Antioxidant activity complexes

Antioxidative activity Complex parameter based on the (bio) chemical reactivity of antioxidants. The antioxidative activity belongs to characteristics typically defined operationally regarding the procedure used. This applies to the utilization of DNA-based biosensor as well. 

The antioxidant activities of carotenoids and other phytochemicals in the human body can be measured, or at least estimated, by a variety of techniques, in vitro, in vivo or ex vivo (Krinsky, 2001). Many studies describe the use of ex vivo methods to measure the oxidisability of low-density lipoprotein (LDL) particles after dietary intervention with carotene-rich foods. However, the difficulty with this approach is that complex plant foods usually also contain other carotenoids, ascorbate, flavonoids, and other compounds that have antioxidant activity, and it is difficult to attribute the results to any particular class of compounds. One study, in which subjects were given additional fruits and vegetables, demonstrated an increase in the resistance of LDL to oxidation (Hininger et al., 1997), but two other showed no effect (Chopra et al, 1996 van het Hof et al., 1999). These differing outcomes may have been due to systematic differences in the experimental protocols or in the populations studied (Krinsky, 2001), but the results do indicate the complexity of the problem, and the hazards of generalising too readily about the putative benefits of dietary antioxidants. 

An interesting development is the combination of HPLC and on-line measurement of reducing capacity or antioxidative activity. This approach allows both direct identification of antioxidative species in complex foods and quantification of the contribution to the overall antioxidative capacity in the absence of synergistic and antagonistic effects. Major advantages are less sample handling and the ability to rim large series of samples in an automated process. 

The on-line measurement of reducing capacity can be performed with either a single or a series of electrochemical detectors, and linear correlations have been demonstrated between total antioxidative activities determined by the electrochemical detection and those determined by DPPH- reduction or by the ORAC assay (Guo et al, 1997 Peyrat-Maillard et al, 2000). The reducing capacity must also be quantified by post-column reactions, either with DPPH- or by the reduction of phosphomolybdenum complexes followed by UV-VIS-detection (Bandoniene and Murkovic, 2002 Cardenosa et al, 2002). A combination of HPLC and semi-automatic ORAC analysis has also been described (Caldwell, 2001). 

In conclusion, oxidation of carotenoids by molecular oxygen, the so-called autoxidation process, is a complex phenomenon that is probably initiated by an external factor (radical, metal, etc.) and for which different mechanisms have been proposed. The autoxidation of a carotenoid is important to take into account when studying antioxidant activity because it can lower the apparent antioxidant activity of a carotenoid. ... 

There are also more complex hydroxycinnamic acids in cereals and other members of the Poaceae. These are cell wall-bound mono-, di- and tri-ferulates. Although they appear to have significant antioxidant activity, their bioavailability (release from cell walls and subsequent absorption in humans) is very low. 

Decomposition of the primary products of lipid oxidation generates a complex mixture including saturated and unsaturated aldehydes such as hexanal. Hexanal is the most commonly measured end product of lipid oxidation, and both sensory and physicochemical methods are used for its determination. Where other antioxidant activity tests may be nonspecific, physicochemical measurement of hexanal offers the advantage of analyzing a single, well-defined end product. 

The photochemiluminiscence (PCL) assay was initially used by Popov and others (1987). Popov and Lewin (1994 1996) have extensively studied this technique to determine water-soluble and lipid-soluble antioxidants. The PCL assay measures the antioxidant capacity, toward the 02 radical, in lipidic and water phase. This method allows the quantification of both the antioxidant capacity of hydrophilic and/or lipophilic substances, either as pure compounds or complex matrices from different origin synthetic, vegetable, animal, human, etc. The PCL method is based on an approximately 1,000-fold acceleration of the oxidative reactions in vitro by the presence of an appropriate photosensitizer. The PCL is a very quick and sensitive method. Chua and others (2008) used this assay to determine the antioxidant potential of Cin-namomum osmophloeum, whereas Kaneh and Wang and others (2006) determined the antioxidant capacity of marigold flowers. The antioxidant activity of tree nut oil extracts was also assessed by this method (Miraliakbari and Shahidi 2008). 

Many methods are available for determining food antioxidant capacity, which is an important topic in food and nutrition research. However, there is a great need to standardize these methods because the frequent lack of an actual substrate in the procedure, the system composition, and the method of inducing oxidation could limit their accuracy. In fact, antioxidant activities in complex systems cannot be evaluated satisfactorily using a single test, and several test procedures may be required. The search for more specific assays that can be more directly related to oxidative deterioration of foods and biological systems should be the objective of future investigations. 

As mentioned above, in contrast to classic antioxidant vitamins E and C, flavonoids are able to inhibit free radical formation as free radical scavengers and the chelators of transition metals. As far as chelators are concerned their inhibitory activity is a consequence of the formation of transition metal complexes incapable of catalyzing the formation of hydroxyl radicals by the Fenton reaction. In addition, as shown below, some of these complexes, for example, iron- and copper-rutin complexes, may acquire additional antioxidant activity. 

In 1989, we showed [142] that the Fe2+(rutin)2 complex is a more effective inhibitor than rutin of asbestos-induced erythrocyte hemolysis and asbestos-stimulated oxygen radical production by rat peritoneal macrophages. Later on, to evaluate the mechanisms of antioxidant activities of iron rutin and copper-rutin complexes, we compared the effects of these complexes on iron-dependent liposomal and microsomal lipid peroxidation [165], It was found that the iron rutin complex was by two to three times a more efficient inhibitor of liposomal peroxidation than the copper-rutin complex, while the opposite tendency was observed in NADPH-dependent microsomal peroxidation. On the other hand, the copper rutin complex was much more effective than the iron rutin complex in the suppression of microsomal superoxide production, indicating that the copper rutin complex indeed acquired additional SOD-dismuting activity because superoxide is an initiator of NADPH-dependent... 

