[99m Tc]Tc-HYNIC-EcgDf21: A defensin short analogue with potential application in infection foci imaging.

Opportunistic infections are a problem of great relevance in public health and the precise detection and localization of infection in the early stages of the disease is of great importance for patient management as well as cost containment. Our proposal seeks to contribute to developing a new agent that meets the needs of diagnosis and follow-up of fungal and bacterial infections, focused on the design of a radiotracer with the potential for recognition of hidden infection foci. Defensins are plant antimicrobial peptides that not only show activity against plant pathogens but also against human ones. A short analogue of EcgDf1 defensin, EcgDf21d (NH2 -ERFTGGHCRGFRRRCFCTKHC-COOH), was labelled through the formation of a 99m Tc-HYNIC complex which was assessed for physicochemical and biological behaviour both in vitro and in vivo. The [99m Tc]Tc-HYNIC-EcgDf21 labelling procedure rendered a single product with remarkably high RCP and stability in the labelling milieu. The Log p value indicated that [99m Tc]Tc-HYNIC-EcgDf21 has a hydrophilic behaviour, confirmed by the biodistribution profiles. The optimal uptake value was obtained for Candida albicans infection model reaching a lesion/muscle ratio of 3, this correlates with in vitro binding studies, and the lesion can be definitely observed in the scintigraphic images.

Flavanones from Erythrina crista-galli Twigs and Their Antioxidant Properties Determined through In Silico and In Vitro Studies.

Flavonoids are a secondary metabolite group with various bioactivities, such as antioxidants. They are rich in the genus Erythrina, such as Erythrina crista-galli. This research aims to isolate and characterize flavonoids from the twigs of E. crista-galli and determine their antioxidant properties through in silico and in vitro assays. The ethyl acetate extract of E. crista-galli twigs were separated by column chromatography and characterized using spectroscopic methods. Density functional theory (DFT) calculations were performed on the isolated flavonoids and the reference compounds (ascorbic acid and quercetin) to obtain global descriptive parameters and a donor-acceptor map (DAM). We successfully isolated lupinifolin (1) and citflavanone (2) for the first time from E. crista-galli, along with lonchocarpol A (3), which has been discovered previously. The DAM suggests that these flavanones are good antiradicals with effective electron donors. However, they tend to be electron acceptors in methanol. The frontier molecular orbital analysis implies that lupinifolin (1) is a better antiradical than the other flavanones. The DPPH assays show that lupinifolin (1) has the highest antioxidant (antiradical) activity, with an IC50 value of 128.64 ppm. The in silico studies showed similar trends to the in vitro assays using the DPPH method.

First report of Colletotrichum siamense causing anthracnose on Erythrina crista-galli in China.

