Luís R. Silva

Can Phlorotannins Purified Extracts Constitute a Novel Pharmacological Alternative for Microbial Infections with Associated Inflammatory Conditions

Bacterial and fungal infections and the emerging multidrug resistance are driving interest in fighting these microorganisms with natural products, which have generally been considered complementary to pharmacological therapies. Phlorotannins are polyphenols restricted to brown seaweeds, recognized for their biological capacity. This study represents the first research on the antibacterial, antifungal, anti-inflammatory and antioxidant activity of phlorotannins purified extracts, which were obtained from ten dominant brown seaweeds of the occidental Portuguese coast. Phlorotannins content was determined by the specific dimethoxybenzaldehyde (DMBA) method and a yield between 75 and 969 mg/Kg phloroglucinol units (dry matter) was obtained. Fucus spiralis ranked first, followed by three Cystoseira species. The anti-inflammatory potential of the purified extracts was assessed via inhibitory effect on nitric oxide (NO) production by lipopolysaccharide-stimulated RAW 264.7 macrophage cells, Cystoseira tamariscifolia being the one showing promising activity for the treatment of inflammation. NO scavenging ability was also addressed in cell free systems, F. spiralis being the species with highest capacity. The antimicrobial potential of the extracts was checked against five Gram-positive and four Gram-negative bacteria and three fungi strains, that commonly colonize skin and mucosa and are responsible for food contamination. The different extracts were more effective against Gram-positive bacteria, Staphylococcus epidermidis being the most susceptible species. Concerning antifungal activity, Trichophyton rubrum was the most sensitive species. Although the molecular mechanisms underlying these properties remain poorly understood, the results obtained turn phlorotannins purified extracts a novel and potent pharmacological alternative for the treatment of a wide range of microbial infections, which usually also present an inflammatory component. In addition to the biological properties demonstrated herein, phlorotannins extracts may also be preferred, in order to avoid side effects and allergic reactions commonly associated with synthetic drugs.

Diversity of Potassium-Solubilizing Microorganisms and Their Interactions with Plants

Potassium is, together with nitrogen and phosphorus, an essential element for plant nutrition, being the third element in the classical chemical fertilizers NPK. Deficiencies in K results in plants with poorly developed roots, small seeds and lower yields, and therefore the availability of this element is crucial for plant growth and development. The plants take this element from soil, but high amounts of K present in soils correspond to insoluble forms from rocks and silicate minerals. The solubilization of K by microorganisms is then a reliable alternative to make available this element for plants. In this chapter, we review the microorganisms reported as K solubilizers and their phylogenetic diversity, including filamentous fungi such as Penicillium or Aspergillus, yeasts such as Torulaspora and bacteria of many different genera from phyla Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. Some of these potassium-solubilizing microorganisms (KSMs) play a positive role in the growth of different plants, which is relevant to sustainable agriculture schemes, and therefore this is a promising field of study in the ‘greening’ era.

Chemical assessment and antioxidant capacity of pepper (Capsicum annuum L.) seeds.

Capsicum annuum L. is reported to be the most widely cultivated species. Recently, waste of vegetable processing, like seeds, has been the subject of many studies as an attempt to find new, alternative and cheap resources of bioactive compounds with application in several industries. Despite their chemical, biological and ecological importance, C. annuum seeds are still poorly studied. To improve the knowledge on the metabolic profile of this matrix, a targeted metabolite analysis was performed in sweet Italian and Reus long pairal pepper seeds. Sterols, triterpenes, organic acids, fatty acids and volatile compounds were determined by different chromatographic methods. The antioxidant activity was assessed against DPPH(·), superoxide and nitric oxide radicals. A concentration-dependent activity was noticed against all radicals. Acetylcholinesterase inhibitory capacity was also evaluated, but no effect was found. Data provide evidence of great similarities between sweet Italian and Reus long pairal pepper seeds. The present study indicates that C. annuum seeds are a potential source of valuable bioactive compounds that could be used in food industry.

