The phenolic characterisation of unknown or not traditionally-used plants are of great interest, due to the potential use as novel sources of bio-functional compounds. Anthocyanins are at the centre of the interest concerning the numerous bioactive effects, observed both in vitro and in vivo: inhibition of growth of human colon cancer cells (1); high antioxidant and anti-inflammatory activity (2,3); protection of neuronal cells from oxidative stress (4); regulation of adipocyte function, preventing thus the risks related to metabolic syndrome (5). Anthocyanins are important also because every days significantly quantity of them are ingested from vegetable sources (6). As described for other foods like dark chocolate, also the proanthocyanidin fraction is correlated to the total antioxidant capacity. P. mahaleb L. ('Mahaleb' or 'St. Lucy' cherry) is a woody species used as rootstock for sweet and sour cherry, only in well-drained soils, as the Apulia (Italy) soils. It produces highly pigmented small stone fruits, that have no importance for crop production, due to their astringent and sour taste. The astringent taste should be probably correlated with a high content on proanthocyanidins. These fruits had never been characterized for their chemical composition, except for the constituents of kernels (7), used finely ground for sweet breads and confectionery. A preliminary phenolic characterization of Prunus mahaleb fruits was done (8). Aim of this study was the chemical characterization of both anthocyanins and proantocyanidins, particularly concerning their qualitative characterization as well as the evaluation of the total antioxidant capacity.

Introduction: Polyphenols of wine have been extensively studied in relation to their health promoting properties. Red wine polyphenols consisted of different class of compounds belonging to flavonoid pathway, such as the resveratrol, quercetin, and anthocyanins which have long been considered to reduce the incidence of mortality and morbidity from cardiovascular diseases (CVD). Endothelial and monocyte activation is a pivotal event in atherosclerotic CVD, in this study was examined the anti-inflammatory effects of Italian red wines polyphenols in human vascular cells, such as endothelial and monocytoid cells. Since the wine polyhenols depend on different factors such as grape cultivar, and wine-making practices, aim of this study was to examine the anti-atherogenic effects of polyphenolic extracts from Italian red wines obtained by two Apulian grape cultivar: Primitivo and Negroamaro. Methods: Six Apulian red wines produced by the cv Negramaro and Primitivo grapes from the 2007-2008 vintage were analyzed. For each wine, triplicate lipophilic fraction analyses were carried out as per Giovinazzo et al., 2005. HPLC wine anthocyanin analysis was performed by direct injection of wine samples as reported by De Villiers et al. 2004. Human Umbilical Vein Endothelial Cells (HUVEC) and human monocytoid cells (U937) were pre-treated with chemically synthesised polyphenols, or with Primitivo and Negroamaro polyphenolic extracts and the corresponding de-alcoholised wines, at 0,2-2 % (v/v), before stimulation with 20 nmol/L phorbol myristate acetate (PMA) or 2 mg/mL lypopolysaccaride (LPS) for 24 h. Then, HUVEC were tested for the expression of Vascular Cell Adhesion Molecule(VCAM)-1 by enzyme immunoassays (EIA), and U937 supernatants were tested for the release of matrix metalloproteinases (MMP)-9 by gelatine zymography. Results: All chemically synthesized polyphenol tested reduced, in a concentration dependent manner, the stimulated expression of VCAM-1 in LPS triggered HUVEC. In the same conditions, lipophilic fraction extracted from both Primitivo and Negroamaro wines as well as the corresponding de-alcoholised wines exhibited an even higher inhibition of VCAM-1 expression (up to 50-70 fold higher than those expected on the base of individual polyphenol content). Similarly, either Primitivo and Negroamaro polyphenolic extracts reduced the PMA stimulated release of MMP-9 by PMA triggered U937.Conclusions: In this study different class of polyphenols from Primitivo and Negramaro red wines were characterised. These fractions, significantly inhibited the monocyte-derived extracellular matrix proteases release as well as the endothelial expression of athero-adhesion molecules. These inhibitory effects could explain, at least in part, the anti-inflammatory and cardio-protective properties of red wine polyphenols.

Prunus mahaleb L. is a rootstock used for sweet cherry crop in Apulia (Italy), an important cherry producing and exporting Italian Region. The hydro-alcoholic extract of P. mahaleb fruits has been analyzed for anthocyanin content and antimutagenic activity. HPLC/DAD/MS analysis indicated that 'Mahaleb' cherry fruit is extremely rich in anthocyanins (1751 mg/100 g D.W, more than 500 mg /100g F.W.), more than sweet and sour cherries. Moreover the protective effect of P. mahaleb was investigated by using D7 strain of yeast Saccharomyces cerevisiae. Results showed that treatments with P. mahaleb fruit extract decreased significantly the mutagenesis induced by hydrogen peroxide in S.cerevisiae.