Erythrina crista-galli L. (Fabaceae) is a popular ornamental plant in tropical and subtropical regions of South Asia. In October 2019, anthracnose-like lesions were observed on the leaves of E. crista-galli planted in Haikou, China. 5-30% of leaves were infected. At first, the circular spots of 1-2 mm in diameter were reddish-brown on the leaves, and then enlarged to circular, subcircular or irregular spots with reddish-brown center and surrounded by a diffuse yellow margin. Neighboring spots sometimes coalesced. Under continuously wet or humid conditions, the lesions expanded quickly, and became gray, subcircular or irregular spots covered by grayish-white mycelium and orange-pink conidial masses. Diseased leaves eventually fell off the trees. To identify the pathogen, diseased leaves were sampled from four gardens. Leaf tissues (5×5 mm) were cut from the margins of typical symptomatic lesions, surface-sterilized in 1% sodium hypochlorite for 1 min, plated on potato dextrose agar (PDA) medium, and incubated at 28.0±0.5℃ in the dark. Similar fungal colonies were obtained from all plated tissues after 3 days. The single-conidium colonies of all isolates were white to pale gray and cottony with visible orange conidial masses. Conidia were one-celled, aseptate, hyaline, straight, cylindrical to fusiform with obtuse ends, and ranged from 14.2-18.6 µm (16.4 µm)× 3.8-5.4 µm (4.7 µm) (n=100). After germination, conidia formed single, brown, oval or slightly irregular appressoria ranging from 8.0 to 11.8 µm (9.6 µm), and from 4.8 to 6.0 µm (5.4 µm). Sexual stage was absent. These characteristics of conidia and appressoria were matched with C. siamense belonging to the C. gloeosporioides complex (Prihastuti et al. 2009; Yang et al. 2009; Weir et al. 20012; Hu et al. 2015). To accurately identify the species, DNA was extracted from four purified isolates (JG-1, JG-3-1, SWS-1-3, SWS-2-1) （Fu et al. 2019）. The internal transcribed spacer of rDNA region (ITS), glyceraldehydes-3-phosphate dehydrogenase (GAPDH), calmodulin (CAL), actin (ACT) and chitin synthase (CHS) genes were amplified and sequenced. The nucleotide sequences were all deposited in GenBank (ITS: MT229427-MT229430, GAPDH: MT250821-MT250824, CAL: MT258893-MT258896, ACT: MT258897-MT258900 and CHS: MT258901-MT258904). Multi-locus phylogenetic analyses (ITS, GAPDH, CAL, ACT and CHS) (Weir et al. 2012) showed that the four isolates were clustered with C. siamense, which was in accordance with BLAST results. Pathogenicity tests of the four isolates were repeated three times on detached leaves (Ji et al. 2019). The conidial suspension (1×106 conidia/mL) was prepared using the conidia from 10-day-old cultures grown on PDA. Two 20-µL drops of conidial suspension were inoculated on non-wounded young healthy leaves, and each isolate was inoculated on 10 leaves. Two 20-µL drops of sterile water were inoculated on non-wounded young healthy leaves as control. The samples were maintained in containers at a relative humidity of 90± 5 per cent inside and 28℃ with a 12-h photoperiod. Gray, subcircular spots similar to the field disease symptoms were observed on the all inoculated leaves after 7 days, whereas no visible symptoms appeared on the non-inoculated leaves. The pathogen was re-isolated from inoculated leaves thus fulfilling Koch's postulates. C. gloeosporioides has been previously reported as a pathogen causing leaf spot on Erythrina (E. indica var. picta, E. variegata var. orientalis) in Guam in 1983 and Brazil in 2012. (Russo et al. 1983; Oliveira et al. 2012). To our knowledge, this is the first report of C. siamense causing leaf spot of E. crista-galli in China.

Gene isolation and structural characterization of a legume tree defensin with a broad spectrum of antimicrobial activity.

MAIN CONCLUSION: The recombinant EcgDf1 defensin has an antimicrobial effect against both plant and human pathogens. In silico analyses predict that EcgDf1 is prone to form dimers capable of interacting with the membranes of microorganisms. Plant defensins comprise a large family of antimicrobial peptides (AMP) with a wide range of biological functions. They are cysteine-rich molecules, highly sequence diverse but with a conserved and stable structure. In this work, a defensin gene (EcgDf1) was isolated from Erythrina crista-galli, a legume tree native from South America. The predicted peptide presents eight cysteines, with a γ-core motif GXCX3-9C and six cysteines distributed like the typical defensin αß motif. The mature EcgDf1 coding sequence was heterologously expressed in Escherichia coli strains and purified by affinity chromatography. Possible dimer and oligomers of EcgDf1 were visible in SDS electrophoresis. Moreover, its 3D structure, determined by homology modeling, docking, and molecular dynamics simulations, was found to be compatible with the formation of homodimers between the ß3 and ß1-loop-α1, leaving the ß2-loop-ß3 free to interact with lipid membranes. The purified recombinant peptide inhibited the growth of several critical plant and human pathogens, like the opportunistic fungi Candida albicans and Aspergillus niger and the plant pathogens Clavibacter michiganensis ssp. michiganensis, Penicillium expansum, Botrytis cinerea, and Alternaria alternata. EcgDf1 is a promising candidate for the development of antimicrobial products for use in agriculture and medicine.

