New Cytotoxic Bibenzyl and Other Constituents from Bauhinia ungulata L. (Fabaceae)

A new bibenzyl, 2′‐hydroxy‐3,5‐dimethoxy‐4‐methylbibenzyl ( 1 ) and four known compounds identified as 2′‐hydroxy‐3,5‐dimethoxybibenzyl ( 2 ), liquiritigenin ( 3 ), guibourtinidol ( 4 ) and fisetinidol ( 5 ) were isolated from the roots of Bauhinia ungulata L. Phytochemical investigations of the stems of B. ungulata led to the isolation of the known compounds identified as liquiritigenin ( 3 ), guibourtinidol ( 4 ), fisetinidol ( 5 ), taraxerol ( 6 ), betulinic acid ( 7 ), taraxerone ( 8 ), glutinol ( 9 ), a mixture of sitosterol ( 10 ) and stigmasterol ( 11 ), pacharin ( 12 ), naringenin ( 13 ) and eriodictyol ( 14 ). The structures of these compounds were elucidated on the basis of their spectral data (IR, MS, 1D‐ and 2D‐NMR). The cytotoxicity of the bibenzyl 1 has been evaluated against four human cancer cell lines, showing the IC₅₀ values of 4.3 and 6.5 μg ml^?1 against pro‐myelocytic leukemia (HL‐60) and cervical adenocarcinoma (HEP‐2) cell lines, respectively. This article also registers for the first time the ¹³C‐NMR data of the known bibenzyl 2 .     

A novel antiproliferative and antifungal lectin from Amaranthus viridis Linn seeds

A lectin from the seeds of Amaranthus viridis Linn has been purified by affinity chromatography on asialofetuin-linked amino activated silica. Amaranthus viridis lectin (AVL) has a native molecular mass of 67 kDa. It is a homodimer composed of two 36.6 kDa subunits. The lectin gave a single band in non-denaturing PAGE at pH 4.5 and pH 8.3 and a single peak on HPLC size exclusion and cation exchange columns. The purified lectin was specific for both T-antigen and N-acetyl-D-lactosamine, markers for various carcinomas, in addition to N-acetyl-D-galactosamine, asialofetuin and fetuin. This lectin reacted strongly with red blood cells (RBCs) from human ABO blood groups and rat. It also reacted with rabbit, sheep, goat and guinea pig RBCs. The lectin is a glycoprotein having no metal ion requirement for its activity. Denaturing agents such as urea, thiourea and guanidine-HCl had no effect on its activity when treated for 15 minutes. AVL showed significant antiproliferative activity towards HB98 and P388D1 murine cancer cell lines. It also exerted antifungal activity against phytopathogenic fungi Botrytis cincerea and Fusarium oxysporum but not against Rhizoctonia solani, Trichoderma reesei, Alternaria solani and Fusarium graminearum.     

Isolation and characterization of a novel lectin with antifungal and antiproliferative activities from Sophora alopecuroides seeds

Sophora alopecuroides lectin (SAL), a novel lectin from the seeds of Sophora alopecuroides, was purified by ion-exchange chromatography on diethylaminoethyl (DEAE)- and carboxymethyl (CM)-Sepharose columns, followed by gel filtration on a Sephadex 75 10/300 GL column. SAL was found to be a monomer of 39916.3 Da, as determined by tricine-sodium dodecyl sulphate-polyacrylamide gel electrophoresis and high-performance liquid chromatography (HPLC). The N-terminal 10-amino acid sequence of SAL, KPWALSFSFG, resembles those of other legume lectins. SAL exhibits hemagglutinating activity against rabbit erythrocytes at 11.9 μg/ml. Its hemagglutinating activity is stable in the pH range 7-11 and in the temperature range 30-90°C, and is stimulated by Mn(2+). The hemagglutinating activity of SAL is most potently inhibited by 50-mM d-galactose. SAL suppresses mycelial growth in Penicillium digitatum and Alternaria alternata; the IC(50) of the antifungal activity toward P. digitatum and A. alternata were found to be 3.125 and 3.338 μM, respectively. SAL suppresses the proliferation of human cervical cancer cells (HeLa) at an IC(50) of 6.25 μM (P< 0.05). But it has no inhibiting effect on bacteria. This is the first report of a lectin from seeds of S. alopecuroides.     

Isolation and properties of a lectin from the seeds ofMimosa invisa L.

