Impatto di rifaximina sul microbiota intestinale: selezione di bifidobatteri antibiotico resistenti

The ideal approach for the long term treatment of intestinal disorders, such as inflammatory bowel disease (IBD), is represented by a safe and well tolerated therapy able to reduce mucosal inflammation and maintain homeostasis of the intestinal microbiota. A combined therapy with antimicrobial agents, to reduce antigenic load, and immunomodulators, to ameliorate the dysregulated responses, followed by probiotic supplementation has been proposed. Because of the complementary mechanisms of action of antibiotics and probiotics, a combined therapeutic approach would give advantages in terms of enlargement of the antimicrobial spectrum, due to the barrier effect of probiotic bacteria, and limitation of some side effects of traditional chemiotherapy (i.e. indiscriminate decrease of aggressive and protective intestinal bacteria, altered absorption of nutrient elements, allergic and inflammatory reactions). Rifaximin (4-deoxy-4’-methylpyrido[1’,2’-1,2]imidazo[5,4-c]rifamycin SV) is a product of synthesis experiments designed to modify the parent compound, rifamycin, in order to achieve low gastrointestinal absorption while retaining good antibacterial activity. Both experimental and clinical pharmacology clearly show that this compound is a non systemic antibiotic with a broad spectrum of antibacterial action, covering Gram-positive and Gram-negative organisms, both aerobes and anaerobes. Being virtually non absorbed, its bioavailability within the gastrointestinal tract is rather high with intraluminal and faecal drug concentrations that largely exceed the MIC values observed in vitro against a wide range of pathogenic microorganisms. The gastrointestinal tract represents therefore the primary therapeutic target and gastrointestinal infections the main indication. The little value of rifaximin outside the enteric area minimizes both antimicrobial resistance and systemic adverse events. Fermented dairy products enriched with probiotic bacteria have developed into one of the most successful categories of functional foods. Probiotics are defined as “live microorganisms which, when administered in adequate amounts, confer a health benefit on the host” (FAO/WHO, 2002), and mainly include Lactobacillus and Bifidobacterium species. Probiotic bacteria exert a direct effect on the intestinal microbiota of the host and contribute to organoleptic, rheological and nutritional properties of food. Administration of pharmaceutical probiotic formula has been associated with therapeutic effects in treatment of diarrhoea, constipation, flatulence, enteropathogens colonization, gastroenteritis, hypercholesterolemia, IBD, such as ulcerative colitis (UC), Crohn’s disease, pouchitis and irritable bowel syndrome. Prerequisites for probiotics are to be effective and safe. The characteristics of an effective probiotic for gastrointestinal tract disorders are tolerance to upper gastrointestinal environment (resistance to digestion by enteric or pancreatic enzymes, gastric acid and bile), adhesion on intestinal surface to lengthen the retention time, ability to prevent the adherence, establishment and/or replication of pathogens, production of antimicrobial substances, degradation of toxic catabolites by bacterial detoxifying enzymatic activities, and modulation of the host immune responses. This study was carried out using a validated three-stage fermentative continuous system and it is aimed to investigate the effect of rifaximin on the colonic microbial flora of a healthy individual, in terms of bacterial composition and production of fermentative metabolic end products. Moreover, this is the first study that investigates in vitro the impact of the simultaneous administration of the antibiotic rifaximin and the probiotic B. lactis BI07 on the intestinal microbiota. Bacterial groups of interest were evaluated using culture-based methods and molecular culture-independent techniques (FISH, PCR-DGGE). Metabolic outputs in terms of SCFA profiles were determined by HPLC analysis. Collected data demonstrated that rifaximin as well as antibiotic and probiotic treatment did not change drastically the intestinal microflora, whereas bacteria belonging to Bifidobacterium and Lactobacillus significantly increase over the course of the treatment, suggesting a spontaneous upsurge of rifaximin resistance. These results are in agreement with a previous study, in which it has been demonstrated that rifaximin administration in patients with UC, affects the host with minor variations of the intestinal microflora, and that the microbiota is restored over a wash-out period. In particular, several Bifidobacterium rifaximin resistant mutants could be isolated during the