Antimicrobial Evaluation of Erythrinan Alkaloids from Erythrina cristagalli L.

BACKGROUND: Several species of the genus Erythrina have been used as sedative, antidepressant, and anticonvulsant. Erythrina crista-galli is native to the Pampa Biome and is widely used for medicinal purposes. Erythrinan alkaloids exhibit a range of pharmacological properties. OBJECTIVE: The aim of this study was to evaluate the basic fractions and the alkaloids isolated from E. crista-galli bark against a collection of bacteria and fungi for the first time. METHODS: Erythrina crista-galli stem bark was extracted with MeOH under reflux. The crude extract was dissolved in water, acidified and extracted with diethyl ether. Basification of the aqueous solution followed by diethyl ether and ethyl acetate extractions gave the basic ether and basic ethyl acetate fractions. Chromatographic purification of these fractions afforded five known alkaloids: erytharbine (1), erysotrine (2), erysotramidine (3), erysotrine N-oxide (4) and erythratidine (5) along with a new alkaloid named here erythratidine N-oxide (6). Alkaloids 1-6 were investigated against a collection of bacteria and fungi using the broth micro dilution method. RESULTS: In this work, a new alkaloid was isolated from E. crista-galli. The most significant bacterial inhibitory effect of alkaloidal fractions was observed against the Gram-negative Pseudomas aeroginosa (MIC values of 31.25 µg.mL-1). Basic ether fraction displayed good antimicrobial activity against Shigella sonnei with MIC= 62.5 µg.mL-1. Isolated alkaloids 1-6 showed inhibitory activity against all bacteria tested (MIC values of 50-100 µg.mL-1). In addition, the crude extract and alkaloids 1, 2, and 5 also showed good antifungal potential against Candida krusei (MICs between 12.5 and 31.25 µg.mL-1). The previously undescribed alkaloid 6 presented MIC values between 50 and 100 µg.mL-1 against all tested microorganisms. CONCLUSION: In general, as with a considerable number of phytochemicals with antimicrobial activity, alkaloids 1-6 may be considered with potential as antibacterial/antifungal agents. The MIC values of the extract, alkaloidal fractions and compounds 1-6 indicate that, at least in part, the isolates were responsible for the antimicrobial activity observed.

Bioassay-guided isolation of potent aphicidal Erythrina alkaloids against Aphis gossypii from the seed of Erythrina crista-galli L.

BACKGROUND: The cotton aphid (Aphis gossypii Glover) is one of the most invasive pests of cotton. Many botanical phytochemicals have a long history as a source of insecticides, and as templates for new insecticides. This study was undertaken to isolate aphicidal compounds from the seeds of Erythrina crista-galli L. using the bioassay-guided isolation method. RESULTS: Three novel and 11 known Erythrina alkaloids were isolated. Erysodine (9), erysovine (10), erysotrine (8) and erythraline (11) showed moderate to excellent aphicidal activity with LD50 values of 7.48, 6.68, 5.13 and 4.67 ng aphid-1 , respectively. The Potter spray tower bioassay gave corresponding LC50 values of 186.81, 165.35, 163.74 and 112.78 µg ml-1 . A unique substructure, which presents an sp3 methylene at C-8, a non-oxygenated site at N-9 and a conjugated dienes group (Δ1,2 and Δ6,7 ), plays a crucial role in the aphicidal activity. Application of erythraline (11) led to different increases in the activities of superoxide dismutase, catalase and glutathione S-transferase. CONCLUSION: The study demonstrated that the Erythrina alkaloids erysodine (9), erysovine (10), erysotrine (8) and erythraline (11) have potential use as botanical aphicides for commercial application, or as templates for the development of new insecticides. © 2017 Society of Chemical Industry.

A New Erythrinan Alkaloid Glycoside from the Seeds of Erythrina crista-galli.

A new Erythrina alkaloid glycoside, named erythraline-11ß-O-glucopyranoside, was isolated from the seeds of Erythrina crista-galli L., together with five known Erythrina alkaloids and an indole alkaloid. The structure of the new alkaloid glycoside was elucidated by spectroscopic methods, and all of the compounds were evaluated for their antiviral activity against tobacco mosaic virus.

