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 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.

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.

Artemisinin is a sesquiterpene antimalarialcompound produced, though at low levels (0.1-1% dryweight), in Artemisia annua in which it accumulates in theglandular trichomes of the plant. Due to its antimalarialproperties and short supply, efforts are being made toimprove our understanding of artemisinin biosynthesis andits production. Native ?-cyclodextrins, as well as thechemically modified heptakis(2,6-di-O-methyl)-?-cyclodextrin(DIMEB) and 2-hydroxypropyl-?-cyclodextrins,were added to the culture medium of A. annua suspensioncultures, and their effects on artemisinin production wereanalysed. The effects of a joint cyclodextrin and methyljasmonate treatment were also investigated. Fifty millimolarDIMEB, as well as a combination of 50 mMDIMEB and 100 ?M methyl jasmonate, was highlyeffective in increasing the artemisinin levels in the culturemedium. The observed artemisinin level (27 ?mol g-1 dryweight) was about 300-fold higher than that observed inuntreated suspensions. The influence of ?-cyclodextrinsand methyl jasmonate on the expression

The increased consumption of whole wheat grains and whole wheat products has been associated with reduced risk of developing chronic diseases, such as cardiovascular disease, type 2 diabetes and colon cancer. These beneficial effects have been ascribed to the presence in whole wheat kernels of bioactive compounds which may vary for total content and composition among different wheat species and wheat varieties. In this work we present the profile of hydrophilic and lipophilic bioactive compounds of whole wheat semolina from five durum wheat elite cultivars. Whole semolina samples were analyzed to evaluate the total content and composition of phenolic acids (hydrophilic extract) and the total content and composition of carotenoids, tocopherols and tocotrienols (lipophilic extract). The total phenolic acid content was variable among the cultivars and ranged from 488 ?g/g to 1490 ?g/g whole flour. Among the detected compounds, ferulic acid was the most abundant, followed by sinapic acid and p-coumaric acid. Total carotenoid content varied from 2.64 µg/g whole flour and 4.75 µg/g whole flour and were mostly represented by lutein and zeaxanthin, while ?- e ?-carotene were present in trace amounts. Three different homologues of tocotrienols were detected (?, ? and ?), varying in a range between 18.3 and 28.6 µg/g whole flour, while tocopherols were detected in trace amounts. Duilio and Svevo cultivars, exhibited the highest content of hydrophilic and lipophilic bioactive compounds, respectively, and were selected to test the anti-inflammatory activity of extracts on human intestinal HT-29 cells. Preliminary experiments were carried out in order to assess the highest not cytotoxic concentration of lipophilic and hydrophilic extracts by using MTT test. Both extracts will be used to assess the anti-inflammatory activity on HT-29 cells stimulated by LPS mitogen, by quantification of IL-8. This research shows that whole wheat semolina flours of these five cultivars varied significantly in their contents of bioactive compounds and differences in their anti-inflammatory potential might suggest the possibility that durum wheat varieties could be selected based on potential health benefits.

Higher plants synthesize an amazing diversity of phenolic secondary metabolites.Phenolics are defined secondary metabolites or natural products because, originally, they wereconsidered not essential for plant growth and development. Plant phenolics, like other naturalcompounds, provide the plant with specific adaptations to changing environmental conditions and,therefore, they are essential for plant defense mechanisms. Plant defensive traits are costly for plantsdue to the energy drain from growth toward defensive metabolite production. Being limited withenvironmental resources, plants have to decide how allocate these resources to various competingfunctions. This decision brings about trade-offs, i.e., promoting some functions by neglecting othersas an inverse relationship. Many studies have been carried out in order to link an evaluation of plantperformance (in terms of growth rate) with levels of defense-related metabolites. Available resultssuggest that environmental stresses and stress-induced phenolics could be linked by a transductionpathway that involves: (i) the proline redox cycle; (ii) the stimulated oxidative pentose phosphatepathway; and, in turn, (iii) the reduced growth of plant tissues.

