J.A. Del Río

Antioxidant activity of phenolics extracted from Olea europaea L. leaves

The purpose of this study was to identify the main phenolic compounds present in an olive leaf extract (OL) in order to delineate the differential antioxidant activities of these compounds through the extent of their abilities to scavenge the ABTS+ radical cation and to clarify the structural elements conferring antioxidant capacity in aqueous systems. The results show that the relative abilities of the flavonoids from olive leaf to scavenge the ABTS+ radical cation are influenced by the presence of functional groups in their structure, mainly the B-ring catechol, the 3-hydroxyl group and the 2,3-double bond conjugated with the 4-oxo function. For the other phenolic compounds present in OL, their relative abilities to scavenge the ABTS+ radical cation are mainly influenced by the number and position of free hydroxyl groups in their structure. Also, both groups of compounds show synergic behaviour when mixed, as occurs in the OL.

A kinetic study on the suicide inactivation of peroxidase by hydrogen peroxide

In the absence of reductant substrates, and with excess H2O2, peroxidase (donor: hydrogen-peroxide oxidoreductase, EC 1.11.1.7) shows the kinetic behaviour of a suicide inactivation, H2O2 being the suicide substrate. From the complex (compound I-H2O2), a competition is established between two catalytic pathways (the catalase pathway and the compound III-forming pathway), and the suicide inactivation pathway (formation of inactive enzyme). A kinetic analysis of this system allows us to obtain a value for the inactivation constant, ki = (3.92 +/- 0.06) x 10(-3) x s-1. Two partition ratios (r), defined as the number of turnovers given by one mol of enzyme before its inactivation, can be calculated: (a) one for the catalase pathway, rc = 449 +/- 47; (b) the other for the compound III-forming pathway, rCoIII = 2.00 +/- 0.07. Thus, the catalase activity of the enzyme and, also, the protective role of compound III against an H2O2-dependent peroxidase inactivation are both shown to be important.

Inactivation of peroxidase by hydrogen peroxide and its protection by a reductant agent

Hydrogen peroxide, the oxidant substrate of peroxidase, is also an inactivating agent of this enzyme. The reductant substrates protect the enzyme from the inactivating process. A reaction mechanism is proposed, in which two competitive routes exist for Compound I of peroxidase; one catalytic and one inactivating. The analytical solution produced at the end of the reaction supports the proposed mechanism and shows the dependence between the number of turnovers of the enzyme (r) and the ratio of both substrates.

Enhancement of phenolic compounds in olive plants (Olea europaea L.) and their influence on resistance against Phytophthora sp.

Abstract The total phenol levels in different olive organs and tissues are studied. The HPLC-MS studies point to the presence of oleuropein, catechin and tyrosol as some of the main phenolic compounds in these extracts. The effect of Brotomax treatment on phenolic compound levels in this plant and the possible role of these compounds as antifungal agents against Phytophthora sp. are also studied. An increase in the total phenol content of leaves and stems was observed 120 days after treatment with 0.3% Brotomax. The cortex was the stem tissue which showed the greatest accumulation of these secondary compounds. An in vitro study of the inhibitory effect of these compounds on fungal growth revealed that tyrosol was the most active agent, followed by catechin and oleuropein, their fungitoxic effect being greater when they acted synergically.

Citrus limon: a source of flavonoids of pharmaceutical interest

Some of the medicinal properties of lemons are due to the flavonoids they contain since they are involved in many biological activities and have many health-related functions. The levels of the principal flavanones and flavone found in different cultivars of Citrus limon, are analysed in an attempt to identify the most interesting as regards the content of such secondary compounds. The results show that the immature fruits from cultivars Lisbon and Fino-49 are ideal for obtaining the flavanone hesperidin, while the mature fruits of cultivar Fino-49 and the leaves of cultivar Eureka are the most interesting for obtaining the flavone diosmin and the flavanone eriocitrin.

UV irradiation alters the levels of flavonoids involved in the defence mechanism of Citrus aurantium fruits against Penicillium digitatum

The effect of UV irradiation on the levels of the flavanone, naringin, and the polymethoxyflavone, tangeretin, in the peel of Citrus aurantium fruits is described, as changes in the synthesis and/or accumulation of these compounds after infection with Penicillium digitatum. The growth of P. digitatum on previously irradiated fruit was reduced by up to 45%. Changes in flavonoid levels were detected, associated with inhibition of fungus growth, the naringin content falling by 69% and tangeretin levels increasing by 70%. The possible participation of naringin and tangeretin in the defence mechanism of this Citrus species is discussed.

