damage; Hot Temperature; In Situ Nick-End Labeling; Lipid Peroxidation; Male; Melatonin; Membrane Potential, Mitochondrial; Oxidative Stress; Reactive Oxygen Species; Semen Preservation; Sperm Motility; Spermatozoa; Thioctic Acid

Examination of the crocin bleaching assay performance and in-house validation were focused on probe and test compound characteristics, conditions for peroxyl radical generation, reaction monitoring, and expression of results. HPLC and spectrometric examination showed that any authentic commercial saffron (origin, grade) can be used for probe preparation given that (a) interferences, such as tocopherols, are removed, (b) working solution concentration is adequately adjusted, and (c) stock probe solution changes during storage are not neglected. As suggested by log P values, calculated for a great number of radical scavengers (AHs), any AH more polar than Trolox (common reference compound) can be tested in the aqueous environment of the assay. AH activities order obeyed principles of structure-activity relationships. The assay was robust toward preheating of the azo-initiator (2,2'-azobis(2-aminopropane) dihydrochloride). Reaction monitoring through periodic UV-vis spectra recording was very informative. An alternative expression of results as "percent inhibition of crocin bleaching value", % Inh = [(DeltaA(0) - DeltaA)/DeltaA(0))] x 100, is proposed for [AH]/[crocin] = 1, instead of the so far used k(rel) values. The above findings also lead to analysis cost and time reduction.

Topics: Carotenoids; Chemical Phenomena; Chemistry, Physical; Chromans; Chromatography, High Pressure Liquid; Crocus; Free Radical Scavengers; Kinetics; Peroxides; Reproducibility of Results; Spectrophotometry

The antioxidant properties of S-nitrosoglutathione, a nitric oxide-derived product were studied in different experimental systems. By using the crocin bleaching test, S-nitrosoglutathione, in the presence of copper ions, shows an antioxidant capacity about six times higher than that of Trolox c and referable to the interception of peroxyl radicals by nitric oxide. Copper alone shows a modest inhibitory action, which is about seven times lower than that of Trolox c. S-nitrosoglutathione prevents lipid peroxidation induced by the well-known Fe2+/ascorbate system (IC50 = 450 microM) and the inhibitory effect is strongly reinforced by the presence of copper ions (IC50 = 6.5 microM). In addition, cumene hydroperoxide-induced lipid peroxidation is markedly decreased by S-nitrosoglutathione, provided that copper ions, maintained reduced by ascorbate, are present. Decomposition of S-nitrosoglutathione through metal catalysis and/or the presence of reducing agents and the consequent release of nitric oxide are of crucial importance for eliciting the antioxidant power. In this way, copper ions and/or reducing species with low antioxidant potency are able to promote the formation of an extremely strong antioxidant species such as nitric oxide.

Topics: Animals; Antioxidants; Ascorbic Acid; Benzene Derivatives; Carotenoids; Chromans; Copper; Ferrous Compounds; Lipid Peroxidation; Microsomes, Liver; Nitric Oxide; Peroxides; Rats; Reducing Agents; S-Nitrosoglutathione; Thiobarbituric Acid Reactive Substances

A competition kinetics procedure for measuring total antioxidant capacity in wine is described. This procedure is based on the "crocin bleaching test" [28] as modified for analyzing the antioxidant capacity of complex mixtures [24]. The antioxidant capacity of white wines ranged from 0.08 to 1.2 mM equivalents of the reference antioxidant (Trolox C), while for red wines values ranging from 6.4 to 41.9 mM have been obtained. Although a correlation exists between antioxidant capacity and total phenol content of wines, due to the variable reactivity of different phenol groups, the analysis of the phenol content provides only a crude indication of the actual antioxidant capacity. The analysis of antioxidant capacity on different polyphenol classes, separated by solid phase extraction, indicated that anthocyanins are the major antioxidants of young red wines, and tannins of old red wines and white wines. Several vintages of the same grape have been analyzed and the expected decrease of antioxidant capacity upon ageing was not observed, although spectrophotometric analysis clearly demonstrated the shift from anthocyanin monomers to polymers. Artificial ageing by stirring under air produced a rapid decrease (30 min) of antioxidant capacity followed by an increase (up to two weeks), but not any significant modification of the spectrophotometric chemical age factor. Since neither natural nor artificial ageing present a clear-cut relationship with a decrease of antioxidant capacity, we could conclude that the a priori assumption that an old wine contains less antioxidant capacity, although popular, is not fully correct.

Topics: Air; Anthocyanins; Antioxidants; Binding, Competitive; Carotenoids; Chromans; Color; Fermentation; Flavonoids; Flavonols; Free Radicals; Italy; Kinetics; Oxygen; Phenols; Polymers; Rosales; Tannins; Time Factors; Vitamin E; Wine

The antioxidant effects of manganese and other transition metals were studied as the inhibition of microsomal lipid peroxidation and crocin bleaching by peroxyl radicals. The peroxyl radical scavenging capacity was measured by competition kinetics analysis. While Zn(II), Ni(II), and Fe(II) were almost completely ineffective, Mn(II) and Co(II) showed a free radical scavenging capacity, exhibiting relative rate constant ratios respectively of 0.513 and 0.287. This indicates that Mn(II) is by far the most active. Therefore, the chain-breaking antioxidant capacity of Mn(II) seems to be related to the rapid quenching of peroxyl radicals according to the reaction R-OO. + Mn(II) + H(+)-->ROOH+Mn(III). The antioxidant mechanism is discussed considering the different reduction potentials of the examined cations.

Topics: Antioxidants; Carotenoids; Cations, Divalent; Chromans; Free Radicals; In Vitro Techniques; Kinetics; Lipid Peroxides; Manganese; Microsomes; Oxygen Consumption; Spectrum Analysis