Plants Probiotics as a Tool to Produce Highly Functional Fruits: The Case of Phyllobacterium and Vitamin C in Strawberries

The increasing interest in the preservation of the environment and the health of consumers is changing production methods and food consumption habits. Functional foods are increasingly demanded by consumers because they contain bioactive compounds involved in health protection. In this sense biofertilization using plant probiotics is a reliable alternative to the use of chemical fertilizers, but there are few studies about the effects of plant probiotics on the yield of functional fruits and, especially, on the content of bioactive compounds. In the present work we reported that a strain of genus Phyllobacterium able to produce biofilms and to colonize strawberry roots is able to increase the yield and quality of strawberry plants. In addition, the fruits from plants inoculated with this strain have significantly higher content in vitamin C, one of the most interesting bioactive compounds in strawberries. Therefore the use of selected plant probiotics benefits the environment and human health without agronomical losses, allowing the production of highly functional foods.

Integrated Analysis of COX-2 and iNOS Derived Inflammatory Mediators in LPS-Stimulated RAW Macrophages Pre-Exposed to Echium plantagineum L. Bee Pollen Extract

Oxidative stress and inflammation play important roles in disease development. This study intended to evaluate the anti-inflammatory and antioxidant potential of Echium plantagineum L. bee pollen to support its claimed health beneficial effects. The hydromethanol extract efficiently scavenged nitric oxide (•NO) although against superoxide (O2 •−) it behaved as antioxidant at lower concentrations and as pro-oxidant at higher concentrations. The anti-inflammatory potential was evaluated in LPS-stimulated macrophages. The levels of •NO and L-citrulline decreased for all extract concentrations tested, while the levels of prostaglandins, their metabolites and isoprostanes, evaluated by UPLC-MS, decreased with low extract concentrations. So, E. plantagineum bee pollen extract can exert anti-inflammatory activity by reducing •NO and prostaglandins. The extract is able to scavenge the reactive species •NO and O2 •− and reduce markers of oxidative stress in cells at low concentrations.

Inoculation with Bradyrhizobium japonicum enhances the organic and fatty acids content of soybean (Glycine max (L.) Merrill) seeds

Soybean (Glycine max (L.) Merrill) is one of the most important food crops for human and animal consumption, providing oil and protein at relatively low cost. The least expensive source of nitrogen for soybean is the biological fixation of atmospheric nitrogen by the symbiotic association with soil bacteria, belonging mainly to the genus Bradyrhizobium. This study was conducted to assess the effect of the inoculation of G. max with Bradyrhizobium japonicum on the metabolite profile and antioxidant potential of its seeds. Phenolic compounds, sterols, triterpenes, organic acids, fatty acids and volatiles profiles were characterised by different chromatographic techniques. The antioxidant activity was evaluated against DPPH, superoxide and nitric oxide radicals. Inoculation with B. japonicum induced changes in the profiles of primary and secondary metabolites of G. max seeds, without affecting their antioxidant capacity. The increase of organic and fatty acids and volatiles suggest a positive effect of the inoculation process. These findings indicate that the inoculation with nodulating B. japonicum is a beneficial agricultural practice, increasing the content of bioactive metabolites in G. max seeds owing to the establishment of symbiosis between plant and microorganism, with direct effects on seed quality.

Phytochemical investigations and biological potential screening with cellular and non-cellular models of globe amaranth (Gomphrena globosaL.) inflorescences.

Gomphrena globosaL. is a popular edible plant used as food colourant and in traditional medicine. In this work, 24 phenolic compounds and eight betacyanins were determined by HPLC-DAD in three different extracts of G. globosa inflorescences. The decoction presented the highest amount of phenolic compounds, kaempferol-3-O-(6-rhamnosyl)hexoside plus kaempferol-3-O-hexoside being the main compounds. The rich betacyanins extract showed isogomphrenin III and gomphrenin III as major metabolites. Decreases in the nitric oxide amounts were observed in LPS-stimulated RAW 264.7 cells. In vitro scavenging micro assays against several reactive species revealed a great antioxidant capacity, particularly against superoxide radical. In general, the best results were obtained with decoction. Some relationships between activity and composition were established. The data provide evidence of the G. globosa inflorescences potential as a source of anti-inflammatory compounds, with relevance for the treatment of acute or chronic inflammatory conditions, and health-promoting antioxidants for use by both food and pharmaceutical industries.