A lectin has been purified from the seeds ofMimosa invisa L. by gel filtration and preparative Polyacrylamide gel electrophoresis. The purified lectin was homogeneous as judged by analytical Polyacrylamide gel electrophoresis, immunodiffusion and Immunoelectrophoresis. The apparent molecular weight is 100,000; the protein is a tetramer with two types of subunits (molecular weight 35,000 and 15,000). The lectin is a glycoprotein with approximately 21% carbohydrate and interacts with Sephadex and concanavalin A-Sepharose. It agglutinates erthrocytes non-specifically, does not agglutinate leucocytes and is not mitogenic, agglutinates Mimosa-nodulatingRhizobium and is a panagglutinin; the agglutination is not inhibited by several simple sugars. It is thermo-stable and has no metal ions.     

洋紫荆凝集素的分离纯化和性质

利用酸化处理的Ｓｅｐｈａｒｏｓｅ ６Ｂ亲和柱从洋紫荆种子中分离纯化出了洋紫荆凝集素（ＢＶＬ）,其比活性比抽提液提高了１５９倍,活力回收率为４９．０％。ＢＶＬ分子量为８１０００,由两个相同的亚基组成。等电聚焦凝胶电注测得其等电点为４．９５。其紫外吸收高峰在２７６ｎｍ处。ＢＶＬ具一定的的热稳定性和酸碱稳定性。Ｎ－乙酰－Ｄ氨基半乳能强烈地抑制ＢＶＬ对兔红细胞的凝集作用。乳糖、半乳糖也有较强的抑制定性。Ｎ     

Isolation of a novel legumin-like lectin with potent hemagglutinating activity from seeds of the Chinese chestnut Castanea mollisima

A novel mannose- and glucose-specific lectin with high hemagglutinating activity was isolated from seeds of the Chinese chestnut Castanea mollisima. The lectin possessed a molecular mass of 140 kDa and was made up of two subunits, one with a molecular mass of 31 kDa and another with a molecular mass of 32 kDa. They exhibited substantial homology in N-terminal sequence to the storage protein legumin. The lectin was unstable in the presence of acid and alkali and at temperatures above 50 degrees C, but it was unaffected by various salts. The lectin was purified with a procedure involving ion exchange chromatography on CM-Sepharose, Q-Sepharose and Resource Q and gel filtration on Superose 12.     

Purification,primary structure and potential functions of a novel lectin from Bauhinia forficata seeds

A new lectin, BfL, was purified from Bauhinia forficata seeds by ammonium sulfate fractionation, DEAE-Sephadex ion exchange chromatography, Sepharose-4B and chitin affinity chromatographies and Superdex 75 size exclusion chromatography. The molecular homogeneity and purity of BfL were assessed by reversed-phase HPLC. BfL appeared as a single band of approximately 27.0 kDa on SDS-PAGE under non-reducing and reducing conditions, and its molecular weight was determined to be 27,850 Da by LC/ESI-MS. BfL is a glycoprotein with a carbohydrate content of 6.24% determined by the phenol–sulfuric acid method. Fetuin, asialofetuin, thyroglobulin and azocasein inhibited the hemagglutinating activity of BfL, whereas saccharides did not. BfL hemagglutinating activity was stable at 100 °C for 30 min, pH-dependent, with the highest activity at pH 6.0, and metal-independent. The primary structure of BfL shows similarity with other lectins from the genus Bauhinia. Deconvolution of the BfL circular dichroism (CD) spectrum indicated the presence of α-helix and β structures. BfL increases coagulation time, but this effect is not related to human plasma kallikrein or human factor Xa inhibition. BfL also inhibits ADP- and epinephrine-induced platelet aggregation in a dose-dependent manner and is the only currently described lectin from Bauhinia that exhibits anticoagulant and antiplatelet aggregating properties.     

A novel sialic acid-specific lectin from Phaseolus coccineus seeds with potent antineoplastic and antifungal activities

A novel lectin (PCL) with specificity towards sialic acid was purified from Phaseolus coccineus L. (P. multiflorus willd) seeds using ion exchange chromatography on CM and DEAE-Sepharose, and gel filtration on Sephacryl S-200 column. PCL was a homodimer consisting of 29,831.265 Da subunits as determined by gel filtration and MS. Also, PCL was a non-metaloprotein and its N-terminal 23-amino acid sequence, ATETSFSFQRLNLANLVLNKESS, was determined. Subsequently, MTT method, cell morphological analysis and LDH activity-based cytotoxicity assays demonstrated that PCL was highly cytotoxic to L929 cells and induced apoptosis in a dose-dependent manner. Using caspase inhibitors, a typical caspase-dependent pathway was confirmed. PCL also showed remarkable antifungal activity towards some plant pathogenic fungi. Furthermore, when sialic acid-specific activity was fully inhibited, cytotoxicity and antifungal activity were abruptly decreased, respectively, suggesting a significant correlation between sialic acid-specific site and its bi-functional bioactivities.     

