Identification of bioactive compounds in Phyllenthus emblica L. fruit and their free radical scavenging activities

Emblica has been used as an important traditional herbal medicine in southeast Asia since ancient times. In this study, the air-dried hulls of emblica fruit were extracted with 95% ethanol, and then the extract was partitioned by diethyl ether and ethyl acetate (EA). The EA fraction was purified by silica gel column and thin layer chromatography (TLC) to obtain six compounds. They were identified as cinnamic acid (C1), quercetin (C2), 5-hydroxymethylfurfural (C3), gallic acid (C4), β-daucosterol (C5) and ellagic acid (C6) using mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy. Cinnamic acid and 5-hydroxymethylfurfural were identified as components of emblica fruit for the first time. The DPPH and ABTS⁺ radical scavenging activities of components were evaluated. All the compounds showed significant DPPH and ABTS⁺ radical scavenging activity except for cinnamic acid. Gallic acid showed the highest DPPH radical scavenging activity while ellagic acid showed the highest ABTS⁺ scavenging activity amongst all the compounds tested.     

Essential oil composition of Pistacia lentiscus L. and Myrtus communis L.: Evaluation of antioxidant capacity of methanolic extracts

The seasonal variation of the essential oil composition, the antioxidant activity (DPPH, FRAP assays) and the total phenolic content (Folin-Ciocalteu assay) of two aromatic wild plants, Pistacia lentiscus L. (Anacardiaceae) and Myrtus communis L. (Myrtaceae), grown in Zakynthos, a Greek island, was investigated. The essential oil was obtained by hydrodistillation and subsequently analysed by GC–MS.     

Antioxidant and antiacetylcholinesterase activities of essential oils from Cymbopogon schoenanthus L. Spreng. Determination of chemical composition by GC–mass spectrometry and C NMR

Cymbopogon schoenanthus L. Spreng, is an aromatic herb consumed in salads and used to prepare traditional meat recipes in Tunisia. The chemical composition, antioxidant activities and acetylcholinesterase inhibitory properties of the essential oils from fresh leaves, dried leaves and roots collected from three different locations in southern Tunisia, were evaluated. Essential oils were analysed by GC–mass spectrometry and ¹³C NMR. The major components were limonene (10.5–27.3%), β-phellandrene (8.2–16.3%), δ-terpinene (4.3–21.2%) and α-terpineol (6.8–11.0%). Antioxidant activity was measured by DPPH assay. The results ranged from 36.0% to 73.8% (2 μl of essential oil per mL of test solution).     

Activity-guided isolation and identification of free radical-scavenging components from an aqueous extract of Coleus aromaticus

An activity-directed fractionation and purification process was used to identify the DPPH (l,l-diphenyl-2-picrylhydrazyl) free radical-scavenging components of Coleus aromaticus Benth. Fresh leaves of C. aromaticus were extracted with water and then separated into hexane, ethyl acetate, and water fractions. Among these, only the ethyl acetate phase showed strong DPPH radical-scavenging activity in vitro, when compared with water and hexane phases. The ethyl acetate fraction was then subjected to separation and purification using Sephadex LH-20 chromatography. Three compounds showing strong DPPH radical-scavenging activity were shown, by spectral methods (¹H NMR, ¹³C NMR, and MS) and by comparison with literature values, to be rosmarinic acid, chlorogenic acid and caffeic acid. In addition, HPLC identification and quantification of isolated compounds were also performed. Rosmarinic acid was found as a major component and principally responsible for the radical-scavenging activity of C. aromaticus.     

Phenolics from hull of Garcinia mangostana fruit and their antioxidant activities

