Long-chain aliphatic beta-diketones from epicuticular wax of Vanilla bean species. Synthesis of nervonoylacetone

Analysis of the neutral lipids from Vanilla fragrans and Vanilla tahitensis (Orchidaceae) without saponification resulted in the isolation and identification of a new product family in this plant: beta-dicarbonyl compounds. The compound structures are composed of a long aliphatic chain with 2,4-dicarbonyl carbons and a cis double bond at the n-9 position. They represent approximately 28% of the neutral lipids, that is, 1.5%, in immature beans, and approximately 10% of the neutral lipids, that is, 0.9%, in mature beans. Using retention indices, gas chromatography-mass spectrometry, derivatization reactions, and chemical degradation, five beta-dicarbonyl compounds have been identified including 16-pentacosene-2,4-dione, 18-heptacosene-2,4-dione, 20-nonacosene-2, 4-dione, 22-hentriacontene-2,4-dione, and 24-tritriacontene-2, 4-dione. Among them (Z)-18-heptacosene-2,4-dione, or nervonoylacetone, has been synthesized in two steps starting from nervonic acid. The major constituent, nervonoylacetone, represented 74.5% of the beta-dicarbonyl fraction. The range of these compounds has been studied in relation with bean maturity for V. fragrans and V. tahitensis species. This compound family has not been found in the leaves or stems of any of the three vanilla species studied and is markedly absent in the beans of V. madagascariensis.     

Characterization of key chromophores formed by nonenzymatic browning of hexoses and L-alanine by using the color activity concept

Thermal treatment of an aqueous solution of D-glucose and L-alanine in the presence of the carbohydrate degradation product furan-2-aldehyde resulted in the formation of a variety of colored compounds, among which (Z)-2-[(2-furyl)methylidene]-5, 6-di(2-furyl)-6H-pyran-3-one (I), [E]- and [Z]-1, 2-bis(2-furyl)-1-pentene-3,4-dione (IIa/IIb), 4, 5-bis(2-furyl)-2-methyl-3H-furan-2-one (III), and (S,S)- and (S, R)-2-[4, 5-bis(2-furyl)-2-hydroxy-2-methyl-3(2H)-pyrrol-1-yl]propionic acid (IVa/IVb) as well as 2-[(2-furyl)methylidene]-4-hydroxy-5-[(E)-(2-furyl)methylidene]methyl -2H-furan-3-one (V) were successfully identified as the most intense by application of the color dilution analysis. To measure the contribution of these colorants to the overall color of the browned Maillard mixture, color activity values were calculated as the ratio of the concentration to the visual detection threshold of each colorant. By application of this color activity concept, 16.0% of the overall color of the Maillard mixture accounted for these five types of colorants, thus confirming them as key chromophores. On the basis of synthetic model experiments, the formation pathways leading to the chromophores IIa/IIb, III, and IVa/IVb were proposed.     

Analysis of furanone, pyranone, and new heterocyclic colored compounds from sugar-glycine model Maillard systems

Aqueous sugar (xylose or glucose)-glycine model systems were refluxed for 2 h with the pH maintained at 5. Reverse-phase HPLC of the total reaction products gave two resolved peaks (one of which was colored) for the xylose system and five resolved peaks (two of which were colored) for the glucose system. The components responsible for these peaks were isolated from the ethyl acetate extracts by semipreparative HPLC. Using mainly NMR, the colored compound from the xylose system was identified as the new 2-acetyl-6-(hydroxymethyl)-5,6-dihydro-4H-pyridinone. The colored compounds from the glucose system were most likely to be two novel cis/trans ring isomers of the related new compound 2-acetyl-6-hydroxy-7-(hydroxymethyl)-1,5,6,7-tetrahydro-4H-azepinone+ ++. These compounds are the first one-ring structures isolated from sugar-amino acid model systems that are reported to be colored. Two of the colorless components of the glucose system were identified, mainly by NMR experiments, as the related compounds 4-hydroxy-2-(hydroxymethyl)-5-methyl-3(2H)-furanone and 2, 3-dihydro-3,5-dihydroxy-6-methyl-4H-pyranone. The remaining compound from the glucose system and the colorless compound from the xylose system were identified as 5-(hydroxymethyl)furfural and 4-hydroxy-5-methyl-3(2H)-furanone, respectively.     

