A Quantitative Human Red Blood Cell Agglutination Assay for Characterisation of Galectin Inhibitors.

Galectins are a family of beta-galactoside-binding proteins that are characterised by their carbohydrate recognition domain (CRD) and include galectin-1 and galectin-3. These galectins have been implicated in numerous diseases due to their pleiotropic nature, including cancer and fibrosis, with therapeutic inhibitors being clinically developed to block the CRD. One of the early methods developed to characterise these galectins was the hemagglutination of red blood cells. Although it is insightful, this approach has been hampered by a lack of sensitivity and accurate quantification of the agglutination observed. In this study, we aimed to validate a more precise and quantitative method to enable the further investigation of differences between galectins in respect to agglutination induction in different blood groups, as well as the characterisation of small molecule inhibitors. Quantification of hemagglutination was shown to be optimal using U-bottom plates imaged and analysed with FIJI ImageJ rather than flat-bottom plates read for absorbance on an optical density plate reader. Galectin-3-induced red blood cell agglutination efficacy increased significantly from blood group O to A to B. However, for both the galectin-1 monomer and concatemer, a more comparable effect was observed between blood group B and O, but with more potent effects than in blood group A. Inhibition assays for both galectin-3 and galectin-1 induced-hemagglutination were able to demonstrate clear concentration responses and expected selectivity profiles for a set of small-molecule glycomimetics, confirming the historical profiles obtained in biochemical binding and functional cellular assays.

Selection and Characterization of FD164, a High-Affinity Signal Regulatory Protein α Variant with Balanced Safety and Effectiveness, from a Targeted Epitope Mammalian Cell-Displayed Antibody Library.

Phagocytic resistance plays a key role in tumor-mediated immune escape, so phagocytosis immune checkpoints are a potential target for cancer immunotherapy. CD47 is one of the important phagocytosis immune checkpoints; thus, blocking the interaction between CD47 and signal regulatory protein α (SIRPα) may provide new options for cancer treatment. Using computer-aided targeted epitope mammalian cell-displayed antibody library, we screened and obtained an engineered SIRPα variant fragment crystallizable fusion protein, FD164, with higher CD47-binding activity than wild-type SIRPα Compared with wild-type SIRPα, FD164 has approximately 3-fold higher affinity for binding to CD47, which further enhanced its phagocytic effect in vitro and tumor suppressor activity in vivo. FD164 maintains the similar antitumor activity of the clinical research drug Hu5F9 in the mouse xenograft model. Furthermore, FD164 combined with rituximab can significantly improve the effect of single-agent therapy. On the other hand, compared with Hu5F9, FD164 does not cause hemagglutination, and its ability to bind to red blood cells or white blood cells is weaker at the same concentration. Finally, it was confirmed by computer structure prediction and alanine scanning experiments that the N45, E47, 52TEVYVK58, K60, 115EVTELTRE122, and E124 residues of CD47 are important for SIRPα or FD164 recognition. Briefly, we obtained a high-affinity SIRPα variant FD164 with balanced safety and effectiveness. SIGNIFICANCE STATEMENT: Up to now, few clinically marketed drugs targeting CD47 have been determined to be effective and safe. FD164, a potential signal regulatory protein α variant fragment crystallizable protein with balanced safety and effectiveness, could provide a reference for the development of antitumor drugs.

Cytotoxicity of Frutalin on Distinct Cancer Cells Is Independent of Its Glycosylation.

