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.     

Evidence for surfactant solubilization of plant epicuticular wax

The solubilization of isolated, reconstituted tomato (Lycopersicon esculentum Mill.) fruit and broccoli (Brassica oleracaea var. botrytis L.) leaf epicuticular waxes (ECW) by nonionic octylphenoxypolyethoxy ethanol surfactant (Triton X-100) was demonstrated in a model system by TLC and fluorescence analysis using pyrene as a fluorescent probe. ECW was solubilized at or above the surfactant critical micelle concentration; solubilization increased with an increase in micelle concentration. As shown by the fluorescence quenching of pyrene, surfactant solubilization of the ECW increased rapidly for the first 12 h, then approached a plateau, increased linearly with an increase in temperature (22--32 degrees C), and decreased linearly with the log of the polyoxyethylene chain length (range 5--40 oxyethylenes). These data are discussed in relation to surfactant effects on phytotoxicity and performance of foliar spray application of agrochemicals.     

Novel fatty acid esters of p-coumaryl alcohol in epicuticular wax of apple fruit

Hexane extracts of epicuticular wax from cv. Gala apples were noted to have an unusual, broad absorbance maximum at approximately 258 nm, which led us to isolate and identify the primary UV-absorbing compounds. Column and thin-layer chromatography yielded a fraction that gave a series of paired, 260-nm-absorbing peaks on C(18) HPLC. These were shown to be a family of phenolic fatty acid esters, for which retention times increased with increasing fatty acid chain length, and paired peaks were esters of two related phenolics with the same fatty acid moiety. Alkaline hydrolysis of the esters released two water-soluble phenolics separable by C(18) HPLC. Electrospray ionization mass spectrometry gave a molecular mass of 150 for both, and (1)H NMR plus UV absorbance spectra identified them as E and Z isomers of p-coumaryl alcohol. Alkaline cleavage of the fatty acid esters in the presence of methanol or ethanol resulted in partial derivatization of E-p-coumaryl alcohol to the corresponding gamma-O-methyl or O-ethyl ether. Gradient HMQC NMR of the HPLC-purified stearate ester of E-p-coumaryl alcohol indicated that fatty acid esterification occurs at the gamma-OH rather than at the 4-OH on the phenyl ring. This is the first report of fatty acid esters of monolignols as a natural plant product.     

Purification and characterization of vanilla bean (Vanilla planifolia Andrews) beta-D-glucosidase

Vanilla bean beta-D-glucosidase was purified to apparent homogeneity by successive anion exchange, hydrophobic interaction, and size-exclusion chromatography. The enzyme is a tetramer (201 kDa) made up of four identical subunits (50 kDa). The optimum pH was 6.5, and the optimum temperature was 40 degrees C at pH 7.0. K(m) values for p-nitrophenyl-beta-D-glucopyranoside and glucovanillin were 1.1 and 20.0 mM, respectively; V(max) values were 4.5 and 5.0 microkat.mg(-1). The beta-D-glucosidase was competitively inhibited by glucono-delta-lactone and 1-deoxynojirimycin, with respective K(i) values of 670 and 152 microM, and not inhibited by 2 M glucose. The beta-D-glucosidase was not inhibited by N-ethylmaleimide and DTNB and fully inhibited by 1.5-2 M 2-mercaptoethanol and 1,4-dithiothreitol. The enzyme showed decreasing activity on p-nitrophenyl-beta-D-fucopyranoside, p-nitrophenyl-beta-D-glucopyranoside, p-nitrophenyl-beta-D-galactopyranoside, and p-nitrophenyl-beta-D-xylopyranoside. The enzyme was also active on prunasin, esculin, and salicin and inactive on cellobiose, gentiobiose, amygdalin, phloridzin, indoxyl-beta-D-glucopyranoside, and quercetin-3-beta-D-glucopyranoside.     

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.     

