Total Phenolic Fraction Research Articles

Management of Rumex dentatus L. in Vicia faba L. cultivations via Ononis vaginalis Vahl. As a potential bioherbicide

Biological control of undesirable weeds associated with crop cultivation is a sustainable approach that can reduce chemical herbicide dependence. The current study aimed to assess the bio-herbicidal potential of the donor species Ononis vaginalis Vahl. on germination efficiency as well as various growth and physiological parameters of the recipient species Rumex dentatus L., a major broad bean pest (Vicia faba L.). To assess the greatest inhibitory allelopathic effect on the recipient species in mixed (Rumex dentatus L. and Vicia faba L.) and pure cultures (each one separately), two experiments were conducted under laboratory conditions. A Petri dish experiment using O. vaginalis shoot aqueous extract (5%, 10%, 20%, and 40%) and a pot experiment using O. vaginalis shoot crude powder (1%, 2%, 5%, and 10%) were conducted to investigate its biological activity on some growth and physiological parameters of both crop and weed species. O. vaginalis underwent a general phytochemical screening that revealed a high production of allelochemicals, which are secondary metabolites and may have a function like that of natural herbicides. The result showed that the germination of V. faba seeds in both pure and mixed cultures was not significantly affected by low levels of O. vaginalis shoot aqueous extract treatments in pure and mixed cultures, in contrast, those recorded for R. dentatus gradually dropped as levels of O. vaginalis increased in both cultures. Results recorded a significant increase in total phenolics of V. faba shoots and roots under different treatments, except at the high concentrations of crude powder at the donor species level (5 and10%). A reduction in the total phenolic and flavonoid fractions was observed in R. dentatus roots under varying concentration treatments. Conversely, under high concentration treatments, flavonoids decreased in the roots of the mixed culture of R. dentatus but increased in the shoots. In conclusion, allelopathy can be used to suppress weeds in field crops. The study confirmed the use of O. vaginalis into current weed control techniques. O. vaginalis could be explored further for weed suppression in the field.

Application of Cellulase for Contributing Phenolic Release and Conversion in Oats (Avena sativa L.) During Microbial Fermentation.

In this work, Monascus fermentation and cellulase hydrolysis (MCF) of oats (Avena sativa L.) to release and convert phenolic fraction was investigated. Results showed the fungus Monascus grew well with a biomass of 27.03mg/g glucosamine equivalent in MCF, following the destruction of oat cellular structures. SDS-PAGE revealed lots of enzymes were regulated with the α-amylase and FPase activity achieved 139.25 U/g and 1.84 U/g in MCF, respectively. Compared with unfermented oats, content of the total phenolic fractions was increased by 19.2 times in MCF, suggesting a phenolic release process occurred during fermentation. Moreover, the soluble-free chlorogenic acid upregulated to 510.00mg/kg whereas the insoluble-bound ferulic acid downregulated to 193.36mg/kg in MCF, indicating a transformation process of chlorogenic acid from ferulic acid in oats was enhanced. Based on this, a possible pathway of phenolic release and conversion in oats during fermentation with Monascus spp. was revealed. This study was helpful to enrich the theory of microbial metabolism and transformation in grain materials.

Investigation on Nutritional, Phytochemical, and Antioxidant Abilities of Various Traditional Rice Varieties.

Rice (Oryza sativa L.) is a popular grain that is consumed by almost half of the world's population. Rice has several distinct breeds classified as traditional and ancient grains. The consumption of rice by 60% of the world's population is a staple food. Entire grain rice, often known as brown rice, is the unpolished counterpart of white rice, containing bran, germ, and endosperm. Eating whole-grain rice is linked to many health benefits; thus, scientists have focused on identifying and quantifying bioactive chemicals in this meal. The present study assessed the physio-chemical, cooking attributes, mineral contents, phytochemicals, and free radical scavenging capabilities of conventional rice cultivars using known approaches according to particle size analysis. Rice grain lengths varied between 4.10 ± 16 and 6.20 ± 007. The length and thickness ratios of rice were not substantially different (p > 0.05). Kattuyanam has the maximum protein content (9.99 ± 06%) of all the rice varieties that were physio-chemical investigated. It also has highest phenolic content and antioxidant properties compared to gallic acid equivalent (334.900 ± 61g) (GAE)/100g, followed by anthocyanins and flavonoids. Total phenolic content and phenolic fractions of the HPLC profile in tested landraces indicate the presence of bioactive substances derived from traditional rice cultivars and a range of phytonutrients, including phenol, revealing their healthy potentials in the present study.

