Crude Fraction Research Articles

EVALUACIÓN DE UNA UNIDAD HPHT ACOPLADA CON SONDA NIR PARA DETERMINAR EL INICIO DE LA PRECIPITACIÓN DE ASFALTENOS BAJO DIFERENTES PRESIONES

Asphaltenes are characterized as the crude oil fraction with the highest molar mass and polarity, presetting mainly (poly) aromatic groups. The flocculation and deposition of asphaltenes causes large losses to the oil industry. Understanding the phase behavior of asphaltenes under conditions closer to those found in reservoirs is important. Therefore, LMCP/UFRJ started operating a high-pressure high-temperature (HPHT) unit coupled to a near-infrared spectrometer probe that can use different flocculants. This work describes the development of a procedure as well as the validation of the results obtained from this unit. Due to the complex composition of crude oil, model systems (MS) were prepared with asphaltenes extracted with n-pentane (C5I) and n-heptane (C7I). The experiments were carried out at atmospheric pressure, titrated with n-heptane, and at 100 and 300 bar titrated with propane. As expected, C7I asphaltenes were more unstable, presenting lower precipitation onset than C5I asphaltenes under ambient conditions and higher pressures. However, for both MS, the stability increased with rising pressure when using propane as a solvent. The proposed method to evaluate asphaltenes precipitation onset was effective for MS in toluene and dead crude oil, and is a promising alternative for investigation of different types of crude oil.

Antioxidant Activity, Cytotoxicity, and Identification of Secondary Metabolites of Kigelia africana from Waterpark Platinum Riau

Antioxidants can prevent reactive oxygen-associated diseases, which trigger carcinogenesis, cardiovascular disease, and premature aging. India and Africa have mostly practiced sausage trees (Kigelia africana) as traditional medicine, whereas Indonesia is still limited. The research aims to determine the phytochemistry and bioactivity of the n-hexane fractions from crude methanol extract of the leaves and bark of the sausage tree as antioxidants and their toxicity. The sausage tree is derived from Waterpark Platinum Riau. The sausage trees were macerated with methanol and fractionated by n-hexane and ethyl acetate. All samples were tested for their antioxidant to 1,1-diphenyl-2-picrylhydrazyl (DPPH) and cytotoxicity to Artemia salina Leach larvae. Phytochemical results of methanol crude extract, n-hexane, and ethyl acetate fractions of leaves and sausage bark showed the presence of all secondary metabolites except alkaloids in the n-hexane fraction. Antioxidant and cytotoxic activity of stem and leaf bark ethyl acetate fraction was stronger than crude methanol extract and n-hexane fraction. Based on LC-MS/MS data, the secondary metabolite components that have contributed strongly antioxidant activity of this study are flavonoid compounds such as kaempferol and the derivatives, lignans (cubebin), and steroids (pregnant).

Prediction and Rationalization of Abundant C-N-H Molecules in Different Environments.

The extreme chemical diversity of CmNnHk molecules is at the same time very important (central in organic chemistry) and difficult to rationalize in the sense that some molecules are abundant and easy to synthesize, while others are rare and difficult to make. Using the recently developed criteria of molecular "magicity", combined with evolutionary structure prediction and quantum-chemical calculations, we study these molecules in a wide range of compositions (0 ≤ m ≤ 13, 0 ≤ n ≤ 4, and 0 ≤ k ≤ 14). "Magic" molecules are defined as those that are lower in energy than any isochemical mixture of molecules with the nearest compositions. The predicted "magic" molecules are in good agreement with compounds found in versatile environments (interstellar and circumstellar media, Titan's lower atmosphere, and crude oil fractions) and in experimental chemistry. This work shows the predictive power of our approach, capable of predicting and explaining stable molecules in complex systems.

Использование бактериальной инокуляции для улучшения вегетативного роста и нутрицевтических качеств китайской капусты в условиях регулируемой агроэкосистемы