Leaf epicuticular wax as a factor of antixenotic resistance of cabbage to cabbage flea beetles and cabbage stink bugs attack

The aim of present research was to establish the role of epicuticular wax content in eight cabbage genotypes (four white hybrids and one red hybrid, two red varieties and one white variety) in the context of its natural resistance to attack cabbage flea beetles (Phyllotreta spp.) and cabbage stink bugs (Eurydema spp.), which are among the most important cabbage pests in southern Europe. For this reason and for the purpose of diminishing the use of synthetic insecticides against the cabbage pests the field experiments in 2006 and 2008 were conducted. We found out that individual cabbage genotypes – they had different epicuticular wax content – differ in regard to their susceptibility to attacks by the studied groups of harmful insect pests. The highest susceptibility to attacks by Phyllotreta spp. was confirmed for the hybrid ‘Cheers F1’, in the first year (1.68 ± 0.05), as well as in the second year of the experiment (2.87 ± 0.13). Cabbage stink bugs in both years of the experiment caused the highest extent of injuries on the hybrids ‘Destiny F1’, ‘Cheers F1’, and ‘Vestri F1’. In both years we found higher epicuticular wax content in red cabbage genotypes. In almost all studied genotypes we found a pronounced negative correlation between the content of epicuticular wax and the extent of injuries done by both groups of harmful pests. We have established that epicuticular wax is an important factor of cabbage's antixenotic resistance to attacks by cabbage flea beetles and cabbage stink bugs, and that the cabbage genotypes with higher content of this substance are consequently more suitable for environmentally acceptable manners of cabbage production.     

Biological effects of epicuticular flavonoids from Primula denticulata on human leukemia cells

The biological effects of epicuticular substances in farinose exudates accumulated on inflorescence shafts and calyces of Primula denticulata on human acute myeloid leukemia cells (HL-60) were analyzed. The crude material possessed little antioxidative capacity but strong cytostatic properties. Some of its known components (5-hydroxyflavone, 2'-hydroxyflavone, 5,2'-dihydroxyflavone, and 5,8-dihydroxyflavone) were further tested to identify the biologically active compounds. The effects of these flavones on cell cycle progression, mitochondrial membrane potential, and reactive oxygen species have been investigated by flow cytometry. The flavonol quercetin was included in the study as reference compound because of its known cytostatic properties and its activity as radical scavenger. Compared to quercetin the flavones induced little apoptosis (up to 40 microM), but despite their low toxicity, the Primula flavonoids possessed strong cytostatic properties even at low concentrations. The cell cycle distribution showed a characteristic time-dependent shift, giving evidence of a generally short-lived effect of the test compounds in the exposed cells. The antioxidative properties quantified according to two different methods correlated with the number of hydroxyl groups. Whereas quercetin strongly affected the mitochondrial membrane potential, none of the Primula flavones showed a comparable effect.     

Properties of diphenolase from Vanilla planifolia (Andr.) shoot primordia cultured in vitro.

Properties of diphenolase (PPO, EC1.10.3.1) from vanilla (Vanilla planifolia Andr.) shoot primordia culture were investigated. Two pH optima of the enzyme extraction at pH 6 and 8 were found. Nevertheless, the enzymes shared the same optimum pH of activity-between pH 3 and 4. Sodium dodecyl sulfate slightly improved diphenolase extraction but caused a 3-fold increase in its specific activity. The extracts of pH 6 and 8.0 revealed three isozyme bands after polyacrylamide gel electrophoresis-two of them were similar in both extracts and two distinct. The enzyme showed high thermal stability-no loss was observed after 120 min at 50 degrees C. Diethyldithiocarbamic acid, ethylenediaminetetracetic acid disodium salt, ethylene glycol bis(beta-aminoethyl ether) N,N,N',N'-tetraacetic acid, L-ascorbic acid, dithiothreitol, glutathione (reduced), and beta-mercaptoethanol were found to be potent inhibitors of the diphenolase studied. The enzyme showed also monophenolase activity. Km and Vmax were calculated with monophenols [p-coumaric acid, 3-(p-hydroxyphenyl)propionic acid, 4-hydroxybenzyl alcohol, 4-hydroxybenzaldehyde, 4-hydroxybenzoic acid] and with diphenols (caffeic acid, hydrocaffeic acid, chlorogenic acid, 4-methylcatechol, protocatechuic aldehyde and acid, and 3,4-dihydroxyphenylalanine). The highest Vmax was found with 4-hydroxybenzyl alcohol and the greatest affinity to protocatechuic acid, respectively-the most abundant monophenol and one of the least abundant o-diphenols in the studied Vanilla tissue.     

