Phenolic composition of four sage species: salvia farinacea, salvia mexico, salvia greggii and salvia officinalis

Salvia species are used worldwide for medicine purposes. In general, these medicinal plants have high amounts of flavonoids and phenolic acids, that are thought to be closely related to their health properties [1,2]. In this work, the aerial parts of Salvia farinacea, Salvia mexico, Salvia greggii and Salvia officinalis were extracted with hot water [3]. Extracts were evaluated for their total phenolic content by an adaptation of the Folin-Ciocalteu method and further analysed by high performance liquid chromatography associated with electrospray mass spectrometry (HPLC-DAD-ESI-MSn) in the negative ion mode [4], in order to identify their individual phenolic constituents. The aqueous extracts of S. farinacea, S. mexico, S. officinalis and S. greggii contained, respectively, 106±13, 159±38, 175±46 and 136±1 μg GAE/mg of total phenolics. These four species were characterized by a clear prevalence of caffeic acid derivatives, in particular of rosmarinic acid (MW 360), that is generally the most abundant phenolic compound in Salvia species [2,3]. In addition, S. mexico and S. officinalis contained moderate amounts of salvianolic acid B (MW 718). Among these two, S. mexico was richer in O-caffeoylquinic acid (MW 354), while the latter presented high amounts of salvianolic acid K (MW 556) and moderate amounts of its structural isomer. All the extracts were enriched in flavones: S. farinacea and S. officinalis contained high amounts of luteolin-O-glucuronide while S. mexico contained luteolin-C-glucoside with respective characteristic mass spectrometry fragmentation pattern m/z at 461→285 and m/z at 447→357, 327. Similarly, S. greggii extract presented high content of luteolin-7-O-glucoside ([M-H]− at m/z 447→ 285) and luteolin-C-glucoside and moderate quantities of apigenin-C-hexoside ([M-H]− at m/z 431→341, 311). Further studies are being undertaken in order to understand the contribution of these phenolic constituents in the biological activities of Salvia plants.

Phenolic composition of four sage species: salvia farinacea, salvia mexico, salvia greggii and salvia officinalis

Salvia species are used worldwide for medicine purposes. In general, these medicinal plants have high amounts of flavonoids and phenolic acids, that are thought to be closely related to their health properties [1,2]. In this work, the aerial parts of Salvia farinacea, Salvia mexico, Salvia greggii and Salvia officinalis were extracted with hot water [3]. Extracts were evaluated for their total phenolic content by an adaptation of the Folin-Ciocalteu method and further analysed by high performance liquid chromatography associated with electrospray mass spectrometry (HPLC-DAD-ESI-MSn) in the negative ion mode [4], in order to identify their individual phenolic constituents. The aqueous extracts of S. farinacea, S. mexico, S. officinalis and S. greggii contained, respectively, 106±13, 159±38, 175±46 and 136±1 μg GAE/mg of total phenolics. These four species were characterized by a clear prevalence of caffeic acid derivatives, in particular of rosmarinic acid (MW 360), that is generally the most abundant phenolic compound in Salvia species [2,3]. In addition, S. mexico and S. officinalis contained moderate amounts of salvianolic acid B (MW 718). Among these two, S. mexico was richer in O-caffeoylquinic acid (MW 354), while the latter presented high amounts of salvianolic acid K (MW 556) and moderate amounts of its structural isomer. All the extracts were enriched in flavones: S. farinacea and S. officinalis contained high amounts of luteolin-O-glucuronide while S. mexico contained luteolin-C-glucoside with respective characteristic mass spectrometry fragmentation pattern m/z at 461→285 and m/z at 447→357, 327. Similarly, S. greggii extract presented high content of luteolin-7-O-glucoside ([M-H]− at m/z 447→ 285) and luteolin-C-glucoside and moderate quantities of apigenin-C-hexoside ([M-H]− at m/z 431→341, 311). Further studies are being undertaken in order to understand the contribution of these phenolic constituents in the biological activities of Salvia plants.

