Laburnum Alpinum Research Articles

Determination of Cytisine and N-Methylcytisine from Selected Plant Extracts by High-Performance Liquid Chromatography and Comparison of Their Cytotoxic Activity

Quinolizidine alkaloids exhibit various forms of biological activity. A lot of them were found in the Leguminosae family, including Laburnum and Genista. The aim of the study was the optimization of a chromatographic system for the analysis of cytisine and N-methylcytisine in various plant extracts as well as an investigation of the cytotoxic activities of selected alkaloids and plant extracts obtained from Laburnum anagyroides, Laburnum anagyroides L. quercifolium, Laburnum alpinum, Laburnum watereri, Genista germanica, and Genista tinctoria against various cancer cell lines. The determination of investigated compounds was performed by High Performance Liquid Chromatography with Diode Array Detection (HPLC-DAD), while High Performance Liquid Chromatography coupled with Quadrupole Time-of-Flight–Mass Spectrometry (HPLC-QTOF-MS) was applied for the qualitative analysis of plant extracts. The retention, separation selectivity, peaks shape, and systems efficiency obtained for cytisine and N-methylcytisine in different chromatographic systems were compared. The application of columns with alkylbonded and phenyl stationary phases led to a very weak retention of cytisine and N-methylcytisine, even when the mobile phases containing only 5% of organic modifiers were used. The strongest retention was observed when hydrophilic interaction chromatography (HILIC) or especially when ion exchange chromatography (IEC) were applied. The most optimal system in terms of alkaloid retention, peak shape, and system efficiency containing an strong cation exchange (SCX) stationary phase and a mobile phase consisted of 25% acetonitrile and formic buffer at pH 4.0 was applied for investigating alkaloids analysis in plant extracts. Cytotoxic properties of the investigated plant extracts as well as cytisine and N-methylcytisine were examined using human tongue squamous carcinoma cells (SCC-25), human pharyngeal squamous carcinoma cells (FaDu), human triple-negative breast adenocarcinoma cell line (MDA-MB-231), and human breast adenocarcinoma cell line (MCF-7). The highest cytotoxic activity against FaDu, MCF-7, and MDA-MB cancer cell lines was observed after applying the Genista germanica leaves extract. In contrast, the highest cytotoxic activity against SCC-25 cell line was obtained after treating with the seed extract of Laburnum watereri. The investigated plant extracts exhibit significant cytotoxicity against the tested human cancer cell lines and seem to be promising for further research on its anticancer activity.

Genetic vs. non‐genetic responses of leaf morphology and growth to elevation in temperate tree species

SummaryAt high elevation, temperate trees generally exhibit adaptive genetic differentiation in their morphological and physiological traits. On account of this directional selection, we hypothesized that tree populations growing near their upper cold elevational limits exhibit lower phenotypic plasticity of growth and leaf morphological traits in response to temperature changes than populations growing at lower elevations.Seedlings of six common deciduous tree species originating from low and high elevations were transplanted into eight common gardens along two elevational gradients in the Swiss Alps. The aim of the experiment was to evaluate the genetic differentiation in growth and leaf morphology between populations from low and high elevations and to quantify the phenotypic plasticity of these traits to temperature changes.In contrast to growth that decreased with increasing elevation, leaf mass per area (LMA) showed no significant change with elevation of common garden, except for a decrease inLaburnum alpinumfor both low‐ and high‐elevation provenances. Interestingly, leaf density was found to decrease with elevation of the gardens for all species. Genetic differentiation between low‐ and high‐elevation populations was found in both leaf morphology and growth: high‐elevation populations tended to have slower growth rate than low‐elevation populations, while no consistent trend was found forLMAacross species. Interestingly, forAcer pseudoplatanusandFraxinus excelsiorhigh‐elevation populations exhibited a lower phenotypic plasticity of growth in response to temperature compared to lower populations, whereas no interactions between the elevation of a provenance and the elevation of the garden was detected for the four other species.Hence, during young life stages, the expected increase in tree growth in future warmer climates might be lower in populations living in the coldest part of the species distribution range in temperate species such asAcer pseudoplatanusandFraxinus excelsior, but similar in other tree species, disregarding other environmental changes.