antibiotic treatment, but they disappeared after the antibiotic suspension. Furthermore, bacteria belonging to Atopobium spp. and E. rectale/Clostridium cluster XIVa increased significantly after rifaximin and probiotic treatment. Atopobium genus and E. rectale/Clostridium cluster XIVa are saccharolytic, butyrate-producing bacteria, and for these characteristics they are widely considered health-promoting microorganisms. The absence of major variations in the intestinal microflora of a healthy individual and the significant increase in probiotic and health-promoting bacteria concentrations support the rationale of the administration of rifaximin as efficacious and non-dysbiosis promoting therapy and suggest the efficacy of an antibiotic/probiotic combined treatment in several gut pathologies, such as IBD. To assess the use of an antibiotic/probiotic combination for clinical management of intestinal disorders, genetic, proteomic and physiologic approaches were employed to elucidate molecular mechanisms determining rifaximin resistance in Bifidobacterium, and the expected interactions occurring in the gut between these bacteria and the drug. The ability of an antimicrobial agent to select resistance is a relevant factor that affects its usefulness and may diminish its useful life. Rifaximin resistance phenotype was easily acquired by all bifidobacteria analyzed [type strains of the most representative intestinal bifidobacterial species (B. infantis, B. breve, B. longum, B. adolescentis and B. bifidum) and three bifidobacteria included in a pharmaceutical probiotic preparation (B. lactis BI07, B. breve BBSF and B. longum BL04)] and persisted for more than 400 bacterial generations in the absence of selective pressure. Exclusion of any reversion phenomenon suggested two hypotheses: (i) stable and immobile genetic elements encode resistance; (ii) the drug moiety does not act as an inducer of the resistance phenotype, but enables selection of resistant mutants. Since point mutations in rpoB have been indicated as representing the principal factor determining rifampicin resistance in E. coli and M. tuberculosis, whether a similar mechanism also occurs in Bifidobacterium was verified. The analysis of a 129 bp rpoB core region of several wild-type and resistant bifidobacteria revealed five different types of miss-sense mutations in codons 513, 516, 522 and 529. Position 529 was a novel mutation site, not previously described, and position 522 appeared interesting for both the double point substitutions and the heterogeneous profile of nucleotide changes. The sequence heterogeneity of codon 522 in Bifidobacterium leads to hypothesize an indirect role of its encoded amino acid in the binding with the rifaximin moiety. These results demonstrated the chromosomal nature of rifaximin resistance in Bifidobacterium, minimizing risk factors for horizontal transmission of resistance elements between intestinal microbial species. Further proteomic and physiologic investigations were carried out using B. lactis BI07, component of a pharmaceutical probiotic preparation, as a model strain. The choice of this strain was determined based on the following elements: (i) B. lactis BI07 is able to survive and persist in the gut; (ii) a proteomic overview of this strain has been recently reported. The involvement of metabolic changes associated with rifaximin resistance was investigated by proteomic analysis performed with two-dimensional electrophoresis and mass spectrometry. Comparative proteomic mapping of BI07-wt and BI07-res revealed that most differences in protein expression patterns were genetically encoded rather than induced by antibiotic exposure. In particular, rifaximin resistance phenotype was characterized by increased expression levels of stress proteins. Overexpression of stress proteins was expected, as they represent a common non specific response by bacteria when stimulated by different shock conditions, including exposure to toxic agents like heavy metals, oxidants, acids, bile salts and antibiotics. Also, positive transcription regulators were found to be overexpressed in BI07-res, suggesting that bacteria could activate compensatory mechanisms to assist the transcription process in the presence of RNA polymerase inhibitors. Other differences in expression profiles were related to proteins involved in central metabolism; these modifications suggest metabolic disadvantages of resistant mutants in comparison with sensitive bifidobacteria in the gut environment, without selective pressure, explaining their disappearance from faeces of patients with UC after interruption of antibiotic treatment. The differences observed between BI07-wt e BI07-res proteomic patterns, as well as the high frequency of silent mutations reported for resistant