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The modern two-phase technology of the Leopard series for olive oil industrial extraction developed by the Pieralisi Group (Pieralisi Maip S.p.A., Jesi, AN, Italy) generates large quantities of a novel by-product (olive paste - OP) constituted of the partially defatted wet drupe pulp without any traces of the kernel. OP is rich in lipophilic and hydrophilic molecules able to exert beneficial effects on human health, including triterpenic acids, phytosterols, tocopherols, carotenoids, polyphenols, minerals and fibers [1]. In the perspective of by-product valorization through a modern biorefinery approach, it is worth noting that dried OP contains more than 10% oil characterized by a well-balanced lipid profile, rich in mono and polyunsaturated fatty acids, and a very good oxidative stability, due to the high concentration of fat-soluble antioxidants. This makes OP oil particularly suitable as functional ingredient for food/feed industry, as well as for the formulation of nutraceutical, cosmeceutical and pharmaceutical products. Supercritical carbon dioxide (SC-CO2) is a green and environmentally friendly technology for the effective extraction of high-value natural molecules. It is gaining a foothold in industrial production of solvent-free vegetable oils and has been also applied to concentrate oil products of lipophilic micronutrients (e.g. vitamin E in soybean and olive oils) [2]. Nevertheless, the industrial application of SC-CO2 technology requires a careful optimization of the operative parameters to make the process efficient from both an economic and a productive point of view. In this work, a response surface methodology approach, based on the Box-Behnken Design, was used to determine the optimal parameters of pressure, temperature and time to simultaneously maximize oil production from the freeze-dried OP of cultivar Leccino and concentrate the most abundant lipophilic bioactives. The experimental data were fitted to a second-order polynomial equation by multiple regression analysis, and examined using appropriate statistical methods. The 3-D response surface plots derived from the mathematical models were applied to determine the optimal extraction parameters, which resulted: temperature 70 °C, pressure 35.5 MPa and time 62 min. Under these conditions, the oil experimental yield was 14.0±0.7%, in close agreement with the predicted value. Compared to the freeze-dried OP, SC-CO2 extracted oil was much concentrated in phytosterols (~13 fold), tocopherols (~6 fold) and squalene (~8 fold); total carotenoid concentration remained almost unchanged, while triterpenic acids, being polar, were substantially reduced (- 79.5%). However, their total content was much higher than in extra virgin olive oil. The biological effects produced by OP oil intake were examined on 3 groups of healthy BALB/c mice (n=5/group) orally administered (for 4 weeks) with 20 or 40 mg/die of OP oil. Untreated animals were used as control. OP oil supplementation had no significant effects neither on body weight, nor on the activity of most liver cytoprotective enzymes (glutathione-disulfide reductase, glutathione S-transferase and glucose-6-phosphate dehydrogenase) with the exception of NAD(P)H:Quinone Oxidoreductase 1, whose activity was significantly increased (P

Nell'ambito dei Programmi di Sviluppo Rurale, P.S.R. Puglia 2007/2013, destinati al miglioramento del Sistema Regionale della biodiversità (Reg. CE n. 1698/2005, PSR Puglia 2007/2013, Misura 214 - Azione 4 Sub-azione a; e successivo P.S.R. Puglia 2014-2020, Misura 10 pagamenti-agro-climatico- ambientali, Art. 28 Reg. UE n. 1305/2013 Sottomisura 10.2) è stato finanziato il progetto: "Recupero, caratterizzazione, salvaguardia e valorizzazione di leguminose e cereali da granella e foraggio in Puglia" (SaVeGraINPuglia). Questo volume include schede descrittive delle leguminose, cereali e foraggere reperite in Puglia acquisite in regione e caratterizzate con approccio multidisciplinare nell'ambito del proggetto SaVeGraINPuglia.