Direct In Vitro Comparison of the Anti-Leishmanial Activity of Different Olive Oil Total Polyphenolic Fractions and Assessment of Their Combined Effects with Miltefosine.

The bioactive compounds present in the edible products of the olive tree have been extensively studied and their favorable effects on various disease risk factors have been demonstrated. The aim of this study was to perform a comparative analysis of the anti-leishmanial effects of total phenolic fractions (TPFs) derived from extra virgin olive oil with different phenolic contents and diverse quantitative patterns. Moreover, the present study investigated their association with miltefosine, a standard anti-leishmanial drug, against both extracellular promastigotes and intracellular amastigotes of a viscerotropic and a dermotropic Leishmania strain. The chemical compositions of TPFs were determined by high performance liquid chromatography with diode array detection (HPLC-DAD). Analysis of parasite growth kinetics, reactive oxygen species production and apoptotic events were determined by microscopy and flow cytometry. Our results revealed that the presence of oleacein (OLEA) and oleocanthal (OLEO) secoiridoids enhances the anti-leishmanial effect of TPF. The association between TPFs and miltefosine was suggested as being additive in Leishmania infantum and Leishmania major promastigotes, and as antagonistic in intracellular amastigotes, as was evaluated with the modified isobologram method. The obtained data verified that TPFs are bioactive dietary extracts with a strong anti-leishmanial activity and highlighted that fractions that are richer in OLEA and OLEO phenolic compounds possess stronger inhibitory effects against parasites. This study may contribute to improving the therapeutic approaches against leishmaniasis.

Antiradical activity, total phenolic, and total flavonoids extract and fractions of pumpkin (Cucurbita moshata Duch) leaves

Pumpkin (Cucurbita moshata Duch) is one of the Cucurbitaceae plants, which traditionally used to treat skin diseases, measles, jaundice, insomnia, cancer and enhances endurance. Therefore, it was necessary to explore the potential of pumpkin leaves as antiradical. This research aim was to examine the antiradical activity and total phenolic and total flavonoids of pumpkin leaves extract and its fractions using the DPPH method and determined the phenolic and flavonoid contents. Pumpkin leave powder was extracted with methanol. Furthermore, water was added into methanol extract, and be partitioned using n-hexane and ethyl acetate to obtain n-hexane, ethyl acetate, and water fractions. The antiradical activities of pumpkin leave extract and fractions were determined using DPPH (2,2-diphenyl-1-picryhidrazyl) method. Ethyl acetate fraction obtained higher antiradical activities (IC50 6.737±0.196 µg/mL). Correlation of total phenolic and flavonoid contents to inhibit DPPH radical showed that phenolic and flavonoid contents on pumpkin leaves could be inhibited DPPH radical R2 = 0.8994 and R2 = 0.9061, respectively. Extracts and fraction pumpkin leaves show strong antiradical activity with DPPH methods, so their potential as antiradical can developed and can be used as a functional food.

Total Phenolic Fraction (TPF) from Extra Virgin Olive Oil: Induction of apoptotic-like cell death in Leishmania spp. promastigotes and in vivo potential of therapeutic immunomodulation.

