Ontogenic profiling of glucosinolates, flavonoids, and other secondary metabolites in Eruca sativa (salad rocket), Diplotaxis erucoides (wall rocket), Diplotaxis tenuifolia (wild rocket), and Bunias orientalis (Turkish rocket)

As an influence of the Mediterranean diet, rocket species such as Eruca sativa L., Diplotaxis species, and Bunias orientalis L. are eaten all over the world at different ontogenic stages in salads and soups. They are all species within the plant order Capparales (glucosinolate-containing species), and all are from the family Brassicaceae. Predominantly, the leaves of these species are eaten raw or cooked, although Eruca flowers are also consumed. There is considerable potential with raw plant material for a higher exposure to bioactive phytochemicals such as glucosinolates, their hydrolysis products, and also phenolics, flavonoids, and vitamins such as vitamin C. These compounds are susceptible to ontogenic variation, and the few published studies that have addressed this topic have been inconsistent. Thus, an ontogenic study was performed and all samples were analyzed using a previously developed robust liquid chromatography/mass spectrometry method for the identification and quantification of the major phytochemicals in all tissues of the rocket species. Seeds and roots of both Eruca and Diplotaxis contained predominantly 4-methylthiobutylglucosinolate. Leaves of Eruca and Diplotaxis contained high amounts of 4-mercaptobutylglucosinolate with lower levels of 4-methylthiobutlyglucosinolate and 4-methylsulfinylbutylglucosinolate. Flowers of Eruca and Diplotaxiscontained predominantly 4-methylsulfinylbutyl-glucosinolate. In addition, roots of both Diplotaxisspecies contained 4-hydroxybenzylglucosinolate but 4-hydroxybenzylglucosinolate was absent from roots of Eruca. Seeds and seedlings of all Eruca contained N-heterocyclic compounds but no sinapine, whereas Diplotaxis contained sinapine but not the N-heterocycles. In all tissues of B. orientalis, 4-hydroxybenzylglucosinolate and 4-methylsulfinyl-3-butenylglucosinolate were predominant. All rocket tissues, except roots, contained significant levels of polyglycosylated flavonoids, with/without hydroxycinnamoyl acylation. The core aglycones were kaempferol, quercetin, and isorhamnetin. The exception was B. orientalis, which had a negligible seed flavonoid content as compared with the other species. Anthocyanins were only detected in Eruca flowers and consisted of a complex pattern of at least 16 different anthocyanins.     

Multivariate correlation between color and mineral composition of honeys and by their botanical origin

The mineral content and color characteristics of 77 honey samples were analyzed. Eighteen minerals were quantified for each honey. Multiple linear regression (MLR) was used to establish equations relating the colorimetric CIELAB coordinates to the mineral data. The results obtained shown that lightness (L) was significantly correlated with S, Ca, Fe, As, Pb, and Cd for the dark honey types (avocado, heather, chestnut, and honeydew). For the light and brown honey types (citrus, rosemary, lavender, eucalyptus, and thyme), C(ab) and b showed the lower correlation with the mineral content of the honeys; their regression functions involve a few independent variables (Mg and Al for b and only Al for C(ab)). Furthermore, by means of application of linear discriminant analysis to the mineral content, it was possible to obtain a model that classifies the honeys by their lightness. The prediction ability of the built model, determined with the test set method, was 85%.     

Quantitative high-performance liquid chromatography analyses of flavonoids in Australian Eucalyptus honeys