Frutalin is a plant lectin with beneficial immunobiological action, although the access to its active form is still restricted. Moreover, there is a knowledge gap on isoform activity and glycosylation impact on its bioactivity, and recombinant production protocols were seen as ineffective. Here, a simpler and faster production and purification protocol was developed, attaining a yield of purified frutalin 3.3-fold higher than that obtained previously. Hemagglutination assays confirmed that this frutalin isoform could not agglutinate rabbit erythrocytes, while maintaining the native tetrameric structure, as indicated by DLS analysis, and strong interaction with methyl-alpha-galactose, in fluorescence spectroscopy studies. The cytotoxicity of the recombinant frutalin isoform was shown in a broad panel of human cancer cells: colon (HCT116), melanoma (A375), triple-negative breast cancer (MDA-MB-231), and ovarian (IGROV-1). Treatment with 8.5-11.8 µM TrxFTL reduced proliferation of all cancer cells to half in 48 h. This anti-proliferative effect encompasses the p53 pathway since it was significantly reduced in p53-null colon cancer cells (HCT116 p53-/-; GI50 of 25.0 ± 3.0 µM), when compared to the isogenic p53-positive cells (HCT116 p53+/+; GI50 of 8.7 ± 1.8 µM; p < 0.002). This recombinantly produced frutalin isoform has relevant cytotoxic effect and its biological activity is not dependent on glycosylation. The developed E. coli production and purification protocol generates high yield of non-glycosylated frutalin isoform with potent cytotoxic activity, enabling the development of novel anticancer p53-targeting therapies.

Agglutination Activity of Fasciola gigantica DM9-1, a Mannose-Binding Lectin.

The DM9 domain is a protein unit of 60-75 amino acids that has been first detected in the fruit fly Drosophila as a repeated motif of unknown function. Recent research on proteins carrying DM9 domains in the mosquito Anopheles gambiae and the oyster Crassostrea gigas indicated an association with the uptake of microbial organisms. Likewise, in the trematode Fasciola gigantica DM9-1 showed intracellular relocalization following microbial, heat and drug stress. In the present research, we show that FgDM9-1 is a lectin with a novel mannose-binding site that has been recently described for the protein CGL1 of Crassostrea gigas. This property allowed FgDM9-1 to agglutinate gram-positive and -negative bacteria with appropriate cell surface glycosylation patterns. Furthermore, FgDM9-1 caused hemagglutination across all ABO blood group phenotypes. It is speculated that the parenchymal located FgDM9-1 has a role in cellular processes that involve the transport of mannose-carrying molecules in the parenchymal cells of the parasite.

Glycoprotein Mimics with Tunable Functionalization through Global Amino Acid Substitution and Copper Click Chemistry.

Glycoproteins and their mimics are challenging to produce because of their large number of polysaccharide side chains that form a densely grafted protein-polysaccharide brush architecture. Herein a new approach to protein bioconjugate synthesis is demonstrated that can approach the functionalization densities of natural glycoproteins through oligosaccharide grafting. Global amino acid substitution is used to replace the methionine residues in a methionine-enriched elastin-like polypeptide with homopropargylglycine (HPG); the substitution was found to replace 93% of the 41 methionines in the protein sequence as well as broaden and increase the thermoresponsive transition. A series of saccharides were conjugated to the recombinant protein backbones through copper(I)-catalyzed alkyne-azide cycloaddition to determine reactivity trends, with 83-100% glycosylation of HPGs. Only an acetyl-protected sialyllactose moiety showed a lower level of 42% HPG glycosylation that is attributed to steric hindrance. The recombinant glycoproteins reproduced the key biofunctional properties of their natural counterparts such as viral inhibition and lectin binding.

A novel mannose-binding lectin from Liparis nervosa with anti-fungal and anti-tumor activities.

Plant lectins are carbohydrate-binding proteins with nonimmune origin, which can reversibly bind with carbohydrates, agglutinate cells, and precipitate polysaccharides and glycoconjugates. Plant lectins have attracted much attention for their anti-virus, anti-proliferation, and pro-apoptosis properties. Thus the exploration of new lectins has received special attention. Here we purified a mannose-binding lectin from the rhizomes of Liparis nervosa by ion exchange chromatography on DEAE-Sepharose, affinity chromatography on Mannose-Sepharose 4B, and gel filtration chromatography on Sephacryl S-100. The purified L. nervosa lectin (LNL) was identified to be a monomeric protein with a molecular mass of 13 kDa. LNL exhibited hemagglutinating activity towards rabbit erythrocytes, and its activity could be strongly inhibited by D-mannose, N-acetyl glucosamine and thyroglobulin. In vitro experiments showed that LNL exhibited a comparable anti-fungal activity against Piricularia oryzae (Cavara), Bipolaris maydis, Fusarium graminearum, and Sclerotium rolfsii, and anti-proliferation activity against tumor cells by inducing apoptosis. The full-length cDNA sequence of LNL is 715 bp in length and contains a 525 bp open reading frame (ORF) encoding a 110-residue mature protein. It was predicted to have three mannose-binding conserved motifs 'QXDXNXVXY'. The binding pattern of LNL was further revealed by homology modeling and molecular docking. We demonstrated that LNL is not only a potential therapeutic candidate against tumor but also a new anti-fungal agent.