Leishmaniasis is a serious multifactorial parasitic disease with limited treatment options. Current chemotherapy is mainly consisted of drugs with serious drawbacks such as toxicity, variable efficacy and resistance. Alternative bioactive phytocompounds may provide a promising source for discovering new anti-leishmanial drugs. Extra Virgin Olive Oil (EVOO), a key-product in the Mediterranean diet, is rich in phenols which are associated with anti-inflammatory, anti-cancer and anti-microbial effects. In this study, we investigate the anti-leishmanial effect of Total Phenolic Fraction (TPF) derived from EVOO in both in vitro and in vivo systems by investigating the contributing mechanism of action. We tested the ability of TPF to cause apoptotic-like programmed cell death in L. infantum and L. major exponential-phase promastigotes by evaluating several apoptotic indices, such as reduction of proliferation rate, sub-G0/G1 phase cell cycle arrest, phosphatidylserine externalization, mitochondrial transmembrane potential disruption and increased ROS production, by using flow cytometry and microscopy techniques. Moreover, we assessed the therapeutic effect of TPF in L. major-infected BALB/c mice by determining skin lesions, parasite burden in popliteal lymph nodes, Leishmania-specific antibodies and biomarkers of tissue site cellular immune response, five weeks post-treatment termination. Our results show that TPF triggers cell-cycle arrest and apoptotic-like changes in Leishmania spp. promastigotes. Moreover, TPF treatment induces significant reduction of parasite burden in draining lymph nodes together with an antibody profile indicative of the polarization of Th1/Th2 immune balance towards the protective Th1-type response, characterized by the presence of IFN-γ-producing CD4+ T-cells and increased Tbx21/GATA-3 gene expression ratio in splenocytes. TPF exhibits chemotherapeutic anti-leishmanial activity by inducing programmed cell death on cell-free promastigotes and immunomodulatory properties that induce in vivo T cell-mediated responses towards the protective Th1 response in experimental cutaneous leishmaniasis. These findings enable deeper understanding of TPF's dual mode of action that encourages further studies.

Bamboo Tar as a Novel Fungicide: Its Chemical Components, Laboratory Evaluation, and Field Efficacy Against False Smut and Sheath Blight of Rice and Powdery Mildew and Fusarium Wilt of Cucumber.

The application of agricultural and forest residues can benefit the environment and the economy; however, they also generate a large amount of byproducts. In this study, bamboo tar (BT), a waste product of bamboo charcoal production, was dissolved in natural ethanol and the surfactant alkyl glucoside to manufacture a 50% (wt/wt) BT emulsifiable concentrate (BTEC) biopesticide. BTEC was screened for fungicidal activity against pathogens. The greatest activity was seen against Ustilaginoidea virens with a half-maximal effective concentration (EC50) value of 6 mg/liter. Four phytopathogenic fungi, Podosphaera xanthii, Rhizoctonia solani, Fusarium oxysporum, and Botrytis cinerea, showed EC50 values of <60 mg/liter. Greenhouse tests in vivo showed 2,000 mg/liter BTEC had a 78.4% protective effect against U. virens, and replicated treatments had an 80.6% protective effect. In addition, replicated 2-year field trials were conducted in two geographic locations with four plant diseases: false smut (U. virens), rice sheath blight (Thanatephorus cucumeris [Frank] Donk), cucumber powdery mildew (P. xanthii), and cucumber Fusarium wilt (F. oxysporum). Results showed that 1,000 to 2,000 mg/liter BTEC significantly inhibited these diseases. Gas chromatography-mass spectrometry analysis showed that the total phenolic mass fractions of two BT samples were 45.39 and 48.26%. Eleven components were detected, and their percentage content was as follows (from high to low): 2,6-dimethoxyphenol > 2- or 4-ethylphenol > 2- or 4-methylphenol > phenol > 4-ethylguaiacol > dimethoxyphenol > 4-methylguaiacol > 4-propenyl-2,6-dimethoxyphenol > 2,4-dimethylphenol. Some of the phenolic compounds identified from the tar might be fungicidally active components. BT is a biochar waste, which has potential as a biofungicide and has promise in organic agriculture. The value of this tar may not be because of any fundamental physical differences from other synthetic fungicides but rather caused by reduced production expenses and more efficient use of waste products.

Pilot continuous centrifugal liquid-liquid extraction of extra virgin olive oil biophenols and gram-scale recovery of pure oleocanthal, oleacein, MFOA, MFLA and hydroxytyrosol