General food semiochemicals attract omnivorous German cockroaches, Blattella germanica

Stale beer and peanut butter are effective baits for the German cockroach (GCRs), Blattella germanica (L.) (Dictyoptera: Blattellidae). In still-air arena olfactometer experiments it was previously shown that headspace volatile extracts of peanut butter and solvent extract of beer attract male GCRs. The objective of this study was to identify the semiochemicals that mediate attraction of GCRs to these sources. Coupled gas chromatographic-electroantennographic detection (GC-EAD) and GC-mass spectrometric (MS) analyses of these attractive extracts, or fractions thereof, and of synthetic standards revealed many candidate semiochemicals. Elaborate olfactometer experiments determined that 1-hexanol from peanut butter, and ethanol and 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP) from beer, are the key semiochemicals of these food sources. 1-Hexanol is a well-known headspace volatile of decomposing lipids, ethanol conveys food fermentation, and DDMP with a caramel-type flavor has been found in several types of food. By responding to these rather general food-derived compounds, the omnivorous GCRs appear to exploit semiochemicals that indicate the presence of various food components, such as lipids and carbohydrates. Synthetic equivalents of these semiochemicals may be formulated as baits or be added to, and thus enhance the attractiveness of, natural food sources as trap or insecticidal baits.     

Volatile flavor components of stored nonfat dry milk.

Nonfat dry milk (NDM) is widely used both as an ingredient in other preparations and for direct consumption. Flavor quality of NDM is a critical parameter because it can directly impact final product quality. Flavors can be formed in NDM during subsequent storage. Identification of compounds responsible for storage-induced flavors is necessary to correlate sensory quality with potential sources of the flavors. Six NDM samples were selected for volatile flavor analysis based on sensory analysis and storage time. Volatile components were extracted by direct solvent extraction/high vacuum distillation. Volatile extracts were separated into neutral/basic and acidic fractions and analyzed by gas chromatography-olfactometry (GCO) and aroma extract dilution analysis (AEDA). A variety of aldehydes, ketones, and free fatty acids were responsible for generation of flavors in stored NDM. The following compounds exhibited high aroma impact by AEDA: 3-(methylthio)propanal (boiled potato); o-aminoacetophenone (corn tortilla); 2,5-dimethyl-4-hydroxy-3(2H)-furanone and 2-methyl-3-hydroxy-4H-pyran-4-one (burnt sugar); butanoic acid (cheesy); pentanoic acid (sweaty); acetic and hexanoic acids (sour/vinegar); octanoic, decanoic, and dodecanoic acids (waxy); p-cresol (cowy/barny); 3-methylindole (fecal); dimethyl trisulfide (cabbage); (E,E)-2,4-decadienal (fried/fatty); furfuryl alcohol (rubber/vitamin); phenylacetic acid (rose-like); and 1-octen-3-one (mushroom).     

Quantitative model studies on the efficiency of precursors in the formation of cooling-active 1-pyrrolidinyl-2-cyclopenten-1-ones and bitter-tasting cyclopenta-[b]azepin-8(1H)-ones

The yields of the cooling-active compounds 3-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one (1) and 5-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one (2) as well as the bitter tastants 7-methyl-2,3,6,7-tetrahydrocyclopenta-[b]azepin-8(1H)-one (3) and 7-methyl-2,3,4,5,6,7-hexahydrocyclopenta-[b]azepin-8(1H)-one (4) obtained by heating mixtures of possible Maillard-type precursors in model systems varying in temperature, pH value, or water content were determined quantitatively. The results showed that hexose-derived cyclotene is the common precursor for all four tastants and that the formation of each individual tastant is strongly determined by the structure of the nitrogen-containing precursor, e.g., reaction of cyclotene with pyrrolidine formed by thermal decarboxylation of L-proline produced the cooling compounds 1 and 2 only, whereas in the presence of 1-pyrroline formed upon Strecker reactions of L-proline, the bitter tasting azepinone 3 was produced exclusively. In contrast, the structure of the secondary amino acid L-proline enabled the formation of compound 4, whereas the pyrrolidine and 1-pyrroline, respectively, do not generate this tastant. In addition, a nonvolatile, tasteless intermediate, (S)-3-methyl-2-[(2'-carboxy)-1-pyrrolidinyl]-2-cyclopenten-1-one (5), was isolated from the cyclotene/L-proline reaction mixture and could be confirmed as an efficient precursor for the cooling compound 1. The data, obtained by these studies, are the scientific basis to tailor the desired overall flavor of foods by means of a more controlled Maillard-type technology.     