Chemotypical variation in Vanilla planifolia Jack. (Orchidaceae) from the Puebla-Veracruz Totonacapan region

One of the threats in the diversity loss of the primary gene pool of Vanilla planifolia is the lack of information on existing level of polymorphism in cultivated germplasm, and the different expressions of this polymorphism. For this reason, it is proposed to study the chemical polymorphism of the four phytochemicals that define the vanilla aroma quality in fruits (vanillin, vanillic acid, p-hydroxybenzaldehyde, p-hydroxybenzoic acid) by HPLC analysis (High Performance Liquid Chromatography) of 25 collections of unknown genotype, grown in the region Totonacapan Puebla-Veracruz, Mexico. The results identified a selection process, domestication in fruit aroma of vanilla, during which increased the participation of vanillin and reduced the presence of three minor compounds (vanillic acid, p-hydroxybenzaldehyde and p-hydroxybenzoic acid) in the global aroma. We distinguished a total of six chemotypes of V. planifolia in the Totonacapan region, some chemotypes with wild aromatic characteristics (low participation of vanillin) related to the material less cultivated in the region and domesticated chemotypes with high participation of vanillin, for the most cultivated material. The results show that the diversification of the chemotypes of V. planifolia is not related to environmental variation. The data indicate that in the possible center of origin of vanilla, there is phytochemical polymorphism, which indirectly suggests the existence of genetic polymorphism, essential for the design of a breeding program for optimizing the use and conservation of diversity of the primary gene pool of Vanilla planifolia.     

Preparation of bean curds from protein fractions of six legumes.

Chickpeas, lentils, smooth peas, mung beans, and faba beans were milled into flours and fractionated to protein and starch fractions. Compositions of the seeds, cotyledons, and flours were compared for each legume and the weight and protein recovery of each fraction analyzed. Bean curds were prepared from the protein fractions through heat denaturation of protein milk, followed by coagulation with calcium sulfate or magnesium sulfate. The effect of chickpea protein concentration and coagulant dosage on the texture of bean curds was evaluated using a texture analyzer. Textural analysis indicated that curd prepared at 2.3-3.0% protein concentration and 1.5% CaSO(4) dosage had better yield and better texture than curds prepared under other conditions. Bean curds prepared from chickpeas and faba beans exhibited the second highest springiness and cohesiveness after those from soybeans. Curds of mung beans and smooth peas, on the other hand, had the highest yields and the highest moisture contents. The protein yield of the first and second soluble extracts used for curd preparation accounted for approximately 90% of the total protein of the seeds.     

Biotransformation of common bean (Phaseolus vulgaris L.) flavonoid glycosides by bifidobacterium species from human intestinal origin

Bifidobacteria strains from human origin were screened for the specific activity (beta-glucosidase activity) involved in the metabolism of dietary flavonoids. Five strains with high beta-glucosidase activity were selected for further metabolism analyses (high-performance liquid chromatography separations) of flavonoid glycosides occurring in Phaseolus vulgaris L. (common bean) seeds and seedlings. All selected strains were found to be active in the conversion of kaempferol 3-O-glucoside, daidzin, genistin, and glycitin into their aglyconic forms. No metabolites were detected after the fermentation tests with the diglucosidic compound kaempferol 3-O-xylosylglucoside. In addition, to verify the effective bioavailability of flavonoid aglycones, the degradation rates of daidzein, genistein, glycitein, and kaempferol, following incubation with selected strains, were monitored. The results showed that the five selected strains of bifidobacteria, being active in the biotranformation of flavonoid glycosides occurring in common bean seeds and seedlings, could be considered as probiotic dietary adjuncts to improve the nutritional and health properties of flavonoid-based products, comprising hypothetical common bean food derivatives.     