This study provides an efficient pilot-scale liquid-liquid extraction process for the recovery of total phenolic fraction (TPF) from extra virgin olive oil (EVOO) and the further purification of the major TPF constituents (oleocanthal, oleacein, monoaldehydic form of oleuropein aglycone, monoaldehydic form of ligstroside aglycone and hydroxytyrosol) using successively centrifugal partition chromatography and reversed phase high performance liquid chromatography. Special attention was paid for the selection of the appropriate raw material, which should be rich in the targeted high-value compounds. For the extraction of the selected EVOOs, a green technique based on liquid-liquid partitioning using the biphasic system n-Hept/EVOO/EtOH/H2O 3:2:3:2 v/v was carried out, first on pioneering laboratory-scale annular centrifugal extractor (ACE) and then on pilot device. 240 L from three different EVOOs (3x80 L of each EVOO) were extracted yielding 285 g of TPF in approximately 2 h. The second step of this procedure was the purification of the target compounds. A preparative centrifugal partition chromatography (CPC) process was firstly applied for the initial TPF fractionation using a step gradient elution-extrusion method with a series of 4 biphasic systems composed of n-Hept/EtOAc/EtOH/H2O in ratios 4:1:3:2, 3:2:3:2, 2:3:3:2 and 1:4:3:2 (v/v/v/v). Preparative reversed phase high performance liquid chromatography (prep-RP-HPLC) was then carried out to polish the target compounds from the enriched CPC fractions resulting in purity higher than 97%. In parallel, the above purification analysis led to the isolation of four minor TPF constituents of which three are new natural products (EDA lactone, (1R, 8E)-1-ethoxy-ligstroside aglycon and (1S, 8E)-1-ethoxy-ligstroside aglycon) and one ((9E, 11E)-13-oxotrideca-9,11-diencoic acid) is described for the first time as EVOO constituent.

In vitro antioxidant analysis and quantitative determination of phenolic and flavonoid contents of Emilia sonchifolia (L) D.C (Asteraceae) leaf extract and fractions

Emilia sonchifolia (L.) D.C. is a medicinal plant from the family Asteraceae known for a wide range of ethnomedicinal uses such as management of inflammatory diseases, pains, cancer, diabetes, cataract, asthma and liver disease. This research was aimed at assessing the antioxidant potential and quantitative determination of phenolic and flavonoid contents of the extract and fractions of Emilia sonchifolia leaves. The leaves of the plant were processed, extracted and partitioned successively and exhaustively with dichloromethane (DCM) and ethyl acetate. Phytochemical screening was carried out on the methanol extract using standard methods. Antioxidant evaluation of the methanol extract and fractions of Emilia sonchifolia leaves was carried out using Ferric Reducing Antioxidant Power (FRAP) assay and 2, 2, diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay. Quantitative determination of total phenolic content and total flavonoid content was done. The extraction and partitioning yielded the crude extract and respective fractions. The phytochemical screening revealed the presence of tannins, saponins, flavonoids, alkaloids and cardiac glycosides. Antioxidant evaluation of the crude extract and fractions of the plant exhibited significant (p<0.001) antioxidant activity in both assay though not comparable with the standard agent, ascorbic acid. In DPPH assay, the highest percentage inhibition was observed in ethyl acetate fraction (62%) followed by the extract (61%), DCM fraction (55%) and aqueous fractions (54%) at 100 µg/mL. In FRAP assay, the highest reducing power was exhibited by ethyl acetate fraction (0.457 nm) followed by the extract (0.444 nm), DCM fraction (0.441 nm) and aqueous fraction (0.439 nm). Although these activities were significant, they were not comparable to that of the standard drug ascorbic acid (0.914 nm). In total phenolic content assay ethyl acetate fraction had the highest phenolic content (5.804 mg/g), followed by crude extract (2.500 mg/g). The total flavonoid content was observed to be highest in ethyl acetate fraction (10.556 mg/g), followed by crude extract (4.444 mg/g). From this research work it was observed that Emilia sonchifolia leaves have a good antioxidant activity which could be responsible for its wide range of ethnomedicinal activities and this lends scientific credence for the use of this plant in the management of disease conditions.

Phenolic Amides with Immunomodulatory Activity from the Nonpolysaccharide Fraction of Lycium barbarum Fruits.

The fruits of Lycium barbarum have a long history as an edible and medicinal food in Asian regions and have multiple consumption methods; the polysaccharides (LBPs) are commonly considered as their major immunological constituents. The current study revealed that the total phenolic amide moieties from L. barbarum fruits showed greater potential immunomodulatory activity in vivo than did LBPs. Through subsequent investigation on the immunological bioactive phenolic amides, three new phenolic amides, lyciumamides L-N (1-3), as well as 12 analogues, were obtained from the total phenolic amide fraction. Extensive spectroscopic methods were used to elucidate the new structures. Compounds 4-6 and 15 significantly promoted LPS-stimulated B splenocyte, while compounds 4-6 displayed accelerative effects on the proliferation of Con A-stimulated T lymphocytes at a concentration of 20.0 μg/mL. These data indicated that extracts from L. barbarum fruits enriched with phenolic amides could be developed as a nutritional dietary supplement for immunocompromised individuals.