In plants, an increased production of toxic oxygen species is commonly observed under low oxygen stress, but cellular responses still have to be fully investigated. Plant cell cultures can be a valuable tool to study plant metabolic responses to various environmental stresses including low oxygen condition. Arabidopsis suspension cultures growing in shake flasks were subjected to hypoxia by stopping shaking for different intervals, showing an increase of the antioxidant metabolite a-tocopherol. In order to obtain a more controlled condition, cultivation of Arabidopsis suspension cultures was established in a 5-L stirred bioreactor. A constant aeration of 20 % dissolved oxygen was found to be the most suitable for cell growth. A 4-hour anoxic shock was induced by suspending the aeration and flushing into the vessel with nitrogen. During the anoxic stress, tocopherol levels resulted increased at the end of the treatment, indicating that the complete oxygen deprivation indeed induced a defence response involving antioxidant metabolism. The presence of an oxidative stress as a consequence of anoxic condition was also confirmed by the increased levels of H2O2. Overall, these results indicate that Arabidopsis suspension cultures grown in a stirred bioreactor can be a useful in vitro system for investigating low oxygen stress.

This review reports the use of wheat milling by-products for the extraction of high quality oil and vitamin E including our results on the exploitation of durum wheat bran as a valuable source of important healthful compounds. Wheat oil can be used as an ingredient in food, pharmaceutical or cosmetic preparations because it contains important bioactive compounds such as vitamin E, carotenoids and unsaturated fatty acids. Different methods are used for oil recovery from plant materials, such as solvent extraction, mechanical pressing or the ecofriendly supercritical carbon dioxide (SC-CO2) extraction technology. By using SC-CO2, we obtained an oil from durum wheat (Triticum durum Desf.) bran and optimized the extraction conditions to increase oil and vitamin E yields. Wheat bran, which is composed of pericarp, aleurone layer and germ, is discarded during the early stages of durum wheat milling processes to obtain a final product (semolina) that is stable over time. Maximum oil and vitamin E yields were obtained when a durum wheat bran matrix with particle size of *30 mesh and a moisture content of 2.6 % was used. The optimal conditions for oil extraction were: 300-350 bar, 60-70 °C, and 4 l min-1 gaseous CO2 flow rate for 1 h. The chemical composition (vitamin E forms, carotenoids, quinones, lipids and fatty acids) of the SC-CO2 extracted oil was analyzed and compared to that of the oil extracted by Soxhlet using hexane as solvent. The findings here reported highlight the importance of durum wheat bran as a rich source of valuable natural nutrients.

Artemisinin is an effective antimalarial compound obtained by thè aerial parts of thè herbalplant Artemisia annua L. Unfortunately, thè production of artemisinin in plants is very low (0.1-1% on a dry weight basis) and its chemical synthesis is very difficult. In recent yearsbiotechnological attempts to improve artemisinin production either in plant or geneticallyengineered microrganisms were made. A. annua celi and tissue cultures were also exploredfor thè production of artemisinin, although thè yieids obtained have been so far quite low. Wehave recently established A. annua celi cultures able to biosynthesize artemisinin and torespond to thè elicitation by methyl jasmonate (MeJA). Interestingly, amounts of artemisininproduced by these cultures were also found in thè spent culture medium. Cyclodextrins (CDs)are known to increase thè water solubility of various compounds, by forming host-guestinclusion complexes. Chemically modified CDs, such as 2,6 dimethyl-p-cyclodextrins (DIMEB)have been successfully used to improve thè production of health-promoting plant metabolitessuch as resveratrol in grape celi cultures. The aim of this work was to evaluate thè ability ofDIMEB to enhance artemisinin production in A. annua suspension celi cultures. HPLCanalysis of artemisinin and Real Time PCR gene expression analysis were carried out inDIMEB treated and untreated cultures. DIMEB induced a 300-fold increase of artemisininlevels in thè spent culture medium after a three-day-treatment. The addition of MeJA induceda further increase of thè artemisinin amounts. The expression levels of artemisininbiosynthetic genes (cytochrome P450 monoxygenase, cytochrome P450 reductase andartemisinic aldehyde A11 reductase) were not altered by thè DIMEB treatment. These resultssuggest that DIMEB improved artemisinin yieids of A. annua celi cultures, probably notaffecting thè biosynthetic pathway but enhancing thè solubility and stability of artemisininmolecules.

The aim of this study was to investigate the influence of encapsulation on the storage stability of oil extracted by supercritical carbon dioxide from a micronized durum wheat bran fraction. Wheat bran oil was encapsulated in 2% (w/v) sodium alginate beads. Encapsulated and unencapsulated oil samples were stored at 4°C or 25°C, daylight or darkness, over 90 days and, at defined time points, subjected to stability evaluation based on fatty acid hydroperoxide production, and tocopherol (?, ? and ? forms), tocotrienol (?, ? and ? forms) and carotenoid (lutein, zeaxanthin and ?-carotene) degradation. The encapsulation of the oil into alginate beads significantly increased stability, optimally when stored at 4°C, maintaining high levels of isoprenoids and low content of fatty acid hydroperoxides over 30 days of storage.