Búsqueda de compuestos bioactivos en Erythrina crista-galli infectada con Phomopsis sp / Search for bioactive compounds in Erythrina crista-galli infected with Phomopsis sp

Una de las estrategias alternativas utilizadas para encontrar nuevas moléculas bioactivas es investigar otros organismos que habitan en los biotopos, y se presta especial atención a las interacciones ambientales entre sí y las plantas que podrían producir aún una mayor cantidad de metabolitos secundarios. Dentro de estos microorganismos, los endófitos son considerados una importante fuente de nuevos metabolitos secundarios de explotación potencial para usos médicos, agrícolas o por otras áreas industriales. En este trabajo la investigación se centró en la búsqueda de compuestos bioactivos de Erythrina crista-galli infectada con un hongo endofítico. Se recolectaron muestras de E. crista-galli de los alrededores de la Provincia de Buenos Aires y de todas estas muestras se aisló e identificó al endófito como perteneciente al género Phomopsis. Se estudiaron las actividades antimicrobiana y antiinflamatoria de los extractos de E. crista-galli relacionadas con algunos de sus usos tradicionales. El extracto acetónico de las ramas jóvenes presentó actividad antibacteriana sobre Bacillus brevis y B. subtilis y antiinflamatoria in vivo (en edema de oreja de ratón) e in vitro (en macrófagos activados por lipopolisacáridos). Del extracto acetónico con actividad antimicrobiana se aislaron e identificaron dos compuestos, daidzeína y coumestrol, y se identificó genisteína por cromatografía contra testigos. La concentración inhibitoria mínima de daidzeína, coumestrol y genisteína frente a B. brevis fue de 137,81, 16,33 y 64,81 µM, respectivamente. En el extracto acetónico con actividad antiinflamatoria se identificaron, por cromatografía contra testigos, daidzeína, genisteína, coumestrol, biochanina A y formononetina. Genisteína, coumestrol y biochanina A presentaron una dosis efectiva 50, de 0,14, 0,54 y 0.75 µg/ml, respectivamente, en el modelo de edema en oreja de ratón. Para evaluar la influencia del endófito en el metabolismo secundario de E. crista-galli se obtuvieron plantines de cultivo. No se pudieron obtener plantines libres de hongo con los tratamientos aplicados. Se sobreinfectó un lote de plantines con Phomopsis sp. y se cuantificó su contenido de coumestrol (0,04 %) y daidzeína (0,05 %) en comparación con el de un grupo control (0,01 % y 0,02 %, respectivamente). Del extracto acetónico de los plantines sobreinfectados se aisló e identificó un glicósido de isorhamnetina. (AU)

Polyphenols from Erythrina crista-galli: Structures, Molecular Docking and Phytoestrogenic Activity.

OBJECTIVES: The current study aimed at exploring the secondary metabolites content of Erythrina crista-galli aqueous methanol extract and assessing its phytoestrogenic and cytoprotective activities. METHODS: Isolation of the compounds was carried out using conventional chromatographic techniques. The structures of the isolated compounds were elucidated based on the UV, NMR spectral data along with their mass-spectrometric analyses. The phytoestrogenic activity was evaluated in-silico and in vitro using the Arabidopsis thaliana pER8: GUS reporter assay and the proliferation-enhancing activity of MCF-7 cells. KEY FINDINGS: Phytochemical investigation of E. crista-galli aqueous methanol extract resulted in the isolation and identification of five flavonoids. The plant extract and its fractions showed significant estrogenic activities compared to controls. CONCLUSION: Five flavonoids were identified from E. crista-galli aqueous methanol extract. To the best of our knowledge, among these flavonoids, apigenin-7-O-rhamnosyl-6-C-glucoside was isolated for the first time from nature. Moreover, luteolin-6-C-glucoside was isolated for the first time from this plant. The plant revealed promising phytoestrogenic activities. This gives rationale to some of its pharmacological properties and suggests additional phytoestrogenic effects, which have not been reported yet.