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Unusually severe food poisoning from vanilla slices.

Thirty six people suffered from severe vomiting and diarrhoea 15 min to 3 h after eating vanilla slices from the same bakery. Five patients were admitted to hospital, and one developed unusual skin lesions after admission. Staphylococcus aureus was isolated in large numbers from vanilla slices of the same batch as those giving rise to symptoms, and from five faecal specimens obtained from affected persons. Bacillus cereus and Bacillus subtilis were also isolated from the slices. Unbaked custard provides an ideal environment for bacterial multiplication, especially when (as on this occasion) the ambient temperature is persistently high.

Adult↗

Bioactive aromatic compounds from leaves and stems of Vanilla fragrans.

Alcoholic extracts of leaves and stems of Vanilla fragrans were fractionated with ethyl acetate and aqueous butanol. All three fractions of ethyl acetate, butanol, and water were screened for toxic bioactivity against mosquito larvae. The results of these experiments showed that the fractions from the ethyl acetate and butanol phases were both active in the bioassay. Bioactivity of the ethyl acetate fraction was found to be much greater than that from the butanol fraction in mosquito larvae toxicity. The water phase appeared to contain no substances that impaired mosquito larval growth. Repeated column chromatography of the ethyl acetate fraction on silica gel led to the isolation of 4-ethoxymethylphenol (1), 4-butoxymethylphenol (2), vanillin (3), 4-hydroxy-2-methoxycinnamaldehyde (4), and 3,4-dihydroxyphenylacetic acid (5). Compounds 4 and 5 were isolated from Vanilla species for the first time and 2 has not been reported to have been found in a natural form. 4-Ethoxymethylphenol (1) was the predominant compound, but 4-butoxymethylphenol (2) showed the strongest toxicity to mosquito larvae. The structures of the compounds were determined on the basis of their mass spectra and (1)H or (13)C NMR data.

Animals↗

Novel chemistry of abdominal defensive glands of nymphalid butterfly Agraulis vanillae.

Abdominal defensive glands of both sexes of the Gulf fritillary butterfly, Agraulis vanillae (Linnaeus) (Nymphalidae:Heliconiinae) emit a pronounced odor when disturbed. We have identified 6-methyl-5-hepten-2-one; oleic, palmitic, and stearic esters of the corresponding alcohol 6-methyl-5-hepten-2-ol; hexadecyl acetate; 1,16-hexadecanediol diacetate; and 1,15-hexade-canediol diacetate in the glandular exudate. Since we have determined that free-flying birds or birds in a butterfly conservatory discriminate against A. vanillae as prey, we suggest that the constituents in the glands may play a defensive role against potential avian predators.

Abdomen↗

Enhancement of sweetness ratings of aspartame by a vanilla odor presented either by orthonasal or retronasal routes.

When taste stimuli are presented with specific odor stimuli, the perceived intensity of taste is enhanced, a phenomenon called odor-induced taste enhancement. There is a possibility, however, that the odor substances might have stimulated the taste receptors in the oral cavity as well as odor receptors in the nasal cavity because the odor substances were dissolved in the taste solutions in some preceding studies. Schifferstein and Verlegh (1996) found that the odor-induced taste enhancement effect was not found when the subjects wore a nose clip to prevent the olfactory perception. Thus, it was suggested that the odor-induced taste enhancement did not result from the stimulation of receptors in the oral cavity. To confirm and extend their study, we presented the odor stimuli simultaneously with, but not dissolved in, the taste stimuli with a more advanced approach to stimulus presentation. The participants reported enhancement of sweetness ratings for aspartame when the taste stimuli were presented with a vanilla odor. This odor induced taste enhancement was found when the gaseous odor stimuli were presented either by the retronasal route or by the orthonasal route. There was little possibility that the vanilla odor stimulated the taste receptors during the orthonasal stimulation because the odor stimuli were presented directly into the nasal cavity. Thus, we could show that the odor-induced taste enlancement is elicited by olfactory perception. These results also suggested that there is little functional difference between retronasal and orthonasal olfaction.

Administration, Oral↗

Characterization of a multifunctional methyltransferase from the orchid Vanilla planifolia.

The final enzymatic step in the synthesis of the flavor compound vanillin (4-hydroxy-3-methoxybenzaldehyde) is believed to be methylation of 3,4-dihydroxybenzaldehyde. We have isolated and functionally characterized a cDNA that encodes a multifunctional methyltransferase from Vanilla planifolia tissue cultures that can catalyze the conversion of 3,4-dihydroxybenzaldehyde to vanillin, although 3,4-dihydroxybenzaldehyde is not the preferred substrate. The higher catalytic efficiency of the purified recombinant enzyme with the substrates caffeoyl aldehyde and 5-OH-coniferaldehyde, and its tissue distribution, suggest this methyltransferase may primarily function in lignin biosynthesis. However, since the enzyme characterized here does have 3,4-dihydroxybenzaldehyde-O-methyltransferase activity, it may be useful in engineering strategies for the synthesis of natural vanillin from alternate sources.