Influence of fungicide residues and in vitro gastrointestinal digestion on total antioxidant capacity and phenolic fraction of Graciano and Tempranillo red wines

The effect of fenhexamid, mepanipyrim and cyazofamid fungicides on in vitro bioavailability of antioxidant activity and phenolic compounds of Tempranillo and Graciano red wine was studied by simulating the digestive process by dialysis in semipermeable membranes. Determination of antioxidant activity was through reaction with the DPPH • radical and the measurement of phenolic compounds was made with liquid chromatography with diode detector (HPLC-DAD). Fenhexamid, mepanipyrim and cyazofamid reduce the total polyphenol content in both wines. During dialysis there was a large loss of total polyphenols (80–90%) and of antioxidant activity (> 90%). The bioavailability of the phenolic compounds is lower than that for non-treated wines and the highest dialization percentages were found for stilbenes > 50%. While for the remaining phenolic fraction the order is the following hydroxycinnamic derivatives > anthocyanins > flavonols.

Oleocanthal as a Promising Natural Therapeutic Agent: the Experimental Leishmaniasis Paradigm

Oleocanthal (OC) is a phenolic compound of Extra Virgin Olive Oil (EVOO), the main source of fatty acids in the Mediterranean diet that is associated with beneficial health effects such as reduced incidence of cardiovascular mortality and cancer. OC is responsible for the localized irritation in the oropharangeal region, is homologous with the non‐steroidal anti‐inflammatory drug ibuprofen and several studies evidenced relevant pharmacological properties of OC in different pathogenic processes. Our group has previously elucidated the anti‐leishmanial effect of Total Phenolic Fraction derived from EVOO and of the phenolic compound Oleuropein. Current study aimed to evaluate the effect of OC in murine experimental models of cutaneous and visceral leishmaniasis since there is no effective vaccine and the chemotherapeutic treatments have serious drawbacks such as toxicity, parasite resistance and high cost. BALB/c mice were infected either subcutaneously in the footpad with 106 L. major promastigotes (Cutaneous Model ‐ CM) or intravenously in the tail vein with 107 L. infantum promastigotes (Visceral Model ‐ VM), according to EC regulation conformed by FASEB Statement of Principles for the use of Experimental Animals. At 7 days post infection, mice were intraperitonealy treated with 5 and 10 mg/kg body weight of OC, 3 days per week for 5 weeks. Mice of the positive control group received orally 15 mg/kg b.w of the reference drug miltefosine (HePC), for 25 days. In the CM, results demonstrate significant reduction of the footpad swelling in OC‐treated mice along with the reduction of parasite load in the popliteal lymph nodes, 1 month post‐treatment termination. Correspondingly, in the VM, OC promoted a severe reduction of parasite burden in target organs, liver and spleen at 10 days post‐treatment termination. Moreover, in order to validate the profile of cytokines induced by OC treatment, intracellular cytokine production in splenocytes of all experimental groups was determined for IL‐4 (Th2‐type cytokine) and IL‐12 (Th1‐type cytokine) at 48h post Soluble Leishmania Antigen in vitro re‐stimulation and revealed that OC was able to stimulate important reduction in IL‐4 production by CD4+ T cells in both CM and VM, while the IL‐12 production was equal in all experimental groups. Furthermore, specific antibodies of both IgG1 and IgG2α isotypes, were detected in the plasma of OC‐treated mice. These data indicate that OC treatment resulted in the induction of a mixed Th1/Th2 immune response moderately polarized towards Th1 type that effectively mediated the elimination of parasitic spread in both disease models. Therefore, OC appears as a potential therapeutic agent into the leishmaniasis drug discovery pipeline.Support or Funding InformationThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Identification of bioactivity markers of Sorbus domestica leaves in chromatographic, spectroscopic and biological capacity tests: Application for the quality control