Valorization of the neglected Prunus mahaleb L., fruits as a source of functional molecules Gerardi C.1, Frassinetti S.3, Leone A.1, Calabriso N.2, Carluccio M.A.2, F. Blando1 and Mita G. 1 1 Institute of Sciences of Food Production, CNR, Lecce, Italy 2 Institute of Clinical Physiology, CNR, Lecce, Italy 3 Institute of Biology and Agricultural Biotechnology, CNR, Pisa, Italy Abstract: 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.

Table olives have been a component of the Mediterranean diet for centuries and their consumption is increasing worldwide. They contain several nutritional components such as unsaturated fatty acids, vitamin E, triterpenic acids, sterols, polyphenols and carotenoids [1]. The content of these bioactive molecules depends on the olive variety, the ripening stage of the olive fruit, the cultivation conditions and the processing method [2]. There are numerous commercial preparations of table olives and in a recent work we have developed and validated a new procedure for table olive production based on the use of selected autochthonous microbial starters and a productive process optimized to mimic the microbial evolution observed during spontaneous fermentations. [3]. In this work, we characterized some bioactive compounds and coloring pigments of fermented black table olives, belonging to two Italian (Cellina di Nardò and Leccino) and two Greek (Kalamàta and Conservolea) cultivars. Greek table olives resulted richer in carotenoids, vitamin E and triterpenic acids (maslinic and oleanoic acids) than Italian cv, contents ranged from 2.28 to 6.91 µg/g fw (edible portion), 53.64 to 59.15 µg/g fw and 2.19 to 3.28 mg/g respectively. Polyphenol compounds are known to be responsible for the bitter taste of the olive fruit and most of them are involved in table olives colour changes. In all tested black table olive cultivars, polyphenols content ranged from 0.87 mg/g fw (in Conservolea) to 23.49 mg/g fw (Cellina di Nardò). Moreover, we analysed the content of tyrosol, hydroxytyrosol, oleuropein aglycone, verbascoside, isoverbascoside, rutin, pyrocathecol, luteolin glucoside, quercetin, cyanidine rutinoside and cyniadine glucoside. In particular, Cellina di Nardò is characterized by the highest levels of anthocyanin pigments such as cyanidin 3-O-glucoside (14.77 mg/g fw) and cyanidin-3-O-rutinoside (3.17 mg/g fw) responsible for the characteristic black color. The results indicated that fermented black table olives are an excellent natural source of bioactive compounds and can represent a potential important functional food.

Lycopene is a carotenoid pigment found in a wide variety of vegetables and fruits. More than six hundred carotenoids have been identified in nature, but only twenty are present in human blood and tissues, including lycopene (the most abundant in plasma), ?-carotene, lutein and zeaxanthin [1]. Several exogenous pigmented antioxidants act on cancer progression and have immune-modulating effects by free radical scavenging activity and/or still unknown mechanisms. However, the controversy recently arisen for some of them requires a careful investigation in relation to their source and synergistic effects [2]. The purpose of this work was: i) to optimize the extraction of lycopene from ripe fruits of tomato, watermelon and GAC by supercritical CO2 in order to obtain natural, solvent-free lycopene containing oleoresins suitable as ingredients for the formulation of innovative functional foods and/or nutraceuticals; ii) to characterize each oleoresin with respect to carotenoids, tocochromanols, lipids, lipophilic and hydrophilic antioxidant activities, micro and macro elements; iii) the preparation of inclusion complexes (ICs) of the oleoresins in food grade ?-cyclodextrins (?-CDs) as hydrophilic nanocarriers [3]; iv) to assess, in-vitro, the effects of administration of each oleoresin/?-CD IC at different concentrations (0.5 to 10 µM) on cell apoptosis and on Interleukin-8 (IL-8) release by A549 human lung adenocarcinoma cells, for 24, 48 and 72 hours. The results showed that all oleoresins represent a healthy and high added-value source of lycopene characterized by: i) the absence of toxic impurities such as solvents, pesticides and heavy metals; ii) a high lycopene bioavailability, increased by the presence of co-extracted natural lipophilic components; iii) a high lipophilic antioxidant activity, improved by synergy with other biomolecules. When administered at a concentration of 10 µM for 72 hours, the three lycopene extracts showed conflicting behaviors: watermelon oleoresin increased cell apoptosis and decreased the release of IL-8, a pro-inflammatory cytokine involved in cancer progression; GAC and tomato oleoresins, instead, decreased cell apoptosis and increased IL-8 release. This discrepancy is likely related to a different lycopene/vitamin E ratio, with watermelon oleoresin showing the highest value among the extracts. Shorter incubation time and lower lycopene concentrations did not affect cell viability and IL-8 release. Further studies are required to assess the possible use of the lycopene containing oleoresin/?-CD ICs in health promoting functional foods and nutraceuticals.