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.

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.

Crocus sativus L., mostly famous as saffron, has gained more attention due to its crocin (crocetin ester) pigment responsible for its extensive pharmaceutical properties. In this study, we established two different callus cultures from corm and style explants of saffron to find out the best explant as a suitable source for crocin production. Comparative analyses of total phenolic, flavonoid, carotenoid and anthocyanin contents were also performed in the two callus cultures. For callus induction, different combinations of MS medium with name thidiazuron (TDZ), benzylaminopurine (BA), 1-naphthaleneacetic acid (NAA), and 2,4-dichlorophenoxyacetic acid (2,4-D) alone or in combination were tested. Of the used media, all the combinations containing TDZ and NAA gave 100% callus induction. HPLC-DAD and HPLC-ESI-MS analysis were used for identification of crocin esters in established callus cultures. The highest amount of 0.35 mg g(-1) DW crocin was detected in style originated calli grown on the medium containing 3 mg L-1 NAA +1 mg L-1 TDZ while the corm calli showed the most abundant total carotenoid (0.73 mg g(-1) DW), phenolic (15.04 mg gallic acid equivalent g(-1) DW) and flavonoid (0.76 mg rutin equivalent g(-1) DW) contents. In general, style-derived calli showed longer time survival with a fine texture and good quality compared to corm-derived calli.

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.

Safer and more environmentally friendly methods, including the use of natural substances, would be favorably considered for weed management, particularly in those habitats where the use of chemicals is restricted or banned. Ascochyta caulina, a proposed mycoherbicide for biocontrol of the weed Chenopodiumalbum, produces in liquid culture three main metabolites proposed as possible natural herbicides. Recently a research project,namedECO-VIA, initiated a series of studies aimed at developing the technologies to obtain a natural herbicide based on these bioactive metabolites. Particular attention was given to: maximizing toxin production; lowering the production costs; scaling up the production in fermentation systems; setting up a large scale purification method and identifying fast and inexpensive chemical methods to quantify toxins yields. The fungus proved to grow well and to produce up to 230 mgof toxins l_1 culture when grown for 5-10 days in shaken conditions, provided the initial inoculum was at least 105-106 conidia ml_1 of culture. Toxin production was improved by the addition of yeast extract to the medium. Given suitable inoculum and growth conditions, a stirred fermenter could be suitable for mass production of toxin.

In this study, the quali-quantitative composition of hydrophilic (phenolic acids)and lipophilic (isoprenoids) extracts from whole-meal flour of five elite Italian durum wheatcultivars was determined. Significant differences in the content of bioactive compoundswere observed among the wheat extracts, in particular concerning the content of boundphenolic acids, lutein and ?-tocotrienols. The cultivars Duilio and Svevo showed the highestamount of phenolic acids and isoprenoids, respectively. Extracts were evaluated for theiranti-inflammatory activity on HT-29 human colon cells by measuring the levels ofinterleukin 8 (IL-8) and transforming growth factor ?1 (TGF-?1). Durum wheat extractssignificantly inhibited the secretion of the pro-inflammatory IL-8 mediator at 66 ?g/mLof phenolic acids and at 0.2 ?g/mL of isoprenoids. Conversely, the secretion of theanti-inflammatory mediator TGF-?1 was not modified by neither hydrophilic nor lipophilicextracts. These results provide further insight into the potential of durum wheat on humanhealth suggesting the significance of varieties with elevated contents of bioactive components.

This review is dedicated to the memory of Prof. Domenico Mariotti, who significantly contributed to establishing the Italian research community in Agricultural Genetics and carried out the first experiments of Agrobacterium-mediated plant genetic transformation and regeneration in Italy during the 1980s. Following his scientific interests as guiding principles, this review summarizes the recent advances obtained in plant biotechnology and fundamental research aiming to: (i) Exploit in vitro plant cell and tissue cultures to induce genetic variability and to produce useful metabolites; (ii) gain new insights into the biochemical function of Agrobacterium rhizogenes rol genes and their application to metabolite production, fruit tree transformation, and reverse genetics; (iii) improve genetic transformation in legume species, most of them recalcitrant to regeneration; (iv) untangle the potential of KNOTTED1-like homeobox (KNOX) transcription factors in plant morphogenesis as key regulators of hormonal homeostasis; and (v) elucidate the molecular mechanisms of the transition from juvenility to the adult phase in Prunus tree species.