mutants of Bifidobacterium could be the consequences of an increased mutation rate, mechanism which may lead to persistence of resistant bacteria in the population. However, the in vivo disappearance of resistant mutants in absence of selective pressure, allows excluding the upsurge of compensatory mutations without loss of resistance. Furthermore, the proteomic characterization of the resistant phenotype suggests that rifaximin resistance is associated with a reduced bacterial fitness in B. lactis BI07-res, supporting the hypothesis of a biological cost of antibiotic resistance in Bifidobacterium. The hypothesis of rifaximin inactivation by bacterial enzymatic activities was verified by using liquid chromatography coupled with tandem mass spectrometry. Neither chemical modifications nor degradation derivatives of the rifaximin moiety were detected. The exclusion of a biodegradation pattern for the drug was further supported by the quantitative recovery in BI07-res culture fractions of the total rifaximin amount (100 μg/ml) added to the culture medium. To confirm the main role of the mutation on the β chain of RNA polymerase in rifaximin resistance acquisition, transcription activity of crude enzymatic extracts of BI07-res cells was evaluated. Although the inhibition effects of rifaximin on in vitro transcription were definitely higher for BI07-wt than for BI07-res, a partial resistance of the mutated RNA polymerase at rifaximin concentrations > 10 μg/ml was supposed, on the basis of the calculated differences in inhibition percentages between BI07-wt and BI07-res. By considering the resistance of entire BI07-res cells to rifaximin concentrations > 100 μg/ml, supplementary resistance mechanisms may take place in vivo. A barrier for the rifaximin uptake in BI07-res cells was suggested in this study, on the basis of the major portion of the antibiotic found to be bound to the cellular pellet respect to the portion recovered in the cellular lysate. Related to this finding, a resistance mechanism involving changes of membrane permeability was supposed. A previous study supports this hypothesis, demonstrating the involvement of surface properties and permeability in natural resistance to rifampicin in mycobacteria, isolated from cases of human infection, which possessed a rifampicin-susceptible RNA polymerase. To understand the mechanism of membrane barrier, variations in percentage of saturated and unsaturated FAs and their methylation products in BI07-wt and BI07-res membranes were investigated. While saturated FAs confer rigidity to membrane and resistance to stress agents, such as antibiotics, a high level of lipid unsaturation is associated with high fluidity and susceptibility to stresses. Thus, the higher percentage of saturated FAs during the stationary phase of BI07-res could represent a defence mechanism of mutant cells to prevent the antibiotic uptake. Furthermore, the increase of CFAs such as dihydrosterculic acid during the stationary phase of BI07-res suggests that this CFA could be more suitable than its isomer lactobacillic acid to interact with and prevent the penetration of exogenous molecules including rifaximin. Finally, the impact of rifaximin on immune regulatory functions of the gut was evaluated. It has been suggested a potential anti-inflammatory effect of rifaximin, with reduced secretion of IFN-γ in a rodent model of colitis. Analogously, it has been reported a significant decrease in IL-8, MCP-1, MCP-3 e IL-10 levels in patients affected by pouchitis, treated with a combined therapy of rifaximin and ciprofloxacin. Since rifaximin enables in vivo and in vitro selection of Bifidobacterium resistant mutants with high frequency, the immunomodulation activities of rifaximin associated with a B. lactis resistant mutant were also taken into account. Data obtained from PBMC stimulation experiments suggest the following conclusions: (i) rifaximin does not exert any effect on production of IL-1β, IL-6 and IL-10, whereas it weakly stimulates production of TNF-α; (ii) B. lactis appears as a good inducer of IL-1β, IL-6 and TNF-α; (iii) combination of BI07-res and rifaximin exhibits a lower stimulation effect than BI07-res alone, especially for IL-6. These results confirm the potential anti-inflammatory effect of rifaximin, and are in agreement with several studies that report a transient pro-inflammatory response associated with probiotic administration. The understanding of the molecular factors determining rifaximin resistance in the genus Bifidobacterium assumes an applicative significance at pharmaceutical and medical level, as it represents the scientific basis to justify the simultaneous use of the antibiotic rifaximin and probiotic bifidobacteria in the clinical treatment of intestinal disorders.

