Senescense: association of synthesis of Acid phosphatase with banana ripening

During ripening of banana (Musa sapientum L., var. Gros Michel or Valery) acid phosphatase activity increases 13-to 26-fold in the precipitate and 2- to 4-fold in the supernatant fraction of tissue homogenates. These increases are closely correlated with the onset and peak of the climacteric. The precipitate enzyme may be extracted with Triton X-100, CaCl₂ or NaCl; about 80% of it is in a 500g precipitate. Studies on effect of tonicity of the grinding medium indicate that the precipitate enzyme is desorbed from membrane or cell wall surfaces, and is not released as a result of lysis of membranes. The development of acid phosphatase during aging of tissue slices is the same as in intact fruit. Short term studies of tissue slices with cycloheximide and actinomycin D indicate that the increase in activity is owed to new enzyme synthesis, which is dependent upon synthesis of RNA. The possible effects of the increase in acid phosphatase on ripening are discussed.     

Effect of gibberellic Acid on the plasticity and elasticity of Avena stem segments

Extensibility characteristics of Avena stem segments treated with gibberellic acid (GA) were investigated in living internodes using a microgrowth method and in partially extracted cell walls subjected to Instron extensometer analysis. Both techniques showed that treatment with GA greatly increases internodal plasticity, but has virtually no effect on internodal elasticity. The increase in plasticity occurred 1 to 2 hours after the initiation of hormone treatment, which is similar to the time of onset of GA-enhanced growth and cell wall synthesis. Cycloheximide was shown to inhibit the effect of GA on plasticity.     

Abscisic Acid and stomatal regulation

The closure of stomata by abscisic acid was examined in several species of plants through measurements of CO₂ and H₂O exchange by the leaf. The onset of closure was very rapid, beginning at 3 minutes from the time of abscisic acid application to the cut base of the leaf of corn, or at 8 or 9 minutes for bean, Rumex and sugarbeet; rose leaves were relatively slow at 32 minutes. The timing and the concentration of abscisic acid needed to cause closure were related to the amounts of endogenous abscisic acid in the leaf. Closure was obtained in bean leaves with 8.9 picomoles/cm². (+)-Abscisic acid had approximately twice the activity of the racemic material. The methyl ester of abscisic acid was inactive, and trans-abscisic acid was likewise inactive. The effects of stress on levels of endogenous abscisic acid, and the ability of very small amounts of abscisic acid to cause rapid closure suggests that stomatal control is a regulatory function of this hormone.     

Induction of Excessive Deoxyribonucleic Acid Synthesis in Escherichia coli by Nalidixic Acid

Prior treatment of Escherichia coli with nalidixic acid in nutritionally complete medium altered the subsequent pattern of deoxyribonucleic acid (DNA) synthesis normally observed in nutritionally deficient medium. Transfer of E. coli 15 TAU to an amino acid- and pyrimidine-deficient medium usually resulted in a 40 to 50% increase in DNA content. Previous treatment with nalidixic acid caused a 200 to 300% increase in DNA content under these conditions. The extent of this DNA synthesis depended on the duration of prior exposure to nalidixic acid. The maximal rate of synthesis was obtained after a 40- to 60-min exposure to nalidixic acid and was two to three times that of the control. The induction of this excessive DNA synthesis was prevented by chloramphenicol or phenethyl alcohol, but the synthesis of this DNA was only partially sensitive to these agents. With E. coli TAU-bar, the rate of DNA synthesis, after removal of nalidixic acid, was similar to that of E. coli 15 TAU, but the maximal amount of DNA synthesized was 180 to 185% of that initially present. Cesium chloride density gradient analysis demonstrated that DNA synthesis after removal of nalidixic acid occurs by a semiconservative mode of replication. The density distribution of this DNA was similar to that obtained after thymine starvation. These results suggest that nalidixic acid treatment may induce additional sites for DNA synthesis in E.coli.     

