Direct characterisation of phenolic antioxidants in infusions from four Mapuche medicinal plants by liquid chromatography with diode array detection (HPLC-DAD) and electrospray ionisation tandem mass spectrometry (HPLC-ESI-MS)

A simple, fast and direct method was developed for the qualitative analysis of phenolic constituents from infusions of Mapuche medicinal plants. Teas made of Linum chamissonis Schiede, Quinchamalium chilensis Mol., Adesmia emarginata Clos. and Escallonia illinita K. Presl. were analysed by high-performance liquid chromatography with diode array detector (HPLC-DAD) and electrospray mass spectrometry (ESI-MS). This technique allowed for the first time the tentative identification of 16 phenolic compounds in E. illinita, 27 in Q. chilensis, 10 in L. chamissonis and 19 in A. emarginata. The compounds were mainly phenolic acids, flavonoid glycosides, anthocyanins and tannins. The total phenolic and flavonoid content of the infusions was assessed as well as the free radical scavenging capacity measured by the bleaching of a solution of the 2,2-diphenyl-1-picrylhydrazyl (DPPH⁻) radical. From the four species, Q. chilensis exhibited the strongest antioxidant activity with highest total phenolic and flavonoid content.     

Chemical analysis of edible aromatic plants growing in Tanzania

The volatiles from the aerial parts of edible plants growing in Tanzania, Leucas glabrata, Plectranthus laxiflorus, Salvia nilotica and Vernonia smithiana, were investigated by GC and GC/MS. Thirty-five compounds were identified from L. glabrata, representing 80.4% of the total oil; forty-three from P. laxiflorus (86.7%); twenty-four from S. nilotica (94.3%); and thirty-nine compounds from V. smithiana (92.9%). Among the identified components, menthone, (p + o)-cymene, trans-caryophyllene and caryophyllene oxide were found as the main ones. Furthermore, the essential oils were investigated for their antimicrobial activity as well as for their antiradical activity, through the DPPH method. Upon antimicrobial assays, the oil of V. smithiana showed very strong antimicrobial activity against Gram-positive bacteria, oral pathogens and pathogenic fungi; the oil of P. laxiflorus also exhibited strong activity, mostly against Gram-positive bacteria and especially oral pathogens, while L. glabrata showed strong activity against all assayed bacteria. The essential oil of S. nilotica appeared to have the most antioxidant activity but was almost inactive against all tested microorganisms.     

Toxigenic microorganisms in medicinal plants used for ritual protection of infants

Plants used as a part of infant protective rituals in some countries of South Eastern Africa and which use has been associated to food poisoning episodes were submitted to a microbiological analysis to investigate potential microbiological hazards. This characterization led to the detection of a high load of moulds and aerobic spore-formers microorganisms. Isolates of Bacillus cereus and Bacillus thuringiensis were observed to contained different toxin-encoding genes, and the production of diarrheal enterotoxin was confirmed in some of them. The production of aflatoxin B₁ and cyclopiazonic acid by strains of Aspergillus flavus, and citrinin and penicillic acid by Penicillium citrinum was revealed by HPLC. The toxicity of these isolates was also showed by the Artemia salina lethality test. The results indicate the presence of microorganisms with toxigenic potential in plants used as folk medicine in South Eastern Africa. The traditional use of these preparations should be carefully reconsidered due to the microbiological risks associated with their ingestion.     

Flavan-3-ol-cysteine and acetylcysteine conjugates from edible reagents and the stems of Cynomorium songaricum as potent antioxidants

Flavan-3-ol derivatives, including 3 cysteine conjugates and 3 acetylcysteine conjugates, were prepared using Cynomorium songaricum and edible reagents. The structures were determined, based on NMR and MS spectra. All compounds had stronger radical-scavenging activity than catechin and epicatechin. Moreover, the cysteine conjugates were active against α-glucosidase, whereas catechin and epicatechin were not. Two acetylcysteine flavan-3-ol conjugates, 4β-(l-acetylcysteinyl)-epicatechin 3-O-gallate and 4β-(l-acetylcysteinyl)-epiafzelechin, are novel compounds with better log P values than their cysteine counterparts. These conjugates can be prepared from purified flavan-3-ol polymers or directly from herbal material.     