Supercritical carbon dioxide (SC-CO2) represents an effective, non-toxic technology for extracting food-grade oleoresins from a range of edible plant materials [1]. The obtained oleoresins are a rich source of bioactive molecules, mainly carotenoids, tocochromanols and polyunsaturated fatty acids (PUFAs). Most bioactives are very reactive to light, oxygen and free radical species, this greatly limits their stability and reduce their shelf-life and health benefits [2,3]. Encapsulation into micro or nano particles is the most promising technique to increase their stability over time. Cyclodextrins (CDs) are biocompatible, non-toxic cyclic oligosaccharides obtained by enzymatic conversion of starch. They can form inclusion complexes with specific guest molecules, contributing to their stabilization against the adverse effects of physicochemical agents. CDs are approved in Europe as soluble dietary fiber and novel food ingredient. In this work is reported the encapsulation in CDs of pumpkin and tomato oleoresins, extracted by SC-CO2. The stability of tocochromanols, carotenoids and fatty acids in the oleoresin/CD complexes, compared to the corresponding free oleoresins, was monitored over time in different storage conditions (25°C, dark; 25°C light; 4°C, dark). Differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR) analyses, demonstrated that oleoresins/CDs have different physical characteristics from free oleoresins. Regardless of light, storage at 25°C of free oleoresins determined a rapid decrease in carotenoids, tocochromanols and PUFAs. CD encapsulation improved the stability of most bioactive compounds. Upon storage, the retention of tocochromanols, carotenoids and fatty acids was higher in oleoresin/CD complexes stored at 4°C than in the corresponding free oleoresins, indicating that encapsulation and low temperature have a synergistic effect in preventing the degradation against oxidation of bioactives. Unlike all other analyzed antioxidants, lycopene in tomato oleoresin/CD complex resulted to be more susceptible to oxidation than in free oleoresin, likely due to its selective sequestration from the interaction with other lipophilic molecules of the oleoresin. The results suggest that oleoresin/complexes, being generally more stable than free oleoresins, can be used as ready-to-mix high-quality ingredients in the preparation of innovative functional foods simultaneously enriched with carotenoids tocochromanols, PUFAs and fibers.

DNA-based approaches were used to characterize a strain (Salento-1) of Xylella fastidiosa obtained from an olive plant suffering from the syndrome of quick decline in Apulia (South Italy). Salento-1 was indistinguishable from strain CoDiRO previously isolated from olive in Apulia and assigned to X. fastidiosa subsp. pauca. Based on our results and comparative analysis with reported data, the subspecies pauca, multiplex, and fastidiosa may invade olive throughout the world (California, Italy, Argentina and Brazil). The strain Salento-1 has been deposited in the National Collection of Plant Pathogenic Bacteria (NCPPB), England, and in the Belgian Coordinated Collections of Microorganisms (BCCM), Belgium.