The Weak Acid Preservative Sorbic Acid Inhibits Conidial Germination and Mycelial Growth of Aspergillus niger through Intracellular Acidification

The growth of the filamentous fungus Aspergillus niger, a common food spoilage organism, is inhibited by the weak acid preservative sorbic acid (trans-trans-2,4-hexadienoic acid). Conidia inoculated at 10⁵/ml of medium showed a sorbic acid MIC of 4.5 mM at pH 4.0, whereas the MIC for the amount of mycelia at 24 h developed from the same spore inoculum was threefold lower. The MIC for conidia and, to a lesser extent, mycelia was shown to be dependent on the inoculum size. A. niger is capable of degrading sorbic acid, and this ability has consequences for food preservation strategies. The mechanism of action of sorbic acid was investigated using ³¹P nuclear magnetic resonance (NMR) spectroscopy. We show that a rapid decline in cytosolic pH (pH_cyt) by more than 1 pH unit and a depression of vacuolar pH (pH_vac) in A. niger occurs in the presence of sorbic acid. The pH gradient over the vacuole completely collapsed as a result of the decline in pH_cyt. NMR spectra also revealed that sorbic acid (3.0 mM at pH 4.0) caused intracellular ATP pools and levels of sugar-phosphomonoesters and -phosphodiesters of A. niger mycelia to decrease dramatically, and they did not recover. The disruption of pH homeostasis by sorbic acid at concentrations below the MIC could account for the delay in spore germination and retardation of the onset of subsequent mycelial growth.     

Endogenous levels of abscisic Acid and decanoic Acid in dutch iris bulbs and the influence of abscisic Acid and decanoic Acid on iris meristems cultured in vitro

Abscisic acid (ABA) and decanoic acid inhibited shoot elongation and floral development of Dutch iris (Iris hollandica Hoog. cv Ideal) meristems cultured in vitro. No synergism with respect to inhibition of leaf growth between ABA and decanoic acid was observed. With monthly harvest dates, from July 10, 1981 to October 10, 1981, there was a progressive decrease in endogenous level of free ABA in `Ideal' iris bulbs. Bulbs subjected to a full set of the usual preplanting storage conditions flowered, on average, 46 days after planting versus 194 days after planting for bulbs planted directly after harvest. ABA levels at harvest were 4- to 5-fold those after the preplanting storage treatment. In general, ABA levels did not correlate well with the length of time from planting until flowering of iris bulbs. Endogenous decanoic acid levels did not follow any pattern with respect to harvest date or postharvest treatment. After the postharvest high temperature treatment, there was about a 3-fold increase in nonscale decanoic acid concentration. Decanoic acid levels, in nonscale tissue, remained high after each of the other postharvest treatments. It is concluded that there is no good evidence to support the contention that either ABA or decanoic acid is directly involved in iris bulb dormancy.     

Reduction of Lactic Acid, Nonprotein Nitrogen, and Ash in Lactic Acid Whey by Candida ingens Culture

A simple, efficient procedure for removing lactic acid and for reducing nonprotein nitrogen and ash in lactic acid whey has been developed. The procedure consists of culturing Candida ingens on the whey. This organism could assimilate >98% of the lactic acid and approximately 40% of the nonprotein nitrogen. Ash reduction of up to 45% resulted from precipitation of calcium apatite due to the increase in pH from 4.4 to approximately 8.0 which occurred during growth of C. ingens. Improved fluxes during laboratory-scale ultrafiltration were obtained for the treated lactic acid whey. C. ingens treatment of lactic acid whey appears to facilitate processing of this material to a more useful product.     

Effect of nitrate on the organic Acid and amino Acid composition of legume nodules