Two Early Nodulation Genes are not Markers for the Capacity of Leguminous Nursery Crops to Form Root Nodules

Abstract Little is known about mechanisms that permit or prevent formation of root nodules in which nitrogen gas (N2) is fixed by rhizobial bacteria in association with nursery crops in the legume family. We tested the hypothesis that two genes associated with the fixation of N2 could be used as markers of nodulation capacity among legumes because they occur only in species that can form functional nodules. The presence of ENOD2 and ENOD12 was tested in genomic DNA from three non-nodulating legumes [Cercis canadensis L. (eastern redbud), Gleditsia triacanthos L. var. inermis Willd. (thornless honeylocust), Gymnocladus dioicus (L.) K. Koch (Kentucky coffeetree)] and two nodulating legumes [Albizia julibrissin Durazz. (silk-tree), Laburnum alpinum (Mill.) Bercht. & Presl. (Scotch laburnum)]. Southern analyses indicated that ENOD2 is present in thornless honeylocust, Kentucky coffeetree, and Scotch laburnum, and that ENOD12 is present in eastern redbud, thornless honeylocust, and Scotch laburnum. These results diversify the group of nodulating legumes in which ENOD2 and ENOD12 have been found and show these genes also occur in legumes considered incapable of nodulation. We conclude that neither gene can be used to screen existing or new leguminous nursery crops for the capacity to form N2-fixing nodules.

Early Nodulin Genes are Not Markers of the Capacity of Woody Legumes to Nodulate

Knowing whether trees can develop nodules in which nitrogen is fixed is important for managing inputs during culture. Early nodulin genes, such as ENOD2 and ENOD12, could function as molecular markers for the capacity to nodulate if they are conserved only in species that nodulate. Several nodulating species are known to possess these genes; but, their occurrence has not been studied among non-nodulating taxa, including Cercis canadensis L. (redbud), Gleditsia triacanthos L. var. inermis Willd. (honey locust), and Gymnocladus dioica (L.) C. Koch (Kentucky coffee tree). Our objective was to determine the relationship between the capacity to nodulate and the occurrence of putative ENOD2 or ENOD12 genes by probing the genomes of these non-nodulating species and the genomes of two legumes that nodulate, Albizia julibrissin Durazz. (silk tree) and Laburnum alpinum (Mill.) Bercht. & J. Presl (Scotch laburnum). ENOD2 and ENOD12 cDNA clones from Glycine max (L.) Merill (soybean) and Pisum sativum L. (pea), respectively, and cloned ENOD2 PCR fragments from Maackia amurensis Rupr. & Maxim. (Amur maackia) and Styphnolobium japonicum (L.) Schott (Japanese pagodatree) were used as probes for Southern hybridizations. Sequences from genomes of silk tree, Scotch laburnum, honey locust, and Kentucky coffee tree hybridized to ENOD2 probes on Southern blots. Putative ENOD12 sequences were detected in the genomes of Scotch laburnum, redbud, and honey locust. Thus, we conclude that ENOD2 and ENOD12 can not be used as markers for the capacity to nodulate.

Leaching Fraction and Fertilization Effects on Growth of Three Woody Legumes Inoculated with Rhizobia

Scotch laburnum [Laburnum alpinum (Mill.) Bercht.], Amur maackia (Maackia amurensis Rupr. & Maxim.), and Chinese wisteria [Wisteria sinensis (Sims) Sweet] were inoculated with compatible rhizobia and treated with leaching fractions (LF) of 0, 0.2, and 0.4 using fertilizer solutions with 3.6 and 10.7 mol N/m3 for 10 weeks. LF did not affect plant dry mass, leaf area, or stem length. Growth was higher among plants provided 10.7 mol N/m3, but only plants provided 3.6 mol N/m3 formed root nodules. We conclude that growth is not reduced by eliminating leaching during the first 10 weeks of seedling development, and that application of 10.7 mol N/m3 prevents nodulation of these species.

