Contribution of plant phytochemicals to organic homeostasis: focus on the aging process and opportunistic infections

Aging process is conceived as a normal stage during human life cycle, but it is also considered a hot topic among scientists and medical community. Alarming rates of premature aging and oxidative stress-related diseases have increasingly affect human individuals. Stress, pollution and exposition to chemical substances are considered the main triggering factors for those conditions; in addition, they also suppress the immune system and, therefore, improve organic vulnerability and occurrence of opportunistic infections [I]. Apart from the associated morbidity and mortality, the increasing rates of antimicrobial resistance improve the severity of the clinical conditions [2]. Botanical preparations possess a multitude of bioactive properties, namely acting as antimicrobials, antioxidants, and homeostasis modulators. Thus, upcoming alternatives, mainly based in plant phytochemicals, are necessary to improve the wellbeing as also life expectancy of individuals. The present study aims to evaluate and to compare both antioxidant and antimicrobial properties of plant extracts rich in phenolic compounds. Among the tested plants, Glycyrrhiza glabra L. (licorice) evidenced the most pronounced free radicals scavenging and antimicrobial effects, followed by Salvia officina/is L. (sage), Thymus vulgaris L. (thyme) and Origanum vulgare L. (oregano). Eucalyptus globulus Labill. (blue gum) and Juglans regia L. (walnut) also showed a high effect, while Pterospartum tridentatum (L.) Willk. (carqueja) and Rubus ulmifolius Schott (elm leaf blackberry) displayed moderate effects, and lastly, Tabebuia impetigirwsa (Mart. ex DC) Standley (pau d'arco), Foeniculum vulgare Miller (fennel), Rosa canina L. (rose hips) and Matricaria recutita L. (chamomile) gave only slight effects. In general, the most pronounced bioactivities were observed in the plant preparations (infusion>decoction>hydromethanolic extract) with higher levels of phenolic compounds (both flavonoids and phenolic acids). The observed synergisms between the phenolic compounds present in the extracts highlight the use of phytochemicals as future health promoters. However, further studies are necessary to understand the effective mode of action of individual phenolic constituents as also the existence of polyvalence relationships between them.

Dietary supplements based on borututu: wich is the most suitable choice for bioactive purposes?

Borututu ( Cochlospermum angolensis Welw.) is a widespread tree in Angola used since antiquity by traditional healers for the prevention and treatment of hepatic diseases and for the prophylaxis of malaria [1]. This plant is mostly consumed as infusions but is also available as dietary supplements, such as piiis, capsules, and syrups, among others. In the present study, the aim was to evaluate the proximate composition and energetic contribution of borututu as weii as its composition in hydrophilic (sugars and organic acids) and lipophilic (fatty acids and tocopherols) compounds, given the fact that this plant is directly introduced in some dietary supplements. Furthermore, the bioactivity (antioxidant, hepatoprotective and antimicrobial activities) of three different formulations of borututu (infusion, pills, and syrup) was assessed and compared, and since plant beneficial properties are often ascribed to phenolic compounds [2], the phenolic profile of the formulations was also analysed. Carbohydrates (88 g/100 g) and fat (2.5 g/100 g) were the major and tl1e minor components of the studied borututu dry barks, respectively, with an energetic contribution of 384 kcal/100 g. Fructose was the most abundant sugar (1.3 g/100 g), foilowed by sucrose, trehalose and glucose (1.1, 0.98 and 0.79 g/100 g, respectively). Oxalic (0.70 g/100 g), malic (0.63 g/100 g) and citric (0.57 g/100 g) acids were present in higher amounts but shikimic and fumaric acids were also detected. Among the fatty acids found in borututu, a prevalence of saturated fatty acids (SF A; 48.2%) was observed, whereas polyunsaturated (PUFA) and monounsaturated (MUFA) fatty acids were detected in relative percentages of 30.9% and 20.8%, respectively. P-tocopherol was the most abundant of the four isoforms found in the sample, foiiowed by o-, a- and y-tocopherol, present in concentrations of 597,43, 3.7 and 2.0 g/100 g, respectively. Borututu infusion revealed the highest antioxidant activity, with EC50 values ranging from 20 to 600 J.lg/mL and was the only formulation inhibiting the growth of an HepG2 ceii line, with a Gl5o value of 146 J.lg/mL. This formulation.also revealed the best antimicrobial capacity and proved to be able to inhibit the growth of Escherichia coli, E. coli ESBL, Staphylococcus aureus and Pseudomonas aeruginosa, with MIC values of 50, 6.2, 1.6 and 25 mg!mL, respectively. Pills revealed activity against some of the studied bacterial strains and the syrup did not reveal antimicrobial activity at the studied concentration. Eilagic acids, methyl ellagic acids, eucaglobulinlglobulusin B and (epi)gaiiocatechin-0-gallate were the compounds present in all the different formulations. The highest concentration of phenolic compounds was found in the infusion extract. Protocatechuic acid was the most abundant phenolic compound in the infusions, the only preparation where it was detected, whereas ( epi)gaiiocatechin- 0-gallate was the main phenolic in the pills and eucaglobulinlglobulusin in the syrup. In a general way, borututu proved to be a good source of phytochemicals such as phenolic compounds, with the infusions revealing the best bioactive properties.