Nitrate supplied to legume plants inhibits the activity of nitrogenase in Rhizobium bacteroids in root nodules. The accumulation of amino N which is known to occur in Glycine max (L.) Merr. nodules as nitrogenase activity declines was studied in more detail by analysis of changes in free amino acid composition in response to high nitrate supply. A 6-fold increase in asparagine concentration in Bradyrhizobium japonicum bacteroids was found about the time of maximum nitrogenase inhibition. However, the accumulation of amino acids in soybean nodules lagged behind the inhibition of nitrogenase. Furthermore, in studies of a second legume, Phaseolus vulgaris (L.) inoculated with two different strains of Rhizobium phaseoli, a high nitrate treatment inhibited nitrogenase but had no significant effect on amino acid composition of nodules. The possibility that nitrate may interfere with the supply of carbon substrates to bacteroids was examined by the analysis of organic acids in legume nodules supplied with nitrate. Nitrate had a small (10-20%) negative effect on the concentration of tricarboxylic acid cycle acids in P. vulgaris nodules. However, in G. max nodules, high nitrate treatment resulted in significant increases in the concentration of malate, succinate, fumarate, and citrate. Thus, carbon deprivation of bacteroids also seems unlikely as a cause of the inhibition of nitrogenase by nitrate. There was a transient increase in ammonium concentration in P. vulgaris nodules in response to high nitrate treatment. This effect was rapid relative to other effects of nitrate on nodule composition and was roughly coincident with the rapid decline in acetylene reduction activity.     

Onset of deoxyribonucleic Acid synthesis in germinating wheat embryos

Germinating wheat embryos (Triticum vulgare var. Florence) synthesize proteins before the onset of DNA synthesis. The onset of DNA replication occurs at about 15 hours of germination and was shown to depend on proteins synthesized before 9 hours of germination with the use of blasticidin S, a specific inhibitor of protein synthesis. A 10-fold increase in the activity of DNA-dependent DNA polymerase was found in extracts derived from germinated embryos, as compared to the activity found in extracts from ungerminated embryos.     

Abscission: role of abscisic Acid

The effect of abscisic acid on cotton (Gossypium hirsutum L. cv. Acala 4-42) and bean (Phaseolus vulgaris L. cv. Red Kidney) explants was 2-fold. It increased ethylene production from the explants, which was found to account for some of its ability to accelerate abscission. Absci is acid also increased the activity of cellulase. Increased synthesis of cellulase was not du to an increase in aging of the explants but rather was an effect of abscisic acid on the processes that lead to cellulase synthesis or activity.     

Initiation of Ripening in Bartlett Pear with an Antiauxin alpha(p-Chlorophenoxy)isobutyric Acid

A vacuum infiltration technique was used to apply an anti-auxin, α-(p-chlorophenoxy) isobutyric acid to mature green pears (Pyrus communis var. Bartlett). Application of α-(p-chlorophenoxy) isobutyric acid, at 0.02, 0.2, and 2.0 mm progressively accelerated the onset of chlorophyll degradation, softening, and CO₂ evolution. The action of α(p-chlorophenoxy) isobutyric acid is apparently independent of ethylene, since the auxin analogue depressed ethylene evolution and could overcome ethylene deficiency in fruit ripening under hypobaric conditions.     

Escherichia coli and Salmonella enterica Are Protected against Acetic Acid,but Not Hydrochloric Acid,by Hypertonicity