Agaricus bisporus mannose binding protein is not an agglutinating protein.

Agaricus bisporus mannose binding protein (Abmb) demonstrates permeability to epithelial monolayer barrier of the intestine, resistance to gastrointestinal tract conditions and to proteolysis therefore it holds potential as a drug carrier for oral route administration. Abmb also display antiproliferative activity to breast cancer cells and stimulation of immune system thus could potentially be also developed for therapeutic purpose. It is not immunogenic or toxic thereby safe for use. In this paper we further provide evidence that Abmb also lacks of agglutinating activity despite sharing high structural homology to lectins. Abmb is thereby the only mannose specific binding protein that is not member of lectin family. This evidence provides further support on the use of Abmb as pharmaceutical or medicinal agent. Its molecular globularity that may contribute to its lack of agglutination capacity was also evaluated.

Pharmacological activities of Justicia adhatoda.

The current study focused on the pharmacological activities of Justicia adhatoda; including antibacterial, antifungal, phytotoxic, cytotoxic, haemagglutination, insecticidal, in vitro antiglycation, DPPH antioxidant and anti-termite. The crude methanolic extract (Crd. Met. Ext) showed 46.4 % antibacterial activity against M. morganii while the n-hexane fraction showed good (71.4%) and moderate (55.1%) activity against M. morganii and A. baumannii respectively. The EtOAc and aqueous fractions, in most of the cases, showed low to no activity against the selected bacterial pathogens, against A. niger, T. harzianum, A. parasiticus and V. dahliae. The Crd. Met. Ext and fractions showed low activity, against P. notatum and P. digitatum, Crd. Met. Ext. and all fractions were inactive. The percent growth regulation, in case of phytotoxic activity, by Crd. Met. Ext was 25 and 16.6, n-hexane fraction 16.6, 16.6 and 0, CHCl3 25, 8.33 and 0 % and EtOAc fractions 8.33, 8.33 and 0% at 1000 and 100 and 10µg/ml respectively. The aqueous fraction was inactive at all the test concentrations. The results of brine shrimp cytotoxic activity for Crd. Met. Ext was 13.33% and n-hexane fraction 20% at 1000, µg/ml respectively. All of the other fractions showed low to no activity at different test concentrations. All of the test samples were inactive against RBC's of the blood groups at all concentration indicating that the selected plant lack phytolectins and haemagglutination activity. The Crd. Met. Ext and various fraction showed low activity against the test insects i.e. C. pulicaria, C. chinensis and T. castaneum. The absorbance value of plant extract for anti-glycation activity at various concentration were: 0.08, 0.067, 0.053 and 0.04 in comparison with Aminoguanidine 0.04, 0.035, 0.03 and 0.02 respectively at 10, 50, 90 and 130µl. The DPPH radical scavenging activities were proportional to the concentration of the fractions, as the concentration of these increased, the percent scavenging activity also increased. The CHCl3 and EtOAc fractions killed all the termites in 24 hours while Crd. Met. Ext, n-hexane and aqueous fractions took 2-3 days.

Chemoenzymatic synthesis and characterization of N-glycolylneuraminic acid-carrying sialoglycopolypeptides as effective inhibitors against equine influenza virus hemagglutination.