Use of inert gas-enriched atmospheres and post-packaging irradiation in shelf-life extension of watercress

Watercress (Nasturtium officinale R. Br.) is a semi-aquatic plant of the Brassicaceae family highly appreciated in the Mediterranean cuisine. It features sharp, peppery and slightly tangy taste and contains health-promoting phytochemicals. Its consumption as a fresh-cut product has increased in recent years, as well as the global market of minimally processed vegetables. This demand is driven by the growing interest in the role of food in promoting the human health and wellbeing and to meet consumer needs for fresh-like and more convenient foods. Due to the reduced shelf-life of this plant, the suitability of inert gas-enriched atmospheres and ionizing irradiation for preserving visual, nutritional and functional quality attributes during cold storage was studied. Watercress samples were gathered in the Northeast region of Portugal, rinsed in tap water and a portion was immediately analyzed (non-stored control). The remaining fresh material was packaged in polyethylene bags under N2- and Ar-enriched atmospheres, conventional atmosphere (air) and vacuum (no atmosphere). Samples under conventional atmosphere were irradiated at 1, 2 and 5 kGy of gamma-rays (predicted doses) in a 60Co experimental chamber. A non-irradiated control followed all the experiment. Then, all packaged samples were stored at 4 ºC for 7 days. The studied quality parameters included the colour that was measured with a Konica Minolta colorimeter, and total soluble solids and pH determined in squeezed juice. The proximate composition (moisture, proteins, fat, ash, carbohydrates and energy) was evaluated using the AOA C procedures. Organic acids, free sugars, fatty acids and tocopherols were analyzed by chromatographic techniques. Samples were also evaluated for its DPPH• scavenging activity, reducing power, and lipid peroxidation inhibition capacity trough the inhibition of the β-carotene bleaching and thiobarbituric acid reactive substances (TBAR S) formation. Differences among treatments were analyzed using the one-way analysis of variance (ANO VA) and a linear discriminant analysis (LDA ) was used to evaluate the effects on the overall postharvest quality. After evaluating the effect on the individual quality parameters, the LDA revealed that the Ar-enriched atmosphere and the irradiation at 2 kGy were suitable processing choices for preserving the integrity of the non-stored control samples during cold storage. Thus, these non-thermal treatments were highlighted for shelf-life extension of fresh-cut watercress.

HPLC-DAD-ESI/MS phenolic characterization and biological activity of Equisetum giganteum L.

Naturally-occurring phytochemicals have received a pivotal attention in the last years, due to the increasing evidences of biological activities. Equisetum giganteum L., commonly known as “giant horsetail”, is a native plant from Central and South America, being largely used in dietary supplements as diuretic, hemostatic, antiinflammatory and anti-rheumatic agents [1,2]. The aim of the present study was to evaluate the antioxidant (scavenging effects on 2,2-diphenyl-1-picrylhydrazyl radicals- RSA, reducing power- RP, β-carotene bleaching inhibition- CBI and lipid peroxidation inhibition- LPI), anti-inflammatory (inhibition of NO production in lipopolysaccharidestimulated RAW 264.7 macrophages) and cytotoxic (in a panel of four human tumor cell lines: MCF-7- breast adenocarcinoma, NCI-H460- non-small cell lung cancer, HeLa- cervical carcinoma and HepG2- hepatocellular carcinoma; and in non-tumor porcine liver primary cells- PLP2) properties of E. giganteum, providing a phytochemical characterization of its extract (ethanol/water, 80:20, v/v), by using highperformance liquid chromatography coupled to diode array detection and electrospray ionisation mass spectrometry (HPLC-DAD–ESI/MS). E. giganteum presented fourteen phenolic compounds, two phenolic acids and twelve flavonol glycoside derivatives, mainly kaempferol derivatives, accounting to 81% of the total phenolic content, being kaempferol-O-glucoside-O-rutinoside, the most abundant molecule (7.6 mg/g extract). The extract exhibited antioxidant (EC50 values = 123, 136, 202 and 57.4 μg/mL for RSA, RP, CBI and LPI, respectively), anti-inflammatory (EC50 value = 239 μg/mL) and cytotoxic (GI50 values = 250, 258, 268 and 239 μg/mL for MCF-7, NCI-H460, HeLa and HepG2, respectively) properties, which were positively correlated with its concentration in phenolic compounds. Furthermore, up to 400 μg/mL, it did not revealed toxicity in non-tumor liver cells. Thus, this study highlights the potential of E. giganteum extracts as rich sources of phenolic compounds that can be used in the food, pharmaceutical and cosmetic fields.