Phenolic compounds and antioxidant capacity of three thymus species plants

Thymus plants comprise distinct species with claimed health properties [1], commonly associated to their essential oils and phenolic compounds. Albeit that, the phenolic composition and the biological activities of many Thymus species remain unclear. This work aimed to elucidate the phenolic composition and antioxidant properties of aqueous extracts from Thymus herba barona, Thymus caespetitus and Thymus fragrantissimus. The aqueous extracts of the three Thymus species were evaluated for their total phenolic compounds by an adaptation of the Folin-Ciocalteu method [2], and individual phenolic compounds were identified by high performance liquid chromatography associated with electrospray mass spectrometry (HPLC-DAD-ESI-MSn) in the negative mode. The antioxidant activity of each extract was carried out by DPPH● scavenging assay and ferric reducing antioxidant power assays [3]. Total phenolic compounds in the three extracts ranged from 236±27 (T. caespetitus) to 273±17 μg GAE/mg (T. fragrantissimus). Similarly to other Thymus species [1,4], these extracts were rich in caffeic acid derivatives (characteristic UV spectra maxima at 290 and 328 nm) and mainly composed of rosmarinic acid (MW 360). Other caffeic acid derivatives included salvianolic acid K (MW 556) and 3′-O-(8″-Z-caffeoyl)rosmarinic acid (MW 538). High amounts of the flavone luteolin-O-glucuronide ([M-H]− at m/z 461→285) were found in T. caespetitus while the others species contained moderate amounts of this compound. T. herba barona, T. caespetitus and T. fragrantissimus extracts showed high DPPH radical scavenge ability (EC50 values 11.6±0.9, 13.8±0.6 and 10.9±1.2 μg/mL respectively), as well as high reducing power (EC50 values of 35.1±4.5, 39.3±2.7 and 32.4±4.3 μg/mL, respectively), that were comparable to those of reference compounds. This work is an important contribution for the phytochemical characterization and the antioxidant capacity of these three Thymus species.

Phenolic compounds and antioxidant capacity of three thymus species plants

Thymus plants comprise distinct species with claimed health properties [1], commonly associated to their essential oils and phenolic compounds. Albeit that, the phenolic composition and the biological activities of many Thymus species remain unclear. This work aimed to elucidate the phenolic composition and antioxidant properties of aqueous extracts from Thymus herba barona, Thymus caespetitus and Thymus fragrantissimus. The aqueous extracts of the three Thymus species were evaluated for their total phenolic compounds by an adaptation of the Folin-Ciocalteu method [2], and individual phenolic compounds were identified by high performance liquid chromatography associated with electrospray mass spectrometry (HPLC-DAD-ESI-MSn) in the negative mode. The antioxidant activity of each extract was carried out by DPPH● scavenging assay and ferric reducing antioxidant power assays [3]. Total phenolic compounds in the three extracts ranged from 236±27 (T. caespetitus) to 273±17 μg GAE/mg (T. fragrantissimus). Similarly to other Thymus species [1,4], these extracts were rich in caffeic acid derivatives (characteristic UV spectra maxima at 290 and 328 nm) and mainly composed of rosmarinic acid (MW 360). Other caffeic acid derivatives included salvianolic acid K (MW 556) and 3′-O-(8″-Z-caffeoyl)rosmarinic acid (MW 538). High amounts of the flavone luteolin-O-glucuronide ([M-H]− at m/z 461→285) were found in T. caespetitus while the others species contained moderate amounts of this compound. T. herba barona, T. caespetitus and T. fragrantissimus extracts showed high DPPH radical scavenge ability (EC50 values 11.6±0.9, 13.8±0.6 and 10.9±1.2 μg/mL respectively), as well as high reducing power (EC50 values of 35.1±4.5, 39.3±2.7 and 32.4±4.3 μg/mL, respectively), that were comparable to those of reference compounds. This work is an important contribution for the phytochemical characterization and the antioxidant capacity of these three Thymus species.

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.

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.

