The contribution of phenolic acids to the anti-inflammatory activity of mushrooms: Screening in phenolic extracts, individual parent molecules and synthesized glucuronated and methylated derivatives

In the present study, the ethanolic extracts of fourteen edible mushrooms were investigated for their anti-inflammatory potential in LPS (lipopolysaccharide) activated RAW 264.7 macrophages. Furthermore the extracts were chemically characterized in terms of phenolic acids and related compounds. The identified molecules (p-hydroxybenzoic, p-coumaric and cinnamic acids) and their glucuronated and methylated derivatives obtained by chemical synthesis were also evaluated for the same bioactivity, in order to establish structure-activity relationships and to comprehend the effects of in vivo metabolism reactions in the activity of the compounds. The extracts of Pleurotus ostreatus, Macrolepiota procera, Boletus impolitus and Agaricus bisporus revealed the strongest anti-inflammatory potential (EC50 values 96 ± 1 to 190 ± 6 µg/mL, and also the highest concentration of cinnamic acid (656 to 156 µg/g), which was also the individual compound with the highest anti-inflammatory activity. The derivatives of p-coumaric acid revealed the strongest properties, specially the derivative methylated in the carboxylic group (CoA-M1) that exhibited similar activity to the one showed by dexamethaxone used as anti-inflammatory standard; by contrast, the derivatives of p-hydroxybenzoic revealed the lowest inhibition of NO production. All in all, whereas the conjugation reactions change the chemical structure of phenolic acids and may increase or decrease their activity, the glucuronated and methylated derivatives of the studied compounds are still displaying anti-inflammatory activity.

Use of green fluorescent protein-based reporters to monitor balanced production of antifungal compounds in the biocontrol agent Pseudomonas fluorescens CHA0.

AIMS: To develop reporter constructs based on stable and unstable variants of the green fluorescent protein (GFP) for monitoring balanced production of antifungal compounds that are crucial for the capacity of the root-colonizing Pseudomonas fluorescens strain CHA0 to control plant diseases caused by soil-borne pathogenic fungi. METHODS AND RESULTS: Pseudomonas fluorescens CHA0 produces the three antifungal metabolites 2,4-diacetylphloroglucinol (DAPG), pyoluteorin (PLT) and pyrrolnitrin (PRN). The gfp[mut3] and gfp[AAV] reporter genes were fused to the promoter regions of the DAPG, PLT and PRN biosynthetic genes. The reporter fusions were then used to follow the kinetics of expression of the three antifungal metabolites in a microplate assay. DAPG and PLT were found to display an inverse relationship in which each metabolite activates its own biosynthesis while repressing the synthesis of the other metabolite. PRN appears not to be involved in this balance. However, the microbial and plant phenolic metabolite salicylate was found to interfere with the expression of both DAPG and PLT. CONCLUSIONS: The results obtained provide evidence that P. fluorescens CHA0 may keep the antifungal compounds DAPG and PLT at a fine-tuned balance that can be affected by certain microbial and plant phenolics. SIGNIFICANCE AND IMPACT OF THE STUDY: To our knowledge, the present study is the first to use stable and unstable GFP variants to study antibiotic gene expression in a biocontrol pseudomonad. The developed reporter fusions will be a highly valuable tool to study in situ expression of this bacterial biocontrol trait on plant roots, i.e. at the site of pathogen suppression.

Bioactive compounds and acceptance of cookies made with Guava peel flour

The use of fruit industrial waste in the processing new foods represents an important new step for the food industry. This study aimed to develop cookie recipes using different amounts of guava peel flour (GPF) levels (30%, 50%, and 70%) to evaluate the proximate composition, and the phenolic compound, lycopene, and β-carotene levels in the cookies and flour and to evaluate the cookie sensory acceptance. The results demonstrated low moisture, lipid and carbohydrate contents in the flour and cookies. GPF was considered rich in fiber, ash, polyphenols, and β-carotene. The sensory analysis showed satisfactory acceptance of the cookies containing 30% GPF regarding the aroma, flavor, and texture attributes. The cookies containing 50% and 70% GPF received satisfactory acceptance regarding to aroma only. In conclusion, GPF can be used to partially replace wheat flour in the preparation of cookies to improve the nutritional quality without affecting the product sensory quality.

Nutritional quality and bioactive compounds of partially defatted baru almond flour

This study aimed to investigate the nutritional quality and bioactive potential of partially defatted baru (Dipteryx alataVog.) almond flour (BAF). The flour’s proximate and mineral compositions, total phenolic, tocopherols and carotenoids contents, antioxidant capacity, trypsin inhibitor and amino acid analyses were performed. An experiment was conducted with 24 male Wistar rats in order to evaluate the flour’s protein quality. BAF has high protein, fiber and mineral contents (iron, zinc, magnesium and copper), and it is a source of calcium. BAF presented relevant amounts of total phenolics (625 mg/100g) and good antioxidant capacity (130 µmol/Trolox eq). Autoclaved BAF showed essential amino acids profile, digestibility and protein quality better than in natura BAF. Autoclaved BAF might be used for human consumption as a source of quality protein and bioactive compounds, in healthy diets and processed foods.

