Thin Layer Chromatography Fractionation Research Articles

Bioguided fractionation of procyanidin B2 as potent anti coxsackie virus B and Herpes simplex from cypress (Cupressus sempervirens L.)

ABSTRACT The methanolic extracts of cypress (Cupressus sempervirens L.) collected at three phenological stages were evaluated for their cytotoxicity on Vero cells by MTT test as well as on Herpes simplex (HSV-2) and coxsackie (CVB-3) viruses by plaque reduction assay. The methanolic extract exhibited the highest cytotoxicity against HSV-2 (IC50 = 20.40 µg/mL) and CVB-3 (IC50 = 47.50 µg/mL) at the flowering stage. This extract also exhibited a virucidal action both during the entry of viruses and the release of newly formed virions. The methanolic extract bioguided purification showed that the ethyl-acetate fraction was responsible for virucidal activity. This fraction was endowed with more important selectivity index of 8.15 for HSV-2 and 4.40 for CVB-3. The ethyl acetate fraction was subjected to thin layer chromatography fractionation and identification by HPLC-DAD-ESI-MSn. Results showed that the condensed tannin procyanidin B2 was identified for the first time responsible of the antiviral activity of cypress.

In vitro antimicrobial activities of several extracts endophytic Pseudomonas azotoformans UICC B-91

Pseudomonas azotoformans known for the benefit on agriculture area as soil fertility enhancer. Pseudomonas azotoformans UICC B-91 was successfully isolated from tissues of N. altissima which traditionally known for treating infectious diseases. Therefore, endophytic P. azotoformans might contain antimicrobial compound. This research aims on exploring the antimicrobial activities of several extracts P. azotoformans UICC B-91 against pathogenic microbes. Isolates were extracted with dichloromethane, chloroform, and ethyl acetate. Each crude extract was observed for its potential antimicrobial activity with disc diffusion method against several pathogen microbes. Bioactive compound was assayed by thin layer chromatography (TLC) and screened by bioautography. Results indicated that all crude extract have inhibitory effect on Escherichia coli ATCC 8739, Bacillus cereus ATCC 10876, Staphylococcus aureus ATCC 6583, Staphylococcus aureus ATCC 25923, Salmonella typhimurium ATCC 25241, Pseudomonas aeruginosa ATCC 15442, Bacillus subtilis ATCC 19659 and Candida albicans ATCC 10231. Highest clear zone was on dichloromethane extract. TLC fraction of dichloromethane extract was able to inhibit growth on pathogenic bacteria (except for B. subtilis ATCC 19659 and S. aureus ATCC 6583) and fungi at minimum concentration in the range 125-1,000 μg/mL and 125 μg/mL, respectively. Bioactive compound from endophytic P. azotoformans extracted with dichloromethane have antibacterial and anticandidal effect.

Translating the diatom-grazer defense mechanism to antiparasitic treatment for monogenean infection in guppies

Monogenean parasites impose a major constraint on sustainable aquacultural production and trade due to fish morbidity and mortality. Traditional chemical treatments carry environmental and human health risks and are frequently banned in aquaculture; associated development of pathogen resistance is also common. These highlight the need for alternative. The pennate diatom Phaeodactylum tricornutum is a potent source for food additives and pharmaceuticals due to its robustness and content of high-value compounds. Its extracts also have antibacterial properties. The use of P. tricornutum to develop a treatment against the monogenean parasite Gyrodactylus turnbulli affecting Poecilia reticulate (guppies) was investigated. Ethanolic extracts of freshly broken P. tricornutum biomass were effective against the parasite in vitro and in vivo. Sonication and Kolliphor EL (food-grade nonionic solubilizer and emulsifier) increased the efficacy of the ethanolic extract in vitro; complete detachment and 97% mortality of the parasite were achieved within 240 min at a concentration of 5 μL mL−1. Complete infection clearance from fish was achieved following a bath treatment with 2.5 μL mL−1 ethanolic extract for 24 h. The diatom extract's antibacterial properties, tested against the Gram-positive Streptococcus iniae and Gram-negative Vibrio harveyi, were used to calibrate P. tricornutum culture conditions and extract preparation. Maximal antibacterial effect was obtained with a two-week-old culture, following cell breakage of the wet biomass and extraction in 100% ethanol. Fatty acid composition analysis of thin-layer chromatography fractions of the ethanolic extracts revealed that the free fatty acids liberated during cell breakage, enriched in 16:0, 16:1n7, and eicosapentaenoic acid (20:5n3), are the major active compounds. In-vitro antiparasitic assays and gas chromatography–mass spectrometry analysis confirmed that free 16:1n7 and 20:5n3 are responsible for most of the antiparasitic activity. Results support the use of broken P. tricornutum ethanolic extracts as a potential treatment against monogenean infection in cultured fish.