Phenolic compounds in Coleostephus myconis (L.) Rchb.f.: characterization by HPLC-DAD-ESI/MS and phenology effects

The Asteraceae family is spread worldwide. In Portugal, there are more than 300 species, standing out as one of the botanical families with largest representation in the Portuguese flora. Coleostephus myconis (L.) Rchb.f. is a scarcely studied Asteraceae species, characterized as having ruderal growth and persistence in abandoned soils (an expanding problem due to the desertification phenomena in rural areas). In this work, the flowers of C. myconis were collected in three different flowering stages (i: flower bud; ii: flower in anthesis; iii: senescent flower) from the Northwestern area of the Portuguese territory. Powdered samples (1 g) were extracted twice with ethanol:water 50:50 (v/v). After removing solvents, the combined extracts were re-dissolved, filtered through 0.22-μm disposable LC filter disks and analyzed by high performance liquid chromatography coupled to a diode array detector and electrospray ionization-mass spectrometry (HPLC-DAD/ESI-MS). The phenolic compounds were characterized according to their UV and mass spectra, and retention times. For the quantitative analysis, calibration curves of standard compounds were used. According to the UV spectra (λmax = 314-330 nm) and pseudomolecular ions ([M-H]-) at m/z 353 and 515, all producing an m/z 191 ion, four compounds derived from quinic acid were detected: 3-O-caffeoylquinic acid (Figure 1A), 5-O-caffeoylquinic acid (Figure 1B), 3,5-O-dicaffeoylquinic acid (Figure 1C) and 4,5-O-dicaffeoylquinic acid (Figure 1D), as also supported by the literature [1,2]. A fifth phenolic acid was identified as protocatechuic acid. The detected flavonoid were quercetin-O-glucuronide, quercetin-3-Oglucoside, myricetin-O-methyl-hexoside and a second glycosylated myricetin (not possible to identify completely). Some statistically significant changes were detected among the different assayed flowering stages; nevertheless, 3,5-O-dicaffeoylquinic acid was the major compound, independently of the phenologic stage. According to the previous results, C. myconis might be considered as a potential natural source of these valuable bioactive compounds, especially considering the high botanical representativeness of this plant and its inexpensiveness.

HPLC-DAD-ESI/MS analysis of phenolic compounds in four tomato varieties and evaluation of the antioxidant capacity

Tomato (Lycopersicon esculentum L.) is the second most important vegetable crop worldwide and a key component in the so-called “Mediterranean diet”. In the Northeastern region of Portugal, local populations still prefer to consume traditional tomato varieties which they find very tasty and healthy, as they are grown using extensive farming techniques. A previous study of our research team described the nutritional value of the round (batateiro), long (comprido), heart (coração) and yellow (amarelo) tomato varieties [1], but the phenolic profile was unknown until now. Thus, the objective of this study was to characterize the phenolic profiles of these four tomato farmers’ varieties by using HPLC-DAD-ESI/MS and evaluate its antioxidant capacity through four in vitro assays based on different reaction mechanisms. A cis p-coumaric acid derivative was the most abundant compound in yellow and round tomato varieties, while 4-O-caffeolyquinic acid was the most abundant in long and heart varieties. The most abundant flavonoid was quercetin pentosylrutinoside in the four tomato varieties. Yellow tomato presented the highest levels of phenolic compounds, including phenolic acids and flavonoids, but the lowest antioxidant activity. In turn, the round tomato gave the best results in all the antioxidant activity assays. This study demonstrated that these tomato farmers’ varieties are a source of phenolic compounds, mainly phenolic acid derivatives [2], and possess high antioxidant capacity [1]; being thus key elements in the diet to prevent chronic degenerative diseases associated to oxidative stress, such as cancer and coronary artery disease.

Gomphrena globosa L. como fonte alternativa de pigmentos naturais: caracterização de betacianidinas por HPLC-PDA-ESI/MS

A perceção, as opiniões e os desejos dos consumidores têm um enorme impacto na indústria alimentar. Na perceção visual, a cor torna-se um fator fundamental e, neste campo, os corantes alimentares assumem uma extrema importância. A cor pode ser considerada um dos atributos mais impressionantes dos géneros alimentícios, que influencia diretamente a preferência e a seleção dos consumidores[1]. Existem muitos corantes naturais utilizados na indústria alimentar, tais como carotenóides, antocianinas e betalaínas. As betalaínas incluem compostos com cores que vão do vermelho-violeta (betacianidinas) ao amarelo-laranja (betaxantinas). As betalaínas não têm sido tão extensamente estudadas como as antocianinas, mas possuem uma capacidade corante três-vezes maior. A única betalaína autorizada como corante natural deriva da beterraba(E-162)[2], mas existem outras fontes alternativas de betacianidinas ,como a que se apresenta neste trabalho: Gomphrenaglobosa L., vulgarmente designada por perpétua roxa.