Phenolic profile of Cynara scolymus L. and Silybum marianum(L.) Gaertn and related antimicrobial properties

Cynara scolymus L. (artichoke) and Silybum marianum (L.) Gaertn (milk thistle), belonging to the Asteraceae family, are medicinal plants vo.ith well-reported antioxidant and hepatoprotective effects. Widely consumed as infusions, these plants can also be found in several formulations to allow an easier consumption. The bioactivity of infusions, pills, and syrups based on artichoke and milk thistle was previously reported by our research group [1 ,2] and among the various phytochemicals present in these dietary supplements, phenolic compounds are pointed out as the most responsible for their beneficial properties. With the aim of studying the antimicrobial activity and possible relation vo.ith the phenolic composition, two different formulations of each plant were assessed (pills and syrups). The phenolic profiles were obtained by HPLC-DAD-ESIIMS, and the antimicrobial activity was performed with clinical isolates from hospitalized patients, namely Escherichia coli, Escherichia coli spectrum extended producer of P-lactarnases (ESBL), Proteus mirabilis, Pseudomonas aeruginosa, and methicillin-resistant Staphylococcus aureus (MRSA). Vanillic acid (5.58 J.tg/g) and luteolin-7-0-glucoside (2.2 J.tg/g) were the most abundant compounds in artichoke syrup, that did not reveal antimicrobial activity against the studied strains, which could be due to their low concentrations. On the other hand, artichoke pills presented a prevalence of 5-0-caffeoylquinic (28.2 J.tg/g), 1,3-dicaffeoylquinic (24 J.tg/g), and 4-0-Caffeoylquinic acids (13.3 J.tg/g); revealing the capacity to inhibit MRSA vo.ith a MIC value of 1.9 mg!g. Regarding milk thistle, isorhamnetin-0-deoxyhexoside-0-hexoside, isorhamnetin-3-0-rutinoside, and isorhamnetin-0-deoxyhexoside-0-dihexoside were the major compounds detected in the syrup, in concentrations of 7.26, 5. 75, and 3.64 J.tg/g, respectively. This formulation proved to be able to inhibit the growth of E. coli, ESBL, MRSA and P. aeruginosa, with MIC values ranging from 0.2 to 1.3 mg!mL. Hydroxylated silibinin (1.565 J.!g/g) was the major flavonoid found in the pills, that revealed antimicrobial activity against ESBL, with a MIC value of 15 mg!mL, but did not inhibit the growth of the remaining bacteria None of the studied samples was able to inhibit P. mirabilis at the studied concentrations (1000 and 26.4 mg!mL for the syrups of artichoke and milk thistle, respectively; 150 mg/mL for both pills). Overall, the studied syrups and pills of artichoke and milk thistle revealed to be a good source of phenolic compounds, with some of these formulations revealing antimicrobial activity.

Effects of gamma and electron beam irradiation in the antioxidante potential of methanolic extracts and infusions of Arenaria Montana L

Irradiation is a methodology qualified for dry ingredients preservation or decontamination and can be performed using various radiation sources and energy levels in accordance with the objectives to be achieved [1]. Electron beam irradiation is used mainly for food products with low density, while gamma irradiation is mainly used for large volumes [2]. Arenaria Montana L. has a high antioxidant potential and richness in bioactive phytochemicals. It is used in Portuguese traditional medicine, acting therapeutically as an anti-inflammatory and diuretic plant [3]. The aim of this work was to evaluate the effects of gamma and electron beam irradiation at different doses (I and 10 kGy) in the antioxidant activity of A. montana. Free radicals scavenging activity, reducing power and lipid peroxidation inhibition properties of its methanolic extracts and infusions were evaluated. Through a global analysis, it was concluded that the antioxidant activity proved to be higher in methanolic extracts in comparison with the infusions, where it decreased with increasing irradiation dose regardless of the technology used (gamma or electron beam). For methanolic extracts, electron beam resulted in increased antioxidant activity while gamma irradiation caused a decrease in these extracts. Thus, the antioxidant potential is variable depending not only on the type of radiation and the dose applied, but also on the solvent used in the preparation of the extracts (methanol or water).