The air-dried fruit hulls of Garcinia mangostana Linn. were extracted with 70% MeOH, and then partitioned into the n-BuOH fractions. Furthermore, three major phenolic components related to their antioxidant activities were purified by silica gel column chromatography and Sephadex LH-20 and then identified as P₁ (1,3,6,7-tetrahydroxy-2,8-(3-methyl-2-butenyl)), P₂ [1,3,6-trihydroxy-7-methoxy-2,8-(3-methyl-2-butenyl) xanthone] and P₃ (epicatechin) using UV–visible spectrophotometry, IR spectrophotometry and NMR spectroscopy, respectively. The antioxidant activities of three major phenolics were evaluated using different tests, including the free radical scavenging capability and total antioxidant activity in a linoleic acid peroxidation. These three phenolic compounds exhibited different antioxidant activities in these antioxidant tests. The hydroxyl radical and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capabilities and the activity against linoleic acid peroxidation of P₁ were greater than those of P₂ and P₃, while the superoxide anion radical scavenging activity of P₃ was greater than that of P₁, but was close to that of P₂ or α-tocopherol. An activity-guided isolation and purification process was used to identify the components showing the strong DPPH radical scavenging activity from G. mangostana Linn.     

Isolation and evaluation of the antioxidant activity of phenolic constituents of the Garcinia brasiliensis epicarp

A new glycosylated biflavonone, morelloflavone-4′″-O-β-d-glycosyl, and the known compounds 1,3,6,7-tetrahydroxyxanthone, morelloflavone (fukugetin) and morelloflavone-7″-O-β-d-glycosyl (fukugeside) were isolated from the epicarp of Garcinia brasiliensis collected in Brazil. The structures of these compounds were established using ¹H and ¹³C NMR, COSY, gHMQC and gHMBC spectroscopy. The compounds exhibited antioxidant activity. The greatest potency was displayed by morelloflavone (2), with IC₅₀ = 49.5 mM against DPPH and absorbance of 0.583 at 400 μg/mL for the reduction of Fe³⁺. The weakest potency was displayed by 1,3,6,7-tetrahydroxyxanthone (1), with IC₅₀ = 148 mM against DPPH and absorbance of 0.194 at 400 μg/mL for the reduction of Fe³⁺.     

Free radical scavenging properties and phenolic characterization of some edible plants

Antioxidant activity of the ethanolic extracts of the fruits of Fritillaria pontica Wahlenb. (Liliaceae), Euonymus latifolius (L.) Mill. ssp. latifolius (Celastraceae), and Vicia sativa L. ssp. nigra (L.) Ehrh. var. nigra L. (Fabaceae), the aerial parts of Turritis laxa (Sibth & Sm.) Hayek (Brassicaceae), Vicia cracca L. (Fabaceae), and Oxyria digyna (L.) Hill. (Polygonaceae) was screened by 2,2-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and ferric-reducing antioxidant power (FRAP) assays at 0.5, 1.0, and 2.0 mg ml⁻¹ concentrations. Total phenolic contents of the extracts were determined using Folin-Ciocalteau reagent. T. laxa was also tested for its anti-acetylcholinesterase activity. The extracts were analyzed by LC–DAD–MS for their flavonoid content and the ethanolic extract of T. laxa has been found to contain rutin in appreciable amounts (7.63 ± 0.2%). Rutin and hyperoside were detected qualitatively in F. pontica, where vitexin was identified in O. digyna. It was also the most active in the antioxidant tests.     

Antioxidant and antimicrobial activities of Laetiporus sulphureus (Bull.) Murrill

Antioxidant capacity and antimicrobial activities of Laetiporus sulphureus (Bull.) Murrill. extracts obtained with ethanol were investigated in this study. The study was aimed at determining the antioxidant activity (DPPH free radical-scavenging, β-carotene/linoleic acid systems), total phenolic content and total flavonoid concentration of L. sulphureus. Inhibition values both of L. sulphureus ethanol and the standards increased parallel with the elevation of concentration in the linoleic acid system. Inhibition values of L. sulphureus (LS) extract, BHA and α-tocopherol standards were found to be 82.2%, 96.4% and 98.6%, respectively, at a concentration of 160 μg/ml. DPPH free radical-scavenging activity was found to exhibit 14%, 26%, 55% and 86% inhibition, respectively, at concentrations of 100, 200, 400 and 800 μg/ml. Total flavanoids were 14.2 ± 0.12 μg mg⁻¹ (quercetin equivalent) while the phenolics were 63.8 ± 0.25 μg mg⁻¹ (pyrocatechol equivalent) in the extract. Positive correlations were found between total phenolic content in the mushroom extracts and their antioxidant activities. Edible mushrooms may have potential as natural antioxidants. L. sulphureus showed narrow antibacterial activity against Gram-negative bacteria and strongly inhibited the growth of the Gram-positive bacteria tested. The crude extract exhibited high anticandidal activity on Candida albicans. Therefore, the extracts could be suitable as antimicrobial and antioxidative agents in the food industry.     