Abstract. In recent years, plant inoculation with beneficial plant growth-promoting microorganisms PGPR bacteria in agriculture has been steadily increasing, and they are expected to partially replace chemical fertilizers, pesticides and other growth regulators in the future, providing an environmentally friendly solution for sustainable farming practices. Consequently, the purpose of this study was to investigate the bacterial inoculation of Bacillus cereus to improve the vegetative growth and nutraceutical qualities of Chinese cabbage pak-choi plants grown by hydroponics under regulated agroecosystem conditions. Objects of the study are the plants of Chinese cabbage pak-choy (Brassica campestris ssp. cheninsis L.). Methods. When conducting research, we measured biometric indicators, determined the indicators of crude, dry matter, dietary fiber and mass fraction of ash of pak-choy plants according to GOST. The contents of vitamins, cations, anions and amino acids in plant products were estimated by capillary electrophoresis using “Kapel’-205” device (Russia). The obtained results were processed statistically by means of analysis of variance. Scientific novelty. The efficiency of root inoculation of Bacillus cereus bacteria in the cultivation of pak-choy plants by hydroponics in closed agro-ecosystems has been determined. The dependence of the improvement of quality indicators in the biomass of bacterial inoculation of pak-choi plants has been established. Results. As a result of studies, it was found that the inoculation of B. cereus had a positive effect on the development of the assimilating apparatus of pak-choi plants. The formation of the main morpho-biometric parameters (crude, dry weight, shoot length and number of leaves) of plants was not significantly affected by bacterial inoculation. All obtained values were at the control level. Pak-choi plants inoculated with bacteria showed a significant increase in concentration of mineral elements (K, S and P), amino acids (proline, phenylalanine, valine, histidine and lysine) and vitamins (ascorbic acid and choline).

Evaluation of in vitro Antioxidant, Antimicrobial and Cytotoxic Activities of Crude Ethanol Extract and Fractions of Achillea sintenisii Hub. Mor.

Objective: The Achillea species have been used to treat various ailments due to its anti-inflammatory, hemostatic, spasmolytic and cholagogue effects in the Turkish traditional medicine. However, there is no biological activity studies on some Achillea species except for the well-knowns. This work aimed to determine the antioxidant, antimicrobial and cytotoxic activities of the crude ethanolic extracts and fractions of Achillea sintenisii using in vitro methods. 
 Methods: The antioxidant activity was investigated by DPPH (1,1´-diphenyl- 2-picrylhydrazyl), ABTS (2,2´-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) radical scavenging, total phenol and total flavonoid content, and iron chelating methods. Antimicrobial activity evaluated by micro-plate dilution method against five test organisms. Cytotoxicity was determined by MTT method using MCF-7 breast cancer cell line and PC-3 prostate cancer cell line. Apoptosis was also measured by AO/EB staining. 
 Results: The n-Hexane fractions showed the highest antimicrobial and cytotoxic activities, respectively. Administration of the extracts on the cancer cells showed a concentration dependent inhibition on cell proliferation. The anti-proliferation effect could be via apoptosis and associated with the cell death. 
 Conclusion: The results showed that the extracts demonstrated antioxidant, antimicrobial, and cytotoxic activity, also supports the claims of traditional usage.

Insecticidal Activity of Tannins from Selected Brown Macroalgae against the Cotton Leafhopper Amrasca devastans.

Seaweeds, also known as marine macroalgae, are renewable biological resources that are found worldwide and possess a wide variety of secondary metabolites, including tannins. Drifted brown seaweed (DBSW) is particularly rich in tannins and is regarded as biological trash. The cotton leaf hopper Amrasca devastans (Distant) has caused both quantitative and qualitative losses in cotton production. Drifted brown seaweeds (DBSWs) were used in this study to extract, qualitatively profile, and quantify the levels of total tannins, condensed tannins, hydrolyzable tannins, and phlorotannins in the seaweeds; test their insecticidal activity; and determine the mechanism of action. The largest amount of tannin extract was found in Sargassum wightii Greville (20.62%) using the Soxhlet method (SM). Significantly higher amounts of hydrolyzable tannins (p = 0.005), soluble phlorotannins (p = 0.005), total tannins in the SM (p = 0.003), and total tannins in the cold percolation method (p = 0.005) were recorded in S. wightii. However, high levels of condensed tannins (CTAs) were observed in Turbinaria ornata (Turner) J. Agardh (p = 0.004). A. devastans nymphs and adults were examined for oral toxicity (OT) and contact toxicity (CT) against DBSW tannin crude extract and column chromatographic fractions 1 (Rf = 0.86) and 2 (Rf = 0.88). Stoechospermum polypodioides (J.V. Lamouroux) J. Agardh crude tannin was highly effective against A. devastans using the OT method (LC50, 0.044%) when compared with the standard gallic acid (LC50, 0.044%) and tannic acid (LC50, 0.122%). Similarly, S. wightii fraction 2 (LC50, 0.007%) showed a greater insecticidal effect against A. devastans adults in OT than gallic acid (LC50, 0.034%) and tannic acid (LC50, 0.022%). The mechanism of action results show that A. devastans adults treated with crude tannin of T. ornata had significantly decreased amylase, protease (p = 0.005), and invertase (p = 0.003) levels when compared with the detoxification enzymes. The levels of glycosidase, lactate dehydrogenase, esterase, lipase, invertase, and acid phosphate activities (p = 0.005) of S. wightii were reduced when compared with those of the Vijayneem and chemical pesticide Monocrotophos. In adult insects treated with LC50 concentrations of S. wightii tannin fraction 1, the total body protein (9.00 µg/µL) was significantly reduced (OT, LC50-0.019%). The SDS-PAGE analysis results also show that S. wightii tannin fraction 1 (OT and CT), fraction 2 (OT), and S. polypodioides fraction 2 (CT) had a significant effect on the total body portion level, appearance, and disappearance of some proteins and polypeptides. This study shows that the selected brown macroalgae can be utilized for the safer management of cotton leaf hoppers.