Quantification of the group B soyasaponins by high-performance liquid chromatography

High-performance liquid chromatographic methods were developed for the isolation and quantitative determination of the group B soyasaponins, including 2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP)-conjugated soyasaponins alphag, betag, and betaa, and their non-DDMP counterparts, soyasaponins V, I, and II, respectively, with formononetin used as the internal standard. The limits of quantification for soy products were 0.11-4.86 micromol/g. The within-day and between-days assay coefficients of variation were <9.8 and < 14.3%, respectively. The group B soyasaponin concentrations in 46 soybean varieties ranged from 2.50 to 5.85 micromol/g. Soy ingredients (soybean flour, toasted soy hypocotyls, soy protein isolates, textured vegetable protein, soy protein concentrates, and Novasoy) and soy foods (commercial soy milk, tofu, and tempeh) contained the group B soyasaponins from 0.20 to 114.02 micromol/g. There was no apparent correlation between isoflavone and soyasaponin concentrations in the soy products examined.     

Influence of pyrolytic and aqueous-phase reactions on the mechanism of formation of Maillard products

The influence of the reaction phase on the mechanism of formation of Maillard products was studied by comparison of (13)C-label incorporation patterns of the common products formed in model systems consisting of labeled glycine and D-glucoses subjected to both pyrolysis and heating in aqueous solutions. Pyrolysis experiments were performed at 250 degrees C for 20 s, and aqueous model systems were heated in sealed vials for 3 h at 120 degrees C followed by GC/MS analysis. Label incorporation patterns of the following compounds were analyzed: cyclotene, furanmethanol, acetylpyrrole, 5-methyl-pyrrole, trimethylpyrazine, acetic acid, 3-hydroxy-2-butanone, 2,3-butanedione, and 2-methyl-4, 5-dihydro-3(2H)-furanone. Although pyrolysis reaction produced higher number of products, however, the major pathways of formation of variety of important Maillard products followed the same mechanism under both pyrolytic and aqueous systems. Furthermore, contrary to literature speculations, 2-methyl-4, 5-dihydro-3(2H)-furanone was shown to be formed by ring contraction of 2,3-dihydro-3,5-dihydroxy-6-methyl-4(H)-pyran-4-one, through benzilic acid rearrangement, followed by decarboxylation.     

Identification of new heterocyclic nitrogen compounds from glucose-lysine and xylose-lysine maillard model systems

Aqueous sugar (glucose or xylose)-lysine model systems were heated at 80 degrees C for 6 h with the pH maintained at a predetermined value (3, 4, or 5). Selected compounds were isolated by combinations of solvent extraction and semipreparative HPLC, prior to identification by NMR and mass spectrometry. Two compounds were identified from the pH 5 glucose system and were identified as epsilon-[2-formyl-5-(hydroxymethyl)pyrrole-1-yl]-L-norleucine (pyrraline) and the new compound, 1-(5-carboxy-5-aminopentyl)-2-formyl-3-(1,2,3-trihydroxypropyl)pyrrole. A third compound was partially characterized. 2-Acetyl-5-hydroxymethyl-5,6-dihydro-4H-pyridinone was identified in the pH 3 xylose system, and the new compound, 8-furan-2-yl-methyl-5-hydroxymethyl-5,6-dihydro-indolizine-1,7-dione, was identified in the pH 4 xylose system. 2-Furfurylidene-4-hydroxy-5-methyl-3(2H)-furanone was identified in both xylose systems. Mechanisms of formation are proposed for the novel compounds.     