This work is aimed at establishing analytical and bioactivity markers for the quality control of polyphenol-rich extracts of Sorbus domestica leaves – an antioxidant and anti-inflammatory agent. The potential marker analytes, representative of the main groups of S. domestica polyphenols, were pre-selected by LC–MS/MS analysis of the methanol-water (7:3, v/v) leaf extract (MED). For structural identification, two of them (rare flavonoids) were isolated from MED by preparative HPLC and analysed by 1D and 2D NMR (COSY, HMBC, HMQC) spectroscopy. The in vitro tests revealed that all analytes increase the non-enzymatic antioxidant capacity of plasma, effectively protect human plasma components against oxidative/nitrative damage induced by peroxynitrite (reduce the levels of thiobarbituric acid-reactive substances and 3-nitrotyrosine), and inhibit lipoxygenase and hyaluronidase. Considering the activity parameters and concentration levels, the highest contribution (70–80%) to the activity of the total phenolic fraction of MED was attributed to flavanols represented by (–)-epicatechin, procyanidin B2 and procyanidin C1. However, taking into account the observed synergistic effects, flavonoid and phenolic acid fractions (represented by chlorogenic acid, quercitrin, rutin, quercetin and its 3-O-(2′'-O-β-d-glucopyranosyl)-α-l-rhamnopyranoside and 3-O-(2′'-O-β-d-xylopyranosyl)-α-l-rhamnopyranoside) had also a substantial impact. All nine markers were thus applied as calibration standards, and a simple, accurate, reproducible and fully validated RP-HPLC-PDA method for the standardisation of the target extracts was proposed.

In vitro Screening of Antileishmanial Activity of Natural Product Compounds: Determination of IC50, CC50 and SI Values

Neglected tropical diseases gain the scientific interest of numerous research programs in an attempt to achieve their effective control or elimination. In this attempt, more cutting-edge public health policies and research are needed for the discovery of new, safer and effective drugs originated from natural products. Here, we describe protocols for the in vitro screening of a natural product-derived compound required for the determination of its antileishmanial potency. For this purpose, the Total Phenolic Fraction (TPF) derived from extra virgin olive oil is evaluated through the in vitro cell culture method against extracellular promastigote and intracellular amastigote Leishmania spp. forms. The aim of this article is to describe a step-by-step procedure that can be easily applied to accurately estimate the 50% inhibitory concentration (IC50), the 50% cytotoxic concentration (CC50) and the selectivity index (SI) via the resazurin reduction assay. These protocols are based on the ability of resazurin (oxidized blue form) to be irreversibly reduced by enzymes in viable cells and generate a red fluorescent resorufin product and can be easily expanded to the investigation of the antimicrobial activity in other microorganisms.

Phenolic Composition, Antioxidant Properties, and Inhibition toward Digestive Enzymes with Molecular Docking Analysis of Different Fractions from Prinsepia utilis Royle Fruits.

The present study investigated the phenolic profiles and antioxidant properties of different fractions from Prinsepia utilis Royle fruits using molecular docking analysis to delineate their inhibition toward digestive enzymes. A total of 20 phenolics was identified and quantified. Rutin, quercetin-3-O-glucoside, and isorhamnetin-3-O-rutinoside were the major phenolic compounds in the total phenolic fraction and flavonoid-rich fraction. The anthocyanin-rich fraction mainly contained cyanidin-3-O-glucoside and cyanidin-3-O-rutinoside. All of the fractions exhibited strong radical scavenging activities and good inhibition on cellular reactive oxygen species (ROS) generation in H2O2-induced HepG2 cells, as evaluated by DPPH and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assays. Moreover, the powerful inhibitory effects of those fractions against pancreatic lipase and α-glucosidase were observed. The major phenolic compounds that were found in the three fractions also showed good digestive enzyme inhibitory activities in a dose-dependent manner. Molecular docking analysis revealed the underlying inhibition mechanisms of those phenolic standards against digestive enzymes, and the theoretical analysis data were consistent with the experimental results.

Evaluation of total phenolic fraction derived from extra virgin olive oil for its antileishmanial activity