Prunus mahaleb L. is a marginal fruit crop producing cherry-like dark purple drupes with avery bitter taste, rich in polyphenolic compounds.A mahaleb fruit concentrated extract (mfce)has been assayed for its biological activities. In this work, we report results on the in vitroeffects of mfce including: i) anti-proliferative, gap junction intercellular communication (GJIC)modulation and pro-apoptotic properties in a breast cancer cell line (MCF-7), ii) antiinflammatoryproperties in human vascular endothelial cells (HUVEC), iii) anti-mutageniceffect on yeast Saccharomyces cerevisiae strain D7. Mfce exerted a dose-dependent antiproliferativeeffect on MCF-7 cells together with a significant time-dependent increase inGJIC and a pro-apoptotic effect. Furthermore, mfce significantly reduced levels of endothelialinflammatory antigens in a concentration-dependent manner. Finally, mfce protectedS. cerevisiae cells against H2O2 cytotoxicity by direct radical scavenging activity. This studydemonstrated multiple healthful biological effects of mfce, highlighting it as a potentialnutraceutical product.

Plant cell cultures are valuable bioreactors for the production of bioactive compounds. Artemisia annua L. cell suspension cultures established for producing the antimalarial artemisinin (AN) were exploited to investigate the biosynthesis of other healthful phytochemicals. Culture supplementation with the putative elicitors ß-cyclodextrins (ß-CDs) was successful to increase intracellular and extracellular levels of AN. Other bioactive isoprenoid compounds, known for their antioxidant activity, were also investigated in the supplemented A. annua cell cultures by using metabolomic and transcriptomic analyses. In particular, the application of ß-CDs was useful to induce a considerable significant increase of lutein, quinones (Q9 and Q10) intracellular contents compared to untreated cell cultures. Moreover, the release of carotenoids and quinones into the culture medium of ß-CDs-treated cell cultures was significantly enhanced. Real Time PCR analysis revealed an up-regulation of 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) gene, suggesting that the ß-CDs induced increase of isoprenoid compounds could be due to the enhancement of the plastidial isoprenoid biosynthetic route. In addition, protoplasts derived from Artemisia suspension cultures were explored to engineer subcellular compartmentalization using a modified SNARE protein as a molecular tool for biotechnological metabolite accumulation.

Plant cell cultures are valuable tools for the production of bioactive compounds. Recently, many efforts have been made in order to develop and optimize strategies for increasing the yields of the desired plant metabolites by eliciting their biosynthesis or improving the efficiency of product recovery. We established Artemisia annua L. (Asteraceae) cell suspension cultures to investigate the biosynthesis of the antimalarial artemisinin (AN) [1] and other healthful phytochemicals such as carotenoids and quinones. The use of culture supplementation with cyclic oligomers ?-cyclodextrins was successful to significantly increase intracellular and extracellular levels of AN, lutein, Q9 and Q10 [2, 3]. In order to investigate the mechanism underlying these effects, other oligosaccharides having a linear structure were explored as putative elicitors in A. annua cell suspension cultures. For this purpose, oligogalacturonides (OGA) were obtained by hydrolysis of polygalacturonic acid with pectolyase from Aspergillus japonicus. Various OGA fractions were used for culture medium supplementation at different concentrations. The results showed that the 4-5 OGA fraction induced significant increases of AN and the intermediate dihydroartemisinic acid. These results suggest that the ability of ?-CDs to elicit the production of bioactive compounds in A. annua cell cultures could be due to their chemical similarity to pectic oligosaccharides often occurring in plants after fungal infection.

Numerose evidenze scientifiche incoraggiano il consumo di frutta e verdura per prevenire malattie cardiovascolari e tumori. Tuttavia il loro consumo rimane sotto il limite consigliato dall'Organizzazione Mondiale della Sanità. Per questo motivo i consumatori e le aziende alimentari sono alla ricerca di bevande salutistiche "ready-to-drink".I frutti di P. mahaleb, ricchi di composti salutistici (antocianine, flavonoli e cumarine) non sono commestibili a causa del loro sapore amaro, ma l'impiego biotecnologico della fermentazione controllata, utilizzata per migliorare le qualità organolettiche, salutistiche e di serbevolezza di frutta e verdura, potrebbe consentirne l'utilizzo come nuova fonte di probiotici e phytochemicals.