Amino Acid Sequence↗

Evolution of novel O-methyltransferases from the Vanilla planifolia caffeic acid O-methyltransferase.

The biosynthesis of many plant secondary compounds involves the methylation of one or more hydroxyl groups, catalyzed by O-methyltransferases (OMTs). Here, we report the characterization of two OMTs, Van OMT-2 and Van OMT-3, from the orchid Vanilla planifolia Andrews. These enzymes catalyze the methylation of a single outer hydroxyl group in substrates possessing a 1,2,3-trihydroxybenzene moiety, such as methyl gallate and myricetin. This is a substrate requirement not previously reported for any OMTs. Based on sequence analysis these enzymes are most similar to caffeic acid O-methyltransferases (COMTs), but they have negligible activity with typical COMT substrates. Seven of 12 conserved substrate-binding residues in COMTs are altered in Van OMT-2 and Van OMT-3. Phylogenetic analysis of the sequences suggests that Van OMT-2 and Van OMT-3 evolved from the V. planifolia COMT. These V. planifolia OMTs are new instances of COMT-like enzymes with novel substrate preferences.

Amino Acid Sequence↗

beta-Glucosidase and peroxidase stability in crude enzyme extracts from green beans of Vanilla planifolia Andrews.

The extraction method for beta-glucosidase from green vanilla beans has been studied. The effect of storage of green beans and protein extracts on beta-glucosidase and peroxidase activity was investigated: the best method, resulting in the highest enzyme activities, particularly for glucosidase, was through extraction of very fresh green beans in the presence of BisTris propane buffer at pH 8. The best method for storage of the extracts was at -80 degrees C after addition of 15% glycerol, when over 90% of initial activity was still present. Peroxidase activity did not change in frozen beans or in frozen extracts.

Orchidaceae↗

[A simple method for the detection of staphylococcal enterotoxin type B in vanilla custard using the ELISA (author's transl)].

The ELISA for the detection of staphylococcal enterotoxin type B (SEB) was employed to demonstrate SEB in Dutch Vanilla custard. Due to the sensitivity of the ELISA the extraction procedure, which is necessary when the Ouchterlony test is used, can be abbreviated to a great extent. Two successive extractions at pH 7.4 and pH 4.5 followed by a concentration (1:20) was sufficient to detect 0.1 mcg SEB in 100 g custard.

Enterotoxins↗

Simultaneous determination of the major acidic metabolites of catecholamines and serotonin in urine by liquid chromatography with electrochemical detection after a one-step sample clean-up on Sephadex G-10; influence of vanilla and banana ingestion.

A simple method is described for the simultaneous determination of vanilmandelic acid (VMA), 3,4-dihydroxyphenylacetic acid (DOPAC), 5-hydroxyindole-3-acetic acid (5-HIAA) and homovanillic acid (HVA) in urine. The compounds are isolated by a one-step sample clean-up on Sephadex G-10, separated by ion-pair reversed-phase liquid chromatography and detected electrochemically. A single analysis is completed within 65 min. Sample clean-up did not cause losses of the compounds of interest. The detection limits in urine were 0.4, 0.8, 1.0 and 1.6 mumol/l for VMA, DOPAC, 5-HIAA and HVA, respectively. 3,4-Dihydroxymandelic acid and vanillic acid (VA) were also detectable, but, under the chromatographic conditions used, they were not resolved from interfering components. VA and 5-HIAA could be analysed separately in the Sephadex G-10 eluate if more restrictive sampling conditions were used. Ingestion of bananas caused an increase of VMA, DOPAC, 5-HIAA and HVA in 24-h urine. After ingestion of vanilla an increased excretion of VA was observed, while the excretion of VMA, DOPAC and HVA was unaffected.

3,4-Dihydroxyphenylacetic Acid↗

An investigation of the bacteriological quality of retail vanilla slices.

One hundred and thiry-three vanilla slices, purchased from shops in the West Yorkshire Metropolitan County, were examined to determine the numbers and types of bacteria present at the time of purchase. The surface colony count at 37 degrees C was greater than 10(3)/g in 67/133 (50%) of the samples examined, Bacillus cereus being found at that concentration in 21.8%, coliform bacilli including E. coli in 5.3%, Staphylococcus aureus in 3-0% and Streptococcus faecalis in 0-8%. Thirty-four strains of B. cereus were serotyped and 11 (32%) of these were typable with the sera available. Preparation of custard mixed in the laboratory suggests that the milk or milk powder used in the mix may be the major source of B. cereus in the final product. Many of the present methods of manufacture, distribution and storage allow organisms present in the custard at manufacture the opportunity to multiply and possibly reach numbers which present a risk of food poisoning.