Asparagus represents a nutritious and refined food being very popular to consumers. The effects on the quality of spears cooked by conventional (boiling, steaming and microwaving) and different sous vide (using hot water or microwave) methods were analyzed. Physical, chemical and sensory traits were compared. Microwaving caused the largest weight change, the highest dry weight increase and the highest Total Color Difference in cooked samples compared to raw asparagus spears. Sous vide-microwaving showed an increase in greenness (the highest value of h°), better than the raw spears, and the lowest reduction in chlorophyll contents. After sous vide microwawing violaxanthin increased by about 42%, while after steaming neoaxanthin decreased by about 57%. Following sensory analysis, all cooked samples generally resulted acceptable (scores > 5 in a 1-9 hedonic scale), nevertheless sous vide-microwaved asparagus satisfied consumer acceptability more than the other cooked samples, especially when compared to steamed samples. Results indicate the sous vide-microwaving technique as optimal to preserve several traits, including the organoleptic ones, essential for the quality of cook-chilled asparagus spears. They also provide product-specific information usually required for cooking process strategies in the industrial area of ready-to-eat vegetables.

Plants can frequently experience low oxygen concentrations due to environmental factors such as flooding or waterlogging. It has been reported that both anoxia and the transition from anoxia to re-oxygenation determine a strong imbalance in the cellular redox state involving the production of reactive oxygen species (ROS) and nitric oxide (NO). Plant cell cultures can be a suitable system to study the response to oxygen deprivation stress since a close control of physicochemical parameters is available when using bioreactors. For this purpose, Arabidopsis cell suspension cultures grown in a stirred bioreactor were subjected to a severe anoxic stress and analyzed during anoxia and re-oxygenation for alteration in ROS and NO as well as in antioxidant enzymes and metabolites. The results obtained by confocal microscopy showed the dramatic increase of ROS, H2O2 and NO during the anoxic shock. All the ascorbate-glutathione related parameters were altered during anoxia but restored during re-oxygenation. Anoxia also induced a slight but significant increase of ?-tocopherol levels measured at the end of the treatment. Overall, the evaluation of cell defenses during anoxia and re-oxygenation in Arabidopsis cell cultures revealed that the immediate response involving the overproduction of reactive species activated the antioxidant machinery including ascorbate-glutathione system, ?-tocopherol and the ROS-scavenging enzymes ascorbate peroxidase, catalase and peroxidase making cells able to counteract the stress towards cell survival.

Sea fennel (Crithmum maritimum L.) is aperennial halophyte species typical of coastal ecosystems,used fresh in traditional cuisine and folkmedicine due to its sensory properties and a goodcontent of healthy compounds. Although consideredas a promising biosaline crop, this halophyte isunderutilized for commercial cultivation possiblydue to a shortage of its consumer demand. Forpromoting a full exploitation of this species, a newfood product was obtained by drying sea fennel usingdifferent treatments (air-drying, microwave-drying,microwave-assisted air-drying and freeze-drying).Water activity, essential oil content, chlorophylls,surface colour, colouring power and sensory evaluationwere analyzed. All drying treatments allow toobtain a good water activity but significantly reducedthe content of essential oils and chlorophylls. Freezedryingand microwaving preserved the surface colourparameters more than other drying treatments, whilefreeze-drying gave the product the best colouringpower. Based on sensory analysis, microwave-drying,microwave-assisted air-drying and freeze-dryingshowed the highest scores among the drying methods.Taken together the results indicate that microwavingand freeze-drying are optimal for preserving qualitativetraits, including organoleptic properties, in driedsea fennel for food use. Furthermore, dried sea fennelcan be usefully exploited in human food not only forits aromatic traits but also for its food colouring powerlike other plant derived natural colorants. It could beconcluded that this underutilized crop could play abetter role for making up a sustainable food productionsystem.