Product placement e libertà di espressione

Object of the search is the advertising phenomenon of the "product placement", with reference to that it has been investigated legality’s limits, as well as the relationship with the constitutionally protected liberty of expression. Particularly, it has been analyzed, in first place, the problem of the relationship between the freedom of expression and the liberty of economic initiative, with particular reference to the different circles of guardianship to these prepared: or, larger, the one provided for the first from the 21th article of Costitution, more circumscribed, instead, the one established in the 41th article of Costitution, with reference to the second. This analysis has been made with the purpose to investigate the coordination among such liberties in those forms of communications that, for the proper peculiarities that characterize them, can be qualified, according to the concrete circumstances in which they are spread, so much forms of liberty of expression, how much exercise of an activity of enterprise. Under this last profile, it has been taken attention on the advertising activity and, specially, on the non transparent publicities, or not immediately perceivable as such from their receivers, and, therefore, in contrast with the advertising trasparence’s principle: or, the so-called cases of hidden publicity, what the editorial publicity, both "in narrow sense" both "in general sense", as well as the phenomenon of the product placement (or positioning of product), by now diffused in the commercial routine. Therefore, it has been proceeded to a complete and exhaustive examination of innovations introduced by the recent legislative discipline in subject of “planned placement of marks and products” in the cinema works, appraising, in the specific one, the effects, juridical and no juridical, consequential from the introduction of a first form of regulation of the phenomenon of the product placement and, particularly, from the express provision about the legality of the use to such advertising, if it has realized according to specific requirements or condition. In relationship to such profile, it has been also investigate limits (sub kind of normative gaps) from which the recently introduced discipline in subject would seem characterized. Finally, a further circle of investigation has concerned the possible organization of the phenomenon under a negotiate aspect, as particular contract of advertising, in which the object consists in an promotional activity. Concerning this, the experience of foreign countries (above all the Anglo-Saxon one) has been very important, because of the absence, in our arrangement, of a general normative discipline about advertising contracts. Consequently, I’ve investigated principal characteristics of similar contracts, in first place the atypicalness, because of the lack, in Italy, of a legislative discipline of this contract. Such investigation has also been developed through a comparation between the positioning of product and the other advertising contracts, among which, particularly, the sponsorship, as well as the contracts for the advertising exploitation of the name and other people's image, and, specially, the contract of testimonial and the contract of endorsement.

The effect of adding features on product attractiveness: the role of product perceived congruity

Technological progress has been enabling companies to add disparate features to their existing products. This research investigates the effect of adding more features on consumers’ evaluation of the product, by examining in particular the role of the congruity of the features added with the base product as a variable the moderates the effect of increasing the number of features. Grounding on schema-congruity theory, I propose that the cognitive elaboration associated with the product congruity of the features added explains consumers’ evaluation as the number of new features increases. In particular, it is shown that consumers perceive a benefit from increasing the number of features only when these features are congruent with the product. The underlying mechanisms that explains this finding predicts that when the number of incongruent features increases the cognitive resources necessary to elaborate such incongruities increase and consumers are not willing to spend such resources. However, I further show that when encouraged to consider the new features thoughtfully, consumers do seem able to infer value from increasing the number of moderately incongruent features. Nonetheless, this finding does not apply for those new features that are extremely incongruent with the product. Further evidence for consumers’ ability to resolve the moderate incongruity associated with adding more features is also shown, by studying the moderating role of temporal construal. I propose that consumers perceive an increase in product evaluation as the number of moderately incongruent features increases when consumers consider purchasing the product in the distant future, whereas such an increase is not predicted for the near future scenario. I verify these effect in three experimental studies. Theoretical and managerial implications, and possible avenues of future research are also suggested.