Flavonoids of nine Australian monofloral Eucalyptus honeys have been analyzed and related to their botanical origins. The mean content of total flavonoids varied from 1.90 mg/100 g of honey for stringybark (E. globoidia) honey to 8.15 mg/100 g of honey for narrow-leaved ironbark (E. crebra) honey, suggesting that species-specific differences occur quantitatively among these Eucalyptus honeys. All of the honey samples analyzed in this study have a common flavonoid profile comprising tricetin (5,7,3',4',5'-pentahydroxyflavone), quercetin (3,5,7,3',4'-pentahydroxyflavone), and luteolin (5,7,3',4'-tetrahydroxyflavone), which, together with myricetin (3,5,7,3',4',5'-hexahydroxyflavone) and kaempferol (3,5,7,4'-tetrahydroxyflavone), were previously suggested as floral markers for European Eucalyptus honeys. Thus, flavonoid analysis could be used as an objective method for the authentication of the botanical origin of Eucalyptus honeys. Moreover, species-specific differences can also be found in the composition of honey flavonoid profiles. Among these honeys, bloodwood (E. intermedia) honey contains myricetin and tricetin as the main flavonoid compounds, whereas there is no myricetin detected in yapunyah (E. ochrophloia), narrow-leaved ironbark (E. crebra), and black box (E. largiflorens) honeys. Instead, these types of Eucalyptus honeys may contain tricetin, quercetin, and/or luteolin as their main flavonoid compounds. Compared to honeys from other geographical origins, the absence or minor presence of propolis-derived flavonoids such as pinobanksin, pinocembrin, and chrysin in Australian honeys is significant. In conclusion, these results demonstrate that a common flavonoid profile exists for all of the Eucalyptus honeys, regardless of their geographical origins; the individual species-specific floral types of Eucalyptus honey so common in Australia could be possibly differentiated by their flavonoid profile differences, either qualitatively or quantitatively or both.     

Evaluation of the botanical origin of estonian uni- and polyfloral honeys by amino acid content

The free amino acid content of 61 honey samples from Estonia has been determined by HPLC-UV with precolumn derivatization with diethyl ethoxymethylenemalonate. Analyzed samples were seven types of unifloral honeys and polyfloral honeys. The main amino acids found in Estonian honeys were proline and phenylalanine. The resulting data have been analyzed by t test and principal component analysis (PCA). t Test revealed that some amino acids (alpha-alanine, beta-alanine, asparagine, gamma-aminobutyric acid, glutamine, glycine, histidine, ornithine, phenylalanine, proline, serine, and tryptophan) are more potent for assigning honey botanical origin than others. PCA enabled differentiation of some honey types by their botanical origin. In the space of the two first principal components, heather honeys form a cluster that is clearly separable from, for example, polyfloral honeys. It is concluded that analysis of the free amino acid profile may serve as a useful tool to assess the botanical origin of Estonian honeys.     

Identification and quantification of glucosinolates in sprouts derived from seeds of wild Eruca sativa L. (salad rocket) and Diplotaxis tenuifolia L. (wild rocket) from diverse geographical locations

The Brassicaceae rocket species Eruca sativa L. (salad rocket) and Diplotaxis tenuifolia L. (wild rocket) are consumed throughout the world in salads, predominantly the leaves but also the flowers and more recently the sprouts (seedlings). Ontogenic profiling of glucosinolates and flavonoids in plants derived from commercial seed of these species has previously been done, but no studies have been conducted to determine how geographical origin affects glucosinolate composition in rocket species. Seeds from wild E. sativa L. and D. tenuifolia L. from diverse regions of the world were obtained from gene banks and grown under controlled conditions. Sprouts were harvested when they would normally be harvested for consumption, and glucosinolates were extracted and profiled in these accessions. All of the sprouts from Italian E. sativa L. had consistently high total glucosinolate content, with only a few exceptions, and also the highest percentage contents of 4-mercaptobutylglucosinolate. In contrast, sprouts produced from Central and Eastern European seeds had a much higher percentage of 4-methylthiobutylglucosinolate. With a single exception, Tunisia, all sprouts produced from North African seeds had very high 4-methylthiobutylglucosinolate contents. The single sample from China had a high total glucosinolate content and glucosinolate profile that was very similar to the accessions from Uzbekistan and Pakistan. All of the D. tenuifolia L. sprouts had consistently high total glucosinolate contents, and a high percentage of this was 4-mercaptobutylglucosinolate. This glucosinolate variation in levels and profiles of the rockets can be used for genetic studies, selected breeding, and human intervention studies.     