Antimicrobial Evaluation of Latex and TLC Fractions from the Leaves of Aloe adigratana Reynolds.

Background The highest prevalence and emergence of microbial infections coupled with the threat of antimicrobial resistance constitute a global concern, which entails searching for novel antimicrobial agents. Medicinal plants are among the major sources of medicines for novel drug discovery. Aloe adigratana is one of the endemic Aloe species in Ethiopia where the leaf latex of the plant is traditionally used for the treatment of various pathogenic conditions such as wound, dandruff, malaria, and diabetes. In spite of such claims, there was no scientific study done so far. The aim of the current study was, therefore, to evaluate the antimicrobial effect of leaf latex of A. adigratana and its thin layer chromatography (TLC) fractions. Methods Thin layer chromatography (TLC) separation was employed for isolation of bioactive compounds. Agar well diffusion and microdilution assay method were used to evaluate the antimicrobial actions of the leaf latex and TLC fractions against six bacterial strains and four Candida species of reference and clinical isolate microbial strains. Results Three major fractions, AA01, AA02, and AA03, were identified by TLC. Among the tested microbial strains, the reference strain of Staphylococcus aureus ATCC 29213 (MIC = 0.06 mg/mL) and clinical Candida krusei 242/18 (MIC = 0.14 mg/mL) exhibited higher susceptibility towards AA02, while reference strains of Klebsiella pneumoniae ATCC 700603 (MIC = 0.19 mg/mL) revealed the highest susceptibility towards AA01. The leaf latex displayed the highest activity against Staphylococcus aureus ATCC 29213 and clinical Candida krusei 242/18 with a MIC value of 0.19 mg/mL. Conclusion The leaf latex and TLC fractions were found to be active against the tested bacterial and Candida species. Therefore, this finding supports the traditional claim of Aloe adigratana and the need for characterization of the TLC fractions to provide as lead compounds for further comprehensive antibacterial and antifungal activities.

Sand Dune Streptomyces JB66 Native to the Great Indian Thar Desert Inhibits Multidrug-Resistant Pathogens

The efficacy of an actinomycete strain JB66 recovered from a sand dune soil from the Bikaner district of the Thar desert in inhibiting the growth of various bacterial pathogens was studied. The type strains Staphylococcus aureus, Shigella flexneri, Klebsiella pneumoniae and the clinical isolates Escherichia coli, multidrug-resistant S. aureus and P. vulgaris were included in the antimicrobial assays. Polyphasic characterization of JB66 isolate revealed its identity as Streptomyces (MH762010). It showed 88.99-89.24% sequence similarity with the other members of this genus and share the maximum (88.89%) similarity with Streptomyces sp. ATSC13. The strain JB66 was found to produce a high amount of extracellular L-asparaginase, catalase, gelatinase, protease, tyrosinase and urease enzymes. The partial chemical categorization of the methanolic crude extract of the JB66 strain led to the preliminary identification of various metabolic compounds. The thin-layer chromatography fractionation revealed the presence of prodigiosin pigment or chandramycin, cephalosporin or zeatin, daidzein, demethoxy rapamycin, 4,6-dihydroxy-7-methoxyisoflavone, munumbicins and amiclenomycin like compounds. Bio-autography revealed that the metabolites localized at the Rf values of 0.40, 0.46 and 0.53 in TLC profile had the actual bioactive fractions. UV-VIS spectrum absorbance maxima at 288 nm revealed the presence of an aromatic nucleus.

