Wound-induced green leaf volatiles cause the release of acetylated derivatives and a terpenoid in maize

Green leaf volatiles (GLVs), generally occurring C6 alcohols, aldehydes and acetates from plants, play an important role in plant-plant communication. These compounds induce intact plants to produce jasmonic acid, and induce defense-related gene expression and the release of volatile compounds. Here, we address wound-induced GLVs cause the release of acetylated derivatives and a terpenoid, (E)-4,8-dimethylnona-1,3,7-triene (DMNT) in intact maize, which may be a type of plant-plant interaction mediated by airborne GLVs. Upon exposure of intact maize seedlings to wound-induced GLVs, (Z)-3-hexenyl acetate was consistently the most abundant compound released. Exogenous application of individual alcohols and aldehydes mostly resulted in the release of corresponding acetate esters. C6-alcohols with a double bond between the second and third, or the third and fourth carbon atoms, C5- or C6-aldehydes, and (Z)-3-hexenyl acetate triggered the release of DMNT. When (Z)-3-hexenyl acetate and hexyl acetate were used to treat maize seedlings, they were recovered from the plants. These data demonstrated that: (1) apart from direct adsorption and re-release of acetate esters, absorption and conversion of exogenous alcohols and aldehydes into acetate esters occurred, and (2) DMNT was induced by a range of aldehydes and unsaturated alcohols.     

A detailed identification study on high-temperature degradation products of oleic and linoleic acid methyl esters by GC-MS and GC-FTIR

GC-MS and GC-FTIR were complementarily applied to identify oxidation compounds formed under frying conditions in methyl oleate and linoleate heated at 180°C. The study was focused on the compounds that originated through hydroperoxide scission that remain attached to the glyceridic backbone in fats and oils and form part of non-volatile molecules. Twenty-one short-chain esterified compounds, consisting of 8 aldehydes, 3 methyl ketones, 4 primary alcohols, 5 alkanes and 1 furan, were identified. In addition, twenty non-esterified volatile compounds, consisting of alcohols, aldehydes and acids, were also identified as major non-esterified components. Furanoid compounds of 18 carbon atoms formed by a different route were also identified in this study. Overall, the composition of the small fraction originated from hydroperoxide scission provides a clear idea of the complexity of the new compounds formed during thermoxidation and frying.     

Hydroperoxy-arachidonic acid mediated n-hexanal and (Z)-3- and (E)-2-nonenal formation in Laminaria angustata

In higher plants, C6 and C9 aldehydes are formed from C18 fatty acids, such as linoleic or linolenic acid, through formation of 13- and 9-hydroperoxides, followed by their stereospecific cleavage by fatty acid hydroperoxide lyases (HPL). Some marine algae can also form C6 and C9 aldehydes, but their precise biosynthetic pathway has not been elucidated fully. In this study, we show that Laminaria angustata, a brown alga, formed C6 and C9 aldehydes enzymatically. The alga forms C9 aldehydes exclusively from the C20 fatty acid, arachidonic acid, while C6 aldehydes are derived either from C18 or from C20 fatty acid. The intermediates in the biosynthetic pathway were trapped by using a glutathione/glutathione peroxidase system, and subjected to structural analyses. Formation of (S)-12-, and (S)-15-hydroperoxy arachidonic acids [12(S)HPETE and 15(S)HPETE] from arachidonic acid was confirmed by chiral HPLC analyses. These account respectively for C9 aldehyde and C6 aldehyde formation, respectively. The HPL that catalyzes formation of C9 aldehydes from 12(S)HPETE seems highly specific for hydroperoxides of C20 fatty acids.     

Determination of oil oxidation by an aldehyde-requiring mutant of luminous bacteria

A simple, rapid bioluminescence test (BT) for the determination of lipid oxidation is described. The test utilizes an aldehyde-requiring dark mutant of Vibrio harveyi (M42) that emits light in the presence of long chain (C₈-C₁₆) aliphatic aldehydes. The procedure consists of treating the oil or fat with Co²⁺ ion in ethanolic medium at alkaline pH. This treatment facilitates the decomposition of the hydroperoxides into long-chain aldehydes, part of which is used by the bacteria to produce light. The test was evaluated with corn, soybean and safflower oils, and shows excellent correlation with the commonly used peroxide value assay.     