Chemical composition,antioxidant and antibacterial activities of the essential oils isolated from Tunisian Thymus capitatus Hoff. et Link.

The chemical composition, antioxidant and antibacterial activities of essential oils isolated by hydrodistillation from the aerial parts of Tunisian Thymus capitatus Hoff. et Link. during the different phases of the plant development, and from different locations, were evaluated. The chemical composition was analyzed by gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS). The main components of the essential oils were carvacrol (62–83%), p-cymene (5–17%), γ-terpinene (2–14%) and β-caryophyllene (1–4%). The antioxidant activity of the oils (100–1000 mg l⁻¹) was assessed by measurement of metal chelating activity, the reductive potential, the free radical scavenging (DPPH) and by the TBARS assay. The antioxidant activity was compared with that of synthetic antioxidants: butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT). Both the essential oils and BHA and BHT showed no metal chelating activity. Although with the other methodologies, there was a general increase in the antioxidant activity, with increasing oil concentration, maxima being obtained in the range of 500 and 1000 mg l⁻¹ for flowering and post-flowering phase oils. Major differences were obtained according to the methodology of antioxidant capacity evaluation. Antibacterial ability of Th. capitatus essential oils was tested by disc agar diffusion against Bacillus cereus, Salmonella sp., Listeria innocua, four different strains of Staphylococus aureus (C15, ATCC25923, CFSA-2) and a multi-resistant form of S. aureus (MRSA-2). Antibacterial properties were compared to synthetic antibiotics. Higher antibacterial activity was observed with the flowering and the post-flowering phase essential oils.     

Antioxidant constituents from Lawsonia inermis leaves: Isolation,structure elucidation and antioxidative capacity

The antioxidant potential of different fractions of Lawsonia inermis (Lythraceae) was investigated. The n-butanolic fraction showed the highest yield of extraction; it also exhibited a strong antioxidant activity in the DPPH assay and a potent capacity in preventing linoleic acid oxidation. Five phenolic glycosides were identified in this fraction. The structure of a new compound was established as 1,2,4-trihydroxynaphthalene-1-Ο-β-d-glucopyranoside. In addition, the known 2,4,6-trihydroxyacetophenone-2-Ο-β-d-glucopyranoside was described for the first time in this species. The three other compounds, lalioside (2,3,4,6-tetrahydroxyacetophenone-2-Ο-β-d-glucopyranoside), lawsoniaside (1,2,4-trihydroxynaphthalene-1,4-di-Ο-β-d-glucopyranoside) and luteolin-7-Ο-β-d-glucopyranoside, have been previously reported in L. inermis. The antioxidant activity of these glycosides was evaluated by DPPH and β-carotene assays, and compared to those of commercial standards. 1,2,4-Trihydroxynaphthalene-1-Ο-β-d-glucopyranoside was the most active in the DPPH free-radical scavenging test (EC₅₀ = 6.5 μg/ml) and showed a moderate inhibition in the β-carotene bleaching assay. Chemical components of L. inermis have good antioxidant capacities and this species could be used as a potential source of new natural antioxidants.     

Characterisation of phenolic acid derivatives and flavonoids from different morphological parts of Helichrysum obconicum by a RP-HPLC–DAD-(−)–ESI-MS method

The phenolic composition from different morphological parts of Helichrysum obconicum was investigated for the first time and 50 different phenolic compounds were detected. Phenolic acid conjugates, mainly mono- and di-caffeoylquinic acid derivatives, were the major components; some flavonoid derivatives were also detected in small amounts. Their separation and identification was performed by a high-performance liquid chromatography/electron spray ionisation tandem ion trap mass spectrometry method, with special emphasis on MSⁿ fragmentation. The presence of di- and tricaffeoylshikimic acid isomers in Helichrysum species extracts was reported for the first time, the spectra of these compounds were mainly characterised by the presence of a [caffeoylshikimic acid-H]⁻ ion at m/z 335. A lamiridosins-di-O-hexoside, an unusual component in Asteraceae species, was also detected.     