Assessment of asphaltene and resin fractions in crude oil using laser-induced fluorescence spectroscopy based on modified Beer-Lambert (LIFS-MBL)

Crude oil is one of the most significant petrogenic sources of polycyclic aromatic compounds (PACs). These substances play an essential role in the pollution of the marine environment. Therefore, the rapid identification of this pollutant source and its fractions is vital. For this purpose, a fast and on-site method of laser-induced fluorescence spectroscopy based on modified Beer-Lambert (LIFS-MBL) is proposed here using solvent densitometry. Three optical parameters of the self-quenching (K), the extinction (α), and the peak concentration (Cp) are experimentally extracted from MBL graphs. Note that the parameters above are known to be unique characteristics of various crude oils. The corresponding compounds are generally classified into saturate, aromatic, resin, and asphaltene fractions, abbreviated as SARA. Differentiation among these fractions is achieved using the LIFS-MBL method by selecting the optimal excitation wavelength at 405 nm. This line effectively rules out the light aromatic rings and focuses on heavy fractions. The correlation of optical parameters with heavy oil fractions is verified according to analysis of variance. Statistical relations are proposed to calculate crude oil fractions values. The values of light fractions including saturate and aromatic components can also be determined by the heavy fractions. In this method, the test time is notably reduced from four days using the standard methods to less than half an hour according to the presented LIFS-MBL technique.

Assessment of the Environmental Impact of Solid Oil Materials Based on Pyrolysis Oil.

One method of managing used car tires is decomposition by thermochemical conversion methods. By conducting the process at temperatures of 450-750 °C, three fractions are obtained from tires: oil, gas, and solid. The liquid product of the pyrolysis of used car tires is pyrolysis oil, which consists of aromatic, polyaromatic, and aliphatic hydrocarbons. Unconventional building materials were obtained from tire pyrolysis oil and the environmental impact was evaluated. Blocks made from pyrolysis oil showed mechanical strength of up to about 1700 N. No heavy metals or polycyclic aromatic hydrocarbons, which were found in the crude heavy-PO fraction, were detected in the filtrates after incubation of the block obtained from the heavy-PO fraction at 240 °C. The highest inhibition of Sorghum saccharatum shoot (74.4%) and root (57.5%) growth was observed for solid materials from the medium-PO fraction obtained at 240 °C. The most favorable values of the parameters for the process of obtaining blocks based on post-PO were an annealing temperature of 180 °C, time of 20 h, and mass ratio of catalyst to catalyzed oil of 0.045.

Purifying waxes through selective extraction of triacylglycerols from a bioethanol production side-stream using supercritical carbon dioxide

Agro-industrial sourced bioethanol fermentation produces a lipid slurry as a co-product. Corn/sorghum-based (3:2, w/w) post-fermentation lipid slurry was fractionated into crude oil and wax-rich fractions using supercritical carbon dioxide (SC-CO2). The process was optimized for temperature (35–75 °C), pressure (8–40 MPa), and time (2–6 h) for two responses (crude oil and wax yields) via response surface methodology (Box-Behnken design). The optimum conditions were determined as 35 °C, 40 MPa, and 4.1 h for maximum crude oil extraction (47.5 ± 1.4%, w/w) with minimum wax content (1.0 ± 0.2%, w/w). The crude oil fraction had a melting point of 17 °C, whereas the wax-rich fraction exhibited a melting point of 60 °C. The wax-rich fraction was composed of waxy compounds, wax esters, and aldehyde dimers. Overall, this study optimized SC-CO2 extraction conditions to selectively extract triacylglycerols from a bioethanol lipid slurry to purify waxes for food and non-food applications.