Simultaneous quantitative analysis of maillard reaction precursors and products by high-performance anion exchange chromatography

A new analytical setup allowing the simultaneous analysis of precursors and products of the Maillard reaction is described. It is based on high-performance anion exchange chromatography with electrochemical (ECD) and diode array detectors (DAD) coupled in series. Chromatography and detection were optimized to permit simultaneous monitoring of compounds relevant to the Maillard reaction, such as the sugar, the amino acid, and the corresponding Amadori compound as well as the cyclic intermediates 5-(hydroxymethyl)-2-furaldehyde, maltol, and 2,3-dihydro-3,5-dihydroxy-6-methyl-4(H)-pyran-4-one. Separation was achieved on a CarboPac PA-1 column using a gradient of sodium acetate in aqueous sodium hydroxide. The Amadori compound, glucose, and glycine were monitored by an ECD operating in the integrated amperometry mode. The number of analyzed compounds was further increased by coupling the ECD with a DAD for the analysis of ultraviolet-active constituents. This method was successfully applied to model Maillard reaction mixtures based on glucose and glycine.     

Maillard reaction of D-glucose: identification of a colored product with conjugated pyrrole and furanone rings

Formation of colored compounds during the Maillard reaction of D-glucose with butylammonium acetate in aqueous solution has been investigated. Butylamine was used as a model compound analogous to the lysine side chains of proteins. The previously unknown, yellow product, 4-hydroxy-5-methyl-2-(N-butyl-3-hydroxy-5-(2-hydroxyethyl)pyrrolyl-2-methylidene)-2H-furan-3-one (1a), was isolated from the reaction mixtures and identified by spectroscopic data.     

Effect of selected phytotoxins from Guanomyces polythrix on the calmodulin-dependent activity of the enzymes cAMP phosphodiesterase and NAD-kinase

The effect of a series of phytotoxins isolated from the fungus Guanomyces polytrix on calmodulin (CaM)-dependent nicotinamide adenine dinucleotide kinase (NADK) and CaM-dependent cyclic nucleotide phosphodiesterase (PDE) activities was investigated. The results indicated that (2S,3S)-5-hydroxy-6,8-dimethoxy-2,3-dimethyl-4H-2,3-dihydronaphtho[2,3-b]-pyran-4-one, (2S,3S)-5-hydroxy-6,8,10-trimethoxy-2,3-dimethyl-4H-2,3-dihydro-naphtho[2,3-b]-pyran-4-one, (2S,3R)-5-hydroxy-6,8-dimethoxy-2,3-dimethyl-2,3-dihydro-4H-naphtho[2,3-b]-pyran-4-one, (2S,3R)-5-hydroxy-6,8,10-trime-thoxy-2,3-dimethyl-2,3-dihydro-4H-naphtho[2,3-b]-pyran-4-one, 5-hydro-xy-6,8-dimethoxy-2,3-dimethyl-4H-naphtho[2,3-b]-pyran-4-one, rubrofusarin B, and ergosta-4,6,8(14),22-tetraen-3-one inhibited the activation of both target enzymes in the presence of CaM. On the other hand, (2S)-5-hydroxy-6,8-dimethoxy-2-methyl-4H-2,3-dihydronaphtho[2,3-b]-pyran-4-one and (2S)-5-hydroxy-6,8,10-trimethoxy-2-methyl-4H-2,3-dihydronaphtho-[2,3-b]-pyran-4-one inhibited the activation of PDE and the basal activity of NADK. Thus, these phytotoxins are CaM inhibitors and may exert their phytotoxic action by inhibiting the CaM-dependent process, although they could also interfere with other cellular metabolic phenomena. This is the first report of the use of the NADK assay to detect or quantify CaM inhibitors, and it could be a valuable tool for studying those CaM isoforms regulating NADK.     