Acetic Acid Bacteria: Physiology and Carbon Sources Oxidation

Acetic acid bacteria (AAB) are obligately aerobic bacteria within the family Acetobacteraceae, widespread in sugary, acidic and alcoholic niches. They are known for their ability to partially oxidise a variety of carbohydrates and to release the corresponding metabolites (aldehydes, ketones and organic acids) into the media. Since a long time they are used to perform specific oxidation reactions through processes called “oxidative fermentations”, especially in vinegar production. In the last decades physiology of AAB have been widely studied because of their role in food production, where they act as beneficial or spoiling organisms, and in biotechnological industry, where their oxidation machinery is exploited to produce a number of compounds such as l-ascorbic acid, dihydroxyacetone, gluconic acid and cellulose. The present review aims to provide an overview of AAB physiology focusing carbon sources oxidation and main products of their metabolism.     

Antimicrobial Browning-Inhibitory Effect of Flavor Compounds in Seaweeds

Since ancient times, the antimicrobial properties of seaweeds have been recognized. However, antimicrobial activities of volatile compounds in seaweeds have not been explored so far. Here, essential oils from seaweeds including green, brown and red algae such as Laminaria japonica, Kjellmaniella crassifolia, Gracilaria verrucosa and Ulva pertusa were prepared by using SDE (simultaneous distillation and extraction) apparatus. Volatile compounds in the essential oils were identified as aldehydes, ketones, carboxylic acids, alcohols and hydrocarbons by comparison of GC-retention times and MS data with those of authentic specimens. Flavor compounds such as (3Z)-hexenal, (2E)-hexenal and (2E)-nonenal in some essential oils showed strong antimicrobial activities against Escherichia coli TG-1, and Erwinia carotovora. Inhibition of browning can be achieved during either of two stages, namely, oxidation reaction by tyrosinase or subsequent non-enzymatic polymerization. Tyrosinase activity was measured by monitoring absorbance at 475 nm originating from dopachrome formed from L-DOPA. Many kinds of aliphatic carboxylic acids, aldehydes and alcohols were used as inhibitors for PPO activity. The results indicated that the α,β-unsaturated carbonyl compounds strongly inhibit tyrosinase activity. When seaweeds are damaged or macerated, the α,β-unsaturated aldehydes such as (2E)-hexenal and (2E)-nonenal are biosynthesized via the corresponding (3Z)-unsaturated aldehydes from linolenic acid and arachidonic acid. The flavor compounds that are formed could be valuable as safe antimicrobial browning-inhibitory agents of edible seaweed origin.     

In vitro antifungal, anti-elastase and anti-keratinase activity of essential oils of Cinnamomum-, Syzygium- and Cymbopogon-species against Aspergillus fumigatus and Trichophyton rubrum

This study was aimed to evaluate effects of certain essential oils namely Cinnamomum verum, Syzygium aromaticum, Cymbopogon citratus, Cymbopogon martini and their major components cinnamaldehyde, eugenol, citral and geraniol respectively, on growth, hyphal ultrastructure and virulence factors of Aspergillus fumigatus and Trichophyton rubrum. The antifungal activity of essential oils and their major constituents was in the order of cinnamaldehyde>eugenol>geraniol=C. verum>citral>S. aromaticum>C. citratus>C. martini, both in liquid and solid media against T. rubrum and A. fumigatus. Based on promising antifungal activity of eugenol and cinnamaldehyde, these oils were further tested for their inhibitory activity against ungerminated and germinated conidia in test fungi. Cinnamaldehyde was found to be more active than eugenol. To assess the possible mode of action of cinnamaldehyde, electron microscopic studies were conducted. The observations revealed multiple sites of action of cinnamaldehyde mainly on cell membranes and endomembranous structures of the fungal cell. Further, test oils were also tested for their anti-virulence activity. More than 70% reduction in elastase activity was recorded in A. fumigatus by the oils of C. verum, C. martini, eugenol, cinnamaldehyde and geraniol. Similar reduction in keratinase activity in A. niger was recorded for the oils of C. martini and geraniol. Maximum reduction (96.56%) in elastase activity was produced by cinnamaldehyde whereas; geraniol caused maximum inhibition (97.31%) of keratinase activity. Our findings highlight anti-elastase and anti-keratinase activity of above mentioned essential oils as a novel property to be exploited in controlling invasive and superficial mycoses.     