Flavonoids as nutraceuticals: Structural related antioxidant properties and their role on ascorbic acid preservation

Abstract Chemicals generically referred to as flavonoids belong to the group of phenolic compounds and constitute an important group of secondary metabolites due to their applications as well as their biochemical properties. The similarity of these phenolic structures to other naturally occurring ones, as in some female insect hormones, does not allow for the use of a proper chemical definition based on structure. Therefore, the best way to define these structures is according to their metabolic origin. Hence, the phenols, and as a subclass, the flavonoids, can be defined as those substances derived from the shikimate pathway and phenylpropanoid metabolism. Flavonoids, whichshare a common benzo-γ-pyrone structure, constitute a kind of compound which are highly ubiquitous in the plant kingdom. Over 4, 000 different naturally occurring flavonoids have been discovered, and only in the case of flavones, a specific type of flavonoids, over 36, 000 different chemical structures are possible. Flavonoids are present in a wide variety of edible plant sources, such as fruits, vegetables, nuts, seeds, grains, tea and wine. This finding reports a daily intake of flavonoid that ranges from 23 mg/day in the Netherlands to 170 mg/day in the United States. The numerous health related properties of flavonoids, widely described in epidemiological studies, are mainly based on their antioxidant activities. These properties have been found to include anti-inflammatory and antiviral activities, effects on capillary fragility, inhibition of human platelet aggregation and anticancer activity. The antioxidant capacity of any flavonoid will be determined by a combination of the catechol structure in the B-ring, the 2,3-double bond in conjunction with a 4-oxo function, and the presence of both hydroxyl groups in positions 3 and 5. Due to their excellent antioxidant properties, flavonoids are capable of preventing the ascorbic acid oxidation generally by reverting the ascorbate radical to ascorbic acid and by supporting the level of vitamin C in foodstuffs. Besides this antioxidant property, some flavonoids, such as myricetin and quercetin, show the interesting ability to inhibit the ascorbate oxidase, preventing the enzymatic oxidation of ascorbic acid.

BIOPRODUCTION OF DIOSGENIN IN CALLUS CULTURES OF TRIGONELLA FOENUM-GRAECUM L

The production of the steroidal sapogenin, diosgenin, by callus cultures of Trigonella foenum-graecum L. (fenugreek) is described. The levels of this secondary compound were examined by high performance liquid chromatography with a diode-array detector, and its identity was confirmed by mass spectrometry. The levels of diosgenin detected in leaf callus exceeded the levels detected in stem and root calli. The diosgenin levels accumulated in leaf, stem and root calli at 45 days (maximum production) represent 22, 10 and 27%, respectively, of the levels detected in the corresponding organs of the mother plant at 45 days.

Kinetic characterization of the inactivation process of two peroxidase isoenzymes in the oxidation of indolyl-3-acetic acid

Abstract The inactivation process of two peroxidase (EC 1.11.1.7) isoenzymes (basic and acidic) is studied when they catalyze the oxidation of the plant hormone, indolyl-3-acetic acid (IAA). Using a kinetic approach we can establish that a product of the catalysis (i.e., skatole hydroperoxide derived from IAA), acting as substrate of the enzyme, is also its inactivator when the enzyme operates through a peroxidase pathway. High IAA concentrations protect the enzyme from inactivation because a competition between IAA and the hydroperoxide for compound I of peroxidase is established. Hydroperoxide is a more powerful inactivator for the basic isoenzyme than for the acidic one. In order to verify the kinetic approach, the effect produced by the addition of H2O2 or catalase is described.

Oxygen consumption and enzyme inactivation in the indolyl-3-acetic acid oxidation catalyzed by peroxidase

Abstract Oxidation of indolyl-3-acetic acid (IAA) catalyzed by peroxidase (donor:hydrogen peroxide oxidoreductase, EC 1.11.1.7) can be brought about alternatively by two different pathways that correspond to the oxidase or peroxidase activities of the enzyme. The relative participation of the enzyme in the two pathways depends on the enzyme/substrate ratio and/or the oxygen concentration. Low levels of oxygen favour the oxidase pathway due to the high affinity for oxygen of the ferroperoxidase, the enzymatic form that initiates this pathway. The inactivation of the enzyme estimated either by appearance of P-670 or by measurements of the IAA degraded when the reaction has finished ( P ∞ ) is a good indication that the peroxidase pathway operates. Therefore, the conditions that reduce the enzyme inactivation (for example, high IAA concentrations), also favour the oxidase pathway.