Structure determination of a novel 3(6H)-pyranone chromophore and clarification of its formation from carbohydrates and primary amino acids

An intensely orange compound, which has recently been evaluated as one of the main colored compounds formed in Maillard reactions of hexoses, could be unequivocally identified as (Z)-2-[(2-furyl)methylidene]-5,6-di(2-furyl)-6H-pyran-3-one (1) by application of several NMR and LC-MS experiments. To clarify its formation, the effectiveness of certain carbohydrate degradation products as precursors of 1 was studied in a quantitative experiment demonstrating hydroxy-2-propanone, furan-2-aldehyde, and 3-deoxy-2-hexosulose as precursors of the colorant. Site-specific labeling experiments with D-1-[(13)C]glucose and D-6-[(13)C]glucose, respectively, were performed to elucidate the formation pathway of 1 involving a cleavage of the hexose skeleton between carbon atoms C(5) and C(6). In addition, pentoses could be shown to generate 1 via a similar formation pathway involving the 3-deoxy-2-pentosulose.     

Viridepyronone, a new antifungal 6-substituted 2H-pyran-2-one produced by Trichoderma viride

A new antifungal 6-substituted 2H-pyran-2-one, named viridepyronone, has been isolated from a cultural filtrate of a strain of Trichoderma viride showing antagonistic activity in vitro toward Sclerotium rolfsii, which is the causal agent of crown and stem rot of artichoke. Viridepyronone was characterized as 6-(4-oxopentyl)-2H-pyran-2-one 2 with spectroscopic methods. Bioassays showed that viridepyronone had a good antifungal activity against S. rolfsii, and its minimum inhibitory concentration (over 90% inhibition) was found to be 196 microg/mL. This is the first report of viridepyronone produced by any species of fungi.     

Development of a stable isotope dilution assay for the quantification of 5-methyl-(E)-2-hepten-4-one: application to hazelnut oils and hazelnuts.

A stable isotope dilution assay was developed for the quantitation of the hazelnut odorant 5-methyl-(E)-2-hepten-4-one by mass chromatography using synthesized [(2)H](2)-5-methyl-(E)-2-hepten-4-one as the internal standard. Application of the method on two batches of commercial hazelnut oils, processed from either roasted or unroasted nuts, revealed 6.4 microg 5-methyl-(E)-2-hepten-4-one per kg of unroasted oil whereas 315.8 microg per kg was determined in the roasted nut oil. The about 50-fold higher amount of 5-methyl-(E)-2-hepten-4-one in roasted hazelnut oil suggested the necessity of a thermal treatment to generate the flavor compound. Pan frying of raw hazelnuts (9 to 15 min) or boiling of the crushed nut material for 1 h in water led to an increase of 5-methyl-(E)-2-hepten-4-one by factors of 600 and 800, respectively, thereby corroborating that the major part of the nut flavorant is formed during heat treatment from a yet unknown precursor in hazelnuts.     

Generation of Maillard compounds from inulin during the thermal processing of Agave tequilana Weber Var. azul

During the cooking process of Agave tequilana Weber var. azul to produce tequila, besides the hydrolysis of inulin to generate fermentable sugars, many volatiles, mainly Maillard compounds, are produced, most of which may have a significant impact on the overall flavor of tequila. Exudates (agave juice) from a tequila company were collected periodically, and color, Brix, fructose concentration, and reducing sugars were determined as inulin breakdown took place. Maillard compounds were obtained by extraction with CH(2)Cl(2), and the extracts were analyzed by GC-MS. Increments in color, Brix, and reducing sugars were observed as a function of time, but a decrease in fructose concentration was found. Many Maillard compounds were identified in the exudates, including furans, pyrans, aldehydes, and nitrogen and sulfur compounds. The most abundant Maillard compounds were methyl-2-furoate, 2,3-dihydroxy-3,5-dihydro-6-methyl-4(H)-pyran-4-one, and 5-(hydroxymethyl)furfural. In addition, a series of short- and long-chain fatty acids was also found. A large number of the volatiles in A. tequilana Weber var. azul were also detected in tequila extracts, and most of these have been reported as a powerful odorants, responsible for the unique tequila flavor.     

Systematic studies on structure and physiological activity of cyclic alpha-keto enamines, a novel class of "cooling" compounds.