Chapman et al. (B. Chapman, N. Jensen, T Ross, and M. B. Cole, Appl. Environ. Microbiol. 72:5165-5172, 2006) demonstrated that an increased NaCl concentration prolongs survival of Escherichia coli O157 SERL 2 in a broth model simulating the aqueous phase of a food dressing or sauce containing acetic acid. We examined the responses of five other E. coli strains and four Salmonella enterica strains to increasing concentrations of NaCl under conditions of lethal acidity and observed that the average “lag” time prior to inactivation decreases in the presence of hydrochloric acid but not in the presence of acetic acid. For E. coli in the presence of acetic acid, the lag time increased with increasing NaCl concentrations up to 2 to 4% at pH 4.0, up to 4 to 6% at pH 3.8, and up to 4 to 7% (wt/wt of water) NaCl at pH 3.6. Salmonella was inactivated more rapidly by combined acetic acid and NaCl stresses than E. coli, but increasing NaCl concentrations still decreased the lag time prior to inactivation in the presence of acetic acid; at pH 4.0 up to 1 to 4% NaCl was protective, and at pH 3.8 up to 1 to 2% NaCl delayed the onset of inactivation. Sublethal injury kinetics suggest that this complex response is a balance between the lethal effects of acetic acid, against which NaCl is apparently protective, and the lethal effects of the NaCl itself. Compared against 3% NaCl, 10% (wt/wt of water) sucrose with 0.5% NaCl (which has similar osmotic potential) was found to be equally protective against adverse acetic acid conditions. We propose that hypertonicity may directly affect the rate of diffusion of acetic acid into cells and hence cell survival.We previously observed that inactivation of Escherichia coli O157 SERL 2 by acetic acid at adverse pH in a broth model simulating the aqueous phase of acidic sauces and dressings was reduced by the presence of NaCl (4). Specifically, the time to a 3-log₁₀-unit reduction (t_3D) of E. coli SERL 2 as function of NaCl concentration was significantly nonmonotonic; that is, the t_3D initially increased when NaCl was increased (from 1 to 3% [wt/wt] of solution), but the t_3D decreased upon a further increase in NaCl concentration (to 8% [wt/wt] of solution) (4). The statistical significance of this “nonmonotonic” response increased with increasing exposure time from 24 to 72 h (at 23°C), primarily due to a proportionally greater increase in inactivation at 1% (wt/wt) NaCl with increasing treatment time than that which was observed at higher NaCl concentrations (4).The combination of acid and NaCl is a common example of the food industry''s “hurdle” approach, which is used to preserve a large and diverse range of foods, including acidic dressings and sauces, fermented meats, cheeses, and preserved vegetables. Given the widespread use of this hurdle combination in food manufacturing, the first aim of this study was to determine whether the observed protection of E. coli SERL 2 from acid inactivation by NaCl is common among E. coli and Salmonella enterica and at what NaCl concentration maximum protection is achieved. A second aim was to determine whether NaCl protection is specific against acid pH in general or against acetic acid in particular. Third, possible protection against acid inactivation by another osmolyte, sucrose, was assessed to resolve whether the effect is solute specific.When cells are placed in hypertonic environments, plasmolysis occurs as the cytoplasmic volume reduces due to water loss by osmosis. The thin peptidoglycan layer of gram-negative microorganisms is anchored to the cytoplasmic membrane and can be distended by plasmolysis or even ruptured when plasmolysis is more extreme. Decad and Nikaido (5) observed that the cytoplasmic volume in gram-negative microorganisms was reduced to ∼50% at ∼0.3 M NaCl but that the plasmolysis-induced cell wall damage was minimal. At 0.5 M (2.9%, wt/wt) NaCl, however, they observed cell wall damage in a large fraction of cells. Thus, in the experiments described here we explored the mechanism of the protective effect of NaCl, and specifically cell wall damage in E. coli populations simultaneously exposed to NaCl and either acetic or hydrochloric acid (HCl), by enumeration of both injured and noninjured survivors by culture on media with and without bile salts.     

Fatty Acid Composition and the Oxidation of Low-Density Lipoproteins

The effect of the fatty acid composition of low-density lipoprotein (LDL) on copper-ion-catalyzed oxidation of isolated LDL was examined in 18 normolipidemic men. The decrease in LDL linoleic acid concentration (ΔL) during oxidation was found to be strongly correlated with initial LDL linoleic acid concentration (r = 0.976, n = 18, P < 0.001), whereas the production of thiobarbituric acid reacting substances (TBARS) was not. The concentration of oleic acid in LDL was then increased significantly (mean increase 20%, P < 0.05) in 8 male volunteers by a daily dietary supplement of rapeseed oil/muesli for 4 weeks. The mean delay before copper-ion-catalyzed production of conjugated dienes (the lag phase) was significantly (P < 0.001) greater in LDL isolated after the study period (67 min) than that of before the study period (40 mm). The rate of formation of conjugated dienes, ΔL and TBARS production during oxidation of LDL was not significantly altered by the rapeseed oil/muesli supplement. These results suggest that the linoleic acid content of LDL is a determinant of individual variability in LDL oxidation, and that a rapeseed oil/muesli dietary supplement reduces the susceptibility of LDL to oxidation.     