Phenolic acids are major components of cell walls in wheat and have important implications on human health as antioxidants with anti-tumor activity. Understanding the genetic basis of phenolic acids content and composition of wheat grain, and identifying associated markers provide the basis for improving wheat quality by molecular breeding. The objective of this study was: a) to identify phenolic acids genes in wheat by exploiting genomic resources and SNPs detected within the coding sequences of candidate genes; and b) to identify chromosomal regions associated to single phenolic acids and total soluble phenolic compounds by genome-wide association study (GWAS) and candidate gene approaches using a tetraploid wheat collection coupled with the 90K iSelect SNP genotyping array. A set of candidate genes involved in the biosynthesis of hydroxycinnamic acid derivatives (p-coumaric acid, ferulic acid and sinapic acid) and in the total soluble phenolics have been identified in wheat by comparative genomics. Single nucleotide polymorphisms (SNPs) found in the coding sequences of six candidate genes (PAL1, PAL2, C4H, C3H, COMT1 and COMT2) allowed to determine their chromosomal location and accurate map position on two reference consensus linkage maps. The genome-wide association study based on genotyping a tetraploid wheat collection with 81,587 gene-associated SNPs revealed several significant marker-trait associations, identifying 22 quantitative trait loci (QTLs) associated with individual phenolic acids and total soluble phenol compounds distributed on almost all durum wheat chromosomes. A QTL for p-coumaric acid was found to be coincident with the candidate gene phenylalanine ammonia-lyase (PAL2) on chromosome arm 2AL. The availability of perfect, candidate gene-based markers can allow elucidating the mechanism of phenol acids accumulation in the wheat kernels and exploiting the genetic variability of phenol acids content for the nutritional improvement of wheat end-products.

Antioxidant is a new hype in naturopathy and at present it has huge demand in cosmetic, nutritional supplementation, human health and pharmacological industry. It reduces cell damages caused by free-radical which are responsible for various ailments like cancer, diabetes mellitus, atherosclerosis, cataracts and inflammation. Different parts viz. leaves, fruits and bark of Ficus racemosa were successively extracted and studied of total phenol content in which methanol and water gave good quantity of phenol content. Antioxidant activity was determined and IC50 values were found 53 ug/ml and 59 ug/ml in methanolic extract of leaves and bark, respectively. Elemental analysis using ICPMS was carried out from leaves, fruits and barks of Ficus racemosa and results indicated that B, Fe and Zn were found in good quantity in leaves and bark. Water and methanolic extract of all the parts of plant were identified for their metabolites by Q-TOF LCMS. The bioactive constituents showed important health properties for human and animals like anti-cancer, antioxidant activity, anti-inflammatory and anti-diabetic. Therefore, present study suggested that Ficus racemosa leaves and bark have huge potential in terms of commercial use as source of antioxidants and bioactive constituents.

Introduzione Prunus 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. Obiettivi Caratterizzazione chimica di un estratto concentrato di frutti di P. mahaleb e studio in vitro dei suoi effetti anticancerogeni, antiinfiammatori e antimutagenici. Metodologia I 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 cerevisiae D7. Risultati e Discussione L'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.

In questo lavoro è stata condotta la caratterizzazione genetico-molecolare di circa 100 accessioni di olivo campionate in diverse località del Salento. L'obiettivo è stato la caratterizzazione, mediante marcatori molecolari (microsatelliti), di varietà autoctone del Salento a rischio di erosione genetica, e di altre varietà di olivo che, seppure di diversa origine geografica, risultano coltivate nel territorio salentino.

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.