In vitro antioxidant, antibacterial, cytotoxic, and epigenetic screening of crude extract and fractions of the marine sponge Neopetrosia exigua from Mauritius waters

The marine sponge Neopetrosia exigua is known as a goldmine of novel compounds, yet its pharmacological activities remain poorly characterised. Herein, this study investigates the bioactivities of N. exigua collected from Mauritius waters. The crude extract (dichloromethane: methanol), hexane, ethyl acetate and aqueous fractions obtained from N. exigua were subjected to in vitro antioxidant assays. Their antibacterial activities were evaluated using the broth microdilution method to determine the minimum inhibitory concentration (MIC). The cytotoxic and epigenetic activities were further screened using the MTT assay and a cell-based image system that measures de-repression of a silenced Green Fluorescent Protein (GFP) reporter gene, respectively. Higher antioxidant activity was recorded for the ethyl acetate fraction as demonstrated by its significant ferric reducing antioxidant power, radical scavenging, and metal chelating activities relative to control (p < 0.05). The best antibacterial profile was presented by the ethyl acetate fraction against Cutibacterium acnes (MIC: 0.039 mg/ml), Streptococcus mutans (MIC: 0.078 mg/ml) and Mycobacterium smegmatis (MIC: 0.313 mg/ml). Similarly, the fraction displayed significant cytotoxicity against the human liposarcoma SW872 cells with IC50 value of 44.34 ± 2.64 µg/ml and GFP re-activation capacity of 43.79 ± 3.19% (p < 0.05). This work conveys interesting data on the antioxidant, antimicrobial, and anticancer properties of N. exigua. In particular, this study indicates the promising potential of N. exigua as a reservoir of epigenetically active agents that can modulate transcription of silenced genes involved in carcinogenesis. Hence, further investigations to isolate the active constituents is actively warranted.

Invitro Anticoccidial Effect of Fractions of Chromolaena Odorata Extract on Sporulating Eimeria Tenalla Oocysts

Coccidiosis is an important cause of digestive system disorders in birds, especially the poultry. The aim of this study was to evaluate the in-vitro anticoccidial effects of the different fractions of Chromolaena odorata extracts on sporulated Eimeria tenella oocysts. Sporulation inhibition bioassay was used to evaluate the activity of crude methanol fraction of Chromolaena odorata extracts; n-hexane, ethyl acetate, and methanol fractions of Chromolaena odorata on sporulation of Eimeria tenella, which was used to cause coccidiosis. in the birds following oral inoculation of experimental broiler birds with oocysts. Post-mortem examination of dead birds showed lesions consistent with Eimeria tenella. The fractions of the Chromolaena odorata were examined for the oocysticidal activities. Amprolium an anticoccidial drug was used as control. The fractions were used in 4 doses: 0 mg/ml; 0.1 mg/ml; 0.2 mg/ml and 0.4 mg/ml. The control drug was used in corresponding doses. The highest oocysticidal activity of tested plant extracts and fractions was recorded. The n-hexane fraction of Chromolaena odorata gave the highest oocysticidal activity. This oocysticidal activity was noticed at lower concentrations but activity was found to be dose-dependent. The highest oocysticidal activity for the n-hexane fraction of Chromolaena odorata was at 65.67±2.08%. Amprolium showed no oocysticidal activity at all concentrations. However, other fractions of the Chromolaena odorata showed different levels of oocysticidal activities. The methanol fractions were secondary to the n-hexane while the ethyl acetate was the least. In conclusion, this study provides the basis for using Chromolaena odorata, especially the n-hexane fraction as preventive agents against coccidiosis and suggests their use as a feed additive in the control of coccidiosis.

Oliveria decumbens Vent. (Apiaceae): Biological screening and chemical compositions