A series of novel sialoglycopolypeptides carrying N-glycolylneuraminic acid (Neu5Gc)-containing trisaccharides having α(2 â†’ 3)- and α(2 â†’ 6)-linkages in the side chains of γ-polyglutamic acid (γ-PGA) were designed as competitive inhibitors against equine influenza viruses (EIV), which critically recognize the Neu5Gc residue for receptor binding. Using horse red blood cells (HRBC) we successfully evaluated the binding activity of the multivalent Neu5Gc ligands to both equine and canine influenza viruses in the hemagglutination inhibition (HI) assay. Our findings show the multivalent α2,3-linked Neu5Gc-ligands (3a-c and 7) selectively inhibit hemagglutination mediated by both influenza viruses and display a strong inhibitory activity. Our results indicate that the multivalent Neu5Gc-ligands can be used as novel probes to elucidate the mechanism of infection/adhesion of Neu5Gc-binding influenza viruses.

A Protein Isolate from Moringa oleifera Leaves Has Hypoglycemic and Antioxidant Effects in Alloxan-Induced Diabetic Mice.

Moringa oleifera has been used in traditional medicine to treat diabetes. However, few studies have been conducted to relate its antidiabetic properties to proteins. In this study, a leaf protein isolate was obtained from M. oleifera leaves, named Mo-LPI, and the hypoglycemic and antioxidant effects on alloxan-induced diabetic mice were assessed. Mo-LPI was obtained by aqueous extraction, ammonium sulphate precipitation and dialysis. The electrophoresis profile and proteolytic hydrolysis confirmed its protein nature. Mo-LPI showed hemagglutinating activity, cross-reaction with anti-insulin antibodies and precipitation after zinc addition. Single-dose intraperitoneal (i.p.) administration of Mo-LPI (500 mg/kg·bw) reduced the blood glucose level (reductions of 34.3%, 60.9% and 66.4% after 1, 3 and 5 h, respectively). The effect of Mo-LPI was also evidenced in the repeated dose test with a 56.2% reduction in the blood glucose level on the 7th day after i.p. administration. Mo-LPI did not stimulate insulin secretion in diabetic mice. Mo-LPI was also effective in reducing the oxidative stress in diabetic mice by a decrease in malondialdehyde level and increase in catalase activity. Mo-LPI (2500 mg/kg·bw) did not cause acute toxicity to mice. Mo-LPI is a promising alternative or complementary agent to treat diabetes.

Effect of GlcNAc-, Man- and Gal-Specific Lectins of the Cytosol, Nuclear Membrane, and Nuclear Matrix of Calf Brain Cortex on Proliferation Activity of Peripheral Blood Lymphocyte.

We studied the effect of GlcNAc-, Man-, and Gal-specific lectins isolated from cell cytosol, nuclear membrane, and nuclear matrix of calf brain cortex, lyophilized, and stored for 5 years on proliferation activity of human peripheral blood lymphocytes and on hemagglutination activity of trypsinized rabbit erythrocytes. Human peripheral blood lymphocytes treated lyophilized lectins demonstrated lower proliferation activity than lymphocytes treated with concanavalin A (positive control), but higher than control lymphocytes (incubated with saline). Lectins produced no effect on hemagglutination activity.

Purification and molecular characterization of a novel mannose-specific lectin from Dioclea reflexa hook seeds with inflammatory activity.

A novel lectin present in Dioclea reflexa seeds (DrfL) was discovered and described in this study. DrfL was purified in a single step by affinity chromatography in a Sephadex G-50 column. The lectin strongly agglutinated rabbit erythrocytes and was inhibited by α-methyl-D-mannoside, D-mannose, and D-glucose. The hemagglutinating activity of DrfL is optimum at pH 5.0-7.0, stable up to 50 °C, and dependent on divalent cations. Similar to other lectins of the subtribe Diocleinae, the analysis by mass spectrometry indicated that DrfL has three chains (α, ß, and γ) with masses of 25,562, 12,874, and 12,706 Da, respectively, with no disulfide bonds or glycosylation. DrfL showed inflammatory activity in the paw edema model and exhibited low cytotoxicity against Artemia sp.