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Whole wheat foods are significant source of compounds exhibiting health-promoting properties. One of the most abundant class of phytochemicals in the wheat grain is represented by phenolic acids that are typically localized in the bran and germ portions. The objective of this study was to estimate the extent of genetic variation for phenolic acids in durum wheat (T. turgidum L. ssp. durum). In addition, this study aimed at evaluating the anti-inflammatory activity of phenolic acids contained in whole-meal flour extracts. Phenolic acids were recovered from the whole meal flours of 65 durum cultivars and subsequently identified and quantified by HPLC-DAD analysis. Then, the anti-inflammatory activity of phenolic acids extracts was evaluated on LPS-stimulated HT-29 human colon cells by measuring the levels of interleukin 8 (IL-8) and transforming growth factor ?1 (TGF- ?1). A large variation for the content of phenolic acids was observed among genotypes and, on average, it accounted for 830 ?g/g dry weight. Whole meal flour extract significantly inhibited the secretion of the pro-inflammatory IL-8 mediator at 66 µg/mL of phenolic acids. Conversely, the secretion of the anti-inflammatory mediator TGF-?1 was not modified by addition of phenolic acids to HT-29 cells. Results showed that durum cultivars have different contents of phenolic acids, suggesting that a number of elite varieties could be used for breeding purposes. Moreover, results provide further insight into the health-related benefits of durum wheat foods as depending on the anti-inflammatory activity of phenolic acids.

Wheat is a staple cereal and an important source of minerals in human diets. Mineral content in the wheat grain has a genetic basis, but the trait is also influenced by environmental factors [1]. Many efforts have been devoted to increase the content of some elements such as Fe, Zn, Mn and Cu, which have a physiological function in the organism. In particular, mapping populations and genetic linkage maps have been suggested as a useful tool in breeding programs [2]. The present work was undertaken to evaluate by ICPOES the variation in macronutrients (Ca, Mg, K), micronutrients (Zn, Fe, Cu, Mn) and toxic trace elements (Cd, Pb) in the whole grains of 29 wheat accessions and cultivars, representative of old and modern wheat germplasm. All 29 lines were the parentals of existing mapping populations [3] and were grown in two different sites in Italy, in 2013/14 and in 2014/15 seasons. Multivariate Analysis (MVA) of data showed interesting differences in the mineral content for the considered sites, harvesting seasons and grain cultivars. [1] Gomez-Becerra HF, Yazici A, Ozturk L, Budak H, Peleg Z, Morgounov A, Fahima T, Saranga Y, Cakmak I. Genetic variation and environmental stability of grain mineral nutrient concentrations in Triticum dicoccoides under five environments. Euphytica 2010, 171:39-52. [2] Srinivasa J, Arun B, Mishra VK, Singh GP, Velu G, Babu R, Vasistha NK, Joshi AK. Zinc and iron concentration QTL mapped in a Triticum spelta × T. aestivum cross. Theor Appl Genet 2014, 127:1643-1651 [3] Pasqualone A, Piarulli L, Mangini G, Gadaleta A, Blanco A, Simeone R. Quality characteristics of parental lines of wheat mapping populations. Agric Food Sci 2015, 24: 118-127.

BACKGROUND: 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.

L'Artemisia annua L. è una pianta erbacea annuale nota per la produzione del sesquiterpene antimalarico artemisinina. La produzione di artemisinina da parte della pianta è molto bassa e quasi esclusivamente prodotta nelle foglie e nelle infiorescenze, ed accumulata in specifiche strutture (tricomi ghiandolari) (Duke and Paul, 1993; Duke et al., 2009). Pertanto lo sviluppo di biotecnologie per la produzione di artemisinina in laboratorio è di notevole interesse. L'utilizzo di colture in vitro rappresenta una fonte alternativa alla pianta coltivata in campo. Da qui la necessità di sviluppare un valido protocollo per la messa a punto di sistemi in vitro che svincolino la produzione di artemisinina, nella pianta circoscritta ad alcuni organi e limitata a determinati periodi dell'anno, garantendo in questo modo una produzione costante ed efficace nel tempo. Nel nostro laboratorio sono stati allestiti diversi tipi di colture in vitro di A. annua (calli, sospensioni cellulari e piantine micropropagate), ottimizzando le condizioni di crescita mediante l'utilizzo di terreni di coltura con la stessa composizione chimica in macro- e microelementi, ma con ormoni di diverso tipo e concentrazione.