Bioguided identification of antimicrobial compounds from Chrysopogon zizaniodes (L.) Roberty root essential oil.

Aim: To determine the group of compounds from Chrysopogon zizaniodes root essential oil that have antimicrobial activity. Materials & methods: Thin-layer chromatography coupled to direct bioautography was used to determinate the fraction(s) having antimicrobial activity against methicillin-resistant Staphylococcus aureus(MRSA) and vancomycin-resistant Enterococcus faecalis (VREF). Through GC-MS identification, the fractions with the greatest similarity to the active thin-layer chromatography fraction were used to determinate the MIC. Results: The subfraction 8 from column chromatography was responsible for the best MIC for MRSA(62.5μg/ml) and VREF(125μg/ml). Five compounds possibly responsible for antimicrobial activity were preliminary identified. Conclusion: We suggest that Cedr-8-en-13-ol, could be the more relevant compound involved in the antimicrobial activity in this study.

Comprehensive Petroporphyrin Identification in Crude Oils Using Highly Selective Electron Transfer Reactions in MALDI-FTICR-MS

Petroprophyrins are biomarkers used to extract information about petroleum genesis among other characteristics. Identification of particular types, such as Ni, Cu, Mn, vanadyl (VO), and oxygenated or sulfur-containing porphyrins, typically involves exhaustive isolation and purification processes followed by high-resolution mass spectrometry analysis using atmospheric pressure photoionization [APPI-(+)] or electrospray [ESI-(+)] sources. Simultaneous identification of all porphyrins present in a particular crude oil or organic-matter-rich sediment still remains an analytical challenge. Here, we report a straightforward petroporphyrin isolation and identification methodology based on a single-step liquid–liquid (L–L) extraction (crude oil: acetonitrile) and high-performance thin-layer chromatography fractionation (HPTLC, aminopropyl-bonded silica) followed by selective ionization via electron transfer in matrix-assisted laser desorption ionization (MALDI-FTICR). Mass spectrometric analysis of the extracts r...

Thin-layer chromatography fractionation and densitometric determination of residues of diazinon and chlorpyrifos pesticides in lavender and rosemary leaves

Lavender and rosemary are shrubs that have many medicinal uses. Like any other shrubs, they are susceptible to pest infection which needs pesticides treatment. Residues of pesticides in lavender and rosemary leaves may be hazardous to human health. The main objective of this study was to develop accurate and sensitive methods for the determination of residues of pesticides, namely, diazinon and chlorpyrifos, in lavender and rosemary leaves. Thin-layer chromatography (TLC) fractionation was applied to separate the desired pesticides to be analyzed and to determine the rate of the disappearance of these pesticides from lavender and rosemary leaves. Diazinon and chlorpyrifos were separated from extracts of leaves using silica gel 60 F254 plates. The mobile phase was formed of petroleum ether–ethanol–glacial acetic acid (9.5:0.5:0.1, v/v) and (9.0:1.0:0.1, v/v) as the developing systems for diazinon and chlorpyrifos, respectively, followed by densitometric measurement at 254 nm for both pesticides. The methods were validated over a range of 0.01–l.6 μg band−1 for diazinon and 0.04–2.0 μg band−1 for chlorpyrifos. The detection limits of diazinon and chlorpyrifos were 0.003 and 0.012 μg band−1, respectively. The safe harvest interval (pre-harvest interval; PHI), time in days between the last pesticide application to the crop and the time it can be safely harvested, was suggested to be 21 and 24 days for diazinon and chlorpyrifos, respectively. The developed TLC methods were used for sample cleanup and estimation of the studied pesticides residues in leaves extracts, in addition to the determination of the pre-harvest interval.