Bacillus cereus↗

Long-chain gamma-pyrones in epicuticular wax of two vanilla bean species: V. fragrans and V. tahitensis.

Analyses of the neutral lipids from Vanilla species before saponification resulted in the identification of a new product family in this genus: long-chain gamma-pyrone compounds with an aliphatic chain containing a cis double bond at the n-9 position. These compounds represent 7-8% of the neutral lipids in mature beans. Using NMR and gas chromatography/mass spectrometry, three gamma-pyrones have been identified, including 2-(10-nonadecenyl)-2, 3-dihydro-6-methyl-4H-pyran-4-one, 2-(12-heneicosenyl)-2, 3-dihydro-6-methyl-4H-pyran-4-one, and 2-(14-tricosenyl)-2, 3-dihydro-6-methyl-4H-pyran-4-one. The major constituent was 2-(14-tricosenyl)-2,3-dihydro-6-methyl-4H-pyran-4-one, which represented 70.3% of the gamma-pyrone fraction. The variability of this compound family has been studied in relation to bean maturity in V. fragrans and V. tahitensis beans. This compound family has not been found either in leaves or in stems or in V. madagascariensis beans.

Flavoring Agents↗

Identification of phosphoenolpyruvate carboxylase isoforms in leaf, stem and roots of the obligate CAM plant Vanilla planifolia Salib. (Orchidaceae): a physiological and molecular approach.

This study provides the first comparative analysis of phosphoenolpyruvate carboxylase isoforms (PEPc; EC 4.1.1.31) in an obligate crassulacean acid metabolism (CAM) plant, Vanilla planifolia Salisb. (Orchidaceae). Nocturnal CO2 fixation and malate accumulation by the leaves and the green stem show that these organs perform CAM. The chloroplast-containing aerial roots, however, exhibit C3 photosynthesis. The catalytic activity of PEPc was highest in the leaves compared with the stem and aerial roots. The Km (PEP) and Ki (malate) were similar in the PEPc extracted from leaf and aerial roots, and significant higher in stem. cDNA was obtained from those tissues and also from the soil-grown roots, and various cDNA clones were detected and amplified by means of RT-PCR and RACE-PCR. The amino-acid sequences of the PEPc isoforms deduced from the cDNA showed a great degree of homology, and Southern blot analysis suggests that the encoding genes form a small multigene family of at least two members. One PEPc isoform (PpcV1) is assumed to be related to CAM because, as shown by northern blot analysis, it is mainly expressed in the CAM-performing organs, i.e. in the leaves and the stem. A further isoform (PpcV2) was identified in the soil-grown roots and aerial roots, but northern blots show that to some extent PpcV2 is also expressed in the leaf and the stem tissues. Thus, it is assumed that PpcV2 encodes the housekeeping isoform of PEPc. Altogether, the present study provides support in favour of the view that isoforms of PEPc are related to specific functions.

Amino Acid Sequence↗

Interspecific differentiation and hybridization in vanilla species (Orchidaceae)

Vanilla claviculata, V. barbellata and V. dilloniana are distributed throughout the Caribbean islands and are all found in Puerto Rico. The vegetative parts of the species are similar; however, their conspicuous flowers easily distinguish them. Electrophoresis of seven polymorphic enzymes revealed that the genetic composition of the three species is also very similar: they deviate mainly from each other in allele frequencies rather than by specific alleles. A hierarchical analysis of genetic differentiation showed that the between-species component is slightly higher (FSG=0.237) than the component between populations within species (FPS=0.141). Nevertheless, they are efficiently recognized by their genotypic compositions. In V. barbellata and V. claviculata 97-99% of all individuals were assigned to the correct species. Assignment to a wrong species occurred only with individuals at localities where species coexist. This suggests that the species may hybridize.

Journal Article↗

Phenylpropanoid Metabolism in Suspension Cultures of Vanilla planifolia Andr. : III. Conversion of 4-Methoxycinnamic Acids into 4-Hydroxybenzoic Acids.