Identification and quantification of antioxidant components of honeys from various floral sources

Little is known about the individual components of honey that are responsible for its antioxidant activity. The present study was carried out to characterize the phenolics and other antioxidants present in honeys from seven floral sources. Chromatograms of the phenolic nonpolar fraction of the honeys indicated that most honeys have similar but quantitatively different phenolic profiles. Many of the flavonoids and phenolic acids identified have been previously described as potent antioxidants. A linear correlation between phenolic content and ORAC activity was demonstrated (R(2) = 0.963, p < 0.0001). Honeys were separated by solid-phase extraction into four fractions for sugar removal and separation based on solubility to identify the relative contribution of each fraction to the antioxidant activity of honey. Antioxidant analysis of the different honey fractions suggested that the water-soluble fraction contained most of the antioxidant components. Specific water-soluble antioxidant components were quantified, including protein; gluconic acid; ascorbic acid; hydroxymethylfuraldehyde; and the combined activities of the enzymes glucose oxidase, catalase and peroxidase. Of these components, a significant correlation could be established only between protein content and ORAC activity (R(2) = 0.674, p = 0.024). In general, the antioxidant capacity of honey appeared to be a result of the combined activity of a wide range of compounds including phenolics, peptides, organic acids, enzymes, Maillard reaction products, and possibly other minor components. The phenolic compounds contributed significantly to the antioxidant capacity of honey but were not solely responsible for it.     

Protein analysis of honeys by fast protein liquid chromatography: application to differentiate floral and honeydew honeys

Fast protein liquid chromatography on a Superdex 75 HR column has been applied to analyze the proteins of 29 honeys, 12 of floral origin and 17 from honeydew. The molecular masses were comprised between 13100 and 94000 Da. Seven peaks have been separated; four of them were present in all of the honeys, and three were only present in some honeys. Direct observation of the chromatograms of the floral and honeydew honeys did not reveal any information about their botanical origins. However, both types of honeys can be distinguished with the percentages of the areas of four of the seven chromatographic peaks obtained.     

Profiling glucosinolates, flavonoids, alkaloids, and other secondary metabolites in tissues of Azima tetracantha L. (Salvadoraceae)

Azima tetracantha L. (needle bush; bee sting bush; Salvadoraceae) is used as a food and for various herbal medicines in Africa, India, and Madagascar, but there is very little information on the secondary metabolites in this species. High concentrations of N-methoxy-3-indolylmethyl-glucosinolate, a common glucosinolate of Brassica crops such as Brussels sprouts and broccoli, were found in the roots and seeds of A. tetracantha. Lower concentrations were detected in the stems and young leaves. The roots also contained another indole glucosinolate that was provisionally identified, from MS data and comparison with indole glucosinolate standards, as N-hydroxy-3-indolymethyl-glucosinolate. The roots, stems, and leaves contained neoascorbigen (the condensation product of N-methoxy-indole-3-carbinol and ascorbic acid). The seeds of A. tetracantha contained a complex mixture of 26 flavonoids predominantly as glycosides and acyl-glycosides, with traces of aglycones. The core aglycones of these flavonoids were identified as quercetin, isorhamnetin (3'-O-methylquercetin), rhamnetin (7-O-methylquercetin), and rhamnazin (7, 3'-di-O-methylquercetin). No flavonoids or anthocyanins were detected in other tissues, and procyanidins were undetectable. The dimeric piperidine alkaloids azimine, azcarpine, and carpaine were found in all tissues of A. tetracantha.     