Bioactivity of Antibacterial Compounds Produced by Endophytic Actinomycetes from Neesia altissima

Identification of alternate bioactive compounds isolated from microbial endophytes from plants may lead to better solution against antibiotic resistance. Main objective of this research is to determine the bioactivity of antibacterial compounds produced by endophytic actinomycetes, which was isolated from stem bark of Neesia altissima Bl. at Halimun Salak Mount. Detection of bioactive compounds was assayed using Thin Layer Chromatography (TLC) by employing bioautography. Bioactivity was assayed using crude extract against five microorganisms using agar diffusion methods. Tetracycline was used as a positive control. The result of antibacterial activity from endophytic actinomycetes isolate revealed that the bioactive compounds were effective in inhibiting the growth of bacteria Bacillus cereus (1.25 mm), Salmonella typhimurium (5.75 mm), and Shigella flexneri (4.37 mm). Bioactivity of ethyl acetate extract from bioactive compounds of endophytic actinomycetes has an antibacterial activity against bacteria Bacillus cereus (13.00 mm) and Staphylococcus aureus (15.90 mm), at a concentration of 250,000 ppm. Each of TLC fraction was able to inhibit growth of B. cereus at a minimum concentration of 10,000 ppm. In conclusion, the bioactive compounds produced by endophytic actinomycetes can have inhibitory effect towards B. cereus . The category of that compounds are narrow spectrum. The bioactive compound isolated from endophytic actinomycetes may have other implications such as anticancer, antifungal, and antioxidant.

Correlations between Molecular Composition and Adsorption, Aggregation, and Emulsifying Behaviors of PetroPhase 2017 Asphaltenes and Their Thin-Layer Chromatography Fractions

Asphaltenes are polydisperse, compositionally complex solubility fractions of crude oil, with a range of structural motifs that contain two primary structural classes: molecules with single aromatic cores (island) and bridged aromatic moieties (archipelago), each with varying degrees of alkyl substitution (e.g., varied-length alkyl units) and multi-heteroatom functionalities responsible for aggregate formation through weak and strong intermolecular associations. Understanding the link between asphaltene molecular composition and problematic behavior, such as deposit and emulsion formation, is vital to improve crude oils up- and downstream processes in petrochemical industries. Some reports suggest that asphaltene precipitation occurs primarily through self-association of polar functional groups and highlight direct correlations between aggregate size and emulsion stability. Here, we compare the molecular composition of whole and thin-layer chromatography (TLC)-fractioned PetroPhase 2017 asphaltenes charac...

The Antimicrobial Properties of Extracts Isolated from Lichen Parmelia vagans.

This study was aimed to determine the antibacterial properties of extracts isolated from lichen Parmelia vagans.Many pharmaceutical products today have derivatives from living organisms. Recently, Lichens have been found to have secondary metabolites which have been found to have various physiological effects including antimicrobial activity. A lichen is a composite organism composed of a photobiont such as an alga or cyanobacterium and a mycobiont, a fungus. In this study, the antimicrobial activity of extracts from a lichen identified as Parmelia vagans was evaluated.Extracts of Parmelia vagans were initially obtained by using various solvents to find the most efficient extraction of antimicrobial compound(s). We found that extracts obtained using ethyl acetate have the highest antimicrobial activity and were further used to study the properties of the extracted compounds. The antimicrobial activity of the extracts was evaluated using the Resazurin cell viability assay; the microbes investigated were Staphylococcus aureus, Bacillus subtilis, Streptococcus pneumoniae, Klebsiella pneumoniae, and Escherichia coli. It was observed that extracts had no inhibitory effect on the growth of Escherichia coli, and Klebsiella pneumonia at the tested concentrations, but inhibition of growth was observed in Staphylococcus aureus, Bacillus subtilis, and Streptococcus pneumoniae.The constituents of the crude extracts were separated using Thin Layer Chromatography (TLC). After separation, each fraction was tested to identify which contained the antimicrobial activity. From the several fractions, one was found to contain nearly all of the antimicrobial activity and was further evaluated against different strains of bacteria.Our data indicate that purified TLC fractions containing antibacterial activities that inhibit a growth of both gram positive and gram negative bacterial strains.Support or Funding InformationTouro University Nevada Intramural Research Grant

Evaluation of sensory and in vitro anti-thrombotic properties of traditional Greek yogurts derived from different types of milk.