A single amino acid determines the catalytic efficiency of two alkenal double bond reductases produced by the liverwort Plagiochasma appendiculatum

Alkenal double bond reductases (DBRs) catalyze the NADPH-dependent reduction of the α,β-unsaturated double bond of many secondary metabolites. Two alkenal double bond reductase genes PaDBR1 and PaDBR2 were isolated from the liverwort species Plagiochasma appendiculatum. Recombinant PaDBR2 protein had a higher catalytic activity than PaDBR1 with respect to the reduction of the double bond present in hydroxycinnamyl aldehydes. The residue at position 56 appeared to be responsible for this difference in enzyme activity. The functionality of a C56 to Y56 mutation in PaDBR1 was similar to that of PaDBR2. Further site-directed mutagenesis and structural modeling suggested that the phenol ring stacking between this residue and the substrate was an important determinant of catalytic efficiency.     

Role of K binding residues in stabilization of heme spin state of Leishmania major peroxidase

The endogenous cation in peroxidases may contribute to the type of heme coordination. Here a series of ferric and ferrous derivatives of wild-type Leishmania major peroxidase (LmP) and of engineered K⁺ site mutants of LmP, lacking potassium cation binding site, has been examined by electronic absorption spectroscopy at 25 °C. Using UV–visible spectrophotometry, we show that the removal of K⁺ binding site causes substantial changes in spin states of both the ferric and ferrous forms. The spectral changes are interpreted to be, most likely, due to the formation of a bis-histidine coordination structure in both the ferric and ferrous oxidation states at neutral pH 7.0. Stopped flow spectrophotometric techniques revealed that characteristics of Compound I were not observed in the K⁺ site double mutants in the presence of H₂O₂. Similarly electron donor oxidation rate was two orders less for the K⁺ site double mutants compared to the wild type. These data show that K⁺ functions in preserving the protein structure in the heme surroundings as well as the spin state of the heme iron, in favor of the enzymatically active form of LmP.     

Applications of chromic acid-Celite columns to micro- and semimicro preparations of fatty aldehydes.

In a convenient two-phase procedure, columns of Celite 545 charged with aqueous chromic acid are used to oxidize microgram and milligram amounts of fatty alcohols to the corresponding aldehydes with approximately a 70% yield. The original configuration and position of the double bond is fully maintained during the oxidation of unsaturated alcohols.     

New and general synthesis of beta-C-glycosylformaldehydes from easily available beta-C-glycosylpropanones

A highly effective method for the introduction of a formyl group at the anomeric position of pyranosides was developed via enolisation of beta-C-D-glycopyranosylpropan-2-one using thermodynamic conditions then oxidative cleavage of the more substituted double bond. This sequence affords the desired aldehydes that are conveniently protected as aminals for purification and storage and easily regenerated using Dowex resin H+. In this paper, the syntheses of nine differently protected aldehydes derived from d-glucose, d-galactose, lactose and N-acetyl-d-glucosamine are presented. Our strategy proved to be very efficient in most cases excepted in the D-mannose series.     

Mouse hepatic microsomal oxidation of aliphatic aldehydes (C8 to C11) to carboxylic acids.

Addition of saturated and alpha, beta-unsaturated aliphatic aldehydes (C8 to C11) significantly increased NADPH oxidation with mouse hepatic microsomes, and the aldehydes themselves were oxidized to the corresponding carboxylic acids. When these aldehyde substrates were incubated similarly under oxygen-18 gas and the carboxylic acids formed were analyzed by GC-MS after methylation, it was indicated that oxygen-18 was significantly incorporated into the carboxylic acids formed from alpha, beta-unsaturated aldehydes, but not significantly into the carboxylic acids formed from saturated aldehydes. These results indicate that enzyme and/or mechanism responsible for the oxidation of these two types of aldehydes is different from each other.     

Raman spectroscopic monitoring of Lactarius latex

Lactarius is a genus of Basidiomycotina with mainly agaricoid representatives, which are characterised by the excretion of a typical milky fluid. In particular, the colour, changes and taste of this latex-like milk are often used as a taxonomically important character. When it is exuded several chemical reactions occur. To date, NMR spectroscopy is generally used for chemical investigation of this latex. However, as a vibrational spectroscopic technique Raman spectroscopy has several advantageous properties for this type of research. The aim of this study is to investigate whether Raman spectroscopy can be used as an alternative analytical technique to monitor the chemical reactions in Lactarius latex. Therefore, this paper presents the first Raman spectra of Lactarius latex and provides an interpretation of the Raman bands that are present. L. lacunarum latex spectra are thoroughly investigated by 2D correlation analysis and are compared with latex spectra of other species (L. chrysorrheus, L. deterrimus, L. fluens, L. glyciosmus and L. salmonicolor).     

