Effect of processing conditions on phospholipase D activity of corn kernel subcellular fractions.

The influence of physicochemical conditions on the phospholipase D (PLD) activity of subcellular preparations of sweet corn (Zea mays L. cv. Peaches and Cream) kernels has been studied. The microsomal, mitochondrial, and cytosolic preparations of corn kernels possessed PLD activity albeit at varying proportions. The microsomal and cytosolic PLD activities were stimulated 2-fold between 5 and 15 degrees C. Ethanol had varying modulatory effects on PLD activity. By contrast, acetaldehyde was a potent inhibitor of PLD. As well, a naturally occurring C(6) aldehyde such as hexanal and an alcohol such as hexanol inhibited PLD activity efficiently. Divalent cations such as calcium chloride and magnesium chloride stimulated PLD activity at micromolar levels. Monovalent cations such as KCl and NaCl did not appear to affect PLD activity. Partial purification of PLD from the microsomal, mitochondrial, and cytosolic fractions separately revealed four major isoforms with relative molecular masses of 200, 140-150, 102-108, and 60-66 kDa. The importance of PLD in the maintenance of processed food quality is discussed.

Influence of salicylic acid on H2O2 production, oxidative stress, and H2O2-metabolizing enzymes. Salicylic acid-mediated oxidative damage requires H2O2.

We investigated how salicylic acid (SA) enhances H2O2 and the relative significance of SA-enhanced H2O2 in Arabidopsis thaliana. SA treatments enhanced H2O2 production, lipid peroxidation, and oxidative damage to proteins, and resulted in the formation of chlorophyll and carotene isomers. SA-enhanced H2O2 levels were related to increased activities of Cu,Zn-superoxide dismutase and were independent of changes in catalase and ascorbate peroxidase activities. Prolonging SA treatments inactivated catalase and ascorbate peroxidase and resulted in phytotoxic symptoms, suggesting that inactivation of H2O2-degrading enzymes serves as an indicator of hypersensitive cell death. Treatment of leaves with H2O2 alone failed to invoke SA-mediated events. Although leaves treated with H2O2 accumulated in vivo H2O2 by 2-fold compared with leaves treated with SA, the damage to membranes and proteins was significantly less, indicating that SA can cause greater damage than H2O2. However, pretreatment of leaves with dimethylthiourea, a trap for H2O2, reduced SA-induced lipid peroxidation, indicating that SA requires H2O2 to initiate oxidative damage. The relative significance of the interaction among SA, H2O2, and H2O2-metabolizing enzymes with oxidative damage and cell death is discussed.

Changes in Activities of Antioxidant Enzymes and Their Relationship to Genetic and Paclobutrazol-Induced Chilling Tolerance of Maize Seedlings.

The potential role of antioxidant enzymes in protecting maize (Zea mays L.) seedlings from chilling injury was examined by analyzing enzyme activities and isozyme profiles of chilling-susceptible (CO 316) and chilling-tolerant (CO 328) inbreds. Leaf superoxide dismutase (SOD) activity in CO 316 was nearly one-half that of CO 328, in which the high activity was maintained during the chilling and postchilling periods. Activity of glutathione reductase (GR) was much higher in roots than in leaves. CO 328 also possessed a new GR isozyme that was absent in roots of CO 316. Ascorbate peroxidase (APX) activity was considerably lower in leaves of CO 328 than in CO 316, and nearly similar in roots. Paclobutrazol treatment of CO 316 induced several changes in the antioxidant enzyme profiles and enhanced their activities, especially those of SOD and APX, along with the induction of chilling tolerance. These results suggest that increased activities of SOD in leaves and GR in roots of CO 328, as well as SOD and APX in leaves and roots of paclobutrazol-treated CO 316, contribute to their enhanced chilling tolerance.

Methionine metabolism in apple tissue.

A metabolic intermediate isolated from apple tissue fed either methionine or 5'-methylthioadenosine has been tentatively identified as a methionine-pyridoxal Schiff base. The formation of this compound is discussed in relation to ethylene biosynthesis.

Inhibition of in Vivo Conversion of Methionine to Ethylene by l-Canaline and 2,4-Dinitrophenol.

l-Canaline, a potent inhibitor of pyridoxal phosphate-mediated reactions, markedly inhibited the conversion of methionine to ethylene and carbon dioxide by apple tissue. A 50% inhibition of methionine conversion into ethylene was obtained with 50 mum canaline and almost complete inhibition with 300 mum canaline. When 2,4-dinitrophenol, an oxidative phosphorylation uncoupler, was fed to apple tissue, it inhibited the conversion of radioactive methionine to ethylene by 50% at a concentration of 60 mum and by 90% at a concentration of 100 mum. Production of labeled carbon dioxide from acetate-1-(14)C was increased by 2,4-dinitrophenol, indicating that the inhibition of ethylene production was due to uncoupling of phosphorylation. Auxin-induced ethylene production by mungbean (Phaseolus mungo L.) hypocotyl sections was similarly inhibited by these inhibitors.These results support the proposal that pyridoxal phosphate is involved in the formation of ethylene from methionine, substantiate the requirement for ATP in ethylene production, and suggest that this ATP requirement occurs in the step (s) between methionine and ethylene. The biosynthetic mechanism probably involves activation of methionine by ATP followed by a pyridoxal phosphate-mediated gamma-elimination.

Effect of 2,4-Dinitrophenol on Auxin-induced Ethylene Production and Auxin Conjugation by Mung Bean Tissue.

Auxin-induced ethylene production by mung bean (Phaseolus mungo L.) hypocotyl segments was markedly inhibited by 2,4-dinitrophenol regardless of whether or not kinetin was present. Uptake of indoleacetic acid-2-(14)C was also inhibited in the presence of 2,4-dinitrophenol. Segments treated only with indoleacetic acid rapidly converted indoleacetic acid into indole-3-acetylaspartic acid with time whereas kinetin suppressed indoleacetic acid conjugation. Formation of indole-3-acetylaspartic acid was significantly reduced when 2,4-dinitrophenol was present. The suppression of indoleacetic acid conjugation by kinetin and 2,4-dinitrophenol appeared to be additive, and the free indoleacetic acid level in segments treated with 2,4-dinitrophenol in the presence of indoleacetic acid or indoleacetic acid plus kinetin was remarkably higher than in corresponding segments which received no 2,4-dinitrophenol.In the absence of 2,4-dinitrophenol, indoleacetic acid-induced ethylene parallels the free indoleacetic acid level within the tissue. However, in the presence of 2,4-dinitrophenol the rate of ethylene production did not correlate with the free indoleacetic acid level. These results indicate that both indoleacetic acid-induced ethylene production and indoleacetic acid conjugation require a continuous supply of ATP, the formation of which was inhibited by 2,4-dinitrophenol.