Ethnopharmacological relevanceOliveria decumbens Vent. (Apiaceae), a single aromatic species in Iran, is traditionally used for healing inflammation, gastrointestinal disorders, and infections. Aim of the studyRegarding the importance of O. decumbens in traditional medicine, we aimed to set out the plant's biological screening and analyze the chemical components of the active fractions. Materials and methodsAir-dried O. decumbens aerial parts were macerated by ethanol:water (70:30). Using a liquid-liquid extraction (LLE) technique, n-hexane, dichloromethane (DCM), ethyl acetate (EtOAc), n-butanol (n-BuOH), and water were successively used to fractionate the crude extract into different portions. Various biological activities were performed on the crude extract, fractions, and some experiments on pure compounds. The bioassays were as follows: antibacterial activity against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, and Salmonella typhi (using microplate alamar blue assay; MABA), antifungal activity against Aspergillus niger, A. fumigatus, Candida albicans, C. glabarate, Fusarium lini, Microspurum canis, and Trichophyton rubrum (using agar tube dilution method), antileishmanial activity against Leishmania major and L. tropica using a 96-well serial dilution protocol, anti-inflammatory activity using the respiratory burst assay, cytotoxicity against HeLa (cervical cancer) and BJ (normal fibroblast) cells using MTT assay, insecticidal activity against Tribolium castaneum, Sitophilus oryzae, and Rhyzopertha dominica (using the contact toxicity method), larvicidal activity against Aedes aegypti, anti-DPPH• activity, and cytotoxicity against brine shrimp (Artemia salina) in a lethality assay. Eventually, the phytochemicals from the active fractions were studied by gas chromatography coupled with mass spectrometry (GC-MS). ResultsInterestingly, the DCM fraction was the most active, followed by the n-hexane fraction in the biological assays, including antibacterial (>80% inhibition), leishmanicidal (IC50 (L.major) = 29.4 μg/mL, and IC50 (L.tropica) = 30.0 μg/mL), anti-inflammatory (IC50 = 15.8 μg/mL), insecticidal (>80% inhibition), and larvicidal (100% inhibition of A. aegypti) assays. Further GC-MS analysis of the DCM and n-hexane fractions resulted in the characterization of 12 and 14 phytoconstituents, respectively, compared with the NIST library. Thymol and carvacrol were abundant in both fractions. To lesser quantities, the presence of monoterpenoids (p-cymen-8-ol, thymoquinone, 3-hydroxy-β-damascone, and 3-hydroxy-7,8-dihydro-β-ionol), phenylpropanoids (methoxyeugenol, elemicin, and 4-[(1E)-3-hydroxy-1-propenyl]-2-methoxyphanol, simple phenolics (salicylic acid and 4-methoxy-2,3,6-trimethyl-phenol), and a coumarin (6,7-dimethoxy-coumarin) were detected in the DCM fraction. On the other hand, besides a coumarin and monoterpenoids, the fatty acids (tetradecanoid acid, n-hexadecanoic acid, and linolenic acid) and a sesquiterpene (spathulenol) were observed in the n-hexane fraction. The EtOAc fraction scavenged the DPPH• radicals better than other fractions (IC50 = 41.4 μg/mL), while in brine shrimp lethality assay, the crude extract was more active than n-hexane and DCM fractions with LD50 = 385.20, 660.28, and 699.74 μg/mL, respectively. Surprisingly, the crude extract and fractions were ineffective against assayed fungal strains and tested cancer and non-cancer cell lines. ConclusionsOur findings showed that O. decumbens deserves to be a multi-bioactive medicinal plant, besides its ability for cereal protection against pests. To understand the principal mechanism of action, in silico, in vitro, and in vivo experiments may clarify the ambiguities and even figure out the synergistic behavior of the minor secondary metabolites.

Potential hemostatic compounds targeting urokinase plasminogen activator explored from three Euphorbiaceae species: Euphorbia maculata, Euphorbia humifusa, and Acalypha australis, with bio-affinity ultrafiltration UPLC-MS.

Numerous species of the Euphorbiaceae family, including Euphorbia maculata, Euphorbia humifusa, and Acalypha australis, have been used to manage bleeding disorders. However, few investigations have demonstrated their hemostatic potential, and their procoagulant compounds remain elusive. This study aimed to determine the most active procoagulant extracts from the three species' crude extract (CE) and fractions in order to screen out the active compounds and to analyze their possible mechanisms of action. An integrative approach, comprising prothrombin time and activated partial thromboplastin time evaluations and urokinase-type plasminogen activator (uPA) inhibitory assessment, followed by bio-affinity ultrafiltration paired with UPLC/QTOF-MS targeting uPA and docking simulations, was used. The extracts with highest procoagulant activity were the CE for both E. maculata (EMCE) and E. humifusa (EHCE) and the n-butanol fraction (NB) for A. australis (AANB). The most promising ligands, namely, isoquercetin, orientin, rutin, and brevifolin carboxylic acid, were selected from these lead extracts. All of these compounds exhibited pronounced specific binding values to the uPA target and showed tight intercalation with the crucial side chains forming the uPA active pocket, which may explain their mode of action. The activity validation substantiated their hemostatic effectivity in inhibiting uPA as they had better inhibition constant (Ki) values than the reference drug tranexamic acid. Collectively, the integrative strategy applied to these three species allowed the elucidation of the mechanisms underlying their therapeutic effects on bleeding disorders, resulting in the fast detection of four potential hemostatic compounds and their mode of action.