Anti-hemolytic, hemagglutination inhibition and bacterial membrane disruptive properties of selected herbal extracts attenuate virulence of Carbapenem Resistant Escherichia coli.

Expression of a multitude of virulence factors by multi-drug resistant microbial strains, e.g., Carbapenem Resistant Escherichia coli (Family: Enterobacteriaceae; Class: Gammaproteobacteria), is responsible for resistance against beta-lactam antibiotics. Hemolysin production and induction of hemagglutination by bacterial surface receptors inflicts direct cytotoxicity by destroying host phagocytic and epithelial cells. We have previously reported that Berberis aristata, Camellia sinensis, Cyperus rotundus Holarrhena antidysenterica and Andrographis paniculata are promising herbal leads for targeting Carbapenem resistant Escherichia coli. These herbal leads were analyzed for their anti-hemolytic potential by employing spectrophotometric assay of hemoglobin liberation. Anti-hemagglutination potential of the extracts was assessed by employing qualitative assay of visible RBC aggregate formation. Camellia sinensis (PTRC-31911-A) exhibited anti-hemolytic potential of 73.97 ± 0.03%, followed by Holarrhena antidysenterica (PTRC-8111-A) i.e., 68.32 ± 0.05%, Berberis aristata (PTRC-2111-A) i.e., 60.26 ± 0.05% and Cyperus rotundus (PTRC-31811-A) i.e., 53.76 ± 0.03%. Comprehensive, visual analysis of hemagglutination inhibition revealed that only Berberis aristata (PTRC-2111-A) and Camellia sinensis (PTRC-31911-A) exhibited anti-hemagglutination activity. However, Andrographis paniculata (PTRC-11611-A) exhibited none of the inhibitory activities. Furthermore, the pair wise correlation analysis of the tested activities with quantitative phytochemical descriptors revealed that an increased content of alkaloid; flavonoids; polyphenols, and decreased content of saponins supported both the activities. Additionally, flow cytometry revealed that cell membrane structures of CRE were damaged by extracts of Berberis aristata (PTRC-2111-A) and Camellia sinensis (PTRC-31911-A) at their respective Minimum Inhibitory Concentrations, thereby confirming noteworthy antibacterial potential of both these extracts targeting bacterial membrane; hemolysin and bacterial hemagglutination.

A novel N-acetyl-glucosamine lectin of Lonchocarpus araripensis attenuates acute cellular inflammation in mice.

OBJECTIVE AND DESIGN: This study had investigated the anti-inflammatory activity of a seed lectin (LAL) isolated from Lonchocarpus araripensis. MATERIAL/METHODS: LAL was purified by affinity chromatography (chitin column) and ion exchange chromatography (DEAE-Sephacel). In vitro LAL was tested for hemagglutinating activity against rabbit erythrocytes. In vivo LAL was assessed for the anti-inflammatory activity via intravenous injection (i.v.) in Swiss mice (25-30 g; n = 6/group) in models of paw edema and peritonitis. STATISTICAL ANALYSIS: ANOVA (p < 0.05). RESULTS: LAL revealed two bands of 30 and 60 kDa (SDS-PAGE) and exhibited hemagglutinating activity. LAL (10 mg/kg) inhibited the paw edema (77%) and vascular permeability (26%) induced by carrageenan, and the paw edema induced by serotonin (80%), bradykinin (49%), sodium nitroprusside (83%), TNF-α (75%) and PGE2 (64%). LAL also inhibited the neutrophil migration induced by fMLP (70%) or carrageenan (69%). The intravital microscopy showed that LAL inhibited rolling (83%) and adhesion (70%) of leukocytes. LAL anti-inflammatory effect was reversed by its association with N-acetyl-glucosamine. The nine-daily treatment with LAL (10 mg/kg; i.v.) showed no toxicity. CONCLUSION: The novel N-acetyl-D-glucosamine-binding lectin isolated from L. araripensis seeds presents anti-inflammatory effect involving the lectin domain and the inhibition of 5-HT, BK, PGE2, NO, TNF-α and leukocyte rolling and adhesion.