A genome-wide assessment of the genetic diversity,evolution and relationships with allied species of the clonally propagated crop Vanilla planifolia Jacks. ex Andrews

Genetic Resources and Crop Evolution - The Vanilla genus is a complex taxonomic group characterized by a vegetative reproduction mode combined with intra- and inter-specific hybridizations, and...     

Synthesis of an eco-friendly nanocomposite fertilizer for common bean based on carbon nanoparticles from agricultural waste biochar

The burning of agricultural waste is a major cause of environmental pollution. In this study, we sought to prepare biochar from agricultural waste as a source material for the preparation of carbon nanoparticles (CNPs). Surface morphology, hydrodynamic particle size, and purity and crystallinity of CNPs were extensively investigated using transmission electron microscopy (TEM), zeta sizing, and X-ray diffraction (XRD) spectroscopy, respectively. The CNPs were subsequently immersed in a solution of potassium nitrate (KNO³) to prepare a CNPs/NK nanocomposite (CNPs loaded with nitrogen (N) and potassium (K)) as a nanocomposite fertilizer for common bean (Phaseolus vulgaris L.). The CNPs/NK nanocomposite was sprayed as a foliar fertilizer at 0, 10, 20, 30, and 40 mg L^-1 on common bean plants 25 d after sowing on a farm in Shebin El-Kom, El-Monifia, Egypt. The growth, yield, and quality of common bean were investigated during two successive growing seasons (2017 and 2018). The highest seed yields of 2.04 and 2.01 t ha^-1 and the highest values of growth parameters including plant height of 61.5 and 59.2 cm, number of leaves per plant of 35 and 35, number of flowers per plant of 83.3 and 82.7, and plant fresh weight of 148.7 and 152.8 g plant^-1 were obtained when using the CNPs/NK nanocomposite at a concentration of 20 mg L^-1 during the 2017 and 2018 growing seasons, respectively.     

粉丝生产中提高绿豆淀粉收率的浸泡工艺研究

为了提高粉丝生产中豆类淀粉的收率,应用正交试验设计,研究不同浸泡剂在不同浸泡条件下对粉丝生产中绿豆淀粉分离、提取的影响,得到了提高淀粉收率的新方法和新工艺。研究表明：浸泡温度是影响淀粉收率的主要因素,实际生产宜取20～30℃的温度进行浸泡;≤0.2%的HCl溶液浸泡与传统的水浸泡(酸浆沉淀法)没有显著差异;0.2%NaHSO₃在30℃温度下浸泡48 h,可使淀粉收率从83.7%提高到91.6%～96.3%;0.04%NaOH液在室温(20℃左右)下浸泡48 h,可使淀粉收率提高到93.4%～98.3%;浸泡时间也是影响绿豆淀粉分离、提取的重要因素,以24～48 h为宜,亦可根据浸泡温度(T)与浸泡时间(t)的关系式t=905T-1.12(10℃≤T≤30℃)控制浸泡温度和时间。     

Nodulation of African yam bean (Sphenostylis stenocarpa) by Bradyrhizobium sp. isolated from Erythrina brucei

African yam bean (Sphenostylis stenocarpa), which is widely cultivated in Africa because of its growth capability on marginal soils, was nodulated by an endosymbiont (characterized and designed Bradyrhizobium sp. AUEB20) isolated from the Ethiopian tree Erythrina brucei with the formation of a small number of large, indeterminate N₂-fixing nodules. In contrast, 24 other isolates from Ethiopian woody legumes were ineffective. Strain AUEB20 promiscuously nodulated a number of tropical legumes, but none out of five European crop plants tested. Received: 17 September 1996     