Extract of Chenopodium album lowers blood pressure in rats through endothelium-dependent and -independent vasorelaxation

To investigate the antihypertensive effect of crude extract of Chenopodium album (Ca.Cr), based on its medicinal use in hypertension. Ca.Cr and its fractions were tested in-vivo in normotensive anesthetized rats for blood pressure-lowering effect. In-vitro experiments were performed on isolated rat aortae to explore the vascular mechanism(s). In normotensive anesthetized rats, Ca.Cr produced a dose-dependent (1-300mg/kg) fall (30%mmHg) in mean arterial pressure (MAP). Among the fractions, nHexane was the most potent (46% fall). In rat aortic rings precontracted with phenylephrine (PE), Ca.Cr and its fractions (except Ca.Aq) produced endothelium-dependent vasorelaxation, which was partially reversed with endothelium removal and by pretreating intact aortic rings with L-NAME (10μM) and atropine (1μM). This relaxation to Ca.Cr and fractions (nHexane, ethylacetate and chloroform) was also eliminated with indomethacin pretreatment, however, it unmasked a vasoconstriction effect with Ca.Cr only. Surprisingly, the aqueous fraction produced a calcium sensitive strong vasoconstriction instead of vasorelaxation. The crude extract and its fractions (except Ca.Aq) also antagonized vasoconstriction induced with high K+ (80mM), suggesting calcium antagonistic effect. The aqueous fraction produced mild vasorelaxation against high K+. This effect was further confirmed when pretreatment of the aortic rings with different concentrations of crude extract and fractions suppressed CaCl2 concentration response curves, similar to verapamil. In acute toxicity test, Ca.Cr extract was found safe up to 5g/kg body weight in mice. These findings suggest that crude extract and fractions of C.album produced vasorelaxant effect through muscarinic receptors linked-NO pathway, prostaglandin (endothelium-dependent) and calcium antagonism (endothelium-independent), which explains the blood pressure lowering effect of C.album in rats.

Lipopeptides from an isolate of Bacillus subtilis complex have inhibitory and antibiofilm effects on Fusarium solani.

Bacillus subtilis species complex is known as lipopeptide-producer with biotechnological potential for pharmaceutical developments. This study aimed to identify lipopeptides from a bacterial isolate and evaluate their antifungal effects. Here, we isolated and identified a lipopeptide-producing bacterium as a species of Bacillus subtilis complex (strain UL-1). Twenty lipopeptides (six iturins, six fengycins, and eight surfactins) were identified in the crude extract (CE) and fractions (F1, F2, F3, and F4), and the highest content of total lipopeptides was observed in CE and F2. The chemical quantification data corroborate with the hemolytic and antifungal activities that CE and F2 were the most hemolytic and inhibited the fungal growth at lower concentrations against Fusarium spp. In addition, they caused morphological changes such as shortening and/or atypical branching of hyphae and induction of chlamydospore-like structure formation, especially in Fusarium solani. CE was the most effective in inhibiting the biofilm formation and in disrupting the mature biofilm of F. solani reducing the total biomass and the metabolic activity at concentrations ≥ 2µg/mL. Moreover, CE significantly inhibited the adherence of F. solani conidia on contact lenses and nails as well as disrupted the pre-formed biofilms on nails. CE at 100mg/kg was nontoxic on Galleria mellonella larvae, and it reduced the fungal burden in larvae previously infected by F. solani. Taken together, the lipopeptides obtained from strain UL-1 demonstrated a potent anti-Fusarium effect inducing morphological alterations and antibiofilm activities. Our data open further studies for the biotechnological application of these lipopeptides as potential antifungal agents. KEY POINTS: • Lipopeptides inhibit Fusarium growth and induce chlamydospore-like structures. • Lipopeptides hamper the adherence of conidia and biofilms of Fusarium solani. • Iturins, fengycins, and surfactins were associated with antifungal effects.

In vitro cytotoxic and apoptotic activity of the Mauritian marine sponge Neopetrosia exigua