A unique amino acid sequence involved in the putative carbohydrate-binding domain of a legume lectin specific for sialylated carbohydrate chains: primary sequence determination of Maackia amurensis hemagglutinin (MAH).

The primary sequence of 247 amino acids of Maackia amurensis hemagglutinin (MAH) was determined using a protein sequencer. After digestion with endoproteinase Lys-C, Asp-N, Arg-C, or Glu-C of MAH, the resulting peptides were purified by reversed phase high performance liquid chromatography (HPLC) and then subjected to sequence analysis. The primary sequence of MAH was compared with those of several legume lectins, and it was found that the amino acid sequence of the putative carbohydrate-binding domain of MAH exhibited a high degree of homology with those of di-N-acetylchitobiose-binding Cytisus sessilifolius lectin I (CSA-I), Laburnum alpinum lectin I (LAA-I), and Ulex europaeus lectin II (UEA-II). In the legume lectins whose primary sequences have already been determined several amino acid residues involved in carbohydrate-binding were found to be conserved. Very interestingly, in the primary sequence of MAH, one amino acid residue corresponding to the conserved amino acid, asparagine, in the primary sequences of all other legume lectins was shown to be substituted by aspartic acid. This is the first report of the occurrence of an exceptional amino acid residue among the conserved amino acid residues in the carbohydrate-binding domain of the legume lectins.

Correlation between carbohydrate-binding specificity and amino acid sequence of carbohydrate-binding regions of Cytisus-type anti-H(O) lectins

A carbohydrate-binding peptide of the di- N-acetylchitoblose-binding Cytisus sessilifolius anti-H(O) lectin I (CSA-I) was isolated from the endoproteinase Asp-N digest of CSA-I by affinity chromatography on a column of N-acetyl- d-glucosamine oligomer-Sepharose (GIcNAc oligomer-Sepharose). The amino acid sequence of the carbohydrate-binding peptide of CSA-I was determined to be DTYFGKTYNPW using a gas-phase protein sequencer. This sequence corresponds to the sequence from Asp-129 to Trp-139 based on the primary structure of CSA-I, and shows a high degree of homology to those of the putative carbohydrate-binding peptide of the Laburnum alpinum lectin I (LAA-I) (DTYFGKAYNPW) and of the Ulex europaeus lectin II (UEA-II) (DSYFGKTYNPW). The binding of these three anti-H(O) lectins is known to be inhibited by di- N-acetylchitobiose but not by l-fucose. These results strongly suggest that there is a good correlation between the carbohydrate-binding specificity and the amino acid sequence of the carbohydrate-binding regions of di- N-acetylchitobiose-binding lectins.

Carbohydrate-Binding Peptides from Several Anti-H(O) Lectins

Peptide fragments have been obtained from L-fucose-binding anti-H(O) lectins [Lotus tetragonolobus lectin (LTA) and Ulex europeus lectin I (UEA-I)] and di-N-acetylchitobiose-binding anti-H(O) lectins [Ulex europeus lectin II (UEA-II) and Laburnum alpinum lectin I (LAA-I)] by treatment with endoproteinase Asp-N or Lys-C. The peptide fragments were fractionated by affinity chromatography on a column of Fuc-Gel for LTA and UEA-I, and on a column of N-acetyl-D-glucosamine oligomer-Sepharose for UEA-II and LAA-I. The peptides with affinity for these columns were identified by peptide sequencing. All of these retarded peptides were found to be parts of the metal-binding regions of these lectins. It is strongly suggested that these peptides represent the carbohydrate-binding and metal ion-binding sites of legume lectins, respectively.