Changes in phenolic composition and antioxidant activity of virgin olive oil during frying

The concentration of hydroxytyrosol (3,4-DHPEA) and its secoiridoid derivatives (3,4-DHPEA-EDA and 3,4-DHPEA-EA) in virgin olive oil decreased rapidly when the oil was repeatedly used for preparing french fries in deep-fat frying operations. At the end of the first frying process (10 min at 180 degreesC), the concentration of the dihydroxyphenol components was reduced to 50-60% of the original value, and after six frying operations only about 10% of the initial components remained. However, tyrosol (p-HPEA) and its derivatives (p-HPEA-EDA and p-HPEA-EA) in the oil were much more stable during 12 frying operations. The reduction in their original concentration was much smaller than that for hydroxytyrosol and its derivatives and showed a roughly linear relationship with the number of frying operations. The antioxidant activity of the phenolic extract measured using the DPPH test rapidly diminished during the first six frying processes, from a total antioxidant activity higher than 740,mumol of Trolox/kg down to less than 250 mumol/kg. On the other hand, the concentration of polar compounds, oxidized triacylglycerol monomers (oxTGs), dimeric TGs, and polymerized TGs rapidly increased from the sixth frying operation onward, when the antioxidant activity of the phenolic extract was very low, and as a consequence the oil was much more susceptible to oxidation. The loss of antioxidant activity in the phenolic fraction due to deep-fat frying was confirmed by the storage oil and oil-in-water emulsions containing added extracts from olive oil used for 12 frying operations.

Gut microbial catabolism of grape seed flavan-3-ols by human faecal microbiota. Targetted analysis of precursor compounds, intermediate metabolites and end-products.

In vitro batch culture fermentations were conducted with grape seed polyphenols and human faecal microbiota, in order to monitor both changes in precursor flavan-3-ols and the formation of microbial-derived metabolites. By the application of UPLC-DAD-ESI-TQ MS, monomers, and dimeric and trimeric procyanidins were shown to be degraded during the first 10 h of fermentation, with notable inter-individual differences being observed between fermentations. This period (10 h) also coincided with the maximum formation of intermediate metabolites, such as 5-(3′,4′-dihydroxyphenyl)-γ-valerolactone and 4-hydroxy-5-(3′,4′-dihydroxyphenyl)-valeric acid, and of several phenolic acids, including 3-(3,4-dihydroxyphenyl)-propionic acid, 3,4-dihydroxyphenylacetic acid, 4-hydroxymandelic acid, and gallic acid (5–10 h maximum formation). Later phases of the incubations (10–48 h) were characterised by the appearance of mono- and non-hydroxylated forms of previous metabolites by dehydroxylation reactions. Of particular interest was the detection of γ-valerolactone, which was seen for the first time as a metabolite from the microbial catabolism of flavan-3-ols. Changes registered during fermentation were finally summarised by a principal component analysis (PCA). Results revealed that 5-(3′,4′-dihydroxyphenyl)-γ-valerolactone was a key metabolite in explaining inter-individual differences and delineating the rate and extent of the microbial catabolism of flavan-3-ols, which could finally affect absorption and bioactivity of these compounds.

In vitro fermentation of grape seed flavan-3-ol fractions by human faecal microbiota: changes in microbial groups and phenolic metabolites

With the aim of investigating the potential of flavan-3-ols to influence the growth of intestinal bacterial groups, we have carried out the in vitro fermentation, with human faecal microbiota, of two purified fractions from grape seed extract (GSE): GSE-M (70% monomers and 28% procyanidins) and GSE-O (21% monomers and 78 % procyanidins). Samples were collected at 0, 5, 10, 24, 30 and 48 h of fermentation for bacterial enumeration by fluorescent in situ hybridization and for analysis of phenolic metabolites. Both GSE-M and GSE-O fractions promoted growth of Lactobacillus/Enterococcus and decrease in the Clostridium histolyticum group during fermentation, although the effects were only statistically significant with GSE-M for Lactobacillus/Enterococcus (at 5 and 10 h of fermentation) and GSE-O for C. histolyticum (at 10 h of fermentation). Main changes in polyphenol catabolism also occurred during the first 10 h of fermentation, however no significant correlation coefficients (P>0.05) were found between changes in microbial populations and precursor flavan-3-ols or microbial metabolites. Together these data suggest that the flavan-3-ol profile of a particular food source could affect the microbiota composition and its catabolic activity, inducing changes that could in turn affect the bioavailability and potential bioactivity of these compounds.