Marine sponges belonging to the genus Neopetrosia represent a quasi-inexhaustible source of novel cytotoxic compounds. Yet studies delineating their molecular mechanisms of action in cancer cells remain scarce. We investigated the cytotoxic and apoptosis inducing potential of the Mauritian marine sponge Neopetrosia exigua derived crude extract, hexane and ethyl acetate fraction. Their cytotoxic activity was screened against four cancer cell lines and two non-malignant cell lines via the Alamar Blue metabolic assay. The level of intracellular reactive oxygen species (ROS) production, endogenous antioxidant enzyme activity (catalase and superoxide dismutase) and mitochondrial membrane potential were determined. The ability of the active extract to induce apoptosis in cancer cells and modulate the expression levels of apoptotic markers (caspases and polyADP-ribose polymerase (PARP)) was further evaluated via western blot. The ethyl acetate fraction (NEEAF) displayed the highest inhibitory effect with an IC50 of 6.87 μg/mL against the liver hepatocellular carcinoma cell line (HepG2). Mechanistically, NEEAF induced morphological hallmarks characteristic of apoptosis, increased ROS production, decreased catalase and superoxide dismutase activity and mitochondrial membrane depolarisation in a concentration-dependent manner compared to the control (p&lt;0.05). In addition, NEEAF induced the activation of caspase-9, -7, -3 and cleavage of PARP. Overall, this study provides biochemical evidence for oxidative stress-mediated cytotoxicity and apoptosis in HepG2 cells by NEEAF. Further in-depth investigations are needed to isolate the active constituents, which may potentially lead to the development of novel anticancer therapeutics. Significance: Marine sponges represent an untapped goldmine of structurally unique compounds with interesting anticancer properties. This important initial investigative work will set the stage for more in-depth mechanistic studies and chemical characterisation of potentially novel bioactive compounds from the genus Neopetrosia. This work will also help to strengthen frameworks oriented towards the conservation of Neopetrosia species in the Western Indian Ocean region.

Antibiofilm activity of Equisetum hyemale: interference with quorum sensing

IntroductionBiofilm formation results from successive stages of adhesion and coaggregation/auto aggregation, bacterial motility, and quorum sensing (QS), mediated by chemical signals that determine group microbial behaviour. Thus, it is important to look for alternatives that are capable of interfering with these parameters and, consequently, inhibit the formation of biofilms. Plants are a rich source of research, as they produce secondary metabolites that have important biological activities. In this study, we aimed to evaluate the antimicrobial, antibiofilm, and antiquorum sensing activities of a crude extract, fractions, and major substances produced by Equisetum hyemale against P.Pseudomonas aeruginosa. Methods and resultsThe extracts and the fractions inhibited the bacterial growth of P. aeruginosa PA01 (MIC ranging from 26.2 to 13.1 μg/mL), as did the major substances (minimal inhibitory concentration (MIC) between 0.75 and 0.37 μg/mL). Using the biofilm inhibition technique on polystyrene plates, we observed that the crude extract and fractions were able to inhibit biofilm formation by 29–64%, while the major substances inhibited biofilm formation by 67–79%. Atomic force microscopy allowed us to visualise the decrease in microbial mass. Colony counting also showed that there was a significant difference in the colony counts, in relation to the positive control. Moreover, all extracts and compounds were shown to influence motility (swimming, swarming, and twitching). To evaluate QS inhibition, we assessed violacein, a mediator of QS induced by the CviI autoinducer synthase; the plant extract and the major compounds were able to significantly reduce the production of violacein at subinhibitory concentrations. ConclusionsIn view of this, we have shown that this plant has relevant antibiofilm activity and interferes with biofilm formation factors and QS.

Abstract P2128: Insight Into Glycogen Synthase Kinase 3 Beta Localization Within The Heart: Role On Ischemia Reperfusion Injury

Glycogen synthase kinase 3 beta (GSK3β) is a cytosolic serine-threonine kinase first described for inhibiting glycogen synthase. GSK3β structure shows a small sequence that can work as a mitochondrial target sequence, however mitochondrial GSK3β (mtGSK3β) in the heart, and its role have not been investigated. It is known the enzyme can be activated under ischemia-reperfusion (I/R) injury and modulating mitochondrial permeability transition pore (MPTP) opening. In the nervous system, the truncation of GSK3β activates the enzyme, but in the heart, the truncation function remains unknown. We hypothesized that a small subset of GSK3β is truncated and translocated to the mitochondrial matrix, which might modulate MPTP opening . To test our hypothesis, we studied GSK3β expression by Western blot in mitochondrial fractions isolated from differential centrifugation. We found a percentage of GSK3β present in purified mitochondria (Pmito) truncated at the C-terminal side. To evaluate if mtGSK3β is involved in I/R injury, we study its expression in the fractions isolated from hearts subjected to I/R protocol. We found GSK3β is being truncated and its expression was increased in Pmito from I/R-hearts in comparison to normoxia-hearts (controls). To study the role of mtGSK3β in MPTP regulation we measure the sensitivity of the pore under a GSK3β inhibitor, SB216763 (SB21) by fluorescente of Fura-2-FF in a spectrofluorometer. MPTP Ca 2+ -sensitivity was reduced in crude mitochondria fraction (Cm, mitochondria with other organelles) and increased in Pmito under SB21 treatment. This data suggests mtGSK3β is inactive in Pmito and SB21 is inducing MPTP opening by another mechanism. Since GSK3β can be activated and inhibited by phosphorylation at T390 site (pT390) and S9 site (pS9), respectively, we measure it on mtGSK3β. We found pT390 is not present in Pmito likely missing during truncation, while pS9 is in a low amount. Moreover, pS9 increases on Pmito from hearts subjected to I/R protocol. All the data together showed that a small percentage of GSK3β is truncated and translocated within the mitochondrial matrix. However, this mtGSK3β is not involved in the modulation of MPTP opening, since the lack of a C-terminal side causes its inactivation.