Identification of flavonoid markers for the botanical origin of Eucalyptus honey

European Eucalyptus honeys showed a common and characteristic HPLC profile in which the flavonoids myricetin (3,5,7,3',4', 5'-hexahydroxyflavone), tricetin (5,7,3',4',5'-pentahydroxyflavone), quercetin (3,5,7,3',4'-pentahydroxyflavone), luteolin (5,7,3', 4'-tetrahydroxyflavone), and kaempferol (3,5,7, 4'-tetrahydroxyflavone) were identified. Their contents, and relative amounts, in the analyzed honey samples were quite constant and supported their floral origin. In addition, ellagic acid and the propolis-derived flavonoids pinobanksin, pinocembrin, and chrysin were detected in most samples. The contents of these nonfloral phenolics were much more variable as could be expected for their propolis origin. Myricetin, tricetin, and luteolin had not been identified as floral markers in any other honey sample previously analyzed in our laboratory (chestnut, citrus, rosemary, lavender, acacia, rapeseed, sunflower, heather, lime tree, etc.) or reported in the literature, suggesting that these could be useful markers. Only in some individual heather samples produced in Portugal has tricetin previously been detected in minor amounts. These samples, however, were contaminated with Eucalyptus as revealed by their pollen analysis and the lack of tricetin or their glycosides in heather floral nectar. It remains to be established if myricetin, tricetin, and luteolin originate from Eucalyptus floral nectar where the corresponding glycosides should be present.     

Identification of new flavonoid glycosides and flavonoid profiles to characterize rocket leafy salads (Eruca vesicaria and Diplotaxis tenuifolia)

"Rocket" is a collective name used to term some species within the Eruca and Diplotaxis genera, whose leaves are characterized by a more or less pungent taste. Different approaches have been carried out to differentiate both genera that have similar leaf morphologies. Following our research in flavonoid profiling of the Brassicaceae family using high-performance liquid chromatography/ultraviolet-diode array detection/electrospray ionization mass spectroemtry, we have investigated Eruca vesicaria and Diplotaxis tenuifolia leaf samples as new ingredients of fresh salads. The MS/MS study allowed the identification of new naturally occurring quercetin mono- and diacyl-tri-O-glucosides and the elucidation of the flavonoid glycosylation and acylation patterns. Important differences between flavonoid profiles of E. vesicaria and D. tenuifolia were observed. E. vesicaria contained kaempferol derivatives as principal compounds whereas D. tenuifolia instead accumulated quercetin derivatives. The exhaustive study of the profiling of these species could help further studies concerning the bioavailability of these flavonoids for epidemiological or clinical intervention studies because these species have considerable potential as healthy leafy salads because of the bioactive phytochemicals.     

Volatile composition and contribution to the aroma of spanish honeydew honeys. Identification of a new chemical marker

Honeydew honeys from holm-oak, oak, and forest were isolated for aroma compounds by simultaneous distillation-extraction and analyzed by gas chromatography-mass spectrometry. In all, 66 volatile components were identified and quantified. trans-Oak lactone, a characteristic volatile component of oak wood, is proposed as a new chemical marker for the plant origin of honeydew honeys. Other compounds, such as aminoacetophenone and propylanisol, could be considered characteristic of holm-oak honeydew honeys. A total of 15 volatile compounds presented odor activity values (OAVs) greater than 1, with phenylacetaldehyde and beta-damascenone being those with the highest OAVs.     

Behavior of glucosinolates in pickling cruciferous vegetables

Crucifer species, which include widely consumed vegetables, contain glucosinolates as secondary metabolites. Cruciferous vegetables are consumed in Japan in salt-preserved or pickled form as well as cooked and raw fresh vegetables. In this study, changes in contents of glucosinolates during the pickling process were investigated. 4-Methylthio-3-butenyl glucosinolate, a major glucosinolate in the root of Japanese radish, daikon (Raphanus sativus L.), was detected in pickled products with a short maturation period but not in those with a long maturation period. As a model pickling experiment, fresh watercress (Nasturtium officinale) and blanched watercress were soaked in 3% NaCl solution for 7 days. The results showed that the ratio of indole glucosinolates to total glucosinolates increased during the pickling process, whereas total glucosinolates decreased. Myrosinase digestion of glucosinolates in nozawana (Brassica rapa L.) indicated that indole glucosinolates, especially 4-methoxyglucobrassicin, were relatively resistant to the enzyme. The effect of pickling on glucosinolate content and the possible mechanism are discussed in view of degradation by myrosinase and synthetic reaction in response to salt stress or compression during the pickling process.     