Antioxidant profile of Stevia rebaudiana Bertoni cultivated in Portugal and submitted to different conservation conditions

The demand for natural sweeteners has been gaining more and more importance due to the great controversy associated with the use of some synthetic sweeteners as cyclamates, aspartame and acesulfame-K. The steviol glycosides (E 960) are a group of natural sweeteners of generalized use; these compounds are obtained from Stevia rebaudiana Bertoni, a sweet plant native from South America (Carocho et al., 2015). However, Stevia rebaudiana Bertoni may have other uses to be exploited, in particular due to its antioxidant capacity. This plant is already produced in Portugal but it is important to evaluate if the plant chemical composition is maintained regardless of culture conditions. Therefore, in this study, stevia samples were cultivated in Braganca (northeastern of Portugal) in a field trial with defined culture conditions. After harvesting, the plants were submitted to two different treatments: kept fresh by freezing (-20°C) and oven-dried (30°C). The antioxidant profile of the samples was studied through evaluation of free radicals scavenging activity, reducing power, phenolic compounds (HPLC-DAD-ESI/MS), tocopherols (HPLC-fluorescence) and free sugars (HPLC-RI). Significant differences were observed: while oven-dried samples showed the highest antioxidant activity and phenolic compounds concentration (mainly 5-O-caffeoylquinic acid and 3,5-O-dicaffeoylquinic acid), the frozen fresh samples had the highest values of total tocopherols and total sugars. These results confirm that the plants grown in Bragança have excellent bioactive secondary metabolites responsible for the observed antioxidant capacity.

Degradation of compounds present in cork boiling water by gamma radiation

Cork boiling water is an aqueous and complex dark liquor with high concentration of phenolic compounds such as phenolic acids and tannins [1, 2], which are considered biorecalcitrants [2]. Ionizing radiation has been widely studied as an alternative technology for the degradation of organic contaminants without the addition of any other (e.g.: Fenton technologies). The aim of this work was to identify the compounds present in cork boiling water and further evaluate the resulting stable degradation products after gamma irradiation. The irradiation experiments of standard solutions were carried out at room temperature using a Co-60 experimental equipment. The applied absorbed doses were 20 and 50 kGy at a dose rate of 1.5 kGy/h, determined by routine dosimeters [3]. The identification of radiolytic products was carried out by HPLC-DAD-ESI/MS. The phenolic compounds were identified by comparing their retention times and UV–vis and mass spectra with those obtained from standard compounds, when available, as well as by comparing the obtained information with available data reported in the literature. Concerning the obtained results and the literature review, the main cork wastewater components are: quinic, gallic, protocatechuic, vanillic, syringic and ellagic acids. Based on this, we used protocatechuic, vanillic and syringic acids as model compounds to study their degradation by gamma radiation in order to identify the corresponding radiolytic products. Standard aqueous solutions were irradiated and the derivatives of each model compound are represented in figure 1. The obtained results seem to demonstrate that the derivatives of the parent compounds could also be phenolic acids, since it was observed the loss of 44 u (CO2) from the [M-H]- ions. Gallic and protocatechuic acids are identified as derivatives of vanillic and syringic acids, and gallic acid as a protocatechuic acid derivative. Compound 5 ([M-H]- at m/z 169) was tentatively identified as 2,4,6-trihydroxybenzoic acid, since its fragmentation pattern (m/z 151, 125 and 107) is similar to that previously reported in literature [4]. The structure of compound 7 was proposed based on the molecular ion and its fragmentation and compound 6 remains unknown.