Antioxidant,anticholinesterase and antimicrobial constituents from the essential oil and ethanol extract of Salvia potentillifolia

The essential oil of Salvia potentillifolia was analysed by GC and GC–MS. Totally, 123 components were detected in both hydrodistilled and steam-distilled oils, α- and β-pinenes being major compounds. The antioxidant activities were determined by using complementary tests, namely, DPPH radical-scavenging, β-carotene-linoleic acid and reducing power assays. The ethanol extract also showed better activity (IC₅₀ = 69.4 ± 0.99 μg/ml) than that of BHT in the DPPH system, and showed great lipid peroxidation inhibition in the β-carotene-linoleic acid system (IC₅₀ = 30.4 ± 0.50 μg/ml). The essential oil showed meaningful butyrylcholinesterase activity (65.7 ± 0.21%), and α-pinene showed high acetylcholinesterase inhibitory activity (IC₅₀ = 86.2 ± 0.96 μM) while β-pinene was inactive. Antimicrobial activity was also investigated on several microorganisms, and the essential oil showed high activity against Bacillus subtilis and B. cereus. It also exhibited remarkable anticandidal activity against Candida albicans and C. tropicalis with MIC values of 18.5 and 15.5 μg/ml, respectively, while α- and β-pinenes showed moderate activity.     

Extractable antioxidants and non-extractable phenolics in the total antioxidant activity of selected plum cultivars (Prunus domestica L.): Evolution during on-tree ripening

The purpose of this study was to evaluate the contribution of extractable antioxidants and non-extractable phenolics to the total antioxidant activity (TAA) of plums. Therefore, the antioxidant activity was determined (ABTS assay) in aqueous–organic extracts, as well as in the extraction residues which were a subject of two different acidic treatments to release hydrolysable tannins and non-extractable proanthocyanidins (NEPA). In addition, the changes in TAA during the last week of ripening were investigated. Extractable antioxidants contributed less than 18% to the TAA, considerably higher values of antioxidant activity were associated with hydrolysable tannins and NEPA, suggesting that the antioxidant activity of plums may be underestimated in the literature. The ripening resulted in an increase of TAA up to 38% in excess of the value determined on the first sampling date. TAA showed a similar pattern over the ripening period for all cultivars studied.     

An enhanced preparation and purification of the major antioxidants baicalein and wogonin from Scutellariae radix

A simple, rapid and improved method was developed to isolate high-purity baicalein and wogonin from Scutellariae radix. The method involves endogenous baicalinase-catalysed hydrolysis (EBCH), partition, automated low-pressure preparative chromatography (LPPC) and recrystallisation without multiple and tedious column chromatography. This process was optimised for large scale production of baicalein and wogonin with high yields, low costs and process automation. The transformation ratio of baicalin and wogonoside reached 98.21% and 96.60% after EBCH, leading to an increase of 5.41-fold in baicalein and 3.89-fold in wogonin, compared to a raw sample without hydrolysis. The purity of final products was more than 98% after one-step LPPC and recrystallisation. The experimental results show that EBCH–LPPC is an effective method for preparing high purity antioxidants.     

Antioxidant properties of 1,2,3,4,6-penta-O-galloyl-β-d-glucose from Elaeocarpus sylvestris var. ellipticus

The antioxidant potential of 1,2,3,4,6-penta-O-galloyl-β-d-glucose (PGG), isolated from Elaeocarpus sylvestris var. ellipticus, was investigated by various established systems based on cell-free and cell system experiments, such as radical detection, antioxidant enzyme assay, lipid peroxidation detection, and cell viability assay. PGG was found to quench the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical and intracellular reactive oxygen species. PGG recovered the cellular antioxidant enzyme activities of superoxide dismutase, catalase, and glutathione peroxidase, which were reduced by H₂O₂ treatment, thereby resulting in the inhibition of lipid peroxidation. Cytoprotective effects of PGG were based on the results of DNA fragmentation, mitochondrial membrane potential (Δψ), apoptotic body formation, and caspase-3 activity. The results suggest that PGG protects cells against H₂O₂-induced cell damage via antioxidant properties.     