BackgroundLeishmaniasis is a neglected and emerging disease with varying clinical manifestations. The current treatment options rely on limited chemotherapy with serious drawbacks. Thus, there is an increasing interest in the identification of new candidates for designing potent, less toxic and low-cost drugs. PurposeThe purpose of this study was to evaluate the potential antileishmanial activity of the total phenolic fraction (TPF) derived from extra virgin olive oil (EVOO) when added in in vitro and in vivo experimental models of Leishmania infection. Study designWe investigated the in vitro antileishmanial activity of TPF against two Leishmania species: a viscerotropic (L. infantum) and a dermotropic (L. major) strain. The antileishmanial effect was also tested in vivo in a murine cutaneous leishmaniasis model using L. major-infected BALB/c mice. MethodsSeparation and analytical methodologies were applied in order to extract the olive oil phenols (TPF) and determine the concentration of the major ones, respectively. The in vitro antileishmanial activity of TPF against promastigotes and intracellular amastigotes was determined by the resazurin cell viability assay. The TPF-induced nitric oxide synthesis by L. infantum and L. major -infected J774A.1 macrophages was determined using the Griess reaction, while the respective generation of reactive oxygen species was assessed by flow cytometry. Moreover, L. major-infected BALB/c mice were treated with TPF and its in vivo therapeutic effect was determined as reduction of the footpad swelling. ResultsOur data showed that TPF exhibits inhibitory effect against cell free promastigotes and intracellular amastigotes of both L. infantum and L. major parasite strains. TPF demonstrated to be selectively active against Leishmania amastigotes and its antileishmanial activity was possibly mediated by reactive nitrogen and oxygen intermediates generated from the infected J774A.1 macrophages. Furthermore, administration of TPF in BALB/c mice infected with L. major caused significant reduction of footpad swelling demonstrating in vivo its antileishmanial effect. Based on HPLC-DAD analysis the major components of TPF are tyrosol, hydroxytyrosol, oleacein and oleocanthal. ConclusionThis study brings a new low-cost candidate to the leishmaniasis drug discovery pipeline, upon further pharmacological investigation.

Inhibitory effect of Bryophyllum pinnatum (Lam.) Oken leaf extract and their fractions on α-amylase, α-glucosidase and cholinesterase enzyme

Bryophyllum pinnatum (Lam.) Oken leaves are employed as food and as traditional medicines. This study investigates the antioxidant activity (reducing power, DPPH, ABTS, FRAP, H2 O2 scavenging ability and metal ion chelating), carbohydrate digesting enzymes activity and inhibitory activity of cholinergic enzyme of aqueous extract and fractions (n-hexane, ethyl acetate, n-butanol, residual aqueous fraction) of B. pinnatum leaves were investigated. Results showed that aqueous extract of B. pinnatum exhibited DPPH radical scavenging abilities, iron chelation, hydrogen peroxide scavenging abilities and reducing power (Fe3+- Fe2+). B. pinnatum aqueous extract also had considerably α-amylase and α-glucosidase inhibitory activities with IC50 values 149.20 ± 14.44 μg/mL and 126.15 ± 9.76 μg/mL respectively. Our findings indicated that ethyl acetate fraction contained a considerably higher (p < 0.05) amount of total phenolic, flavonoids, total antioxidant, FRAP, metal ion, ABTS and DPPH radical scavenging activity than other solvent fractions. Furthermore, the ethyl acetate fraction elicited a significantly higher (p < 0.05) inhibitory effects on α-glucosidase (IC50 = 70.90 ± 1.23 μg/ml), α-amylase (IC50 = 62.45 ± 1.22 μg/ml), acetylcholinesterase (AChE) (IC50 = 66.75 μg/mL) and butyrylcholinesterase (BChE) (IC50 = 62.97 μg/mL) activities than other fractions. Hence, B. pinnatum leaves were rich in biologically active components; thus, could be employed to formulate new plant-based pharmaceutical and nutraceutical drugs to improve human health.

Antioxidant effects of an olive oil total polyphenolic fraction from a Greek Olea europaea variety in different cell cultures