Early cellular and synaptic changes in dopaminoceptive forebrain regions of juvenile mice following gestational exposure to valproate.

Gestational exposure of mice to valproic acid (VPA) is one currently used experimental model for the investigation of typical failure symptoms associated with autism spectrum disorder (ASD). In the present study we hypothesized that the reduction of dopaminergic source neurons of the VTA, followed by perturbed growth of the mesotelencephalic dopamine pathway (MT), should also modify pattern formation in the dopaminoceptive target regions (particularly its mesoaccumbens/mesolimbic portion). Here, we investigated VPA-evoked cellular morphological (apoptosis-frequency detected by Caspase-3, abundance of Ca-binding proteins, CaBP), as well as synaptic proteomic (western blotting) changes, in selected dopaminoceptive subpallial, as compared to pallial, regions of mice, born to mothers treated with 500 mg/kg VPA on day 13.5 of pregnancy. We observed a surge of apoptosis on VPA treatment in nearly all investigated subpallial and pallial regions; with a non-significant trend of similar increase the nucleus accumbens (NAc) at P7, the age at which the MT pathway reduction has been reported (also supplemented by current findings). Of the CaBPs, calretinin (CR) expression was decreased in pallial regions, most prominently in retrosplenial cortex, but not in the subpallium of P7 mice. Calbindin-D 28K (CB) was selectively reduced in the caudate-putamen (CPu) of VPA exposed animals at P7 but no longer at P60, pointing to a potency of repairment. The VPA-associated overall increase in apoptosis at P7 did not correlate with the abundance and distribution of CaBPs, except in CPu, in which the marked drop of CB was negatively correlated with increased apoptosis. Abundance of parvalbumin (PV) at P60 showed no significant response to VPA treatment in any of the observed regions we did not find colocalization of apoptotic (Casp3+) cells with CaBP-immunoreactive neurons. The proteomic findings suggest reduction of tyrosine hydroxylase in the crude synaptosome fraction of NAc, but not in the CPu, without simultaneous decrease of the synaptic protein, synaptophysin, indicating selective impairment of dopaminergic synapses. The morpho-functional changes found in forebrain regions of VPA-exposed mice may signify dendritic and synaptic reorganization in dopaminergic target regions, with potential translational value to similar impairments in the pathogenesis of human ASD.

Toxic effects of the crude methanolic extract of &lt;i&gt;Austroplenckia populnea&lt;/i&gt; leaves and fractions on the growth of &lt;i&gt;Spodoptera frugiperda&lt;/i&gt;

The aim of the current study is to assess the toxicity of &lt;i&gt;Austroplenckia populnea&lt;/i&gt; leaf extracts towards the &lt;i&gt;Spodoptera frugiperda&lt;/i&gt; population in laboratory conditions. Initially, a crude methanolic extract was obtained from dried leaves of &lt;i&gt;A. populnea&lt;/i&gt;. This extract was fractionated using hexane, ethyl acetate, and then methanol. Neonate &lt;i&gt;S. frugiperda&lt;/i&gt; larvae were isolated in plastic containers and fed (once) on corn leaf fragments treated with crude and fractions of the investigated plant extracts. The life cycle of the larvae treated with the extracts was monitored. In addition to the intake of leaves treated with the plant extracts, such variables as larval and pupal mortality rate, larval and pupal weight, and larval, pupal, and adult stage duration were evaluated. The crude, hexane, and methanol extract fractions exerted an antixenosis effect on the feeding behaviour of neonate &lt;i&gt;S. frugiperda&lt;/i&gt; larvae. A significant antibiosis effect was observed for the hexane fraction of the &lt;i&gt;A. populnea&lt;/i&gt; extract, since all of its concentrations promoted a &lt;i&gt;S. frugiperda&lt;/i&gt; larval mortality rate differing from that observed for the control. The pupal mortality rate was low, but it did not differ significantly from the mortality rate observed for the control treatment, which was nil. The larvicidal effect of the &lt;i&gt;A. populnea&lt;/i&gt; leaf extract, mainly of its hexane fraction, has indicated its potential to be used in integrated &lt;i&gt;S. frugiperda&lt;/i&gt; management programs.