3-Methyl- and 5-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one were recently identified as intense cooling compounds in roasted dark malt. To gain more insights into the molecular requirements of these compounds for imparting a cooling sensation, 26 cyclic alpha-keto enamine derivatives were synthesized, and their physiological cooling activities were evaluated. Any modification of the amino moiety, the carbocyclic ring size, or incorporation of additional methyl groups led to a significant increase of the cooling threshold. Insertion of an oxygen atom into the 2-cyclopenten-1-one ring, however, increased the cooling activity, e.g., the cooling threshold of the 5-methyl-4-(1-pyrrolidinyl)-3(2H)-furanone was found to be 16-fold below the threshold concentration determined for the 3-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one. Shifting the oxygen atom from the 4- into the 5-position of the cyclopentenone ring resulted in a even more drastic increase in cooling activity, e.g., the 4-methyl-3-(1-pyrrolidinyl)-2(5H)-furanone exhibited the strongest cooling effect at the low oral threshold concentration of 0.02-0.06 mmol/L, which is 35-fold below the value determined for (-)-menthol. In contrast to the minty smelling (-)-menthol, most of the alpha-keto enamines were found to be virtually odorless but impart a sensation of "cooling" to the oral cavity as well as to the skin, thus illustrating that there is no physiological link between cooling activity and mint-like odors.     

Anti-inflammatory activities of new succinic and maleic derivatives from the fruiting body of Antrodia camphorata

Six new compounds, trans-3-isobutyl-4-[4-(3-methyl-2-butenyloxy)phenyl]pyrrolidine-2,5-dione (1), trans-1-hydroxy-3-(4-hydroxyphenyl)-4-isobutylpyrrolidine-2,5-dione (2), cis-3-(4-hydroxyphenyl)-4-isobutyldihydrofuran-2,5-dione (3), 3-(4-hydroxyphenyl)-4-isobutyl-1H-pyrrole-2,5-dione (4), 3-(4-hydroxyphenyl)-4-isobutylfuran-2,5-dione (5), and dimethyl 2-(4-hydroxyphenyl)-3-isobutylmaleate (6), together with one known compound, 3-isobutyl-4-[4-(3-methyl-2-butenyloxy)phenyl]furan-2,5-dione (7), were isolated from the fruiting bodies of Antrodia camphorata. The structures of the compounds were elucidated by analysis of their spectroscopic data. To investigate the immunomodulatory potential of the compounds, RAW264.7 macrophage cells were treated with the compounds. Compound 1 significantly increased spontaneous TNF-alpha secretion from unstimulated RAW264.7 cells but suppressed IL-6 production [50% inhibition concentration value (IC50) = 10 microg/mL] in LPS-stimulated cells. Compounds 3, 4, and 6 also suppressed IL-6 production with IC50 values of 17, 18, and 25 microg/mL, respectively, suggesting that these four compounds may have an anti-inflammatory effect on macrophage-mediated responses. Of the six compounds, compound 1 was the most effective, exerting both immunostimulatory and anti-inflammatory effects.     

Syntheses and herbicidal activities of novel triazolinone derivatives

Protoporphyrinogen oxidase (Protox, EC 1.3.3.4) has been identified as one of the most important action targets of herbicides. To search for novel Protox inhibitors, a series of title compounds 1, 2, and 3 were designed and synthesized by introducing three types of pharmacophores, cyclic imide, phenylurea, and ( E)-methyl 2-methoxyimino-2- o-tolylacetate, into the scaffold of triazolinone. The bioassay results indicated that the resulting cyclic imide-type triazolinones 1 displayed much better herbicidal activities than phenylurea-type triazolinones 2. Most fortunately, compound 3, methyl 2-[3-methyl-(2-fluoro-4-chloro-5-ethylsulfonamidephenyl)-4,5-dihydro-5-oxo-1 H-1,2,4-triazol-4-yl]methylenephenyl-2-( E)-methoxyiminoacetate, was found to be the most promising candidate due to its comparable herbicidal activity at 75-150 g of active ingredient/ha with the commercial product sulfentrazone. On the basis of test results of herbicidal spectrum and crop selectivity, compound 3 could be developed as a postemergent herbicide used for the control of broadleaf weeds in rice fields.