The hydroxycinnamic acid content of barley and brewers’ spent grain (BSG) and the potential to incorporate phenolic extracts of BSG as antioxidants into fruit beverages

The hydroxycinnamic acid (HA) content of starting barley for brewers’ spent grains (BSG), whole BSG and phenolic extracts from BSG was measured using high performance liquid chromatography (HPLC) and correlated with antioxidant potential. The effect of BSG phenolic extracts on antioxidant activity of fruit beverages was also assessed (using the total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays). The concentration of HA present in barley extract and BSG was in the order of ferulic acid (FA), p-coumaric acid (p-CA) derivatives, FA derivatives, p-CA, caffeic acid (CA) and CA derivatives. Results suggested that brewing and roasting decreased the HA content. Antioxidant activity was significantly (P < 0.05) correlated with caffeic acid (R² = 0.8309) and total HA (R² = 0.3942) concentrations. Addition of extracts to fruit beverages resulted in a significant (P < 0.05) increase in antioxidant activity of cranberry juice, measured by the FRAP assay. In vitro digestion significantly (P < 0.05) reduced TPC, DPPH and FRAP activity of the fruit beverages.     

Evaluation of the antioxidant activity of Ruellia tuberosa

The antioxidant activity of Ruellia tuberosa L. (Acanthaceae) was investigated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging assay and the hydrogen peroxide-induced luminol chemiluminescence assay. The methanolic extract (ME) and its four fractions of water (WtF), ethyl acetate (EaF), chloroform (CfF), and n-hexane (HxF) were prepared and then subjected to antioxidant evaluation. The results of both methods revealed that R. tuberosa possesses potent antioxidant activity. The antioxidant activities of the different fractions tested decreased in the order of EaF > CfF > ME > WtF > HxF according to the hydrogen peroxide-induced luminol chemiluminescence assay, and results were the same with the exception of the rank order of HxF and WtF according to the DPPH free radical-scavenging assay. The results provide useful information on the pharmacological activities associated with free radicals of this traditional folk remedy.     

Major chemotypes and antioxidative activity of the leaf essential oils of Cinnamomum osmophloeum Kaneh. from a clonal orchard

Essential oils of 92 cutting clones from a clonal orchard of Cinnamomum osmophloeum Kaneh. were obtained by hydrodistillation and characterised by gas chromatography–mass spectrometry. Our results showed that the yields of essential oils ranged between 0.09% and 2.65% (vol/fresh wt). The constituents of essential oils varied among samples. The major chemotypes classified in the individual cutting clones were cinnamaldehyde (50 plants, representing 50–95% of the total volatiles), linalool (1 plant, 73.3%), β-cubebene (2 plants, 59.4% and 78.7%), and cinnamyl acetate (1 plant, 61.8%). The antioxidant activities of the four chemotypes were determined using a 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. The antioxidant activities of the essential oil decreased in the order of cinnamyl acetate > cinnamaldehyde > β-cubebene > linalool. Indigenous cinnamon oil extract showed a good free radical-scavenging capacity at all concentrations studied, except at 2 μg/ml. The scavenging activity increased with increasing concentration of the extract. The capability of the four essential oil chemotypes to reduce the stable radical, DPPH, to DPPH-H was assayed by a decrease in the IC₅₀ values of 10.4 (cinnamyl acetate type) to 29.7 (linalool type) μg/ml. These results suggest that the leaf essential oil of C. osmophloeum possesses chemical compounds with antioxidant activity which can be used as natural preservatives in food and/or by the pharmaceutical industry. Trees in this plantation which can be used for further propagation for the production of chemotypes of interest were identified.     

Antioxidant activity of hydrolysates obtained from scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle

Scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle were hydrolysed with commercially available food-grade proteases. The resulting hydrolysates showed DPPH and hydroxyl radicals scavenging abilities, reducing power, and ferrous ion chelating capacity. The antioxidant activities of hydrolysate of abalone foot muscle (HAFM) increased with increasing incubation time during the whole hydrolysis process in 180 min. Whereas, the antioxidant activities of hydrolysate of scallop adductor muscle (HSAM) increased at initial stage and peaked after 25-30 min of hydrolysis, and then gradually decreased thereafter. Compared with HAFM, HSAM with comparable hydrolysis time contained more free amino acids (FAA) and small-sized peptides (below 500 Da), which may account for the differences in antioxidant activities versus hydrolysis time curves of the two hydrolysates. The above results indicate that limited hydrolysis of proteins can increase their antioxidant activity, whereas extensive hydrolysis can decrease it.     