In questo lavoro ci siamo occupati del recupero del germoplasma olivicolo salentino ancora presente in provincia di Lecce, ma a forte rischio di erosione genetica. In particolare, abbiamo studiato il profilo molecolare di varietà antiche, quali: Colozzese, Barone di Monteprofico, Ornella, Usciana, Cornola, Uliva grossa, e di diversi cloni delle cultivar autoctone più diffuse sul territorio, ovvero Cellina di Nardò e Ogliarola di Lecce. Tutte le varietà analizzate in questo studio sono conservate in un campo collezione allestito negli anni '90 in agro di Carpignano salentino.

During the last years the interest on the recovery, recycling and upgrading of residues from plant food processing, has increased drastically. Generally, food industries produce large volume of by products, both solid and liquid, which represent a problem for disposal and for environmental pollution. Nevertheless, they are also promising sources of bioactive compounds that can be recovered and used as valuable substances by developing new enriched food, such pasta and bread. Because bread and pasta are foods consumed every day, they can be considered as a carrier of functional substances that have a beneficial role on consumer health. Therefore, the aim of this study was to study the impact of agronomic by-product incorporation, such as olive cake flours, on the sensorial qualities of cereal-based food as pasta and bread. Commercial durum wheat semolina for pasta and commercial soft wheat flour for bread, were mixed with 10% w/w of two different olive cake flours. The bioactive compounds present in olive cake, pasta and bread were identified and characterized, and their stability, after the processing, was evaluated. Pasta and bread based on durum wheat semolina and wheat flour (100%), respectively were produced without olive cake flours and used as controls. Dry spaghetti and bread samples were submitted to a panel of 10 trained tasters in order to evaluate the sensorial attributes, since the incorporation of the olive cake flours, could determined a decline of sensory quality. Besides, the effect of olive cake flours particle size on the quality of food, was also studied, thus demonstrating that samples enriched with fine particle size showed the greatest overall quality score.

Laccases are biotechnologically interesting enzymes belonging to the polyphenol oxidases family. They are widely distributed throughout the phylogenetic scale from bacteria to mammals. In fungi the analysis of three-dimensional crystal structures of laccases indicate that ascomycete laccases are processed at their C-termini, at a conserved cleavage site, resulting in the proteolytic removal of C-terminal residues. We have isolated and cloned in expression vectors the cDNAs encoding two laccase isoforms (Ery3 and Ery4) from the basidiomycete Pleurotus eryngii. The Ery3 enzyme expressed in Saccharomyces cerevisiae, is functional, whereas the recombinant Ery4 protein does not show enzymatic activity. In order to explain this evidence, we investigated the relationship between the structure of the C-terminal extension and laccase enzymatic activity. The tasks of the present study were to determine the biological role of laccase C-terminal, and to validate a "molecular engineering" approach for the production of recombinant laccases with novel biochemical properties. Genetically engineered mutant genes were produced from Ery4 by: i) progressive 3'-terminal deletions, ii) point mutations, iii) Ery3/Ery4 chimeras. The mutant genes were expressed in S. cerevisiae and active recombinant laccase isoforms were produced, exhibiting each a different biological behaviour. The correlations between the structural information deriving from both biochemical and bioinformatic analyses shed light on the role of Cterminal region in determining laccase functions. The obtained data also indicated that our approach could represent an efficient method for laccase genetic engineering. To our knowledge, this study has produced the first evidences obtained by biotechnological approach of the involvement of the C-terminal tail in the inactivation/activation process of a basidiomycete laccase.

Plants respond to environmental stimuli, such as heat shock, by re-programming cellular activity through differential gene expression, mainly controlled at the transcription level. The current study refers to two sunflower small heat shock protein (sHSP) genes arranged in tandem in head-to-head orientation and linked by a 3809 bp region. These genes exhibit only slight structural differences in the coding portion. They code for cytosolic class I sHSPs and are named HaHSP17.6a and HaHSP17.6b according to the molecular weight of the putative proteins. The genomic organization of these genes is consistent with the idea that many HSP genes originate from duplication events; in this case, probably an inversion and duplication occurred. The HaHSP17.6a and HaHSP17.6b genes are characterized by different expression levels under various heat stress conditions; moreover, their expression is differently induced by various elicitors. The differential regulation observed for HaHSP17.6a and HaHSP17.6b genes differs from previous observations on duplicated sHSP genes in plants.