A lectin with antifungal and mitogenic activities from red cluster pepper (<Emphasis Type="Italic">Capsicum frutescens</Emphasis>) seeds

A monomeric mannose/glucose-binding lectin, with a molecular mass of 29.5 kDa and an N-terminal sequence GQRELKL showing resemblance to that of the lectin-like oxidized low-density lipoprotein receptor from the rabbit, has been isolated from the seeds of red cluster pepper Capsium frutescens L. var. fasciculatum. The protocol involved anion exchange chromatography on diethylamino ethanol-cellulose and Q-Sepharose and fast protein liquid chromatography on Mono Q. Its hemagglutinating activity toward rabbit erythrocytes was inhibited by d-mannose and glucose, specifically. The activity was stable from 0 to 40°C, reached a maximum at pH 7 and 8, and was potentiated by Ca²⁺ and Mn²⁺ ions. The lectin showed strong mitogenic activity toward spleen cells isolated from BALB/c mice. The mitogenic activity, which reached a peak at a lectin concentration of 0.27 μM, was inhibited specifically by d(+)-mannose. The lectin was capable of inhibiting the germination of Aspergillus flavus and Fusarium moniliforme spores and hyphal growth in the two fungi.     

A napin-like polypeptide with translation-inhibitory, trypsin-inhibitory, antiproliferative and antibacterial activities from kale seeds.

A heterodimeric napin-like polypeptide with translation-inhibiting and antibacterial activities has been isolated from kale seeds. The purification procedure entailed ion-exchange chromatography on dielthylaminoethyl (DEAE)-cellulose, affinity chromatography on Affi-gel blue gel, ion-exchange chromatography by fast protein liquid chromatography (FPLC) on Mono S, and gel filtration by FPLC on Superdex 75. The napin-like polypeptide was unadsorbed on DEAE-cellulose but adsorbed on Affi-gel blue gel and Mono S. Its 7-kDa large subunit differs in N-terminal amino acid sequence from the 4-kDa small subunit. The polypeptide inhibited translation in the rabbit reticulocyte lysate system with an IC50 of 37.5 nM. This activity was preserved between pH 5 and pH 11, and between 10 and 40 degrees C. It fell to a low level at pH 3 and pH 13 and at 70 degrees C. Antibacterial activity against Bacillus, Megabacterium, and Pseudomonas species and antiproliferative activity against leukemia L1210 cells were observed. However, the polypeptide did not exert antifungal, ribonuclease, or protease activity.     

Purification and characterization of a mannose-binding lectin from the rhizomes of Aspidistra elatior Blume with antiproliferative activity

A lectin with a novel N-terminal amino acid sequence was purified from the rhizomes of Aspidistra elatior Blume by ammonium sulphate precipitation, ion exchange chromatography on diethylaminoethyl-Sepharose and carboxymethyl-Sepharose and gel filtration chromatography on Sephacryl S-100. The A. elatior Blume lectin （AEL） is a heterotetramer with a molecular mass of 56 kDa and composed of two homodimers consisting of two different polypeptides of 13.5 kDa and 14.5 kDa held together by noncovalent interactions. Hapten inhibition assay indicated that hemagglutinating activity of AEL towards rabbit erythrocytes could be inhibited by D-mannose, mannan, thyroglobulin and ovomucoid. The lectin was stable up to 70 ℃ , and showed maximum activity in a narrow pH range of 7.0-8.0. Chemical modification and spectrum analysis indicated that tryptophan, arginine, cysteine and carboxyl group residues were essential for its hemagglutinating activity. However, they might not be present in the active center, except some carboxyl group residues. AEL also showed significant in vitro antiproliferative activity towards Bre-04 （66%）, Lu-04 （60%） and HepG2 （56%） of human cancer cell lines.     

A novel lectin from the wild mushroom Polyporus adusta

A lectin with antiproliferative activity toward tumor cell lines and mitogenic activity toward splenocytes was isolated from the mushroom Polyporus adusta. The lectin was composed of two identical subunits each with a molecular weight of 12 kDa. It was adsorbed on both DEAE-cellulose and Q-Sepharose and unadsorbed on CM-Sepharose. The hemagglutinating activity of the lectin was inhibited by turanose and by a large variety of other carbohydrates. It was adversely affected in the presence of NaOH or HCl at a concentration of 7.5mM and above, and when the ambient temperature was raised above 70 degrees C. All divalent and trivalent metallic chlorides tested at 1.25-10mM including CaCl(2), MgCl(2), ZnCl(2), MnCl(2), and AlCl(3), did not alter the hemagglutinating activity of the lectin. FeCl(3) at 10mM caused the hemagglutinating activity to increase by 100%, but it did not change the lectin activity when tested at lower concentrations up to 5mM.     