Erythrina Lectins Detect the H/HI Blood Groups

The lectin purified from Erythrina corallodendron seeds which binds N-acetyllactosamine greater than N-acetyl-D-galactosamine greater than alpha and beta galactosides greater than D-galactose was examined for its ABO(H) blood group specificity. It has been shown that this lectin causes the strongest hemagglutination of O(H) and weakest of Oh(Bombay) red blood cells, and interacts with the H antigen in association with the I antigen. The reactions of Erythrina corallodendron and Erythrina indica lectins (which are similar in sugar specificity) with erythrocytes of different ABO(H) and Ii blood groups (the I bloods were all from adults and the i from either cord or adult bloods) revealed the following order of activity: O(H)I greater than A2 I greater than O(H)i adult greater than A2BI greater than BI greater than O(H)i cord greater than A1I greater than A1i adult greater than Bi cord greater than A1BI greater than Ai cord greater than ABi cord greater than OhI. The Erythrina indica lectin showed a lower differentiation between the agglutination of O(H) and Oh erythrocytes. Both Erythrina lectins exhibited H/HI blood group preference but were not inhibited by the saliva from ABO(H) "secretors". Thus they may be classified with the Cytisus sessilifolius, Lotus tetragonolobus and Laburnum alpinum lectins which are inhibited by lactose but not by H blood group substances in secretions.

The primary structure of the Laburnum alpinum seed lectin

The complete amino acid sequence of the Laburnum alpinum di-N-acetylchitobiose-binding lectin was determined by using a protein sequencer after digestion with endoproteinases Lys-C and Asp-N, and compared with those of other leguminous plant lectins.

Über die Alkaloidzusammensetzung der oberirdischen Teile von Laburnum watereri (Kirchn.) Dipp.

The alkaloid composition of the overground parts (leaves, petioles, stems, flowers, fruits and seeds) of Laburnum watereri (hybrid of Laburnum alpinum × L. anagyroides ) and its seasonal changes (from march to november) have been studied by different chromatographic (TLC, capillary GC) and spectroscopic (GC/MS, HRMS, 1 H-NMR, 13 C-NMR, IR, UV) methods. Seeds only contain cytisine. Besides cytisine, traces of methylcytisine and ammodendrine 12 further alkaloids could be identified in the fruits, among which β-isosparteine, 14e-hydroxysparteine and epi-baptifoline have been found for the first time in the genus Laburnum . Following the seasonal changes of the alkaloid composition of the fruits, an increase of epibaptifoline and ammodendrine was established during ripening, while the content of the main compound cytisine strongly declined. The highest total amount of alkaloids (with 80 to 90% cytisine) was found in the buds in the spring. During the further development of the leaves, the cystine content decreases and a methylation of this alkaloid begins. In summer the alkaloid concentration reaches the lowest level. In the course of July epi-baptifoline (erroneously reported as methylcytisine in literature) increases and becomes the main alkaloid for the following 6 months.

Sugar binding specificities of anti-H(O) lectins disclosed by use of fucose-containing human milk oligosaccharides as binding inhibitors.

The binding to normal and sialidase-treated human erythrocytes of six 125I-labeled lectins [Ulex europeus lectin I (UEA-1) and II (UEA-II), Laburnum alpinum lectins I (LAA-I) and II (LAA-II), and Cytisus multiflorus lectins I (CMA-I) and II (CMA-II)], was studied in detail. Quantitative inhibition assays of the lectin binding to the cells were also performed with various human milk oligosaccharides as inhibitors. Based on a comparison of the inhibition constants of the inhibitors thus obtained with the association constants of the lectins to the cells, the relative activities of cell surface blood group antigens toward the lectins are discussed.

Lectin binding in meningiomas.