Artemisia annua L. (Asteraceae, Asteroideae, Anthemideae) è una pianta erbacea annuale originaria dell’Asia, molto conosciuta ed utilizzata in Cina con il nome tradizionale di “qinghao”. La pianta risulta ora naturalizzata in diverse parti del mondo, inclusa l’Italia. A. annua è uno degli esempi più noti dell’impiego in medicina moderna di una droga naturale tradizionale. Nella medicina popolare la pianta è stata infatti per secoli utilizzata come febbrifugo e nel trattamento della malaria. Indagini fitochimiche hanno, in tempi più recenti, portato all’isolamento del principio attivo, l’artemisinina (“qinghaosu”), un lattone sesquiterpenico prodotto nelle parti aeree della pianta, capostipite di una serie di importanti farmaci antimalarici. Fatto rive- lante e singolare è che la specie A. annua è praticamente l’unica del genere Artemisia a produrre tale principio attivo. Lo studio chimico-analitico ha portato all’isolamento di diversi composti caratteristici. Dalle parti aeree della pianta si estrag- gono tipiche sostanze aromatiche (artemisia chetone, 1,8-cineolo, canfora, ecc.). Dalle foglie e dalle sommità fiorite, oltre all’artemisinina, sono stati isolati e caratterizzati una serie di altri sesquiterpeni analoghi quali ad esempio l’artemisitene, l’arteannuina B e l’acido arteannuico, dotati tuttavia di una minore bioattività. La produzione di principio attivo in vivo, in piante spontanee, è stimata all’incirca fra lo 0.01 0.4%; la coltivazione di cloni selezionati sembra contribuire all’aumento della resa (1-2%). L’accumulo di artemisinina in vivo è stato studiato in dettaglio, portando tuttavia a risultati talvolta contrastanti. Alcuni Autori descrivono un massimo d’accumulo del principio attivo durante la fioritura, altri durante lo stadio di prefioritura. Le parti della pianta dove si è riscontrata la maggior produzione di artemisinina sono le foglie e le infiorescenze; basse quantità sono state invece trovate nei cauli. Non è stata evidenziata alcuna capacità di sintesi del principio attivo nella radice. Le parti aeree di A. annua sono dotate di tipiche strutture ghian- dolari, la cui presenza è stata correlata con la produzione non solo, come atteso, delle sostanze volatili aromatiche ma anche con la biosintesi dell’artemisinina, e dei composti analoghi, il cui accumulo segue parallelamente lo sviluppo fisiologico delle ghiandole. A livello mondiale, la malaria rappresenta una delle più impor- tanti e diffuse infezioni parassi- tarie e costituisce una delle sfide più ardue che i paesi poveri del mondo affrontano quotidiana- mente nella lotta per lo sviluppo ed il miglioramento delle loro condizioni socio-economiche e sanitarie. E’ una patologia che secondo le stime della Organizzazione Mondiale della Sanità (OMS) determina ogni anno la morte di circa un milione di persone, soprattutto bambini, ed è particolarmente endemica in Africa, nella zona a sud del Sahara. Benché tuttavia l’area geografica di diffusione dell’infezione si sia ridotta rispetto al secolo scorso, la possibilità di trasmissione della malaria sembra invece essere aumentata come conseguenza dell’incapacità di contenere il propagarsi della malattia attraverso il controllo dei vettori animali e di fornire terapie adeguate ad un numero significativo di pazienti, particolarmente a quelli che vivono nelle periferie del mondo.