Identification of a Hemagglutinin from Gallibacterium anatis.

Gallibacterium anatis has the ability to hemagglutinate rabbit erythrocytes; however, no bacterial component has yet been associated with this function. In the present work, a protein of approximately 65 kDa with hemagglutinating activity for glutaraldehyde-fixed chicken erythrocytes was purified by ion interchange chromatography from G. anatis F149(T) secreted proteins. The protein was recognized by a rabbit polyclonal serum against a hemagglutinin from Avibacterium paragallinarum. The 65 kDa purified protein presented identity with a G. anatis filamentous hemagglutinin by mass spectrometric analysis. As well, the bacterial surface of G. anatis was labeled by immune gold assays using a polyclonal serum against the 65-kDa protein. A similar protein was recognized in four other G. anatis strains by immunoblots using the same antiserum. The protein binds sheep or pig biotinylated fibrinogen, suggesting an interaction with basement membrane eukaryotic cells components, and the protein is present in G. anatis biofilms. Overall, the results suggest that the 65 kDa hemagglutinin is a common antigen and a potential virulence factor in G. anatis.

Human galectin-2 interacts with carbohydrates and peptides non-classically: new insight from X-ray crystallography and hemagglutination.

Galectin-2 (Gal-2) plays a role in cancer, myocardial infarction, immune response, and gastrointestinal tract diseases. The only reported crystal structure of Gal-2 shows that it is a dimer in which the monomer subunits have almost identical structures, each binding with one molecule of lactose. In this study, we crystallized Gal-2 under new conditions that produced three crystal structures. In each Gal-2 dimer structure, lactose was shown to be bound to only one of the carbohydrate recognition domain subunits. In solution studies, the thermal shift assay demonstrated that inequivalent monomer subunits in the Gal-2 dimer become equivalent upon ligand binding. In addition, galectin-mediated erythrocyte agglutination assays using lactose and larger complex polysaccharides as inhibitors showed the structural differences between Gal-1 and Gal-2. Overall, our results reveal some novel aspects to the structural differentiation in Gal-2 and expand the potential for different types of molecular interactions that may be specific to this lectin.

Synthesis and Antiviral Activity of Novel Phosphorylated Derivatives of Didanosine Against Newcastle Disease Virus in Chicken.

A series of novel phosphorylated derivatives of didanosine were designed and docking studies were performed with a fusion protein of the Newcastle disease virus (NDV), to develop antiviral compounds against NDV. Based on the docking scores and binding affinities, three derivatives were selected. These compounds were synthesized and characterized by IR, (1) H, (13) C, (31) P, and CHN analysis and mass spectra. They were assessed for their in vitro antiviral activity in DF-1 cells; DDI-10 showed better antiviral activity as evidenced by significant reduction in plaque formation and cytopathic effects. DDI-10 was further evaluated in NDV-infected chicken; the survival rates and antioxidant enzyme levels in brain, liver, and lung tissues were estimated. Superoxide dismutase and catalase were significantly raised, and lipid peroxidation and HA titer levels were decreased upon treatment with 1.5 mg/kg body weight of DDI-10 than with 3 mg/kg body weight of DDI. Further histopathological alterations in NDV-infected tissues were restored in chicken treated with DDI-10. Thus, based on the results from in silico, in vitro, and in vivo assays, the novel phosphorylated DDI-10 might be considered as potent antiviral compound for NDV infection in chicken.

Carissa congesta Wight and Benincasa hispida (Thunb.) Cogn. as budding immunomodulatory agents.