BackgroundNumerous studies have been carried out concerning the advantageous health effects, especially the antioxidant effects, of olive oil's (OO) individual biophenolic compounds, but none until now for its total phenolic fraction (TPF). Plenty of evidence, in research about nutrition and healthiness, points out that it is the complex mixture of nutritional polyphenols, more than each compound separate, which can synergistically act towards a health result. PurposeThe aim of the present study was to examine the antioxidant properties of an extra virgin olive oil (EVOO) total polyphenolic fraction, from a Greek endemic variety of Olea europaea in cell lines. MethodsEVOO from a Greek endemic variety was used for the extraction of a total polyphenolic fraction, using a green CPE‑based method. The redox status [in terms of ROS, GSH, TBARS, protein carbonyls] was assessed at a cellular level, particularly in EA.hy926 endothelial, HeLa, HepG2 hepatic cells and C2C12 myoblasts. Moreover, the levels of glutamate-cysteine ligase catalytic subunit (γ-GCLc) of GSH, one of the most important antioxidant enzymes, were assessed by western blot. ResultsAccording to the results, TPF improves the redox profile of all cell lines, mainly by increasing GSH and its catalytic subunit, while at low, not cytotoxic TPF concentrations there was a decrease in TBARS and carbonyls. Regarding ROS levels a reduction was observed only in the HepG2 cell line, contrary to the other cell lines, that there is no statistically significant difference. ConclusionThe TPF appeared to protect cells from oxidative stress due to the strong antioxidant activity of its polyphenols. This could have interesting implications in development of new products based on this olive oil to provide protection and treatment against harmful effects of free radicals.

Comparative Evaluation of Soluble and Insoluble-Bound Phenolics and Antioxidant Activity of Two Chinese Mistletoes.

Mistletoes are used medicinally in order to treat various human illnesses. Few studies have reported on the phenolic content and antioxidant properties of Chinese mistletoes (CMs). In this work, the total phenolic content (TPC), total flavonoid content (TFC), and antioxidant activities of soluble and insoluble-bound phenolic extracts from CMs hosted by Camellia assamica (Mast.) Chang (CMC) and Pyrus, i, f. (CMP) were compared. Phenolic compounds in CMC and CMP were identified and quantified using high-performance liquid chromatography (HPLC). The results indicated that the TPC of soluble phenolic extracts was higher than insoluble-bound phenolic counterparts in both CMC and CMP. In addition, the TPC of soluble, insoluble-bound and total phenolic fractions (9.91 ± 0.23, 4.59 ± 0.27 and 14.50 ± 0.35 μmol ferulic acid equivalents per gram (FAE/g) dry sample) extracted from CMP were higher than those extracted from CMC. The soluble phenolic extracts in CMP showed higher antioxidant activities than those in CMC. Eighteen phenolic compounds from soluble and insoluble-bound phenolic extracts from the CMs were identified and quantified by HPLC. This study indicates that CMC and CMP, especially the latter, could be sources of antioxidants in human health care.

An integrated process for the recovery of high added-value compounds from olive oil using solid support free liquid-liquid extraction and chromatography techniques

An integrated extraction and purification process for the direct recovery of high added value compounds from extra virgin olive oil (EVOO) is proposed by using solid support free liquid-liquid extraction and chromatography techniques. Two different extraction methods were developed on a laboratory-scale Centrifugal Partition Extractor (CPE): a sequential strategy consisting of several “extraction-recovery” cycles and a continuous strategy based on stationary phase co-current elution. In both cases, EVOO was used as mobile phase diluted in food grade n-hexane (feed mobile phase) and the required biphasic system was obtained by adding ethanol and water as polar solvents. For the sequential process, 17.5L of feed EVOO containing organic phase (i.e. 7L of EVOO treated) were extracted yielding 9.5g of total phenolic fraction corresponding to a productivity of 5.8g/h/L of CPE column. Regarding the second approach, the co-current process, 2L of the feed oil phase (containing to 0.8L of EVOO) were treated at 100mL/min yielding 1.03g of total phenolic fraction corresponding to a productivity of 8.9g/h/L of CPE column. The total phenolic fraction was then fractionated by using stepwise gradient elution Centrifugal Partition Chromatography (CPC). The biphasic solvent systems were composed of n-hexane, ethyl acetate, ethanol and water in different proportions (X/Y/2/3, v/v). In a single run of 4h on a column with a capacity of 1L, 910mg of oleocanthal, 882mg of oleacein, 104mg of hydroxytyrosol were successfully recovered from 5g of phenolic extract with purities of 85%, 92% and 90%, respectively. CPC fractions were then submitted to orthogonal chromatographic steps (adsorption on silica gel or size exclusion chromatography) leading to the isolation of additional eleven compounds belonging to triterpens, phenolic compounds and secoiridoids. Among them, elenolic acid ethylester was found to be new compound. Thin Layer Chromatography (TLC), Nuclear magnetic Resonance (NMR) and High Performance Liquid Chromatography – Diode Array Detector (HPLC-DAD) were used for monitoring and evaluation purposes throughout the entire procedure.