Feeding of 4-methoxycinnamic acid, 3,4-dimethoxycinnamic acid and 3,4,5-trimethoxycinnamic acid to cell suspension cultures of Vanilla planifolia resulted in the formation of 4-hydroxybenzoic acid, vanillic acid, and syringic acid, respectively. The homologous 4-methoxybenzoic acids were demethylated to the same products. It is concluded that the side chain degrading enzyme system accepts the 4-methoxylated substrates while the demethylation occurs at the benzoic acid level. The demethylating enzyme is specific for the 4-position. Feeding of [O-(14)C-methyl]-3,4-dimethoxycinnamic acid revealed that the first step in the conversion is the glycosylation of the cinnamic acid to its glucose ester. A partial purification of a UDP-glucose: trans-cinnamic acid glucosyltransferase is reported. 4-Methoxy substituted cinnamic acids are better substrates for this enzyme than 4-hydroxy substituted cinnamic acid. It is suggested that 4-methoxy substituted cinnamic acids are intermediates in the biosynthetic conversion of cinnamic acids to benzoic acids in cells of V. planifolia.

Journal Article↗

Phenylpropanoid Metabolism in Suspension Cultures of Vanilla planifolia Andr. : II. Effects of Precursor Feeding and Metabolic Inhibitors.

Feeding of cinnamic acid and ferulic acid to non-treated and chitosan-treated cell suspension cultures of Vanilla planifolia resulted in the formation of trace amounts of p-hydroxy benzoic acid (5.2 micrograms per gram fresh weight of cells) and vanillic acid (6.4 micrograms per gram fresh weight of cells), respectively. Addition of a 4-hydroxycinnamate: CoA-ligase inhibitor, 3,4-(methylenedioxy)-cinnamic acid (MDCA), resulted in a reduced biosynthesis of ligneous material with a simultaneous significant increased vanillic acid formation (around 75 micrograms per gram fresh weight of cells). A K(1) of 100 micromolar for 4-hydroxycinnamate: CoA-ligase in a crude preparation was estimated for this inhibitor. It is suggested that the conversion of cinnamic acids into benzoic acids does not involve cinnamoyl CoA esters as intermediates. Feeding of (14)C-cinnamic acid and (14)C-ferulic acid to cells treated with MDCA indicate that cinnamic acid, but not ferulic acid, is a precursor of vanillic acid in these cultivated cells of V. planifolia.

Journal Article↗

Phenylpropanoid Metabolism in Suspension Cultures of Vanilla planifolia Andr. : IV. Induction of Vanillic Acid Formation.

Kinetin is used as an elicitor to induce vanillic acid formation in cell suspension cultures of Vanilla planifolia. Maximal induction is observed at a kinetin concentration of 20 micrograms per gram of fresh weight of cells. Vanillic acid synthesis is observed a few hours after elicitation. The effects of kinetin on the activity of some enzymes of the phenylpropanoid pathway, i.e. phenylalanine ammonia-lyase, 4-hydroxycinnamate:coenzyme A ligase and uridine 5'-diphosphate-glucose:trans-cinnamic acid glucosyltransferase, are reported and compared to the effects of chitosan. The former two enzymes are induced by chitosan with a maximum activity of approximately 25 to 40 hours after elicitation. All three enzymes are induced by kinetin with maximum activities for phenylalanine ammonia lyase and 4-hydroxycinnamate:coenzyme A ligase at approximately 50 hours after induction, whereas maximum glucosyltransferase activity is seen already after 24 hours. Furthermore, both elicitors induced the formation of lignin-like material, whereas only kinetin induced vanillic acid biosynthesis. Finally, kinetin but not chitosan induces catechol-4-O-methyltransferase activity, catalyzing the formation of 4-methoxycinnamic acids, which were shown to be intermediates of hydroxybenzoic acid biosynthesis within cells of V. planifolia. It is suggested that this methyltransferase is directly involved in the biosynthesis of vanillic acid.

Journal Article↗

The "vanilla fudge" cyst.

A white cyst, partially covered by jet black splotches and streaks, had a surface appearance of vanilla fudge ice cream. Histologically, the cyst was lined with two distinctly different types of follicular epithelium--one identical to that of the infundibulum and the other to that of the matrix. The black pigment was melanin, produced by numerous dendritic melanocytes that were situated only in the region of the epithelium-resembling matrix.

Adult↗

[Microbiological quality of vanilla ice cream manufactured in Caracas, Venezuela].

A total of 122 samples of vanilla ice cream, the base product used for all flavors, prepared by eight different large firms at the Metropolitan Area of Caracas, Venezuela, were analyzed for aerobic mesophilic and psicrophilic bacteria, Staphylococcus aureus, Enterobacteriaceae, and Filamentous fungi. Findings revealed that within the sampling, 56.6% complied with the international standards proposed for aerobic mesophilic bacteria, 68% for Staphylococcus aureus, and 23% for Enterobacteriaceae. Three serotypes of enteropathogenic Escherichia coli, one of Salmonella, and one of Shigella were found. Ten genera of Filamentous fungi were isolated and identified.

Bacteria, Aerobic↗