Otimização da extração de ergosterol assistida por microondas a partir de Agaricus bisporus L., aplicando a técnica estatística de superfície e resposta combinada com a técnica de HPLC-UV

O aumento de consumo de cogumelos tem-se verificado em todo o mundo, não só pelo seu valor nutricional, sabor apurado e textura, mas também pelas suas propriedades medicinais. Existem vários estudos científicos que descrevem os benefícios do consumo de cogumelos, que advêm da sua riqueza em compostos bioativos, tais como micosteróis, em particular, ergosterol. Agaricus bisporus L. é o cogumelo mais consumido em todo o mundo, sendo a sua fração de esteróis constituída essencialmente por ergosterol (90%) [1], tornando a sua extração um tópico de elevado interesse já que esta molécula apresenta elevado valor comercial e inúmeras aplicações nas indústrias alimentar, farmacêutica e cosmética. Segundo a literatura, o teor de ergosterol pode variar entre 3 e 9 mg por g de cogumelo seco. Atualmente, os métodos tradicionais tais como a maceração e a extração em Soxhlet estão a ser substituídos por metodologias emergentes, nomeadamente a extração assistida por microondas, visando diminuir a quantidade de solvente utilizado e o tempo de extração e, naturalmente, aumentar o rendimento da mesma. No presente trabalho, utilizou-se A. bisporus como fonte de ergosterol, tendo-se otimizado as seguintes variáveis relevantes para a sua extração pela tecnologia de microondas (MAE): tempo (0-20 min), temperatura (60-210 ºC) e razão sólido-líquido (1-20 g/L). O solvente utilizado foi o etanol tendo-se aplicado a técnica estatística de superfície de resposta por forma a gerar modelos matemáticos que permitissem maximizar a resposta e otimizar as variáveis que afetam a extração de ergosterol. O conteúdo em ergosterol foi monitorizado por HPLC-UV. Os resultados demonstraram que a técnica MAE é promissora para a extração de ergosterol, tendo-se obtido, para as condições ótimas (20,4 min, 121,5ºC e 1,6 g/L), 569,4 mg ergosterol/100 g de massa seca, valor similar ao obtido com extração convencional por Soxhlet (671,5±0,5 mg/100 g de massa seca). Em síntese, a extração assistida por microondas demonstrou ser uma tecnologia eficiente para maximizar o rendimento de extração em ergosterol.

Antiangiogenic potential of yogurts added with extracts rich in apigenin derivatives

Angiogenesis is a biological process through which there is the formation of new blood vessels from preexisting ones [I]. However, in pathological cases, the abnormal growth of new blood vessels promotes the development of various diseases including cancer [2) through the production of atypically large amounts of angiogenesis factors, e.g. the vascular endothelial growth factor (VEGF) [3]. The plant secondary metabolites have been the subject of several studies to evaluate their benefits to human health. In particular, the phenolic compounds have high potential for use in the food industry, including the development of functional foods. Among these, apigenin has been associated with chemopreventive effects related to cancer [4]. In fact, chemoprevention is a present-day concept and contemplates the use of medicines, biological compounds or nutrients as an intervention strategy of cancer prevention. In this work, an Arenaria montana L hydroethanolic extract was prepared and after characterization by HPLC-DAD-ESI/MS showed to be rich in apigenin derivatives. Furthermore, it exhibited ability to inhibit the phosphorylation of VEGFR-2 (vascular endothelium growth factor receptor) through an enzymatic assay. However, for the major protection of bioactive compounds, the extract was microencapsulated by an atomization/coagulation technique with alginate as the matrix material. Posteriorly, the hydroethanolic extract, in free and microencapsulated forms, was incorporated in yogurts in order to develop a novel chemopreventer food in relation to the angiogenesis process. The functionalized yogurts with A. montana extracts (free and microencapsulated) showed a nutritional value similar to the used control (yogurt without extract); however, the samples enriched with extracts revealed added-value regarding the VEGFR-2 phosphorylation inhibition ability. This effect was more effectively preserved over time in the samples functionalized with the protected extract. Overall, this work contributes to the valorization of plants rich in flavonoids, exploring its antiangiogenic potential with VEGFR-2 as target. Moreover, the atomization/coagulation technique allowed the production of viable microspheres enriched with the plant extract. The microspheres were effectively incorporated into yogurts, protecting the extract thus envisaging the development of novel functional foods with chemopreventive effects.