Forty-two meningiomas of different morphological sub-type were examined to determine their pattern of binding to 11 different lectins which characterize cell surface components such as carbohydrate residues. Histiocytic and xanthoma cells within meningiomas could be demonstrated with six different lectins: wheat germ agglutinin (WGA), peanut agglutinin (PNA) Bauhinia purpurea agglutinin (BPA), Helix pomatia agglutinin (HPA), Vicia fava agglutinin (VFA) and Soyabean agglutinin (SBA). Vascular elements including endothelial cells and intimal cells, bound Ulex europaeus agglutinin type 1 (UEA 1), WGA and HPA. The fibrous stroma in fibrous and fibroblastic meningiomas bound PNA, Laburnum alpinum agglutinin (LAA) and SBA. Tumour cells in meningotheliomatous meningiomas and some areas of anaplastic meningiomas bound Concanavalin A, PNA, LAA and VFA whereas tumour cells in fibrous and fibroblastic meningiomas bound BPA, LAA and VFA. Lectin binding has proved to be of value in detecting histiocytic and xanthoma cells together with vascular elements within meningiomas. In addition, the different lectin binding patterns allow different histological sub-types of meningioma to be distinguished although the biological significance of the binding patterns is unclear.

Peronospora cytisi . [Descriptions of Fungi and Bacteria

Abstract A description is provided for Peronospora cytisi . Information is included on the disease caused by the organism, its transmission, geographical distribution, and hosts. HOSTS: Laburnum alpinum, L. anagyroides; Robinia pseudacacia (fide Lindtner, 1957, but not confirmed by cross inoculation). DISEASE: Downy mildew of laburnum. The disease is most severe on young seedlings about 6-8 inches high. Symptoms are first seen on the upper surface of the leaves as pale yellow spots, these areas rapidly become brown, dead and brittle. The down, which is greyish brown, develops in a dense felt on the undersurface of these spots. The infected leaves may become completely withered and drop off and the seedlings then soon die. Infection of older plants has been reported by Magnus (1892) who described a single shrub (no indication of size given) with many leaves fallen and the remainder with numerous spots. Berlese (1903) records that in older plants the parasite develops especially in the upper leaves but the plants are not killed. The following year they can look perfectly healthy unless attacked again by the disease. GEOGRAPHICAL DISTRIBUTION: Europe (Britain, Denmark, France, Germany, Poland, Spain, Sweden ( fide Gaumann (1923) but no specimen seen by Gustavsson (1959)), Yugoslavia. TRANSMISSION: No reports. Contaminated seed was not thought to be the source of the attack reported by Kirchner (1892) on seedlings raised from seed from the local botanic garden where the disease was not present nor did it subsequently appear. Oospores form in considerable numbers in the withered leaves and must form a potential source of infection on fallen debris in the soil. However the sporadic outbreaks of the disease suggests that the fungus may perhaps exist on other hosts such as Robinia pseudacacia or be present in small and undetected quantities on older laburnum plants in most seasons and only cause severe infection, and thus attract attention, when weather conditions favour its rapid development.

Studies on the relationship between lectin binding carbohydrates and different strains of Leishmania from the New World

The culture forms of L. mexicana pifanoi (LRC L-90), L. mexicana mexicana (LRC L-94, M-379); L. braziliensis braziliensis (LRC L-77, L-1, M-2903, H-LSS) and L. mexicana amazonensis (H-JMMO, M-JOF, H-21, H-PLL,M-1696) were tested with the following lectins: Canavalia ensiformis, Ricinus communis-120, Axinella polypoides, Phaseolus vulgaris, Evonymus europaeus, lotus tetragonolobus, Dolichos biflorus, Aaptos papillata II, Laburnum alpinum, Ulex europaeus, Arachis hypogaea and Soja hispida. All examined strains of Leishmania were agglutinated by C. ensiformis, R. communis-120 and A. popypoides. No agglutination reactions were observed with P. vulgaris, D.biflorus, A. papillata II, E. europaeus and L. tetragonolobus. Only L. m. pifanoi and the L. m. amazonensis strains H-JMMO and MJOF showed agglutination reactions with S. hispida, U. europaeus, L. alpinum and A. hypogaea, while L. m. mexicana (LRC L-94; M-379) strains, L. b. braziliensis H. LSS, LRC L-77; L-1; M-2903 and the L. m. amazonensis strains, H-PLL, H-21, M-1696 showed no agglutination reactions with these four lectins.