Lipoic acid (the other names are a-lipoic acid or thioctic acid) (Figure 29.9) is a natural compound, which presents in most kinds of cells. Lipoic acid (LA) is contained in many food products, in particular in meat, but it is also synthesized in human organism from fatty acids. Earlier, it has been shown that in humans lipoic acid functions as a component of the pyruvate dehydrogenase complex. However, later on, attention has been drawn to the possible antioxidant activity of the reduced form of lipoic acid, dihydrolipoic acid (DHLA) (Figure 29.9). 

Ubiquinones (coenzymes Q) Q9 and Qi0 are essential cofactors (electron carriers) in the mitochondrial electron transport chain. They play a key role shuttling electrons from NADH and succinate dehydrogenases to the cytochrome b-c1 complex in the inner mitochondrial membrane. Ubiquinones are lipid-soluble compounds containing a redox active quinoid ring and a tail of 50 (Qio) or 45 (Q9) carbon atoms (Figure 29.10). The predominant ubiquinone in humans is Qio while in rodents it is Q9. Ubiquinones are especially abundant in the mitochondrial respiratory chain where their concentration is about 100 times higher than that of other electron carriers. Ubihydroquinone Q10 is also found in LDL where it supposedly exhibits the antioxidant activity (see Chapter 23). 

Thus, the mechanism of MT antioxidant activity might be connected with the possible antioxidant effect of zinc. Zinc is a nontransition metal and therefore, its participation in redox processes is not really expected. The simplest mechanism of zinc antioxidant activity is the competition with transition metal ions capable of initiating free radical-mediated processes. For example, it has recently been shown [342] that zinc inhibited copper- and iron-initiated liposomal peroxidation but had no effect on peroxidative processes initiated by free radicals and peroxynitrite. These findings contradict the earlier results obtained by Coassin et al. [343] who found no inhibitory effects of zinc on microsomal lipid peroxidation in contrast to the inhibitory effects of manganese and cobalt. Yeomans et al. [344] showed that the zinc-histidine complex is able to inhibit copper-induced LDL oxidation, but the antioxidant effect of this complex obviously depended on histidine and not zinc because zinc sulfate was ineffective. We proposed another mode of possible antioxidant effect of zinc [345], It has been found that Zn and Mg aspartates inhibited oxygen radical production by xanthine oxidase, NADPH oxidase, and human blood leukocytes. The antioxidant effect of these salts supposedly was a consequence of the acceleration of spontaneous superoxide dismutation due to increasing medium acidity. 

The application of fullerene on the surfaces has an essential advantage in the studies with cell cultures as in this case we can obtain the maximum contact of cells with fullerene - cells adhere on the surface and colonize it as a confluent monolayer. That is the basic difference from the water-soluble complexes and micro-dispersed suspensions of fullerene C60. The pro-/antioxidant activities of fullerene were tested in chemical and biological systems. 

Figure 9.2. Mechanisms of aminoglycoside toxicity. This schematic representation summarizes the principles of aminoglycoside toxicity discussed in the text. Treatment with the drugs leads to the formation of reactive oxygen species through a redox-active complex with iron and unsaturated fatty acid or by triggering superoxide production by way of NADPH oxidase. An excess of reactive oxygen species, not balanced by intracellular antioxidant systems, will cause an oxidative imbalance potentially severe enough to initiate cell death pathways. Augmenting cellular defenses by antioxidant therapy can reverse the imbalance and restore homeostasis to protect the cell.

Analysis of antioxidant activity by performing a FRAP assay was proposed by Benzie and Strain [23]. It involves colorimetric determination of the reaction mixture in which the oxidants contained in the sample reduce Fe ions to Fe. At low pH, Fe(in)-TPTZ (ferric-tripyridltria-zine) complex is reduced to the ferrous (Fe ) form and intense blue colour at 593 nm can be observed. The FRAP reagent is prepared by mixing 2.5 ml of TPTZ (2,4,6-tris (l-pyridyl)-5-triazine) solution (10 mM in 40mM HCl), 25 ml acetate buffer, pH 3.6, and 2.5 ml FeCl3 H20 (20 mM). The colour of Fe(II)(TPTZ)2 which appears in the solution is measured colorimetri-cally after incubation at 37°C. The measurement results are compared to those of a blank sample, which contains deionised water instead of the analysed sample. The duration of the assay differs from one study to another 4 min [23, 24], 10 min [25] to 15 min [26]. The analysis results are converted and expressed with reference to a standard substance, which can be ascorbic acid [26], FeS04 [23, 25], Trolox [27,18]. 

There are numerous antioxidant methods and modifications for the evaluation of antioxidant activity [139, 145-151]. Multiple assays in screening work are highly advisable, considering the chemical complexity of EOs [152]. 

Thompson M, Williams CR, Elliot GE. 1976. Stability of flavonoid complexes of copper(II) and flavonoid antioxidant activity. Anal Chim Acta 85 375-381. 

The evidence that a 24-hour fasting and tranquillity have both the ability for a strong antioxidant activity and may create some complexity in the definition of antioxidants. In many instances, subjects suffering from some diseases (hypertension, infection, inflammation) or under particular conditions such as menopause may also have OS, which can be considered as an epiphenomenon of that given condition. Once the disease (or the symptom and/or the condition) is controlled by a therapy, the OS may disappear. This means that a product can be considered as an antioxidant indirectly." These aspects may further complicate the definition of an antioxidant. 

Complexing, particularly of copper and iron, has great significance in terms of antioxidant activity (see Chapter 9). 

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