Teratogenic actions of thermally-stressed culinary oils in rats

Lipid oxidation products (LOPs), generated in culinary oils during episodes of thermal stressing can give rise to cellular damage. The aims of this study were to determine whether orally-administered, LOP-containing thermally-stressed safflower oil exerts teratogenic actions in rats, and whether this effect could be prevented by co-administration of alpha-tocopherol (alpha-TOH). Safflower oil was heated for a period of 20 min according to standard frying practices and stored at -20 degrees C under N2. Four experimental groups of pregnant Wistar rats were employed; two received 0.30 ml of pre-heated oil (HO), one of which was also supplemented with 150 mg of alpha-TOH (HOE), and two served as controls, one treated with the non-heated oil (O) and the other without any treatment (C). The oil was administered daily by gavage from day 1 of pregnancy to day 11.5, when the animals were killed and the embryos examined. LOPs and alpha-TOH were determined both in the heated and non-heated oils. The percentage of embryo malformations and reabsorptions were determined in the above four experimental groups. Heating the oil substantially increased its concentration of LOPs and decreased its alpha-TOH content. The percentage of embryo malformations in the HO group was 21.73%, compared with 5.6 and 7% in the O and C groups, respectively. Supplementation of the pre-heated oil with alpha-TOH was found to decrease the percentage of malformations to 7%. The results obtained from these investigations indicate that LOPs detectable at millimolar levels in the heated cooking oils administered (e.g. saturated and alpha,beta-unsaturated aldehydes, and/or their conjugated hydroperoxydiene precursors) exert potent teratogenic actions in experimental animals which are at least partially circumventable by co-administration of the chain-breaking antioxidant alpha-TOH. Plausible mechanisms for these processes and their health relevance to humans regarding diet and methods of frying/cooking are discussed.     

Lipid Unsaturation Properties Govern the Sensitivity of Membranes to Photoinduced Oxidative Stress

Unsaturated lipid oxidation is a fundamental process involved in different aspects of cellular bioenergetics; dysregulation of lipid oxidation is often associated with cell aging and death. To study how lipid oxidation affects membrane biophysics, we used a chlorin photosensitizer to oxidize vesicles of various lipid compositions and degrees of unsaturation in a controlled manner. We observed different shape transitions that can be interpreted as an increase in the area of the targeted membrane followed by a decrease. These area modifications induced by the chemical modification of the membrane upon oxidation were followed in situ by Raman tweezers microspectroscopy. We found that the membrane area increase corresponds to the lipids’ peroxidation and is initiated by the delocalization of the targeted double bonds in the tails of the lipids. The subsequent decrease of membrane area can be explained by the formation of cleaved secondary products. As a result of these area changes, we observe vesicle permeabilization after a time lag that is characterized in relation with the level of unsaturation. The evolution of photosensitized vesicle radius was measured and yields an estimation of the mechanical changes of the membrane over oxidation time. The membrane is both weakened and permeabilized by the oxidation. Interestingly, the effect of unsaturation level on the dynamics of vesicles undergoing photooxidation is not trivial and thus carefully discussed. Our findings shed light on the fundamental dynamic mechanisms underlying the oxidation of lipid membranes and highlight the role of unsaturations on their physical and chemical properties.     

Raman Spectroscopy Analysis of Botryococcene Hydrocarbons from the Green Microalga Botryococcus braunii

Botryococcus braunii, B race is a unique green microalga that produces large amounts of liquid hydrocarbons known as botryococcenes that can be used as a fuel for internal combustion engines. The simplest botryococcene (C₃₀) is metabolized by methylation to give intermediates of C₃₁, C₃₂, C₃₃, and C₃₄, with C₃₄ being the predominant botryococcene in some strains. In the present work we have used Raman spectroscopy to characterize the structure of botryococcenes in an attempt to identify and localize botryococcenes within B. braunii cells. The spectral region from 1600–1700 cm⁻¹ showed ν(C=C) stretching bands specific for botryococcenes. Distinct botryococcene Raman bands at 1640 and 1647 cm⁻¹ were assigned to the stretching of the C=C bond in the botryococcene branch and the exomethylene C=C bonds produced by the methylations, respectively. A Raman band at 1670 cm⁻¹ was assigned to the backbone C=C bond stretching. Density function theory calculations were used to determine the Raman spectra of all botryococcenes to compare computed theoretical values with those observed. The analysis showed that the ν(C=C) stretching bands at 1647 and 1670 cm⁻¹ are actually composed of several closely spaced bands arising from the six individual C=C bonds in the molecule. We also used confocal Raman microspectroscopy to map the presence and location of methylated botryococcenes within a colony of B. braunii cells based on the methylation-specific 1647 cm⁻¹ botryococcene Raman shift.     