Extraction and determination of glucosinolates from soil

The use of glucosinolate-containing plants as soil-incorporated biofumigants for pest and disease control has raised questions regarding the fate of glucosinolates in soil; however, no method for routine analysis of glucosinolates in soil has been reported. A simple method to extract glucosinolates from soil with quantification as desulfoglucosinolates by HPLC is presented. The method involves two extractions with 70% methanol at room temperature, centrifugation, and filtration prior to the desulfation step. The desulfoglucosinolates are then quantified by HPLC using established protocols for plant tissue analysis. There were no significant interfering peaks from the soil extracts, and the method provided high extraction efficiencies (around 100%) for both aromatic (benzyl) and aliphatic (2-propenyl) glucosinolates when amended at a wide range of realistic field soil concentrations (1.6-120 nmol/g of soil). The method was equally effective in three diverse Australian soils that varied in organic matter, clay content, and pH. The method was effective in air-dried or field-moist soil, although evidence for rapid glucosinolate degradation in field-moist soil indicates that extraction of moist soils should be performed as soon as possible after sampling. The method is compatible with field soil sampling at remote sites and utilizes the same equipment and protocols already established for plant tissue analysis. Extraction of glucosinolates in the field following incorporation of Indian mustard (Brassica juncea) and rape (Brassica napus) green manure crops was also tested. Eight different glucosinolates contained in the plant tissues were identified and quantified in soil extracts at concentrations ranging from 0.11 to 21.7 nmol/g of soil.     

Solid-phase extraction and LC-MS analysis of pyrrolizidine alkaloids in honeys

Strong-cation-exchange, solid-phase extraction of pyrrolizidine alkaloids and their N-oxides from honey samples was followed by reduction of the N-oxides and subsequent analysis of total pyrrolizidine alkaloids using high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry. A limited survey of 63 preprocessing samples of honey, purposefully biased toward honeys attributed to floral sources known to produce pyrrolizidine alkaloids, demonstrated levels of pyrrolizidine alkaloids up to approximately 2000 parts per billion (ppb) in a sample attributed to Echium plantagineum. Up to 800 ppb pyrrolizidine alkaloids was detected in some honeys not attributed by the collector to any pyrrolizidine alkaloid-producing floral source. No pyrrolizidine alkaloids were detected in approximately 30% of the samples in this limited study, while some honeys showed the copresence of pyrrolizidine alkaloids from multiple floral sources such as E. plantagineum and Heliotropium europaeum. In addition, retail samples of blended honeys (with no labeling to suggest that pyrrolizidine alkaloid-producing floral sources were used in the blends) have been shown to contain up to approximately 250 ppb pyrrolizidine alkaloids.     

Morphoagronomic and biochemical variation in an Argentinean landrace of wheat

A sample of an Argentinean landrace of wheat showed considerable variation in most of the evaluated morphological and agronomic characters. However, analyses with high molecular glutenins and two isozyme systems, known to be highly polymorphic among current cultivars, revealed very little or no variation, respectively. The large difference in the observed variation between morphoagronomic and biochemical characters is discussed.     

Amino acid composition and antioxidant capacity of Spanish honeys

The amino acid composition of 53 honey samples from Spain, consisting of 39 floral, 5 honeydew, and 9 blend honeys, has been determined. Physicochemical characteristics, polyphenolic content, amino acid composition, and estimation of the radical scavenging capacity against the stable free radical DPPH of the honey samples were analyzed. The resulting data have been statistically evaluated. The results showed that pH, acidity, net absorbance, electrical conductivity, and total polyphenolic contents of the honeys showed a strong correlation with the radical scavenging capacity. The correlation between the radical scavenging capacity of honey and amino acid contents was high with 18 of the 20 amino acids detected, with correlation values higher than those obtained for polyphenolic content. These results suggest that the amino acid composition of honey is an indicator of the sample's scavenging capacity.     