Lipid peroxidation,protein oxidant and antioxidant status of muscle,intestine and hepatopancreas for juvenile Jian carp (Cyprinus carpio var. Jian) fed graded levels of myo-inositol

Lipid peroxidation, protein oxidant and antioxidant status of muscle, intestine and hepatopancreas in juvenile Jian carp (Cyprinus carpio var. Jian) fed graded levels of myo-inositol (MI) (163.5, 232.7, 384.2, 535.8, 687.3, 838.8 and 990.3 mg/kg diet) for 60 days were investigated. Total tissue malondialdehyde (MDA) and protein carbonyl (PC) content showed a downward trend to a point (P < 0.05). Conversely, total tissue anti-hydroxyl radical (AHR), catalase (CAT), glutathione-S-transferase (GST), glutathione peroxidase (GPx), glutathione reducase (GR) activities and glutathione (GSH) content were generally higher in MI-supplemented diets than MI-unsupplemented diet (P < 0.05). Muscle and intestinal superoxide dismutase (SOD), and intestinal anti-superoxide anion (ASA) were increased by MI supplementation (P < 0.05), whereas these parameters in the other tissue showed no alterations (P > 0.05). These results indicated that antioxidant status was improved, and lipid peroxidation and protein oxidant were depressed in muscle, intestine and hepatopancreas by MI.     

Comparative chemical composition,antioxidant and hypoglycaemic activities of Juniperus oxycedrus ssp. oxycedrus L. berry and wood oils from Lebanon

Juniper (Juniperus oxycedrus) is used in European cuisine for its distinguishing flavour. J. oxycedrus ssp. oxycedrus berry and wood essential oils were tentatively identified by GC and GC/MS. Fifty compounds were identified in the berry oil and 23 compounds were identified in the wood oil. The J. oxycedrus ssp. oxycedrus berry oil was characterised by high contents of α-pinene (27.4%) and β-myrcene (18.9%). Other important compounds were α-phellandrene (7.1%), limonene (6.7%), epi-bicyclosesquiphellandrene (2.3%) and δ-cadinene (2.2%) while, in the wood oil, δ-cadinene (14.5%) is a major main component, together with cis-thujopsene (9.2%) and α-muurolene (4.9%). In vitro evaluation of antioxidant activity by the DPPH method showed a significant activity for both oils with IC₅₀ values of 1.45 μl/ml for wood and 7.42 μl/ml for berries. Hypoglycaemic activity was investigated through the inhibition of α-amylase. The results revealed that oil obtained by hydrodistillation from J. oxycedrus ssp. oxycedrus wood exhibits an interesting activity with IC₅₀ of 3.49 μl/ml.     

Development of a strain for efficient degradation of polychlorinated biphenyls by patchwork assembly of degradation pathways

Rhodococcus jostii RHA1 accumulates chlorobenzoates (CBA) during the degradation of polychlorinated biphenyls (PCBs). CBA degradation is considered one of the rate-limiting steps in the complete degradation of PCBs. To reduce the accumulation of CBAs, the upper pathway enzyme genes for PCB degradation of RHA1 were introduced into a CBA-degrading bacterium, Burkholderia sp. NK8. The resulting recombinant strain exhibited no biphenyl 2,3-dioxygenase (BphA) activity encoded by bphAaAbAcAd genes, which encode the large and small subunits of the terminal oxygenase component and the ferredoxin and reductase subunits responsible for electron transfer from NADH to the large subunit. The remaining enzyme genes involved in the transformation of biphenyl to benzoate, bphB2C1D1, which encode dehydrogenase, ring-cleavage dioxygenase and hydrolase, conferred activities to NK8. To obtain the BphA activity of RHA1 in NK8, sets of BphA genes were constructed by combining the bphAaAbAcAd genes of RHA1 and bphA3A4 of Pseudomonas pseudoalcaligenes KF707, encoding the ferredoxin and reductase subunits. Hybrid derivatives of BphA containing the KF707 bphA3 conferred BphA activity to NK8, and a derivative containing the RHA1 bphAaAb and KF707 bphA3A4 genes exhibited the highest BphA activity. A plasmid containing the RHA1 bphAaAb and KF707 bphA3A4 genes plus the RHA1 bphB2C1D1 genes was constructed and introduced into NK8. The resulting recombinant strain efficiently degraded 2-, 3- and 4-chlorobiphenyls with an apparent reduction in CBA accumulation in comparison to the recombinant mutant strain, which had an insertion in the cbeA gene to inactivate CBA dioxygenase.     