Chemical composition and biological screening of Capsella bursa-pastoris

Capsella bursa-pastoris (L.) Medik. (Brassicaceae) is a wild herb with high nutritional value that can be eaten raw or cooked. A metabolomic study was performed with different extracts of its aerial parts that were tested concerning their antiradical, acetylcholinesterase inhibitory and antibacterial activities. Phenolic compounds were identified and quantified by HPLC-DAD, organic acids and amino acids were determined by HPLC-UV, while free fatty acids and sterols were analysed by GC-ITMS. The vegetal material was rich in kaempferol-3-O-rutinoside (mean value 2247.09 mg/kg of dry plant), quinic acid (95628.00 mg/kg of dry plant), arginine (mean value of 1.18 mg/kg of dry plant), palmitic acid (284.48 mg/kg) and β-sitosterol (28%). The extracts presented a concentration-dependent antiradical activity (against DPPH•, O2•- and LOO•), being most effective against •NO (EC25 0.20 µg/mL). In addition, the extracts were also acetylcholinesterase inhibitors and antibacterial active, revealing that, besides the plants good nutritional value, it presents important biological properties as well.

Rhizobium as plant probiotic for strawberry production under microcosm conditions

There is increasing interest in the use of plant growth-promoting rhizobacteria (PGPR) as environmental-friendly and healthy biofertilizers. Strawberries (Fragraria x ananassa) are mainly consumed fresh and hence any PGPRs used for biofertilization must be safe for humans, which is the case for members of the genus Rhizobium. In this study, the effects of inoculation of strawberry plants with Rhizobium sp. strain PEPV16, which belongs to the phylogenetic group of R. leguminosarum, and whose plant growth promotion ability has been reported previously for lettuce (Lactuca sativa) and carrots (Daucus carota), was examined. The results demonstrated that PEPV16 promotes strawberry growth through significant increases in the number of stolons, flowers and fruits as compared with uninoculated controls. Compared to uninoculated controls, the fruits of the inoculated plants had higher concentrations of Fe, Zn, Mn and Mo, and they also had higher concentrations of organic acids, such as citric and malic acid, and lower amounts of ascorbic acid than fruits. Although decreases in ascorbic acid have previously been described after the inoculation of strawberry with strains from different PGPR genera, this is the first study to report increases in organic acids after PGPR inoculation.

Sweet cherries from Fundão possess antidiabetic potential and protect human erythrocytes against oxidative damage

Cherries are one of the most appreciated summer fruits due to their attractive colour, sweet taste, high water content, low level of calories and composition in bioactive compounds which, in turn, are important to prevent some pathologies like diabetes, cardiovascular diseases and cancer. In this work we evaluated the phenolic profile and biological potential of 5 varieties of sweet cherries from Fundão region (Portugal) (Saco, Sweetheart, Satin, Maring and Hedelfinger). A total of 23 phenolic compounds were identified by LC-DAD and distributed by the several classes: 6 anthocyanins, 1 hydroxybenzoic acid, 8 hydroxycinnamic acids, 3 flavan-3-ols and 5 flavonols. Maring revealed higher contents in anthocyanins, while Hedelfinger was the richest in non-coloured phenolics. The antioxidant capacity was evaluated against DPPH and nitric oxide radicals. Hedelfinger was the most active against DPPH• (IC50=12.1μg/mL) and Maring against nitric oxide (IC50=140.9μg/mL). Afterwards, antidiabetic capacity was evaluated through the inhibition of α-glucosidase activity, pointing Hedelfinger as the most active (IC50=10.3μg/mL). The capacity of Saco extracts to inhibit the hemoglobin oxidation and the hemolysis of human erythrocytes was also evaluated. Both assays revealed a concentration-dependent inhibitory effect (IC50=38.6μg/mL and IC50=73.0μg/mL, respectively). The results obtained in this study allow us to conclude that sweet cherries possess a great biological potential, and further investigation should be done to promote commercialization and encourage its use in food supplements and in new pharmaceutical and nutraceutical applications.