Identification and antioxidant potential of flavonoids and low molecular weight phenols in olive cultivar chemlali growing in Tunisia

Increasing interest in phenolic compounds in olives is due to their antioxidant and health-enhancing properties. In this study the phenolics in fruits of the Tunisian olive cultivar Chemlali were extracted by methanol-water and fractionated using Sephadex LH-20 column chromatography. The identification of phenolic monomers and flavonoids was based on separation by high-performance liquid chromatography equipped with a diode array detector followed by liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry analysis. Oleuropein, a secoiridoid glycoside esterified with a phenolic acid, was the major compound. Eight phenolic monomers and 12 flavonoids were also identified in Chemlali olives. Five flavonoids were isolated and purified using Sephadex LH-20 column chromatography and preparative paper chromatography. The antioxidant activity of the extract and the purified compounds was evaluated by measuring the radical scavenging effect on 1,1-diphenyl-2-picrylhydrazyl and by using the beta-carotene-linoleate model assay. Acid hydrolysis of the extract enhanced its antioxidant activity. Hydroxytyrosol and quercetin showed antioxidant activities similar to that of 2,6-di-tert-butyl-4-methylphenol. A hydroxyl group at the ortho position at 3' on the B ring of the flavonoid nucleus could contribute to the antioxidant activity of the flavonoids.     

Thermal and Functional Characterization of Starch from Argentinean Corn

Screening accessions in a germplasm bank aids in the identification of plants with unusual properties ranging from agronomic traits to functional and compositional traits of the seed itself. Results from this study confirm the presence of a wide variation in the thermal and functional properties of starch from several landraces of corn in the Argentinean germplasm. Thermal properties of starch measured using differential scanning calorimetry (DSC) identified several corn landraces with properties of potential commercial interest. The pasting and textural properties of gels obtained from starch of different corn landraces also exhibited considerable variability. The degree of variation in thermal and functional properties of corn reported in this study is comparable to the thermal properties of starch from several other crop species. These corn races show promise for further regeneration to create inbred lines with unusual traits. The potential for further improvement of corn races exists not only based on thermal properties, but directly for specific functional attributes as well. Correlation analyses suggest that the variability in thermal and functional attributes are a function of amylose content, granule size distribution and, possibly, differences in the structural makeup of amylose and amylopectin. The strong correlation observed between the thermal properties and pasting and textural properties will allow for the estimation of starch properties from small sample sizes.     

油菜硫甙的农业生态学意义与研究进展

论述了油菜琉甙(即琉代葡萄糖甙)的组分、主要生理功能及其含量过高对家畜的不利影响以及引种国际油菜导致的生态问题,指出了影响油菜硫甙含量的主要因素、油菜琉甙对维护十字花科植物抗性的自身保护功能、生态学意义及其发展应用潜力。     

Variation of glucosinolates in vegetable crops of Brassica oleracea

Glucosinolates were evaluated in 5 groups and 65 accessions of Brassica oleracea (50 broccoli, 4 Brussels sprouts, 6 cabbage, 3 cauliflower, and 2 kale) grown under uniform cultural conditions. Glucosinolates and their concentrations varied among the different groups and within each group. The predominant glucosinolates in broccoli were 4-methylsulfinylbutyl glucosinolate (glucoraphanin), 3-butenyl glucosinolate (gluconapin), and 3-indolylmethyl glucosinoate (glucobrassicin). Glucoraphanin concentration in broccoli ranged from 0.8 micromol g(-1) DW in EV6-1 to 21.7 micromol g(-1) DW in Brigadier. Concentrations of the other glucosinolates in broccoli varied similarly over a wide range. In Brussels sprouts, cabbage, cauliflower, and kale, the predominant glucosinolates were sinigrin (8.9, 7.8, 9.3, and 10.4 micromol g(-1) DW, respectively) and glucobrassicin (3.2, 0.9, 1.3, and 1.2 micromol g(-1) DW, respectively). Brussels sprouts also had significant amounts of gluconapin (6.9 micromol g(-1) DW). Wide variations in glucosinolate content among genotypes suggest differences in their health-promoting properties and the opportunity for enhancement of their levels through genetic manipulation.