Stimulated production of secondary phenolic metabolites and proline was studied by using cell cultures of artichoke [Cynara cardunculus L. subsp. scolymus (L.) Hayek] submitted to nutritional stress. Artichoke cell cultures accumulated phenolic secondary metabolites in a pattern similar to that seen in artichoke leaves and heads (capitula). This paper shows that both callus and cell suspension cultures under nutritional stress accumulated phenolic compounds and proline, at the same time their biomass production was negatively affected by nutrient deficiency. The results obtained strongly suggest that plant tissues respond to nutrient deprivation by a defensive costly mechanism, which determines the establishment of a mechanism of trade-off between growth and adaptive response. Furthermore, the results of this research suggest that perception of abiotic stress and increased phenolic metabolites are linked by a sequence of biochemical processes that also involves the intracellular free proline and the oxidative pentose phosphate pathway. The main conclusion of this paper is that, once calli and cell suspension cultures respond to nutrient deficiency, in acclimated cells the establishment of a negative correlation between primary metabolism (growth) and secondary metabolism (defence compounds) is observed.

Plants are ideal bioreactors for the production of macromolecules but transport mechanisms are not fully understood and cannot be easily manipulated. Several attempts to overproduce recombinant proteins or secondary metabolites failed. Because of an independent regulation of the storage compartment, the product may be rapidly degraded or cause self-intoxication. The case of the anti-malarial compound artemisinin produced by Artemisia annua plants is emblematic. The accumulation of artemisinin naturally occurs in the apoplast of glandular trichomes probably involving autophagy and unconventional secretion thus its production by undifferentiated tissues such as cell suspension cultures can be challenging.Here we characterize the subcellular compartmentalization of several known fluorescent markers in protoplasts derived from Artemisia suspension cultures and explore the possibility to modify compartmentalization using a modified SNARE protein as molecular tool to be used in future biotechnological applications. We focused on the observation of the vacuolar organization in vivo and the truncated form of AtSYP51, 51H3, was used to induce a compartment generated by the contribution of membrane from endocytosis and from Endoplasmic Reticulum to Vacuole trafficking.The artificial compartment crossing exocytosis and endocytosis may trap artemisinin stabilizing it until extraction; indeed, it is able to increase total enzymatic activity of a vacuolar marker (RGUSChi), probably increasing its stability. Exploring the 51H3-induced compartment we gained new insights on the function of the SNARE SYP51, recently shown to be an interfering-SNARE, and new hints to engineer eukaryote endomembranes for future biotechnological applications.

Tocopherols, collectively known as vitamin E, are lipophilic antioxidants, essential dietary components for mammals and exclusively synthesised by photosynthetic organisms. Of the four forms (alfa, beta, delta and gamma), alfa-tocopherol is the major vitamin E form present in green plant tissues, and has the highest vitamin E activity. Synthetic alfa-tocopherol, being a racemic mixture of eight different stereoisomers, always results less effective than the natural form (R,R,R)- a-tocopherol. This raises interest in obtaining this molecule from natural sources, such as plant cell cultures. Plant cell and tissue cultures are able to produce and accumulate valuable metabolites which can be used as food additives, nutraceuticals and pharmaceuticals. Sunflower cell cultures, growing under heterotrophic conditions, were exploited to establish a suitable in vitro production system of natural a-tocopherol. Optimization of culture conditions, precursor feeding and elicitor application were used to improve the tocopherol yields of these cultures. Furthermore, these cell cultures were useful to investigate the relationship between a-tocopherol biosynthesis and photomixotrophic culture conditions, revealing the possibility to enhance tocopherol production by favouring sunflower cell photosynthetic properties. The modulation of a-tocopherol levels in plant cell cultures can provide useful hints for a regulatory impact on tocopherol metabolism.

Wheat bran is generally considered a byproduct of the flour milling industry, but it is a great source of fibers, minerals, and antioxidants that are important for human health. Phenolic acids are a specific class of wheat bran components that may act as antioxidants to prevent heart disease and to lower the incidence of colon cancer. Moreover, phenolic acids have anti-inflammatory properties that are potentially significant for the promotion of gastrointestinal health. Evidence on the beneficial effects of phenolic acids as well as of other wheat bran components is encouraging the use of wheat bran as an ingredient of functional foods. After an overview of the chemistry, function, and bioavailability of wheat phenolic acids, the discussion will focus on how technologies can allow the formulation of new, functional whole wheat products with enhanced health-promoting value and safety without renouncing the good-tasting standards that are required by consumers. Finally, this review summarizes the latest studies about the stability of phenolic acids in wheat foods fortified by the addition of wheat bran, pearled fractions, or wheat bran extracts.