Empirical and fundamental mechanical tests in the evaluation of dough and bread rheological properties

Bread dough and particularly wheat dough, due to its viscoelastic behaviour, is probably the most dynamic and complicated rheological system and its characteristics are very important since they highly affect final products’ textural and sensorial properties. The study of dough rheology has been a very challenging task for many researchers since it can provide numerous information about dough formulation, structure and processing. This explains why dough rheology has been a matter of investigation for several decades. In this research rheological assessment of doughs and breads was performed by using empirical and fundamental methods at both small and large deformation, in order to characterize different types of doughs and final products such as bread. In order to study the structural aspects of food products, image analysis techniques was used for the integration of the information coming from empirical and fundamental rheological measurements. Evaluation of dough properties was carried out by texture profile analysis (TPA), dough stickiness (Chen and Hoseney cell) and uniaxial extensibility determination (Kieffer test) by using a Texture Analyser; small deformation rheological measurements, were performed on a controlled stress–strain rheometer; moreover the structure of different doughs was observed by using the image analysis; while bread characteristics were studied by using texture profile analysis (TPA) and image analysis. The objective of this research was to understand if the different rheological measurements were able to characterize and differentiate the different samples analysed. This in order to investigate the effect of different formulation and processing conditions on dough and final product from a structural point of view. For this aim the following different materials were performed and analysed: - frozen dough realized without yeast; - frozen dough and bread made with frozen dough; - doughs obtained by using different fermentation method; - doughs made by Kamut® flour; - dough and bread realized with the addition of ginger powder; - final products coming from different bakeries. The influence of sub-zero storage time on non-fermented and fermented dough viscoelastic performance and on final product (bread) was evaluated by using small deformation and large deformation methods. In general, the longer the sub-zero storage time the lower the positive viscoelastic attributes. The effect of fermentation time and of different type of fermentation (straight-dough method; sponge-and-dough procedure and poolish method) on rheological properties of doughs were investigated using empirical and fundamental analysis and image analysis was used to integrate this information throughout the evaluation of the dough’s structure. The results of fundamental rheological test showed that the incorporation of sourdough (poolish method) provoked changes that were different from those seen in the others type of fermentation. The affirmative action of some ingredients (extra-virgin olive oil and a liposomic lecithin emulsifier) to improve rheological characteristics of Kamut® dough has been confirmed also when subjected to low temperatures (24 hours and 48 hours at 4°C). Small deformation oscillatory measurements and large deformation mechanical tests performed provided useful information on the rheological properties of samples realized by using different amounts of ginger powder, showing that the sample with the highest amount of ginger powder (6%) had worse rheological characteristics compared to the other samples. Moisture content, specific volume, texture and crumb grain characteristics are the major quality attributes of bread products. The different sample analyzed, “Coppia Ferrarese”, “Pane Comune Romagnolo” and “Filone Terra di San Marino”, showed a decrease of crumb moisture and an increase in hardness over the storage time. Parameters such as cohesiveness and springiness, evaluated by TPA that are indicator of quality of fresh bread, decreased during the storage. By using empirical rheological tests we found several differences among the samples, due to the different ingredients used in formulation and the different process adopted to prepare the sample, but since these products are handmade, the differences could be account as a surplus value. In conclusion small deformation (in fundamental units) and large deformation methods showed a significant role in monitoring the influence of different ingredients used in formulation, different processing and storage conditions on dough viscoelastic performance and on final product. Finally the knowledge of formulation, processing and storage conditions together with the evaluation of structural and rheological characteristics is fundamental for the study of complex matrices like bakery products, where numerous variable can influence their final quality (e.g. raw material, bread-making procedure, time and temperature of the fermentation and baking).