Epicuticular Wax Accumulation and Fatty Acid Elongation Activities Are Induced during Leaf Development of Leeks

Epicuticular wax production was evaluated along the length of expanding leek (Allium porrum L.) leaves to gain insight into the regulation of wax production. Leaf segments from the bottom to the top were analyzed for (a) wax composition and load; (b) microsomal fatty acid elongase, plastidial fatty acid synthase, and acyl-acyl carrier protein (ACP) thioesterase activities; and (c) tissue and cellular morphological changes. The level of total wax, which was low at the bottom, increased 23-fold along the length of the leaf, whereas accumulation of the hentriacontan-16-one increased more than 1000-fold. The onset of wax accumulation was not linked to cell elongation but, rather, occurred several centimeters above the leaf base. Peak microsomal fatty acid elongation activity preceded the onset of wax accumulation, and the maximum fatty acid synthase activity was coincident with the onset. The C16:0- and C18:0-ACP-hydrolyzing activities changed relatively little along the leaf, whereas C18:1-ACP-hydrolyzing activity increased slightly prior to the peak elongase activity. Electron micrographic analyses revealed that wax crystal formation was asynchronous among cells in the initial stages of wax deposition, and morphological changes in the cuticle and cell wall preceded the appearance of wax crystals. These studies demonstrated that wax production and microsomal fatty acid elongation activities were induced within a defined and identifiable region of the expanding leek leaf and provide the foundation for future molecular studies.     

Seasonal Variation in the Hormone Content of Willow: II. Effect of Photoperiod on Growth and Abscisic Acid Content of Trees under Field Conditions

Levels of abscisic acid were followed in the xylem sap, mature leaves, and apices of field-grown willow (Salix viminalis L.) during the summer months, under natural and artificially extended photoperiods. Although the long day treatment prevented the general onset of dormancy, the plants grown under natural daylengths showed lower concentration of abscisic acid than those kept under long days.     

Enzymic synthesis of caffeoylglucaric Acid from chlorogenic Acid and glucaric Acid by a protein preparation from tomato cotyledons

The phenylpropane metabolism of tomato (Lycopersicon esculentum Mill) cotyledons was investigated. The HPLC analysis revealed two hydroxycinnamic-acid conjugates as major components, identified as chlorogenic acid (5-O-caffeoylquinic acid) and caffeoylglucaric acid (2-O- or 5-O-caffeoyl-glucaric acid). Quantitative analyses indicated a precursor-product relationship between the chlorogenic and caffeoylglucaric acids. Protein preparations from tomato cotyledons were found to catalyze the formation of caffeoylglucaric acid with chlorogenic acid as acyl donor and free glucaric acid as acceptor molecule. This enzyme activity, possibly to be classified as hydroxycinnamoylquinic acid:glucaric acid hydroxycinnamoyltransferase, acts together with hydroxycinnamoyl-CoA: quinic acid hydroxycinnamoyltransferase.     

Relationship between Mefluidide Treatment and Abscisic Acid Metabolism in Chilled Corn Leaves

Mefluidide, N-(2,4-dimethyl-5[([trifluoromethyl]sulfonyl) amino] phenyl)acetamide, a synthetic plant growth regulator, was capable of triggering an increase in endogenous free abscisic acid content when corn (Zea mays L.) plants were grown in a nonstress, day/night, temperature regime (26°C) with sufficient moisture supply. The relevance of such an abscisic acid increase prior to chilling exposure and the water relations during chilling are discussed in reference to the mefluidide protection of the chilled corn plants.     

Metabolic Engineering of a Novel Muconic Acid Biosynthesis Pathway via 4-Hydroxybenzoic Acid in Escherichia coli