In vitro Trypanocidal Activity of Phenolic Derivatives from Peperomia obtusifolia

The trypanocidal activity of crude extracts and fractions from the leaves and stems of Peperomia obtusifolia (Piperaceae) was evaluated in vitro against the epimastigote forms of Trypanosoma cruzi. Bioactivity-guided fractionation of the most active extracts afforded seven known compounds, including three chromanes, two furofuran lignans and two flavone C-diglycosides. The most active compounds were the chromanes peperobtusin A and 3,4-dihydro-5-hydroxy-2,7-dimethyl-8-(2 ``-methyl-2 ``-butenyl)-2-(4`-methyl-1`,3`-pentadienyl)-2H-1-benzopyran-6-carboxylic acid, with IC(50) values of 3.1 mu M (almost three times more active than the positive control benznidazole, IC(50) 10.4 mu M) and 27.0 mu M, respectively. Cytotoxicity assays using peritoneal murine macrophages indicated that the chromanes were not toxic at the level of the IC(50) for trypanocidal activity. This is the first report on the trypanocidal activity besides unspecific cytotoxicity of chromanes from Peperomia species. Additionally it represents the first time isolation of 3,4-dihydro5-hydroxy-2,7-dimethyl-8-(2 ``-methyl-2 ``-butenyl)-2-(4`-methyl-1`,3`-pentadienyl)-2H-1-benzopyran-6-carboxylic acid from P. obtusifolia.

Development of a novel microextraction by packed sorbent-based approach followed by ultrahigh pressure liquid chromatography as a powerful technique for quantification phenolic constituents of biological interest in wines

A novel analytical approach, based on a miniaturized extraction technique, the microextraction by packed sorbent (MEPS), followed by ultrahigh pressure liquid chromatography (UHPLC) separation combined with a photodiode array (PDA) detection, has been developed and validated for the quantitative determination of sixteen biologically active phenolic constituents of wine. In addition to performing routine experiments to establish the validity of the assay to internationally accepted criteria (linearity, sensitivity, selectivity, precision, accuracy), experiments are included to assess the effect of the important experimental parameters on the MEPS performance such as the type of sorbent material (C2, C8, C18, SIL, and M1), number of extraction cycles (extract-discard), elution volume, sample volume, and ethanol content, were studied. The optimal conditions of MEPS extraction were obtained using C8 sorbent and small sample volumes (250 μL) in five extraction cycle and in a short time period (about 5 min for the entire sample preparation step). The wine bioactive phenolics were eluted by 250 μL of the mixture containing 95% methanol and 5% water, and the separation was carried out on a HSS T3 analytical column (100 mm × 2.1 mm, 1.8 μm particle size) using a binary mobile phase composed of aqueous 0.1% formic acid (eluent A) and methanol (eluent B) in the gradient elution mode (10 min of total analysis). The method gave satisfactory results in terms of linearity with r2-values > 0.9986 within the established concentration range. The LOD varied from 85 ng mL−1 (ferulic acid) to 0.32 μg mL−1 ((+)-catechin), whereas the LOQ values from 0.028 μg mL−1 (ferulic acid) to 1.08 μg mL−1 ((+)-catechin). Typical recoveries ranged between 81.1 and 99.6% for red wines and between 77.1 and 99.3% for white wines, with relative standard deviations (RSD) no larger than 10%. The extraction yields of the MEPSC8/UHPLC–PDA methodology were found between 78.1 (syringic acid) and 99.6% (o-coumaric acid) for red wines and between 76.2 and 99.1% for white wines. The inter-day precision, expressed as the relative standard deviation (RSD%), varied between 0.2% (p-coumaric and o-coumaric acids) and 7.5% (gentisic acid) while the intra-day precision between 0.2% (o-coumaric and cinnamic acids) and 4.7% (gallic acid and (−)-epicatechin). On the basis of analytical validation, it is shown that the MEPSC8/UHPLC–PDA methodology proves to be an improved, reliable, and ultra-fast approach for wine bioactive phenolics analysis, because of its capability for determining simultaneously in a single chromatographic run several bioactive metabolites with high sensitivity, selectivity and resolving power within only 10 min. Preliminary studies have been carried out on 34 real whole wine samples, in order to assess the performance of the described procedure. The new approach offers decreased sample preparation and analysis time, and moreover is cheaper, more environmentally friendly and easier to perform as compared to traditional methodologies.