A novel lectin from the sponge Haliclona cratera: isolation,characterization and biological activity

A lectin from the Adriatic sponge Haliclona cratera was purified by ion-exchange and gel chromatography. The molecular mass of the lectin is approximately 29 kDa. Purified lectin is rich in hydrophobic and basic amino acids and has an isoelectric point at pH 8.6. H. cratera lectin is relatively heat- and pH-stable. It agglutinates native and trypsinized, papainized and neuraminidase-treated human A, B, O, AB and sheep erythrocytes, and the hemagglutinating activity is independent of Ca(2+), Mn(2+) and Mg(2+) ions; D-galactose and N-acetyl-D-galactosamine are found to be moderate inhibitors of the activity. H. cratera lectin displays cytotoxic effect on HeLa and FemX cells and weak mitogenic effect on human T-lymphocytes pretreated with phytohemagglutinin (PHA).     

Synthesis,antiproliferative and antitrypanosomal activities,and DNA binding of novel 6-amidino-2-arylbenzothiazoles

A series of 6-amidinobenzothiazoles, linked via phenoxymethylene or directly to the 1,2,3-triazole ring with a p-substituted phenyl or benzyl moiety, were synthesised and evaluated in vitro against four human tumour cell lines and the protozoan parasite Trypanosoma brucei. The influence of the type of amidino substituent and phenoxymethylene linker on antiproliferative and antitrypanosomal activities was observed, showing that the imidazoline moiety had a major impact on both activities. Benzothiazole imidazoline 14a, which was directly connected to N-1-phenyl-1,2,3-triazole, had the most potent growth-inhibitory effect (IC₅₀ = 0.25 µM) on colorectal adenocarcinoma (SW620), while benzothiazole imidazoline 11b, containing a phenoxymethylene linker, exhibited the best antitrypanosomal potency (IC₉₀ = 0.12 µM). DNA binding assays showed a non-covalent interaction of 6-amidinobenzothiazole ligands, indicating both minor groove binding and intercalation modes of DNA interaction. Our findings encourage further development of novel structurally related 6-amidino-2-arylbenzothiazoles to obtain more selective anticancer and anti-HAT agents.     

Purification and characterization of a lectin from seeds and cotyledonary callus of Zizyphus mauritiana

A lectin from Zizyphus mauritiana lamk. has been purified from the 25–50% (NH₄)₂SO₄ fraction of crude seed and cotyledonary leaf callus extracts. The lectins purified from the two sources had the same structure and properties. Upon specific adsorption on Sephadex G-100, the lectin (ZML) could be displaced with 0.1 m d-glucose. ZML yielded a single band corresponding to a M_r of 66 kDa both in the absence and presence of β-mercaptoethanol on native- as well as in SDS-PAGE. It is thermostable but pH sensitive and agglutinates human erythrocytes only. Lectin activity could also be detected in the cotyledons, leaf, and stem, and in their cultures, as well as in in vitro regenerants. Cotyledons, cotyledonary leaf and its callus showed much higher lectin activity than seeds. Received: 5 April 1997 / Revision received: 30 June 1997 / Accepted: 31 October 1997     

A novel homodimeric lectin from Astragalus mongholicus with antifungal activity

A novel lectin (AMML) was isolated from a Chinese herb, i.e., the roots of Astragalus mongholicus, using a combination of ammonium sulfate fraction and ion exchange chromatographies. The molecular mass of intact AMML was determined to be 66,396 Da by MALDI-TOF mass spectrometry and 61.8 kDa by gel filtration, respectively. AMML was a dimeric protein composed of two identical subunits each with a molecular mass of 29.6 kDa. The lectin was a glycoprotein with a neutral carbohydrate content of 19.6%. The purified lectin hemagglutinated both rabbit and human erythrocytes, and showed preference for blood types O (native) and AB (trypsin-treated). Among various carbohydrates tested, the lectin was best inhibited by D-galactose and its derivatives with pronounced preference for lactose (3.13 mM). N-terminal amino acid sequence of AMML was determined as ESGINLQGDATLANN. The optimal pH range for lectin activity was between pH 4.5 and 7.5, and the lectin was active up to 65 degrees C. It also exerted antifungal activity against Botrytis cincerea, Fusarium oxysporum, Colletorichum sp., and Drechslera turcia but not against Rhizoctonia solani and Mycosphaerella arachidicola.