cis,cis-Muconic acid (MA) is a commercially important raw material used in pharmaceuticals, functional resins, and agrochemicals. MA is also a potential platform chemical for the production of adipic acid (AA), terephthalic acid, caprolactam, and 1,6-hexanediol. A strain of Escherichia coli K-12, BW25113, was genetically modified, and a novel nonnative metabolic pathway was introduced for the synthesis of MA from glucose. The proposed pathway converted chorismate from the aromatic amino acid pathway to MA via 4-hydroxybenzoic acid (PHB). Three nonnative genes, pobA, aroY, and catA, coding for 4-hydroxybenzoate hydrolyase, protocatechuate decarboxylase, and catechol 1,2-dioxygenase, respectively, were functionally expressed in E. coli to establish the MA biosynthetic pathway. E. coli native genes ubiC, aroF^FBR, aroE, and aroL were overexpressed and the genes ptsH, ptsI, crr, and pykF were deleted from the E. coli genome in order to increase the precursors of the proposed MA pathway. The final engineered E. coli strain produced nearly 170 mg/liter of MA from simple carbon sources in shake flask experiments. The proposed pathway was proved to be functionally active, and the strategy can be used for future metabolic engineering efforts for production of MA from renewable sugars.     

Prostate Apoptotic Induction and NFκB Suppression by Dammarolic Acid: Mechanistic Insight into Onco-Therapeutic Action of an Aglycone Asiaticoside

Prostate cancer (PCa) is addressed as the second most common form of onco-threat worldwide and is usually considered as the major cause of mortality in men. Recent times have seen a surge in exploration of plant-derived components for alternative therapeutical interventions against different oncological malignancies. Dammarolic acid or Asiatic acid (AsA) is an aglycone asiaticoside that has been reported for its efficacy in several ailments including cancer. The current study aimed to investigate the anti-proliferative potency of AsA against human prostate cancer PC-3 cells. Purified AsA was diluted and PC-3 cells were exposed to 20, 40, and 80 µM concentration and incubated for 24 h. Post-exposure, PC-3 cells showcased a substantial loss of their viability at 20 µM (p < 0.05), moreover, this reduction in cell viability escalated proportionally with an increase in AsA at concentrations of 40 and 80 µM (p < 0.01; p < 0.001) respectively. AsA-impelled loss of cellular viability was also evident from the acridine orange-stained photomicrographs, which was also used to quantify the viable and apoptotic cells using Image J software. Additionally, quantification of ROS within PC-3 cells also exhibited an increase in DCF-DA-mediated fluorescence intensity post-exposure to AsA in a dose-dependent manner. AsA-induced apoptosis in PC-3 cells was shown to be associated with augmented activity of caspase-3 proportionally to the AsA concentrations. Thus, initially, this exploratory study explicated that AsA treatment leads to anti-proliferative effects in PC-3 cells by enhancing oxidative stress and inciting apoptosis en route to onset of nuclear fragmentation.     

Exogenous Isoleucine and Fatty Acid Shortening Ensure the High Content of Anteiso-C15:0 Fatty Acid Required for Low-Temperature Growth of Listeria monocytogenes

Previous studies have demonstrated that the branched-chain fatty acid anteiso-C_15:0 plays a critical role in the growth of Listeria monocytogenes at low temperatures by ensuring sufficient membrane fluidity. Studies utilizing a chemically defined minimal medium revealed that the anteiso fatty acid precursor isoleucine largely determined the fatty acid profile and fatty acid response of the organism to lowered growth temperature. When isoleucine was sufficient, the fatty acid profile was very uniform, with anteiso fatty acids comprising up to 95% of total fatty acid, and the major fatty acid adjustment to low temperature was fatty acid chain shortening, which resulted in an increase of anteiso-C_15:0 solely at the expense of anteiso-C_17:0. When isoleucine was not supplied, the fatty acid profile became more complex and was readily modified by leucine, which resulted in a significant increase of corresponding iso fatty acids and an inability to grow at 10°C. Under this condition, the increase of anteiso-C_15:0 at low temperature resulted from the combined effect of increasing the anteiso:iso ratio and chain shortening. A branched-chain α-keto acid dehydrogenase-defective strain largely lost the ability to increase the anteiso:iso ratio. Cerulenin, an inhibitor of β-ketoacyl-acyl carrier protein synthase (FabF), induced a similar fatty acid chain shortening as low temperature did. We propose that the anteiso precursor preferences of enzymes in the branched-chain fatty acid biosynthesis pathway ensure a high production of anteiso fatty acids, and cold-regulated chain shortening results in a further increase of anteiso-C_15:0 at the expense of anteiso-C_17:0.