Chemical composition, in vitro cytotoxic and antioxidant activities of the essential oil and major constituents of Cymbopogon jawarancusa (Kashmir)

The chemical composition of the hydrodistillate of aerial parts of Cymbopogon jawarancusa, a natural grass considered as major forage for animal nutrition, used in food because of the presence of sufficient concentration of minerals like calcium and potassium was analysed by capillary GC–FID, GC–MS and ¹³C NMR. Seventeen constituents representing 97.8% of the total oil with piperitone (58.6%) and elemol (18.6%) as major constituents were identified. In vitro cytotoxicity of the oil and its constituents on human cancer cell lines THP-1 (leukemia), A-549 (lung), HEP-2 (liver) and IGR-OV-1 (ovary) was evaluated by Sulphorhodamine-B assay. The oil was found to be more potent than its components against cancer cell lines tested with IC₅₀ of 6.5 μg/ml (THP-1), 6.3 μg/ml (A-549), 7.2 μg/ml (HEP-2) and 34.4 μg/ml (IGR-OV-1). Antioxidant activity of oil and its constituents was evaluated by DPPH assay. In conclusion, the results demonstrate potent cytotoxic and antioxidant effects of oil, and its components like piperitone, α-pinene, β-caryophyllene and β-elemene.     

Antioxidant potential and phenolic constituents of Salvia cedronella

The acetone extract of the aerial parts of the plant Salvia cedronella Boiss. was screened for its total phenolic content and flavonoid content. The antioxidant potential was evaluated, in vitro, by using three different assays; β-carotene–linoleic acid test system for total antioxidant activity, DPPH for free radical scavenging activity, Fe²⁺–ferrozine test system for metal chelation. A high content of phenolics, potent radical scavenging ability and significant iron chelating effect were observed. However, the inhibition of lipid peroxidation was not significant in β-carotene–linoleic acid test system. A phytochemical analysis yielded a new coumarin, 3-methoxy-4-hydroxymethyl coumarin, together with p-hydroxyphenylethyl docosanoate, and two triterpenoids oleanolic acid and betulinic acid.     

Application of response surface methodology and central composite design for the optimisation of supercritical fluid extraction of essential oils from Myrtus communis L. leaves

Essential oils of Myrtus communis L. leaves were obtained using supercritical fluid extraction (SFE) and hydrodistillation methods. The experimental parameters of SFE such as pressure, temperature, modifier volume, static and dynamic extraction time were optimised using a central composite design after a 2ⁿ⁻¹ fractional factorial design. The chemical compositions of the SFE extract were identified by GC–MS and determined by GC–FID. The major components of essential oils obtained by hydrodistillation were α-pinene (31.8%), 1,8-cineole (24.6%), limonene (14.8%), linalool (8.3%) and α-terpinolene (4.8%). However, by using the supercritical carbon dioxide in optimum conditions, only three components represented more than 85% of the extract. Therefore, by using the proper SFE conditions, the supercritical extraction is more selective than the conventional hydrodistillation methods. The oil yields based on the hydrodistillation was 0.47% (v/w). Extraction yields based on the SFE varied in the range of 0.5–6.3% (w/w) under different conditions.     

The inhibitory effect of natural microflora of food on growth of Listeria monocytogenes in enrichment broths