Betacyanin (betanin), total phenolics, vitamin C and antioxidant capacity (by Trolox-equivalent antioxidant capacity, TEAC and oxygen radical absorbance capacity, ORAC assays) were investigated in two differently coloured cactus pear (Opuntia ficus-indica (L.) Mill.) genotypes, one with purple fruit and the other with orange fruit, from the Salento area, in Apulia (South Italy). In order to quantitate betanin in cactus pear fruit extracts (which is difficult by HPLC because of the presence of two isomers, betanin and isobetanin, and the lack of commercial standard with high purity), betanin was purified from Amaranthus retroflexus inflorescence, characterized by the presence of a single isomer. The purple variety showed very high betanin content, with higher levels of phenolics, vitamin C, and antioxidant capacity (TEAC) than the orange variety. These findings confirm the potential for exploiting the autochthonous biodiversity of cactus pear fruits. In particular, the purple variety could be an interesting source of coloured bioactive compounds which not only have colouring potential, but are also an excellent source of dietary antioxidant components which may have beneficial effects on consumers' health.

Tomato (Solanum lycopersicum L.) is one of the most cultivated vegetable in the worldand it is a large source of bioactive compounds, including lycopene and othercarotenoids, flavanols (quercetin) and flavanones (naringenin). However, theconcentration of flavonoids in tomato is considered sub-optimal, particularly becauseanthocyanins are not generally present. Therefore this species has been the object of anintense metabolic engineering in order to obtain anthocyanin-enriched tomatoes byusing either breeding or transgenic strategies.Some tomato-related wild species, such as S. chilense, S. cheesmaniae, S.lycopersicoides and S. habrochaites, biosynthesize anthocyanins in the sub-epidermaltissue, and some allneles from those genotypes have been introgressed into cultivatedgenetics backgrounds, resultig in a 'purple' skin color. The breeding activity pursued atTuscia University (Viterbo, Italy), aimed at combining different alleles leading toanthocyanin production, have been lead to "Sun BlackTM , a trade mark-protectedtomato line with deep purple pigmentation, due to the biosynthesis of anthocyanins inthe peel. "Sun Black" is therefore a breeding product, not a GMO product. This aspect isconsidered an added-value for the market. In fact "Sun Black" tomato has been availablein July 2015 on the Italian market, thanks to an exclusive agreement between the ownerof the trademark and Unicoop Firenze.We report here the chemical characterization and structure elucidation of the uniqueanthocyanins found in the epiderm of "Sun Black" tomato, as well as other bioactivecompounds (carotenoids, polyphenols, vitamin C) of the whole fruit.The results show the unique features of this tomato genotipe as a 'novel food' withfunctional properties.Keyword: tomato, anthocyanins, functional food

Characterization of polyphenols and antioxidant activity (ORAC) in cherry cultivars of Apulia - Extracts of sweet cherries (Prunus avium L.) grown in a suitable area of cherry cultivation of Apulia (the south-east area around Bari) were characterized. High-performance liquid chromatographic methods were used to identify and quantify phenolics. A total of five phenolic compounds were characterised as hydroxycinnamic acid derivatives (neochlorogenic acid, p-coumaroylquinic acid and chlorogenic acid) and anthocyanins (cyanidin 3-glucoside, cyanidin 3-rutinoside). The total anthocyanin content ranged from 0.17 to 8.43 mg keracyanin equivalents/g dry weight. The oxygen radical absorbance capacity (ORAC) assay indicated that fruits of indigenous and rare genotypes possessed considerable antioxidant activity. The results of these studies can allow to identify genotypes with good antioxidant capacity, in order to enhance the cherry production in Apulia through breeding for improved nutraceutical characteristics.

Bioactive compounds, mainly polyphenols, present in berries, are thought to be responsible for the health benefits of these fruit. Therefore, it isworthwhile to define the optimal environmental conditions to maximise their polyphenol content.OBJECTIVE: With the aim to define the optimal conditions for berry cultivation in an innovative environment, red raspberry,wild strawberry and blackberry plants were grown in a traditional greenhouse in comparison with two photovoltaicgreenhouses with different shading area.METHODS: Hydroalcoholic extracts of ripe berries were evaluated by HPLC analysis, for their anthocyanins, organic acids and sugar contents. Moreover, phenolic content (by the Folin-Ciocalteu assay) and antioxidant activity (by the Trolox equivalent antioxidant capacity-TEAC assay) were assayed on the same berry extracts.RESULTS: Total anthocyanins, phenols content and antioxidant capacity tended to increase in berries grown under shading.The sugars content was, mostly, not negatively influenced by the shading. Conversely, the organic acids content, in some instances, increased along with the shading percentage.CONCLUSIONS: It can be concluded that it is possible to combine the greenhouse production of high-quality berries (with a particular focus on polyphenols, bioactive compounds valuable for human health) with the production of renewable energy, in the context of sustainable agriculture