Isozyme analysis of three species ofRoegneria C. Koch from China

To collect and exploitRoegneria genetic resources, isozyme variation on 7 different enzymes encoded by 26 presumptive loci were analyzed in leaf extracts ofR. kamoji, R. ciliaris, R. nakaii andTriticum aestivum cv. Chinese Spring (control species) using polyacrylamide gel electrophoresis. No isozyme polymorphism was detected within accession of the threeRoegneria species, all having self-pollinating reproduction. However, extensive variations at isozyme loci were observed among accessions ofR. kamoji — even among accessions distributed at different altitudes within the same collection area. These results suggested that: (1) a select few individuals collected at a collection site may be enough to represent the genetic variability of that population; and (2) collection and maintenance ofR. kamoji materials from different altitudes may be required to increase genetic diversity. Furthermore, the banded phenotypes at EST 1 and EST-2 loci may be used as a biochemical marker associated with a morphological character, pubescences covered at leaf margin inR. kamoji. The banded characters at EST- 1, SOD-1, SOD-2, and SOD-3a loci may be used as biochemical markers to identify theR. kamoji chromosomes carrying these loci in aT. aestivum ×R. kamoji hybridization program.     

Characterization of peptides formed during fermentation of cocoa bean.

Analysis by SDS-PAGE and GPC-MS of fermented cocoa extracts shows changes in the amount and composition of the major proteins, accompanied by formation of complex distributions of peptides. MS/MS studies and application of SEQUEST sequencing software have allowed identification of two related peptides, a hexapeptide and a nonapeptide, formed from vicilin, one of the cocoa storage proteins. Time course studies of the two peptides show different abundance profiles and indicate, in part, production of the hexapeptide from the nonapeptide.     

Genetic variation of five species of Trifolium L. from south-west Turkey

Following a legume collection mission to south-west Turkey in 1996, five species of Trifolium were analysed for genetic variation within and between species in eleven morphological and flowering characters. The five species included two outcrossing species, T. michelianum and T. resupinatum, and three inbreeding species, T. clypeatum, T. glomeratum and T. tomentosum. The genetic diversity found was related to climate and edaphic factors. All five species showed significant amounts of genetic differentiation between sites and the species could be separated morphologically by principal components analysis and cluster analysis. The most significant source of genetic variation was found to be related to geographical distribution with those species which were widely distributed across south-west Turkey exhibiting much greater amounts of genetic variation between sites, than those which had a narrow distribution. The breeding system was found to be less important, but only the morphology of the outbreeding species showed any environmental clines in relation to climate. A multiple regression analysis was computed to estimate the effect of growing season on the days to flowering of each of the species.     

Observations on the geographic distribution, ecology and conservation status of several Phaseolus bean species in Costa Rica

Interest in bean genetic resources of Central America has resumed because of disease pressures (e.g., web blight, BGMV) and limitations of current bean varieties. As most of the diversity in landraces has been explored, focus is now on the exploration of wild forms of the primary gene pool and wild species of the secondary gene pool. A germplasm collection was carried out in the field and resulted in the collection of 29 wild populations for six Phaseolus species; it complemented field work done in 1987. Nine more populations were found for P. costaricensis, 10 for wild P. lunatus, one for P. oligospermus, one for P. tuerckheimii, four for wild P. vulgaris and four for P. xanthotrichus. Ninety-three herbarium voucher specimens were collected for 19 populations of the six species (deposited at CR). These results confirm the presence of wild P. vulgaris on both slopes of the central valley of Costa Rica, namely in the life zones bh-MB and bmh-P, and of P. costaricensis in the life zone bmh-MB. These life zones of limited range in Costa Rica have been heavily modified, thus fully justifying the germplasm collection for ex situ conservation. For both species the range of distribution in Costa Rica has been almost completely sampled. The life characteristics of each species that are relevant for their conservation in situ are briefly reviewed. Distribution ranges of each wild bean species are compared with the present extension of national parks, protected areas and fauna/flora sanctuaries, and suggestions for expanding such protected areas are made.