The aims of this study were to (i) compare the inhibitory effects of the natural microflora of different foods on the growth of Listeria monocytogenes during enrichment in selective and non-selective broths; (ii) to isolate and identify components of the microflora of the most inhibitory food; and (iii) to determine which of these components was most inhibitory to growth of L. monocytogenes in co-culture studies. Growth of an antibiotic-resistant marker strain of L. monocytogenes was examined during enrichment of a range of different foods in Tryptone Soya Broth (TSB), Half Fraser Broth (HFB) and Oxoid Novel Enrichment (ONE) Broth. Inhibition of L. monocytogenes was greatest in the presence of minced beef, salami and soft cheese and least with prepared fresh salad and chicken pâté. For any particular food the numbers of L. monocytogenes present after 24 h enrichment in different broths increased in the order: TSB, HFB and ONE Broth. Numbers of L. monocytogenes recovered after enrichment in TSB were inversely related to the initial aerobic plate count (APC) in the food but with only a moderate coefficient of determination (R²) of 0.51 implying that microbial numbers and the composition of the microflora both influenced the degree of inhibition of L. monocytogenes. In HFB and ONE Broth the relationship between APC and final L. monocytogenes counts was weaker. The microflora of TSB after 24 h enrichment of minced beef consisted of lactic acid bacteria, Brochothrix thermosphacta, Pseudomonas spp., Enterobacteriaceae, and enterococci. In co-culture studies of L. monocytogenes with different components of the microflora in TSB, the lactic acid bacteria were the most inhibitory followed by the Enterobacteriaceae. The least inhibitory organisms were Pseudomonas sp., enterococci and B. thermosphacta. In HFB and ONE Broth the growth of Gram-negative organisms was inhibited but lactic acid bacteria still reached high numbers after 24 h. A more detailed study of the growth of low numbers of L. monocytogenes during enrichment of minced beef in TSB revealed that growth of L. monocytogenes ceased at a cell concentration of about 10² cfu/ml when lactic acid bacteria entered stationary phase. However in ONE Broth growth of lactic acid bacteria was slower than in TSB with a longer lag time allowing L. monocytogenes to achieve much higher numbers before lactic acid bacteria reached stationary phase. This work has identified the relative inhibitory effects of different components of a natural food microflora and shown that the ability of low numbers of L. monocytogenes to achieve high cell concentrations is highly dependent on the extent to which enrichment media are able to inhibit or delay growth of the more effective competitors.     

Polyphenol contents and in vitro antioxidant activities of lyophilised aqueous extract of kiwifruit (Actinidia deliciosa)

The aim of this study was to determine the antioxidant potency and total phenolic and flavonoid contents of kiwifruit (Actinidia deliciosa) in vitro by analysing the radical scavenging activity of lyophilised water extract from kiwifruit (LEK) for 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), 1,1-diphenyl-2-picryl-hydrazyl (DPPH), N,N-dimethyl-p-phenylenediamine (DMPD), and superoxide anion radical (O₂⁻) as well as the total reducing power by FRAP and CUPRAC assays and the metal chelating activities. LEK showed efficient radical scavenging activity with DPPH, ABTS, DMPD, and O₂⁻ radicals; ferric (Fe³⁺) and cupric (Cu²⁺) ion reducing power and metal chelating activities. Moreover, the amounts of phenolic compounds, such as caffeic acid, ferulic acid, syringic acid, ellagic acid, catechol, pyrogallol, p-hydroxy benzoic acid, vanillin, p-coumaric acid, gallic acid, quercetin, ??-tocopherol and ascorbic acid, in LEK were quantified by LC-MS-MS. The results show that pyrogallol (2070.0 mg/kg LEK) is the main phenolic compound responsible for the antioxidant and radical scavenging activities of LEK. Finally, total phenolic and flavonoid contents were determined as gallic acid (GAE) and quercetin equivalents (QE). The GAE and QE values in LEK were 16.67 ± 2.83 ??g GAE/mg and 12.95 ± 0.52 ??g QE/mg, respectively. The results suggest that consumption of kiwifruit (A. deliciosa) can be beneficial effects due to its antioxidant properties.     

Antioxidant activity of cichoric acid and alkamides from Echinacea purpurea, alone and in combination

The antioxidant activity of extracts of the stems, leaves, and roots of Echinacea purpurea was compared with the antioxidant activity of purified cichoric acid and alkamides, both constituents of Echinacea purpurea. The antioxidant activity was determined using different methods: effect on oxygen consumption rate of a peroxidating lipid emulsion, and scavenging of radicals, i.e. 2,2-diphenyl-1-picrylhydrazyl (DPPH), measured by two different techniques. The efficacy of the extracts in the reaction with DPPH correlated well with the amount of cichoric acid present in the various extracts. The alkamides alone showed no antioxidant activity in any of the tests. Alkamides present in the extract increased, however, the antioxidative effect of cichoric acid in the peroxidating lipid emulsion. The activity was further compared with that of rosmarinic acid, a well-characterised antioxidant, and the extracts as well as cichoric acid were found to be efficient scavengers of radicals with an activity comparable to that of rosmarinic acid. Cichoric acid was found to have a stoichiometric factor of 4.0 in scavenging DPPH and to react in a second-order reaction with DPPH with a rate constant of 40 l/mol/s at 25 °C in methanol.     