Plant cell cultures as valuable tools for the production of specific metabolites can be greatly improved by the application of elicitors including cyclodextrins (CDs) for enhancing the yields of the desired plant compounds. Here the effects of 2,6-dimethyl-?-cyclodextrins (DIMEB) on the production of carotenoids and quinones from Artemisia annua L. cell suspension cultures were investigated. The addition of 50 mM DIMEB induced an early increase of intracellular carotenoid and quinone contents, which could be observed to a higher extent for lutein (10-fold), Q9 (3-fold) and Q10 (2.5-fold). Real Time PCR analysis revealed that the expression of 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR) gene in DIMEB treated cell cultures after three days was 2.5-fold higher than in untreated samples, thus suggesting that the DIMEB induced increase of carotenoids and quinones could be due to the induction of the plastidial isoprenoid biosynthetic route. In addition, the DIMEB treatment induced an enhanced release of carotenoids and quinones into the culture medium of A. annua cell suspension cultures possibly due to the ability of CDs to form inclusion complexes with hydrophobic molecules.

I coloranti alimentari sono additivi che conferiscono una colorazione aggiuntiva adun prodotto alimentare, al fine di renderlo più 'appetibile' alla vista. Si tratta quindidi caratteristiche 'estetiche', che possono però determinare un detrimento dellecaratteristiche di salubrità e sicurezza alimentare, soprattutto nel caso di colorantiartificiali ottenuti per sintesi chimica. A questo proposito, negli ultimi anni si è assistitoad un progressivo interesse, da parte del mondo scientifico e dell'opinione pubblica,nei confronti dei coloranti artificiali e nei tentativi di sostituzione di questi ultimi concoloranti di origine naturale. In questa nota è riportato il percorso, anche storico, di questaprogressiva sostituzione, inquadrando anche il ruolo dell'EFSA (l'organismo europeodi sicurezza alimentare) in questo percorso. Particolare enfasi è stata data ai colorantirossi (artificiali e/o naturali) per l'interesse, sia scientifico che mediatico, associato a talicoloranti. Infine, a titolo esemplificativo, sono riportati i risultati di un'indagine localesvolta nel corso di un paio d'anni, in diversi punti vendita della GDO delle provincie diBrindisi e Lecce.

The human health benefits of fruits and vegetables are ascribed to theirphytochemical content, such as carotenoids and polyphenolics. In the search forantioxidative chemicals from native fruits of the Puglia region of Italy, Prunus cerasusL., an acidic cherry widely used for culinary purposes, and Prunus mahaleb L., a treespecies commonly used as rootstock in cherry crop, were studied. The P. mahalebfruits have a high content of organic acids, fructose and vitamin C, but are notconsumed fresh because of a bitter and sour taste. In this work we obtained the 1HNMR spectra of the two species and from the comparison of these spectra, we foundthat P. mahaleb fruits have an higher concentration of phenolic compounds, such asflavonoids, and organic acids, in comparison to P. cerasus fruits. The same resultswere obtained when we focused on anthocyanins. In this study we identified thesignals of anthocyanin protons in 1H NMR spectra of a mixture of compounds inaqueous extracts of both P. cerasus and P. mahaleb fruits but the latter species showeda higher concentration and a larger number of these compounds. This metabolomicanalysis gave us the data to scientifically revalue traditionally-used plants like P.mahaleb and to identify the potential as source of biofunctional compounds to be usedin food and/or pharmaceutical industry. Moreover, in this study, NMR spectroscopycoupled with multivariate data analysis was applied to Prunus metabolomics in orderto investigate the botanical origins of Prunus cerasus and to identify the compoundsresponsible for differentiation of these two species of Prunus (cerasus and mahaleb)and of two cultivars of Prunus cerasus ('Montmorency' and 'Marasca di Zara').