7.10. A hereditary serum component determined by precipitation with Laburnum alpinum seed extract

7.10. A hereditary serum component determined by precipitation with Laburnum alpinum seed extract

Développement des antigènes ABH et Ii chez les enfants de 0 a 16 ans

The ABH and I, i erythrocyte antigens of 926 children from birth to sixteen years were tested in a semi-quantitative assay in order to follow their development. Results show that total activities of A, B and H erythrocyte antigens are reached within 13 and 24 months, the main modifications being observed within the first three months of life. However, these activities depend on the agglutinins used in the test; notably the results with anti-H lectins from Ulex europaeus and Laburnum alpinum are not totally identical. The decrease of i reactivity and the increase of I reactivity are simultaneous, but at birth the I reactivity of cord red cells is widely distributed, while that of i is more homogeneous. The increase in H activity tested with Laburnum alpinum is directly related to the decrease of foetal hemoglobin in the children's red cells.

A glactose specific agglutinin, a blood-group H active polysaccharide, and a trypsin inhibitor in albumin glands and eggs of Arianta arbustorum (helicidae)

Eggs and albumin glands of the land snail Arianta arbustorum contain a powerful agglutinin which reacts specially with rabbit erythrocytes. The agglutination can be inhibited completely by di-, tri-, and oligosaccharides with α-glycosidically (1 → 6) bound galactose residues. β-Linked sugars do not inhibit the agglutinin. The agglutinin activity is not dependent on Ca 2+ ions. Eggs and albumin glands also contain a blood-group active polysaccharide which, unlike the polysaccharide from the albumin gland of Helix pomatia (Baldo, B. A., and Uhlenbruck, G. 1973. Cross-reactive human blood group H-active polysaccharide from Helix pomatia. I. Detection with catfish anti-H and eel sera. Immunology, 25, 1–13) does not react with anti-H eel, but does react with the agglutinins of Evonymus europaeus and Laburnum alpinum. The Arianta polysaccharide has been purified and shown to be galactogen. Finally, the occurrrence of a strong trypsin inhibitor has been demonstrated in the extracts of eggs and albumin glands. The inhibitor has been separated by column chromatography. The precipitation lines of both substances have been identified in the immunoelectrophoretogram of the extracts of albumin glands and eggs.

The H specificities of butanol extracts of human erythrocytes.

A previous report of other researchers that aqueous phase fractions of erythrocyte stromata of secretors contained more ABH blood group activity than similar fractions of non-secretors was confirmed. The recovery of H activity in the butanol phase extracts of both secretors and non-secretors, which had not been seen previously, is reported here for the first time. Three H specificities were present when the aqueous and butanol phases of group O secretor stromata were examined in inhibition of agglutination studies with anti-H of group Oh and group A1 sera and three lectins. The anti-H (group Oh) sera were inhibited by both aqueous and butanol fractions. Anti-H serum (group A1) and the lectin Ulex europaeus were inhibited by the aqueous fraction only. The lectins Laburnum alpinum and Lotus tetragonolobus were not inhibited by either the aqueous or butanol phase extracts.

The H Specificities of Butanol Extracts of Human Erythrocytes

A previous report of other researchers that aqueous phase fractions of erythrocyte stromata of secretors contained more ABH blood group activity than similar fractions of non-secretors was confirmed. The recovery of H activity in the butanol phase extracts of both secretors and non-secretors, which had not been seen previously, is reported here for the first time. Three H specificities were present when the aqueous and butanol phases of group O secretor stromata were examined in inhibition of agglutination studies with anti-H of group O(h) and group A(1) sera and three lectins. The anti-H (group O(h)) sera were inhibited by both aqueous and butanol fractions. Anti-H serum (group A(1)) and the lectin Ulex europaeus were inhibited by the aqueous fraction only. The lectins Laburnum alpinum and Lotus tetragonolobus were not inhibited by either the aqueous or butanol phase extracts.