Characterization of antibiotic resistant enterococci isolated from untreated waters for human consumption in Portugal

Untreated drinking water is frequently overlooked as a source of antibiotic resistance in developed countries. To gain further insight on this topic, we isolated the indicator bacteria Enterococcus spp. from water samples collected in wells, fountains and natural springs supplying different communities across Portugal, and characterized their antibiotic resistance profile with both phenotypic and genetic approaches. We found various rates of resistance to seven antibiotic families. Over 50% of the isolates were resistant to at least ciprofloxacin, tetracyclines or quinupristin-dalfopristin and 57% were multidrug resistant to ≥ 3 antibiotics from different families. Multiple enterococcal species (E. faecalis, E. faecium, E. hirae, E. casseliflavus and other Enterococcus spp) from different water samples harbored genes encoding resistance to tetracyclines, erythromycin or gentamicin [tet(M)-46%, tet(L)-14%, tet(S)-5%, erm(B)-22%, aac(6´)-Ie-aph(2″)-12%] and putative virulence factors [gel-28%, asa1-16%]. The present study positions untreated drinking water within the spectrum of ecological niches that may be reservoirs of or vehicles for antibiotic resistant enterococci/genes. These findings are worthy of attention as spread of antibiotic resistant enterococci to humans and animals through water ingestion cannot be dismissed.     

Evaluation of the antioxidant activity of five endemic Ligustrum species leaves from Taiwan flora in vitro

Leaves from the plant species belonging to the genus Ligustrum are widely used as tea or herbal medicine in Europe, China, and Japan. The antioxidant properties of five Ligustrum species from Taiwan were compared using in vitro antioxidant methods such as DPPH radical scavenging, TEAC, and FRAP assays. Cell-based antioxidant methods were used, including Fe²⁺/ascorbate-induced lipid peroxidation on brain homogenate and AAPH-induced erythrocyte haemolysis. The amounts of major phenolic compounds from the Ligustrum species, including phenylpropanoids, flavonoids, and iridoids, were determined by spectrophotometric methods. The results showed that all Ligustrum species exhibited antioxidant, radical-scavenging, anti-haemolytic, and lipid peroxidation-inhibiting activities at different magnitudes of potency. A significant correlation was found between antioxidant activity and the amount of antioxidant components, in particular, total phenolics and phenylpropanoids. Among all Ligustrum species from Taiwan, Ligustrum morrisonense is presented as potential source of natural antioxidants.     

Retention and distribution of natural antioxidants (α-tocopherol, polyphenols and terpenic acids) after shallow frying of vegetables in virgin olive oil

Potatoes, green peppers, zucchinis and eggplants were shallow fried in virgin olive oil (VOO) according to the Mediterranean traditional culinary practice. Zucchinis and eggplants were also blanketed with wheat flour or batter prior to frying. Polyphenols and hydroxy pentacyclic triterpene acids (HPTAs) were determined by GC/MS, while α-tocopherol was determined by high-performance liquid chromatography. Among 12 polyphenols determined, tyrosol predominated in frying oils and zucchini samples, while chlorogenic acid was the major phenolic species in the other vegetable samples. The triterpene acids maslinic, oleanolic and ursolic were determined in frying oils and fried vegetables, while α-tocopherol was present in all samples. Besides water loss and oil absorption, shallow frying resulted in partial loss of all the antioxidants studied in frying oils and enrichment of fried vegetables with olive oil antioxidants, which was in some extent affected by the type of vegetable fried and the culinary practice followed. The overall retention of the antioxidants in oil and food ranged from 32% to 64% for α-tocopherol, 25% to 70% for polyphenols and 35% to 83% for HPTA. It appears that vegetables fried in VOO provide an additional intake of α-tocopherol, terpenic acids and polyphenols as tyrosol and chlorogenic acid.     

Stilbene levels and antioxidant activity of Vranec and Merlot wines from Macedonia: Effect of variety and enological practices

The content of the stilbenes trans-resveratrol and piceid as well as the antioxidant activity of Macedonian red wines from the two main grape varieties Vranec and Merlot have been evaluated. Тhe effects of time of maceration, type of yeast and the level of sulphur dioxide applied on stilbene content and antioxidant activity have been studied. The most important factor in winemaking technology is the maceration time since the highest concentrations of trans-resveratrol, piceid and highest antioxidant activity were found following 6 and 10 days of maceration. Concerning the yeast type, higher concentrations of trans-resveratrol and piceid have been obtained with French yeast “Levuline CHP” in comparison to Macedonian yeast “Vinalco”. In contrast, the higher antioxidant activity of wines from both varieties of grapes was observed by application of Macedonian yeast “Vinalco”.