Artemisia annua L., is a herbal plant traditionally used for the treatment of fever. The glandular trichomes of this plant accumulate, although at low levels, artemisinin which is highly effective against malaria. Due to the great importance of this compound many efforts have been made to improve the knowledge on artemisinin production both in plants and in cell cultures. In this study A. annua suspension cultures were established in order to investigate the effects of methyl jasmonate (MeJA) and miconazole treatments on artemisinin biosynthesis. Twenty-two mM MeJA induced a three-fold increase of artemisinin production soon after 30 min. Two hundred mM miconazole induced a 2.5 fold increase of artemisinin production after 24 h treatment but had severe effects on cell viability. The influence of these treatments on the expression of biosynthetic genes was also investigated. MeJA induced an up-regulation of CYP71AV1 while miconazole induced an up-regulation of CPR and DBR2.

Given the significant increase in consumer interest in organic food products, there is a need to determine to what extent there is a scientific basis for claims made for organic products. Apart from the sensory quality and food safety, there is a need to assess the nutritional value of organic vs. conventional (or integrated) food. Measurement of antioxidant activity using biologically relevant assays isimportant in the screening of fruits for potential health benefits. ORAC assay has been used in the assessment of antioxidant activity of strawberry extracts, from organically and Integrated Pest Management (IPM) strawberry plants, grown in the highly suited area of Basilicata (south Italy). Results on two genotypes ('Candonga' and 'Ventana') indicated that strawberries organically cultivated had statistically significant higher antioxidant activity than the IPM strawberries. Particularly, in the methanolic extract of organic 'Candonga', at red stage of ripening, the ORAC value was 31.5% higher than the IPM one. Further investigations on bioactive compound composition (namely anthocyanins and other flavonoids) are needed to identify the specific contribution to antioxidant capacity of organic strawberries.

A novel strategy to induce parthenocarpy in tomato fruits by the induction of resveratrol biosynthesis inflower tissues was exploited. Two transgenic tomato lines were considered: a higher resveratrolproducing(35SS) line, constitutively expressing a grape stilbene synthase cDNA, and a lower resveratrolproducing(LoxS) line, expressing stilbene synthase under a fruit-specific promoter. The expression of thestilbene synthase gene affected flavonoid metabolism in a different manner in the transgenic lines, and inone of these, the 35SS line, resulted in complete male sterility. Resveratrol was synthesised either in 35SSor LoxS tomato flowers, at an even higher extent (about 8e10 times) in the former line. We furtherinvestigated whether stilbene synthase expression may have resulted in impaired naringenin accumulationduring flower development. In the 35SS flowers, naringenin was significantly impaired by about50%, probably due to metabolic competition. Conversely, the amount of glycosylated flavonols increasedin transgenic flowers, thereby excluding the diminished production of flavonols as a reason forparthenocarpy in tomato. We further investigated whether resveratrol synthesis may have resultedchanges to pollen structure. Microscopic observations revealed the presence of few and abnormal flakelikepollen grains in 35SS flowers with no germination capability. Finally, the analysis of coumaric andferulic acids, the precursors of lignin and sporopollenin biosynthesis, revealed significant depletion ofthese compounds, therefore suggesting an impairment in structural compounds as a reason for pollenablation. These overall outcomes, to the best of our knowledge, reveal for the first time the major roledisplayed by resveratrol synthesis on parthenocarpy in tomato fruits.

Eugenia myrtifolia Sims. is an evergreen shrub, native to temperate and tropical rainforests of Australia, which is becoming an important containerized ornamental plant in the U.S. and Mediterranean nursery industry. To satisfy the growing market demands for this new ornamental plant, development of an accelerated propagation method is required.The goal of this study was to investigate the in vitro regeneration potential of E. myrtifolia Sims. seeds at different stages of development towards establishment of in vitro multiplication system. Maximum regeneration of adventitious shoots was achieved from immature seeds cultured in the dark on half-strength Murashige and Skoog (MS) macronutrients and full-strength MS micronutrients and vitamins (MS/2) medium supplemented with 2.5 ?M thidiazuron (TDZ). Induction of regeneration occurred after at least two successive subcultures on TDZ-enriched medium, followed by subcultures on Expression Medium (EM; hormone free MS/2) or Multiplication Medium [MM; MS medium enriched with 4.4 ?M 6-benzyladenine (BA) and 0.05 ?M ?-naphthaleneacetic acid (NAA)], where a complete development of shoots occurred. The regenerated shoots were excised and transferred again onto MM for micropropagation, where a proliferation rate of 1: 4 was achieved, and finally the shoots were transferred to a hormone-free MS medium for rooting. Following ex vitro transplanting, acclimatization over a period of 15 days was sufficient to establish greenhouse plants. The regenerated plants grown in the field for more then two years showed the same phenotype as that of mother plants. The adventitious regeneration and micropropagation carried out in this study can be used for a large-scale propagation and genetic engineering of E. myrtifolia Sims.