The use of wild fruits of Fragaria vesca L. in preparations with bioactive properties: chemical characterization, antioxidant, antibacterial and antibiofilm activities

Wild strawberry, Fragaria vesca L., belongs to Rosaceae family and is commonly found in roadsides and slopes [1]. The most consumed parts of this plant are its sweet small fruits, which constitute a source of vitamins and phenolic compounds, being also used in infusions due to their organoleptic properties and for the treatment of some intestinal disorders [2, 3]. In the present work, F. vesca fruits were evaluated for their nutritional value and further used in the preparation of infusions. The chemical composition of the fruits and corresponding infusions was determined in terms of soluble sugars, organic acids, tocopherols, folates (by HPLC coupled to different detectors), phenolic compounds (by HPLC-DAD/ESI-MS) and mineral elements (atomic absorption spectroscopy). Some of these bioactive compounds were correlated with antioxidant and antibacterial properties evaluated either in infusions as also in hydromethanolic extracts. Carbohydrates were the main macronutrients in the fruits, followed by fat and proteins. Regarding the fatty acids, polyunsaturated fatty acids showed higher prevalence, mainly due to the presence of D-linolenic (Cl8:3n3) and y-linolenic (Cl8:3n6) acids. Sucrose and citric acid were, respectively, the main sugar and organic acid found in the fruits and in its infusions. The microelement found in higher amounts in both samples was manganese, while potassium and calcium were the macroelements present in higher levels in the fruits and infusions, respectively. Both samples presented folates and tocopherols, being ytocopherol the main isoform detected in the fruits, while a-tocopherol was the only isoform quantified in the infusion. The hydromethanolic extract prepared from the fruits gave higher antioxidant and antibacterial activities, namely against Escherichia coli and Pseudomonas aeruginosa, than the infusion; it also showed capacity to inhibit the formation of bacterial biofilm. Both bioactivities are highly correlated with the presence of phenolic compounds, in which the major are ellagic acid derivatives (sanguiin hlO) followed by tlavan 3-ols ((+)catechin) and anthocyanin compounds (pelargonidin-3-glucoside). Although fruits of wild F. vesca are mainly consumed in fresh, this study also proves the potentiality of their infusions as a source of bioactive molecules and properties.

Flowers of Gomphrena globosa L. as an alternatve source of betacyanins: optimization of the extraction using response surface methodology

Betacyanins are betalain pigments that display a red-violet colour which have been reported to be three times stronger than the red-violet dye produced by anthocyanins [1]. The applications of betacyanins cover a wide range of matrices, mainly as additives or ingredients in the food industry, cosmetics, pharmaceuticals and livestock feed. Although, being less commonly used than anthocyanins and carotenoids, betacyanins are stable between pH 3 to 7 and suitable for colouring in low acid matrices. In addition, betacyanins have been reported to display interesting medicinal character as powerful antioxidant and chemopreventive compounds either in vitro or in vivo models [2]. Betacyanins are obtained mainly from the red beet of Beta vulgaris plant (between I 0 to 20 mg per I 00 g pulp) but alternative primary sources are needed [3]. In addition, independently of the source used, the effect of the variables that affect the extraction of betacyanins have not been properly described and quantified. Therefore, the aim of this study was to identifY and optimize the conditions that maximize betacyanins extraction using the tepals of Gomphrena globosa L. flowers as an alternative source. Assisted by the statistical technique of response surface methodology, an experimental design was developed for testing the significant explanatory variables of the extraction (time, temperature, solid-liquid ratio and ethanolwater ratio). The identification was performed using high-performance liquid chromatography coupled with a photodiode array detector and mass spectrometry with electron spray ionization (HPLC-PDAMS/ ESI) and the response was measured by the quantification of these compounds using HPLC-PDA. Afterwards, a response surface analysis was performed to evaluate the results. The major betacyanin compounds identified were gomphrenin 11 and Ill and isogomphrenin IJ and Ill. The highest total betacyanins content was obtained by using the following conditions: 45 min of extraction. time, 35•c, 35 g/L of solid-liquid ratio and 25% of ethanol. These values would not be found without optimizing the conditions of the betacyanins extraction, which moreover showed contrary trends to what it has been described in the scientific bibliography. More specifically, concerning the time and temperature variables, an increase of both values (from the common ones used in the bibliography) showed a considerable improvement on the betacyanins extraction yield without displaying any type of degradation patterns.