The secondary market for life insurance policies in the United States market evolution and product valuation

In this work we discuss the secondary market for life insurance policies in the United States of America. First, we give an overview of the life settlement market: how it came into existence, its growth prospects and the ethical issues it arises. Secondly, we discuss the characteristics of the different life insurance products present in the market and describe how life settlements are originated. Life settlement transactions tend to be long and complex transactions that require the involvement of a number of parties. Also, a direct investment into life insurance policies is fraught with a number of practical issues and entails risks that are not directly related to longevity. This may reduce the efficiency of a direct investment in physical policies. For these reasons, a synthetic longevity market has evolved. The number of parties involved in a synthetic longevity transaction is typically smaller and the broker-dealer transferring the longevity exposure will be retaining most or all of the risks a physical investment entails. Finally, we describe the main methods used in the market to evaluate life settlement investments and the role of life expectancy providers.

Creative process in globally distributed teams: a study of new product development in Volvo

With the business environments no longer confined to geographical borders, the new wave of digital technologies has given organizations an enormous opportunity to bring together their distributed workforce and develop the ability to work together despite being apart (Prasad & Akhilesh, 2002). resupposing creativity to be a social process, the way that this phenomenon occurs when the configuration of the team is substantially modified will be questioned. Very little is known about the impact of interpersonal relationships in the creativity (Kurtzberg & Amabile, 2001). In order to analyse the ways in which the creative process may be developed, we ought to be taken into consideration the fact that participants are dealing with a quite an atypical situation. Firstly, in these cases socialization takes place amongst individuals belonging to a geographically dispersed workplace, where interpersonal relationships are mediated by the computer, and where trust must be developed among persons who have never met one another. Participants not only have multiple addresses and locations, but above all different nationalities, and different cultures, attitudes, thoughts, and working patterns, and languages. Therefore, the central research question of this thesis is as follows: “How does the creative process unfold in globally distributed teams?” With a qualitative approach, we used the case study of the Business Unit of Volvo 3P, an arm of Volvo Group. Throughout this research, we interviewed seven teams engaged in the development of a new product in the chassis and cab areas, for the brands Volvo and Renault Trucks, teams that were geographically distributed in Brazil, Sweden, France and India. Our research suggests that corporate values, alongside with intrinsic motivation and task which lay down the necessary foundations for the development of the creative process in GDT.

Teoria e prassi del product placement. Il caso dei film contemporanei per adolescenti (ITA/USA 2006-2010)

Through this research I have tried to demonstrate how the evolution of the newest marketing strategies - that work towards engaging relationships with consumers, thus building an emotional connection between the brand and the user- can be considered as a response to the evolution of young audiences and consumers. More specifically, I have analized product placement as a cultural and social phenomena above all, and not only as an economical one, thus demonstrating all the social and cultural practices that this tool implies. The approach I have chosen to do so, is historical-analytical, particularly focusing on the evolution of the society and of the consumer, especially for what teenagers (both as audiences and as consumers) are concerned.

The influence of career histories on team perfomance in new product development: a study of the video game industry

The purpose of this research is to contribute to the literature on organizational demography and new product development by investigating how diverse individual career histories impact team performance. Moreover we highlighted the importance of considering also the institutional context and the specific labour market arrangements in which a team is embedded, in order to interpret correctly the effect of career-related diversity measures on performance. The empirical setting of the study is the videogame industry, and the teams in charge of the development of new game titles. Video games development teams are the ideal setting to investigate the influence of career histories on team performance, since the development of videogames is performed by multidisciplinary teams composed by specialists with a wide variety of technical and artistic backgrounds, who execute a significant amounts of creative thinking. We investigate our research question both with quantitative methods and with a case study on the Japanese videogame industry: one of the most innovative in this sector. Our results show how career histories in terms of occupational diversity, prior functional diversity and prior product diversity, usually have a positive influence on team performance. However, when the moderating effect of the institutional setting is taken in to account, career diversity has different or even opposite effect on team performance, according to the specific national context in which a team operates.