The identification of novel plant-based functional foods or nutraceutical ingredients that possess bioactive properties with antioxidant function has recently become important to the food, nutraceutical and cosmetic industries. This study evaluates the polyphenolic composition, identifies bioactive compounds and assays the total antioxidant capacity of Prunus mahaleb L. fruits collected from different populations and sampling years in the countryside around Bari (Apulia Region, Italy).RESULTS: We identified nine polyphenolic compounds including major anthocyanins, coumaric acid derivatives and flavonols from P. mahaleb fruits. The anthocyanin content (in some populations > 500 mg/100 g fresh weight; FW) in the fruit was comparable to that reported for so-called superfruits such as bilberries, chokeberries and blackcurrants. Coumaric acid derivatives comprised a large portion of the total polyphenolic content in the P. mahaleb fruits. Antioxidant activities, assessed using oxygen radical absorbance capacity (ORAC) and Trolox equivalent antioxidant capacity (TEAC) assays, measured up to 15 and 4.5 mmol/100 g FW, respectively. CONCLUSION: Our findings suggest that mahaleb fruit (currently not consumed fresh or used in other ways) could serve as a source of bioactive compounds and therefore find interest from the functional food and nutraceutical industries, as a natural food colorant and antioxidant ingredient in the formulation of functional foods.

There is an increasing consumer and retailer demand for new natural colorants. Anthocyanins are a class of molecules already used as water-soluble natural color- ant and for their health effects. Prunus mahaleb L. is a tree native to the Mediterranean area, producing highly pig- mented small-stone fruits currently not used for fresh con- sumption due to their astringent and sour taste. In the pre- sent study, we set up a protocol to obtain a "mahaleb fruit concentrated extract" (mfce) and characterized it chemi- cally by high-performance liquid chromatography/diode array detection. Anthocyanins, flavonols, flavanols, cou- marin, sugars, ascorbic acid and organic acids were identi- fied and quantified. After characterization, the antioxidant capacity of the mfce was assayed in vitro by three different methods: Trolox equivalent antioxidant capacity, Oxygen Radical Absorbance Capacity and Folin-Ciocalteu assays. The concentrated extract produced and characterized in this study showed a high content of anthocyanins, flavonols and coumarin and a strong antioxidant capacity. Therefore, mfce may be of interest as a natural food colorant and as an ingredient in the formulation of functional foods.

PRUNUS MAHALEB, L. FERMENTED FRUITS: A POTENTIAL SOURCE OF ANTIOXIDANT ACTIVITY AND PROBIOTIC MICRORGANISMSGerardi Carmela1, Tristezza Mariana1, Giordano Luca1, Maiorano Gabriele1, Blando Federica1, de Candia Silvia2, Grieco Francesco11 Institute of Sciences of Food Production, CNR, Territorial Unit of Lecce, Italy2 Institute of Sciences of Food Production, CNR, Territorial Unit of Bari, ItalyFermentation is a process traditionally adopted to preserve food which results in novel fermented products, provided with specific organoleptic properties. Prunus mahaleb L. is an autochthonous tree of the Mediterranean area. Its highly pigmented small-stone fruits are characterized for astringent and sour taste and therefore not used for fresh consumption. Recently, Prunus mahaleb L. fruits were found endowed with high bioactive compound content, especially anthocyanins, and strong antioxidant capacity [1]. With the aim to reduce their sourness, P. mahaleb fruits were dipped in water and fermented for 20 days at 25°C with four starter cultures of Lactobacillus plantarum strains alone or in mixture with a single strain of Saccharomyces cerevisiae. During this period, viable cell count of L. plantarum strains was never found lower than 107CFU/ml that is the lower concentration request to probiotic microorganisms to promote beneficial effects on human health. S. cerevisiae in all trials reached a maximum population three days after inoculation and then, cells counts slightly decreased in a range from 107 to 103 CFU/mL depending on the co-inoculated bacteria strain. Among L. plantarum strains, only the FG68 survived after an in vitro digestion protocol specifically set up for the selection of potentially probioitic lactic acid bacteria [2]. As concerns the antioxidant activity, the fermentation medium always showed higher values when L. plantarum strains were co-coltured with S. cerevisiae. Conversely, no differences were found in fermented fruits even though their antioxidant capacity values were higher than the large part of fruits included in the human diet.References1.Gerardi C, Tommasi N, Albano C, Blando F, Rescio L, Pinthus E, Mita G (2015) Prunus mahaleb L., fruit extracts: a novel source from natural food pigments. European Food Research & Technology 241: 683-695.2.Baruzzi F, Poltronieri P, Quero GM, Morea M. (2011) An in vitro protocol for direct isolation of potential probiotic lactobacilli from raw bovine milk and traditional fermented milks. Applied Microbiology and Biotechnology 90: 331-342.