Characterization of active site residues of nitroalkane oxidase

The flavoenzyme nitroalkane oxidase catalyzes the oxidation of primary and secondary nitroalkanes to the corresponding aldehydes and ketones plus nitrite. The structure of the enzyme shows that Ser171 forms a hydrogen bond to the flavin N5, suggesting that it plays a role in catalysis. Cys397 and Tyr398 were previously identified by chemical modification as potential active site residues. To more directly probe the roles of these residues, the S171A, S171V, S171T, C397S, and Y398F enzymes have been characterized with nitroethane as substrate. The C397S and Y398 enzymes were less stable than the wild-type enzyme, and the C397S enzyme routinely contained a substoichiometric amount of FAD. Analysis of the steady-state kinetic parameters for the mutant enzymes, including deuterium isotope effects, establishes that all of the mutations result in decreases in the rate constants for removal of the substrate proton by ∼5-fold and decreases in the rate constant for product release of ∼2-fold. Only the S171V and S171T mutations alter the rate constant for flavin oxidation. These results establish that these residues are not involved in catalysis, but rather are required for maintaining the protein structure.     

Long-chain bases in the sphingolipids of atherosclerotic human aorta

Long-chain bases were prepared from human aorta sphingomyelin by a combined enzymatic hydrolysis-alkaline hydrolysis procedure and these bases were isolated by thin-layer chromatography. Aldehydes, obtained from the long-chain bases by periodate oxidation, were converted to 1,3-dioxolane derivatives. Dioxolanes were identified and quantified by gas-liquid chromatography before and after catalytic hydrogenation, and before and after separation into saturated, monoene, and diene dioxolane fractions. The monoene dioxolanes were converted to aldehydes by reductive ozonolysis with dimethyl sulfide and these aldehydes were isolated and identified as dioxolane derivatives. The double bond positions in the major diene component were established by reductive ozonolysis and permanganate-periodate oxidation. Sphingenines in the cerebroside-sulfatide and sulfatide fractions of aorta were converted to aldehydes by the reductive ozonolysis of intact sphingolipids and these aldehydes were analyzed as the dioxolanes. Human aorta sphingomyelin contained significant amounts of 4-hexadecasphingenine, 4-heptadecasphingenine, sphinganine, 4-sphingenine, and 4,x14-sphingadienine. Small amounts of hexadecasphinganine, 4-tetradecasphingenine, a sphingadienine isomer, an unknown sphinganine, and two unknown diene long-chain bases were also found in sphingomyelin. The presence of a branched-chain 4-sphingenine was tentatively established and the possible presence of a sphingenine isomer was suggested. The major sphingenines were the same in the sphingomyelin, sulfatide, and cerebroside-sulfatide fractions of human aorta.     

Essential Oils,Silver Nanoparticles and Propolis as Alternative Agents Against Fluconazole Resistant Candida albicans,Candida glabrata and Candida krusei Clinical Isolates

Development of effective and safe therapeutic treatment of fungal infections remains one of the major challenge for modern medicine. The aim of presented investigation was to analyze the in vitro antifungal activity of selected essential oils, ethanolic extracts of propolis and silver nanoparticles dropped on TiO₂ against azole-resistant C. albicans (n = 20), C. glabrata (n = 14) and C. krusei (n = 10) clinical isolates. Among tested essential oils, the highest activity has definitely been found in the case of the oil isolated from the bark of Cinnamomum cassia, with MIC and MFC values for all tested strains in the range of 0.0006–0.0097 % (v/v) and 0.0012–0.019 % (v/v), respectively. High activity was also observed for the Lemon, Basil, Thyme, Geranium and Clove (from buds) essential oils. Significant differences in fungicidal activity have been observed in the case of four tested propolis samples. Only one of them revealed high activity, with MFC values in the range from 0.156 to 1.25 % (v/v). Satisfactory fungicidal activity, against C. albicans and C. glabrata isolates, was also observed in the case of silver nanoparticles, however C. krusei isolates were mostly resistant. We also revealed that constituents of most of essential oils and propolis as well as silver nanoparticles are not substrates for drug transporters, which belong to the most important factors affecting resistance of Candida spp. clinical isolates to many of conventional antimycotics. To conclude, the results of our investigation revealed that essential oils, propolis and silver nanoparticles represent high potential for controlling and prevention candidiasis.