Analysis of phenolic compounds in Cynara scolymus L. and Silybum marianum (L.) Gaertn. by HPLC-DAD-ESI/MS

Cynara scolymus L. (artichoke) and Silybum marianum (L.) Gaertn. (milk thistle) are medicinal plants native to the Mediterranean Basin that belong to the Asteraceae family. The flowers and leaves of milk thistle are used in the treatment of liver, spleen and gallbladder disorders [1] and artichoke leaves are used for their cholagogue, choleretic and choliokinetic actions, and also for treatment of dyspepsia and as antidiabetics [2]. The beneficial properties of medicinal plants can be related to their large diversity of phytochemicals, among which phenolic compounds are outstanding. Thereby, the aim of the present work was to obtain and compare the phenolic profiles of artichoke and milk thistle aqueous (prepared by infusion) and hydromethanolic (maceration in methanol: water 80:20, v/v) extracts, using HPLC-DAD-ESI/MS. The aqueous extract of artichoke presented higher concentration in total phenolic compounds (15.29 mg/g extract) than the hydromethanolic extract (4.37 mg/g) with slight differences between the respective profiles; the major flavonoid found in the aqueous and hydromethanolic extract was luteolin-7-O-glucuronide (5.64 and 0.70 mg/g, respectively), followed by luteolin-7-O-glucoside (2.88 and 0.49 mg/g, respectively). Monocaffeoylquinic acid derivatives were only present in the hydromethanolic extract, being 5-O-caffeoylquinic acid (0.49 mg/g) the most abundant one, while dicaffeoylquinic acid derivatives were mostly identified in the aqueous extract; 1,3-O-dicaffeoylquinic acid was the most abundant one in both extracts (0.90 and 0.37 mg/g in the aqueous and hydromethanolic extract, respectively). Regarding to milk thistle preparations, similar phenolic profiles were observed, with only quantitative differences between them. The aqueous extract revealed a higher phenolic compounds concentration (5.57 mg/g) than the hydromethanolic extract (3.56 mg/g), with apigenin-7-O-glucuronide as the major compound in both preparations (3.14 mg/g in the aqueous extract, and 0.58 mg/g in the hydromethanolic extract). Total flavonoids were higher in the aqueous extract (4.66 mg/g), with apigenin-7-Oglucuronide, luteolin-7-O-glucuronide (1.17 mg/g), and apigenin-O-deoxyhexosylglucuronide (0.36 mg/g) as the main constituents. The phenolic acids found in the hydromethanolic extract (total content 1.65 mg/g), included 5-O-caffeolyquinic and protocatechuic acids (0.56 and 0.44 mg/g, respectively). Besides these phenolic acids, the hydromethanolic extract also revealed high levels of luteolin-7-O-glucuronide (0.58 mg/g). Overall, aqueous extracts presented higher phenolic contents than their hydromethanolic extracts in both species, which could be related with the heat treatment to which infusions were subjected.

Minerals and vitamin B9 in dried plants vs. infusions: assessing absorption dynamics of minerals by membrane dialysis tandem in vitro digestion

Vitamins and mineral elements are among the most important phytochemicals due to their important role in the maintenance of human health. Despite these components had already been studied in different plant species, their full characterization in several wild species is still scarce. In addition, the knowledge regarding the in vivo effects of phytochemicals, particularly their bioaccessibility, is still scarce. Accordingly, a membrane dialysis process was used to simulate gastrointestinal conditions in order to assess the potential bioaccessibility of mineral elements in different preparations of Achillea millefolium (yarrow), Laurus nobilis (laurel) and Taraxacum sect. Ruderalia (dandelion). The retention/passage dynamics was evaluated using a cellulose membrane with 34 mm pore. Dandelion showed the highest levels of all studied mineral elements (except zinc) independently of the used formulations (dried plant or infusion), but yarrow was the only species yielding minerals after the dialysis step, either in dried form, or as infusion. In fact, the ability of each evaluated element to cross the dialysis membrane showed significant differences, being also highly dependent on the plant species. Regarding the potential use of these plants as complementary vitamin B9 sources, the detected values were much lower in the infusions, most likely due to the thermolability effect.

In vivo antioxidant activity of phenolic compounds: facts and gaps

Numerous diseases have been related with free radicals overproduction and oxidative stress. Botanical preparations possess a multitude of bioactive properties, including antioxidant potential, which has been mainly related with the presence of phenolic compounds. However, the mechanisms of action of these phytochemicals, in vivo effects, bioavailability and bio-efficacy still need research. Scope and Approach: The present report aims to provide a critical review on the aspects related with the in vivo antioxidant activity of phenolic extracts and compounds from plant origin. Key findings: Biological functions beyond the human metabolism were discussed, comparing in vivo vs. in vitro studies, as also focusing the conditioning factors for phenolic compounds bioavailability and bio-efficacy. Furthermore, an upcoming perspective about the use of phytochemicals as life expectancy promoters and anti-aging factors in human individuals was provided. Conclusions: Overall, and despite all of those advances, the study of the biological potential of numerous natural matrices still remains a hot topic among the scientific community. In fact, the available knowledge about the responsible phytochemicals for the biological potential, their mechanisms of action, the establishment of therapeutic and prophylactic doses, and even the occurrence of biochemical inter-relations, is considerable scarce.