Using four different chromatographic steps, b-galactosidase was purified from the ripe fruit of sweetcherry to apparent electrophoretic homogeneity with approximately 131-fold purification. The Prunusavium b-galactosidase showed an apparent molecular mass of about 100 kDa and consisted of fourdifferent active polypeptides with pIs of about 7.9, 7.4, 6.9 and 6.4 as estimated by native IEF and bgalactosidase-activity staining. The active polypeptides were individually excised from the gel andsubjected to SDS-PAGE. Each of the four native enzymes showing b-galactosidase activity was composedof two polypeptides with an estimated mass of 54 and 33 kDa. Both of these polypeptides were subjectedto N-terminal amino acid sequence analysis. The 54 kDa polypeptide of sweet cherry b-galactosidaseshowed a 43% identity with the 44 kDa subunit of persimmon and apple b-galactosidases and the 48 kDasubunit of carambola galactosidase I. The sweet cherry b-galactosidase exhibited a strict specificitytowards p-nitrophenyl b-D-galactopyranoside, a pH optimum of 4.0 and Km and Vmax values of 0.42 mMand 4.12 mmol min
1 mg
1 of protein respectively with this substrate. The enzyme was also activetowards complex glycans. Taken together the results of this study prompted a role for this class ofenzymes on sweet cherry fruit ripening and softening.

Anthocyanins, the naturally occurring pigments responsible for most red to blue colours of flowers, fruits and vegetables, have also attracted interest because of their potential health effects. With the aim of contributing to major insights into their structure-activity relationship (SAR), we have evaluated the radical scavenging and biological activities of selected purified anthocyanin samples (PASs) from various anthocyanin-rich plant materials: two fruits (mahaleb cherry and blackcurrant) and two vegetables (black carrot and "Sun Black" tomato), differing in anthocyanin content (ranging from 4.9 to 38.5 mg/g DW) and molecular structure of the predominant anthocyanins. PASs from the abovementioned plant materials have been evaluated for their antioxidant capacity using Trolox Equivalent Antioxidant Capacity (TEAC) and Oxygen Radical Absorbance Capacity (ORAC) assays. In human endothelial cells, we analysed the anti-inflammatory activity of different PASs by measuring their effects on the expression of endothelial adhesion molecules VCAM-1 and ICAM-1. We demonstrated that all the different PASs showed biological activity. They exhibited antioxidant capacity of different magnitude, higher for samples containing non-acylated anthocyanins (typical for fruits) compared to samples containing more complex anthocyanins acylated with cinnamic acid derivatives (typical for vegetables), even though this order was slightly reversed when ORAC assay values were expressed on a molar basis. Concordantly, PASs containing non-acylated anthocyanins reduced the expression of endothelial inflammatory antigens more than samples with aromatic acylated anthocyanins, suggesting the potential beneficial effect of structurally diverse anthocyanins in cardiovascular protection.

Solar photovoltaic greenhouses have become more popular, especially in the countries of southern Europe, due to specific government remuneration policies. However, many agronomic questions need to be addressed. This research was carried out in three types of commercial greenhouses covered with different materials (polycarbonate modules [PCM], traditional [TPM] and innovative semi-transparent [IPM] photovoltaic modules) with the aim to verify the compatibility of solar energy production with the production of high-quality wild rocket (Diplotaxis tenuifolia L.). IPM may satisfy the entire electricity demand of a commercial greenhouse. Yield for rocket grown in TPM was lower than for IPM and PCM. Antioxidant properties and dry weight decreased as a consequence of decreasing cumulative photosynthetic photon flux density. Nitrate content in TPM was higher (about 10,000 mg o kg-1 FW) than the maximum limits allowed by EC Regulation No. 1258/2011, whereas it was lower in IPM and PCM (1,805 and 668 mg o kg-1 FW, respectively).The results suggest that it is possible to combine solar energy production with high-quality wild rocket production, using innovative semi-transparent PV modules.