Evaluation of antiulcer activity of the main phenolic acids found in Brazilian Green Propolis

Aim of the study: In a previous study, our group described the gastric protective effect of the hydroalcoholic extract of Brazilian green propolis. The main compounds found in Brazilian green propolis include phenolic acids, such as: caffeic, ferulic, p-coumaric and cinnamic acids. This study was therefore carried out to evaluate the antiulcerogenic property of the main phenolic acids found in Brazilian Green Propolis.Material and methods: The anti-ulcer assays were performed using the following protocols: nonsteroidal-antinflammatory drug (NSAID)-induced ulcer, ethanol-induced ulcer, and stress-induced ulcer. The effects of the phenolic acids on gastric content volume, pH and total acidity, using the pylorus ligated model, were also evaluated.Results: It was observed that treatment using doses of 50 and 250 mg/kg of caffeic, ferulic, p-coumaric and cinnamic acids and positive controls (omeprazol or cimetidine) significantly diminished the lesion index, the total area of the lesion and the percentage of lesion in comparison with the negative control groups. In addition, the percentage of ulcer inhibition was significantly higher in the groups treated with the different phenolic acids, cimetidine or omeprazol, in all the protocols used, compared with the negative control groups. In the model to determine gastric secretion, using ligated pylorus, treatment with phenolic acids and cimetidine reduced the volume of gastricjuice and total acidity and significantly increased the gastric pH (p < 0.05), compared with the control group, with the exception of the group treated with 50 mg/kg of p-coumaric acid, in which no significant difference was observed, compared with the control. In relation to the acute toxicity, none sign of toxicity was observed when phenolic acids, used in this study, were administered for rats in dose of 2000 mg/kg.Conclusions: In conclusion, the results of this study show that caffeic, ferulic, p-coumaric and cinnamic acids display antiulcer activity. (c) 2008 Elsevier B.V. All rights reserved.

Peroxidase and phenylalanine ammonia-lyase activities, phenolic acid contents, and allelochemicals-inhibited root growth of soybean

The influence of the allelochemicals ferulic (FA) and vanillic (VA) acids on peroxidase (POD, EC 1.11.1.7) and phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) activities and their relationships with phenolic acid (PhAs) contents and root growth of soybean (Glycine max (L.) Merr.) were examined. Three-day-old seedlings were cultivated in nutrient solution containing FA or VA (0.1 to 1 mM) for 48 h. Both compounds (at 0.5 and 1 mM) decreased root length (RL), fresh weight (FW) and dry weight (DW) and increased PhAs contents. At 0.5 and 1 mM, FA increased soluble POD activity (18% and 47%, respectively) and cell wall (CW)-bound POD activity (61% and 34%), while VA increased soluble POD activity (33% and 17%) but did not affect CW-bound POD activity. At I mM, FA increased (82%) while VA reduced (32%) PAL activities. The results are discussed on the basis of the role of these compounds on phenylpropanoid metabolism and root growth and suggest that the effects caused on POD and PAL activities are some of the many mechanisms by which allelochemicals influence plant growth.

In vitro Trypanocidal Activity of Phenolic Derivatives from Peperomia obtusifolia

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Phenolic Derivatives from Fruits of Dipteryx lacunifera DUCKE and Evaluation of Their Antiradical Activities

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Antioxidant phenolic and quinonemethide triterpenes from Cheiloclinium cognatum

The triterpenes, 22beta-hydroxypristimerin and cognatine, were isolated together with the known compounds pristimerin, maytenin, 20alpha-hydroxymaytenin, 22beta-hydroxymaytenin, netzahualcoyol, netzahualcoyondiol and netzahualcoyone from root bark of Chei-Ioclinium cognatum. The structures of the isolated compounds were elucidated by interpretation of their spectral data, including gHMQC and gHMBC experiments. The isolates were investigated for their radical scavenging abilities through a spectrophotometric assay involving reduction of 2,2-diphenyl-picryl hydrazyl (DPPH). (C) 2004 Elsevier Ltd. All rights reserved.

Determination of the relative contribution of phenolic antioxidants in orange juice by voltammetric methods

The electrochemical oxidation of caffeic, chlorogenic, sinapic, ferulic and p-coumaric acids was investigated by cyclic voltammetry on acetate buffer pH 5.6 on glassy carbon electrode and modified glassy carbon electrode. According to their voltammetric behavior, the antioxidant activity of these phenolic acids was evaluated and the results pointed to the following sequence: caffeic acid (E-a = +0.31 V) > chlorogenic acid (+ 0.38 V) > sinapic acid (+ 0.45 V) > ferulic acid (+ 0.53 V) >p-coumaric acid (+ 0.73 V). The results were confirmed by DPPH test, which evidenced the strongest antiradical activity for compounds possessing the cathecol moiety (caffeic and chlorogenic acids). Linear calibration graphs were obtained for their determination at concentrations from 1 x 10(-4) to 1 x 10(-3) mol L-1. The method was applied to orange juice. Selectivity was illustrated by the analysis of caffeic and chlorogenic acids electrodeposited on a glassy carbon electrode previously modified by electrochemical activation in the presence of ascorbic acid. (C) 2003 Elsevier B.V. All rights reserved.