Given that fermented dairy products exhibit high bioactivities against cardiovascular diseases (CVDs), the anti-thrombotic properties, fatty acid profiles and sensory properties of cow, goat and ewe derived Greek yogurts have been assessed and compared. The total lipids (TL), total polar lipids (TPL), total neutral lipids (TNL) were obtained and the polar lipid fractions were further separated by thin layer chromatography (TLC). These lipid samples (TL, TPL and TLC fractions) were subsequently assessed for their biological activity against atherosclerosis based on the in vitro inhibition of Platelet Activating Factor (PAF)-induced platelet aggregation. The fatty acid compositions of all yogurts were analyzed by Gas Chromatography with flame ionization detector (GC-FID). Goat yogurt lipids have been found to exert more potent inhibitory activity (i.e. lower IC50 values in both TL and TPL samples) in contrast to the corresponding fractions of cow and ewe yogurts. The observed sensory data indicates that ewe yogurt was the most palatable of all three Greek yogurts.

Fresh Water Cyanobacteria Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 as an Anticancer Drug Resource.

An increasing number of cancer patients worldwide, especially in third world countries, have raised concern to explore natural drug resources, such as the less explored fresh water filamentous cyanobacteria. Six strains of cyanobacteria (Phormidium sp. CCC727, Geitlerinema sp. CCC728, Arthrospira sp. CCC729, Phormidium sp. CCC731, Phormidium sp. CCC730, and Leptolyngbya sp. CCC732) were isolated (paddy fields and ponds in the Banaras Hindu University, campus) and five strains screened for anticancer potential using human colon adenocarcinoma (HT29) and human kidney adenocarcinoma (A498) cancer cell lines. Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 were the most potent as determined by examination of morphological features and by inhibition of growth by graded concentrations of crude extracts and thin-layer chromatography (TLC) eluates. Cell cycle analysis and multiplex assays using cancer biomarkers also confirmed Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 as cancer drug resources. Apoptotic studies in the cells of A498 (cancer) and MCF-10A (normal human epithelial) exposed to crude extracts and TLC fractions revealed no significant impact on MCF-10A cells emphasizing its importance in the development of anticancer drug. Identification of biomolecules from these extracts are in progress.

Lipid‐soluble fraction of Shakuyaku‐kanzo‐to inhibits myometrial contraction in pregnant women

Shakuyaku-kanzo-to, a Kampo medicine composed equally of shakuyaku and kanzo, is an antispasmodic drug that can inhibit contraction of uterine smooth muscles in pregnant women and rats. We aimed to test the inhibitory effects of water- and lipid-soluble extracts of shakuyaku-kanzo-to, shakuyaku, and kanzo in order to identify the fraction responsible for inhibiting uterine smooth muscle contraction in pregnancy. Myometrial tissues were obtained from pregnant women and rats. The water- and lipid-soluble fractions of shakuyaku-kanzo-to, shakuyaku, and kanzo were obtained using the method of Bligh and Dyer. Lipid-soluble fractions were also partially purified using thin-layer chromatography (TLC) with a chloroform : methanol : water (65:25:4 by volume) solvent system to yield four TLC fractions. The effect of each fraction on oxytocin-induced myometrial contraction was examined in vitro. Lipid-soluble fractions obtained from shakuyaku-kanzo-to and kanzo inhibited myometrial contraction; water-soluble fractions had no effect. Of the four TLC fractions, the inhibitory effect was greatest with TLC fraction 1 (0.75 < Rf value ≤ 1.0). Neither the water-soluble nor the lipid-soluble fraction from shakuyaku inhibited myometrial contraction. These results suggest that lipid-soluble substances with low polarity derived from kanzo are responsible for the inhibitory effect of shakuyaku-kanzo-to on myometrial contraction.