Chemical composition and antioxidant activities of Myrtus communis L. berries extracts

Myrtle (Myrtus communis L.) berries extracts were prepared with solvents at different polarity (water, ethanol, and ethyl acetate) and analysed using different in vitro tests in order to evaluate their antioxidant properties. Antiradical and total antioxidant activities were measured with DPPH and FRAP tests, respectively. Their ability to protect biological molecules was assessed using the cholesterol and LDL oxidation assays. In addition, phenolic compounds and unsaturated fatty acids composition was analysed by HPLC–DAD and HPLC–MS/MS. Ethanol and water extracts showed the highest amount of extracted compounds, but the highest antiradical and antioxidant activities were found in ethanol and ethyl acetate extracts. These extracts were also the ones with the highest content of phenolic compounds. In addition, our results showed a highly significant correlation between the amount of total phenols and antiradical (R² = 0.9993) or antioxidant activities (R² = 0.9985) in these extracts. HPLC–DAD and HPLC–MS analyses showed significant quantitative and qualitative differences among these three extracts. The ethyl acetate extract had the highest protective effect in assays of thermal (140 °C) cholesterol degradation and Cu²⁺-mediated LDL oxidation, inhibiting the reduction of polyunsaturated fatty acids and cholesterol, and the increase of their oxidative products. These results suggest that because of these properties, myrtle berries could be used in dietary supplements preparations or as food additives.     

Purification and identification of bovine cheese whey fatty acids exhibiting in vitro antifungal activity

Milk lipids contain several bioactive factors exhibiting antimicrobial activity against bacteria, viruses, and fungi. In the present study, we demonstrate that free fatty acids (FFA) derived from the saponification of bovine whey cream lipids are active in vitro at inhibiting the germination of Candida albicans, a morphological transition associated with pathogenicity. This activity was found to be significantly increased when bovine FFA were enriched in non-straight-chain FFA. At low cell density, this non-straight-chain FFA-enriched fraction was also found to inhibit in a dose-dependant manner the growth of both developmental forms of C. albicans as well as the growth of Aspergillus fumigatus. Using an assay-guided fractionation, the main components responsible for these activities were isolated. On the basis of mass spectroscopic and gas chromatographic analysis, antifungal compounds were identified as capric acid (C10:0), lauroleic acid (C12:1), 11-methyldodecanoic acid (iso-C13:0), myristoleic acid (C14:1n-5), and γ-linolenic acid (C18:3n-6). The most potent compound was γ-linolenic acid, with minimal inhibitory concentration values of 5.4 mg/L for C. albicans and 1.3 mg/L for A. fumigatus, in standardized conditions. The results of this study indicate that bovine whey contains bioactive fatty acids exhibiting antifungal activity in vitro against 2 important human fungal pathogens.     

Antiradical activity of different parts of Walnut (Juglans regia L.) fruit as a function of genotype

The objective of this work was to analyse phenolic compounds and antiradical capacity of different parts of walnut fruit among six genotypes of Juglans regia L. Therefore, total phenolic and flavonoid content were determined and methanolic extracts of walnut genotypes were considered by the reducing power, DPPH (2,2-diphenyl-1-picrylhydrazyl), superoxide anion and nitric oxide radical scavenging. Significant differences were found in phenolic content and radical scavenging capacity of different parts of fruits and among various genotypes. High correlation coefficient (R² = 0.81) was observed between phenol content and radical scavenging activity, but this was not always true (R² = 0.01). These results demonstrated that walnut genotypes have different phenolic compounds and phenolic compounds have different radical scavenging power. The differences of phenolic compounds were confirmed by using high performance liquid chromatography (HPLC).