This study investigates the effects of tomato puree fortification (ftp) with several anthocyanin-rich food colorants on bioactive compound content (phenolics, isoprenoids), antioxidant capacity, in vitro biological activities and consumer acceptance. Tomato puree (tp) was added with different anthocyanin extracts from Daucus carota L. var. atrorubens (Anthocarrot), Vitis vinifera L. fruit skins (Enocolor), Sambucus nigra L. (Elderberry) or Prunus mahaleb L. (Mahaleb) fruits. The consumer acceptance (colour, flavor, taste, visual appearance) was at high level, except for Mahaleb-added extracts. Compared to the control (tp), the addition of colouring extracts increased significantly the total phenolic content, before pasteurization, in addition to the expected anthocyanin content. However, after pasteurization, only Anthocarrot-ftp preserved increased phenolic (+53%) content, as well as higher antioxidant capacity (50%). Consistently, against tp, Anthocarrot-ftp exhibited an increased anti-inflammatory capacity as showed by the reduced expression of vascular cell adhesion molecule (VCAM)-1 in human cultured endothelial cells, under inflammatory conditions.

The lecture will be on subject-specific topics and will try to offer excellent possibilities for exchange of experiences and source for discussion. In my presentation the importance of the anthocyanin content and other polyphenolic compounds for premium (nutraceutical) value cherries will be focused, in order to give growers the awarness of the fruit quality in the chain production.

IntroduzionePrunus mahaleb L. è un arbusto deciduo che cresce in boschi radi anche su terreni marginali e in climi aridi. L'albero è nativo dei paesi del mediterraneo e dell'Asia centrale e produce dei piccoli frutti simili a ciliegie di un colore quasi nero, non commestibili a causa del loro sapore amaro. Studi precedenti hanno dimostrato un elevato contenuto in antocianine in questi frutti, che suggerisce il loro utilizzo come fonte di coloranti alimentari dotati di proprietà salutistiche.ObiettiviCaratterizzazione chimica di un estratto concentrato di frutti di P. mahaleb e studio in vitro dei suoi effetti anticancerogeni, antiinfiammatori e antimutagenici.MetodologiaI frutti di mahaleb sono stati estratti con solventi consentiti per uso alimentare (etanolo + 1% acido citrico 1M) e l'estratto, concentrato sino a 60 °Brix, analizzato utilizzando metodiche HPLC. Sono stati effettuati saggi in vitro per studiare la capacità dell'estratto di inibire la proliferazione di linee cellulari tumorali umane (MCF-7), le sue proprietà antinfiammatorie su cellule endoteliali vascolari umane e infine i suoi effetti antimutagenici su linee di lievito Saccharomyces cerevisiaeD7.Risultati e DiscussioneL'estratto concentrato ha mostrato un elevato contenuto in antocianine, flavonoli e cumarina ed una elevata attività antiossidante simile o superiore a quella di altri noti estratti concentrati di frutti rossi. Lo studio delle attività biologiche dell'estratto ha mostrato: i) un significativo effetto antiproliferativo e capacità di modulare le GJIC in cellule MCF-7; ii) una inibizione dell'espressione di markers infiammatori tipici dell'attivazione endoteliale (ICAM-1, VCAM-1, E-selectine) in cellule endoteliali ottenute da vena ombelicale umana in coltura; iii) una riduzione di circa tre volte dell'effetto mutageno di H2O2 su cellule di S. cerevisiae. Questi risultati suggeriscono l'utilizzo dei frutti di P. mahaleb processati, come fonte di coloranti alimentari naturali e di composti bioattivi con proprietà salutistiche.

Valorization of the neglected Prunus mahaleb L., fruits as a source of functional moleculesGerardi C.1, Frassinetti S.3, Leone A.1, Calabriso N.2, Carluccio M.A.2, F. Blando1 and Mita G. 11 Institute of Sciences of Food Production, CNR, Lecce, Italy2 Institute of Clinical Physiology, CNR, Lecce, Italy3 Institute of Biology and Agricultural Biotechnology, CNR, Pisa, ItalyAbstract: Prunus mahaleb L. is a tree producing dark-red small stone fruits, not used for fresh consumption due to their astringent and sour taste. In this communication will be described a "mahaleb fruit concentrated extract" (mfce) and its chemical characterization. The extract showed high anthocyanin, flavonol and coumarin content. Moreover, the results of the in vitro assays for mfce biological activities will be reported. Mfce showed strong antioxidant capacity and anti-proliferative, pro-apoptotic, anti-inflammatory and anti-mutagenic effects. This study demonstrated several healthful effects of mfce that may be of interest as natural food colorant and as source of molecules for the formulation of functional foods.