In vitro anthelmintic activity of Millettia auriculata leaves and stems

ObjectiveTo study the anthelmintic activity of phytochemically charaterized chloroform extracts of Millettia auriculata leaves and stems. MethodsChloroform extracts of leaves and stems were prepared. Phytochemical characterisation involved preliminary screening, optimisation of solvent system in thin layer chromatography (TLC) and UV absorption measurement of each bright yellow (in view of a common physical property of flavonoids) TLC fraction for each extract. Anthelmintic activity of each extract was carried out for different concentrations (10, 20 and 40 mg/mL) using Pheretima posthuma. ResultsPhytochemical characterisation of the extracts revealed the presence of steroids/triterpenes, phenolics/flavonoids and carbohydrates with optimum TLC-separation in chloroform-hexane, 19:1. All the investigated extracts possessed significant anthelmintic activity (P<0.001) at a minimal dose of 10 mg/mL when compared with the corresponding concentration of reference drug albendazole. Leaf extract was found to have more efficacy and potency than that of stem and albendazole. ConclusionsThe results of this study indicated that Millettia auriculata can be used as anthelmintic drug. It would be also interesting to find out any novel or existing chemical entities showing anthelmintic activity with mechanism of action.

Bioactive potential of endophytic Myrothecium sp. isolate M1-CA-102, associated with Calophyllum apetalum

Context: Endophytes colonizing medicinal plants are diverse, constituting a rich bioresource for novel natural products.Objective: Myrothecium sp. isolate M1-CA-102 was the most promising among the 16 Myrothecium isolates screened. The bioactive potential of the crude extract from the Calophyllum apetalum Willd. endophytic Myrothecium sp. (Alb. & Schwein.) Ditmar (Incertae sedis) isolate M1-CA-102 and its thin layer chromatography (TLC) fractions were screened based on antioxidant, anti-inflammatory, antimicrobial activities, and cytotoxicity.Materials and methods: The antioxidant activity was measured by 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonic acid (ABTS) radical scavenging capacities. Further, 15-lipoxygenase (15-LOX) and human cyclooxygenase-2 (COX-2) inhibition were assessed at different concentrations (25, 50, and 100 μg/mL for the crude extract, 5, 25, and 50 μg/mL for the TLC fractions). DNA-nicking assay as an indicator of the capacity of extracts to scavenge hydroxyl radical was recorded at a concentration of 50 μg/mL. Cell cytotoxicity was recorded by colorimetric 3-(4,5-dimethylthylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Antibacterial (Bacillus subtilis) and anti-Candida (Candida albicans) assays were performed by the microdilution method.Results: The DPPH and ABTS IC50 values of M1-CA-102 extract were 10 and 6 μg/mL compared with 6.1 and 7.03 μg/mL for the positive control quercetin. The cytotoxicity IC50 value of M1-CA-102 extract was 37 μg/mL, while the M-I TLC fraction was 21 μg/mL. The M1-CA-102 extract gave an IC50 value of 58 and 8 μg/mL for 15-LOX and COX-2, respectively. The MIC values for antimicrobial activity for M1-CA-102 extract ranged from 35 to 54 μg/mL, while for the TLC fractions, it ranged from 91 to 515 μg/mL.Conclusion: The results indicate that Myrothecium M1-CA-102 isolated from C. apetalum is a potential source of natural metabolites of pharmaceutical importance.

Bioactive Lipidic Extracts from Octopus (Paraoctopus limaculatus): Antimutagenicity and Antiproliferative Studies

Fractions from an organic extract from fresh octopus (Paraoctopus limaculatus) were studied for biological activities such as antimutagenic and antiproliferative properties using Salmonella tester strains TA98 and TA100 with metabolic activation (S9) and a cancer cell line (B-cell lymphoma), respectively. A chloroform extract obtained from octopus tentacles was sequentially fractionated using thin layer chromatography (TLC), and each fraction was tested for antimutagenic and antiproliferative activities. Organic extract reduced the number of revertants caused by aflatoxin B1 showing a dose-response type of relationship. Sequential TLC fractionation of the active extracts produced several antimutagenic and/or antiproliferative fractions. Based on the results obtained, the isolated fractions obtained from octopus contain compounds with chemoprotective properties that reduce the mutagenicity of AFB1 and proliferation of cancer cell lines.