Carissa congesta and Benincasa hispida are well-known medicinally important plants associated with diabetes, inflammation, protozal infections and cancer. Here, we emphasized up on the immunomodulatory potential of these plants as the source of lupeol, P-sitosterol and ursolic acid. Petroleum ether extracts of C. congesta roots and B. hispida seeds were subjected to acute toxicity studies. They were screened for its immunomodulatory prospective in rats by Haemagglutination Antibody (HA) titre and Delayed-Type Hypersensitivity (DTH) response using Sheep Red Blood Cells (SRBCs of-0.5x109) as antigens. Carbon Clearance test (Phagocytic Index) was estimated by Indian ink suspension. Complete Freund's Adjuvant (CFA) induced arthritis model interpretation was done by paw edema, kene joint erosion (transverse section), body weights, arthritic index and biochemical levels (RBC, WBC and Hb levels). Both the extracts were found to be therapeutically safe up to 5000 mg/kg. Dosage of 100 mg/kg was not satisfactory; and 500 and 250 mg/kg showed significant immunostimmulation (HA Titre) and immunosuppression (DTH response, 48 h). Benincasa hispida seed and Carissa congesta root extracts showed phagocytic Index of 0.0163±0.003, 0.0145±0.003 and 0.0183±0.003, 0.0176±0.003 at 250 mg/kg and 500 mg/kg, respectively. CFA model revealed that the B. hispida seed and C. congesta root extracts decreased paw volume, knee joint erosion, increased body weights and biochemical parameters with an arthritic index of 1.31±0.12, 1.44±0.15 and 1. 16±0.09, 1.36±0.13 at 250 mg/kg and 500 mg, respectively. The results were interpreted by One- way ANOVA followed by Dunnett test. Extracts showed relevance as promising immunostimulators as compared to control.

Antifungal, phytotoxic and hemagglutination activity of methanolic extracts of Ocimum basilicum.

OBJECTIVE: To report some of the pharmacological features of the medicinal plant Ocimum basilicum. METHODS: In the current studies, Antifungal activity of crude methanolic fraction of Ocimum basilicum was determined against eight pathogenic fungal strains using tube dilution assay. The methanolic fraction was also investigated for phytotoxic and hemagglutination activity. RESULTS: Of the eight strains investigated only Candida albicans and Curvilaria lunata were found to be least affected by plant extract while the rest were significantly inhibited. Moderate phytotoxic activity was observed against lemna minor. Hemagglutination activity showed absence of phytolectins and hence no agglutination of erythrocytes. CONCLUSION: The crude extract of Ocimum basilicum has significant properties against fungi and phytotoxic substances.

Antibacterial, antifungal, phytotoxic, antioxidant and hemagglutination activities of organic fractions of Arisaema tortuosum.

In the current study, the antimicrobial, phytotoxic, haemagglutination and antioxidant potential of crude methanolic extract (Crd. MeOH Ext.) and four organic fractions of Arisaema tortuosum was investigated. All fractions have been screened for antimicrobial properties against eight bacterial pathogens and six fungal pathogens using agar well diffusion and tube dilution method, respectively. Furthermore, the organic fractions were also screened for its phytotoxicity against Lemna minor. Haemagglutination was performed against all human blood groups while free radical scavenging activity was performed to investigate the antioxidant potential of A. tortuosum. Results obtained for antibacterial activity exhibited various degree of zone of inhibition and significant activity was observed for Pseudomonas aeruginosa (27.16±0.60) followed by Bacillus cereus (18.55±0.69) for Crd. MeOH Ext. and chloroform (CHCl3) fraction, respectively while some strains showed resistant at same concentration. Similarly, non-significant antifungal activity was observed for the plant extracts. However, the highest activity among the strains was observed for Alternaria alternata (22±1.24%) and Aspergillus niger (20±1.00%) for ethyl acetate (EtOAc) fraction and Crd. MeOH Ext., respectively. The plant extracts showed good phytotoxic activity with 77.06% inhibition for n-hexane fraction at 1000µg/mL. The result of Nitric Oxide (NO) reducing assay revealed that the plant has less antioxidant activity with 46.06% inhibition for CHCl(3) fraction at 900µg/mL. For haemagglutination assay, the result displayed no agglutination in all the testing concentration. Based on the current results, it can be concluded that A. tortuosum has significant antimicrobial and moderate phytotoxic potential and therefore can leads to antibiotics and herbicide production.