Inhibitory effect of some plant crude extracts against cucumber damping-off agents

This study was aimed to investigate the antifungal effect of the crude extracts derived from 37 Iranian plant species against Phytophthora melonis and Pythium aphanidermatum, the causal agents of cucumber root rot and damping off. The crude extracts were obtained using three different solvents; ethanol, methanol and water. The inhibitory effect of these extracts was examined based on the paper disc diffusion method (5 mg per paper disc) with four replications. The ethanolic extract of Acroptilon repens, that showed a remarkable inhibitory effect on the tested oomycetes was selected for further examination. To determine the most effective component, the ethanolic extract of A. repens was fractionated by thin layer chromatography (TLC) and each fraction was tested for the antifungal activity. The potential of the ethanolic extract of A. repens in the control of cucumber damping off agents was evaluated in a completely randomized design with eight replications, under greenhouse conditions. Based on the results obtained from the screening of TLC fractions, it was found that a fraction with Rf value equal to 0.89 had the most inhibitory effect against the tested oomycetes. Greenhouse experiment showed that the soil treatment by the ethanolic extract of A. repens at all the selected concentrations (0.2, 0.4 and 0.6 g per plastic pot) were significantly effective in reducing the disease incidence caused by P. melonis compared to the control. Moreover, seed treatment by 10% w/w of ethanolic extract of A. repens significantly suppressed damping off symptom caused by P. aphanidermatum, when compared to the control.

Chromatography separation, 1H NMR analysis and bioautography screening of methanol extract of Excoecaria agallocha L. from Bhitarkanika, Orissa, India

ObjectiveTo separate the bioactive compounds from crude methanol extract of leaf of Excoecaria agallocha (E. agallocha), a mangrove plant from Bhitarkanika, Odisha, India. MethodsChromatographic separation was carried out by paper, thin layer chromatography (TLC) and column chromatographic techniques. Bioautography of the column fractionated extract and TLC chromatogram were evaluated in vitro for antibacterial and DPPH free radical scavenging activity. The HPLC and 1H NMR spectroscopy of the extract was also carried out to detect the presence of compounds and possible functional groups in the crude plant extract. Results: Evaluation of solvent system for chromatographic separation revealed that n-butanol: acetic acid: water in the ratio of 4:1:1 was the most appropriate for the separation of phenolic compounds in the crude extract. The antibacterial bioautography screening of the TLC fractionated extract of E. agallocha showed positive activity against Staphylococcus aureus (MTCC-1141) and Pseudomonas aeruginosa (MTCC 1034). Evaluation of antioxidant properties of TLC fraction of E. agallocha showed 23.36% scavenging activity using DPPH assay at 40 μL concentration. HPLC chromatogram of the plant extract produced seven peaks for the leaf and two peaks for the stem/bark with the highest peaks at 2.563 and 2.493 minutes of retention time, respectively. The 1H NMR spectroscopic analysis of the plant extract revealed the presence of acyclic aliphatic and α- mono substituted aliphatic group of compounds in the sample. ConclusionsThus, it is apparent from the present study that methanol crude extract of E. agallocha is rich in aliphatic group of compounds which can be purified and characterized for possible pharmaceutical utilization.

Bioassays guided fractionation of Coronopus didymus for its antifungal activity against Sclerotium rolfsii

Sclerotium rolfsii Sacc. is a pathogen of about 500 plant species. In a laboratory screening bioassay, methanolic extracts of 15 mg mL−1 concentrations of different parts of Coronopus didymus (L.) Sm. (Brassicaceae) namely leaf, stem, inflorescence and root reduced the biomass of S. rolfsii by 67%, 26%, 40% and 58%, respectively. Methanolic root extract was successively fractionated with n-hexane, chloroform, ethyl acetate and n-butanol. All the concentrations (3.125–200 mg mL−1) of ethyl acetate fraction completely inhibited the target fungal growth. Two compounds A and B were separated from this fraction by Thin Layer Chromatography (TLC). TLC fraction A was found highly effective against S. rolfsii with MIC value 15.62 µg mL−1 as compared to MIC value 7.81 µg mL−1 of the commercial fungicide mencozeb. This compound may be used for the synthesis of natural product based fungicide for the control of S. rolfsii.