Pretrattamenti termo-meccanici ed enzimatici su scarti dell'industria enologica per la produzione di Biogas

A fundamental assumption for by-product from winery industy waste-management is their economic and commercial increase in value. High energetic value recovery from winery industry is an attractive economic solution to stimulate new sustainable process. Approach of this work is based about physic and biological treatment with grape stalks and grape marc to increase polysaccharides components of cell wall and energetic availability of this by-products. Grape stalks for example have a high percentage of lignin and cellulose and can’t be used, whitout pretreatment, for an anaerobic digestion process. Our findings show enzymatic and thermo-mechanical pre-treatments in combined application for optimise hydrolytic mechanism on winemaking wastes which represents 0,9 milion ton/year in Italy and on straw, cereal by-products with high lignin content. A screening of specifically industrial enzymatic complex for the hydrolysis lignocellulosic biomass were tested using the principal polysaccharides component of the vegetal cells. Combined thermo-mechanical and enzymatic pretreatment improve substrates conversion in batch test fermentation experiment. The conservation of the grape stalks, at temperature above 0°C, allow the growth of spontaneus fermentation that reduce their polysaccharides content so had investigated anarobic condition of conservation. The other objective of this study was to investigate the capability of a proprietary strain of L.buchneri LN 40177 to enhance the accessibility of fermentable forage constituents during the anaerobic conservation process by releasing the enzyme ferulate esterase. The time sequence study by batch tests showed that the L. buchneri LN-40177 inoculated grape stalk substrate was more readily available in the fermenter. In batch tests with grape stalk, after mechanical treatment, the L. buchneri LN41077 treated substrate yielded on average 70% more biogas per kg/DM. Thermo-mechanical, enzymatic and biological treatment with L. buchneri LN-40177 can increase the biogas production from low fermented biomasses and the consequent their useful in anaerobic biodigesters for agro-bioenergy production.

Patent Strategies and R&D in Complex Product Industries

The study aims at providing a framework conceptualizing patenting activities under the condition of intellectual property rights fragmentation. Such a framework has to deal with the interrelated problems of technological complexity in the modern patent landscape. In that respect, ex-post licensing agreements have been incorporated into the analysis. More precisely, by consolidating the right to use patents required for commercialization of a product, private market solutions, such as cross-licensing agreements and patent pools help firms to overcome problems triggered by the intellectual property rights fragmentation. Thereby, private bargaining between parties as such cannot be isolated from the legal framework. A result of this analysis is that policies ignoring market solutions and only focusing on static gains can mitigate the dynamic efficiency gains as induced by the patent system. The evidence found in this thesis supports the opinion that legal reforms that aim to decrease the degree of patent protection or to lift it all together can hamper the functioning of the current system.

Use of Biotechnology to increase the content of bioactive compounds in fermented foods of plant origin

The objectives of this PhD research were: i) to evaluate the use of bread making process to increase the content of β-glucans, resistant starch, fructans, dietary fibers and phenolic compounds of kamut khorasan and wheat breads made with flours obtained from kernels at different maturation stage (at milky stage and fully ripe) and ii) to study the impact of whole grains consumption in the human gut. The fermentation and the stages of kernel development or maturation had a great impact on the amount of resistant starch, fructans and β-glucans as well as their interactions resulted highly statistically significant. The amount of fructans was high in kamut bread (2.1g/100g) at the fully ripe stage compared to wheat during industrial fermentation (baker’s yeast). The sourdough increases the content of polyphenols more than industrial fermentation especially in bread made by flour at milky stage. From the analysis of volatile compounds it resulted that the sensors of electronic nose perceived more aromatic compound in kamut products, as well as the SPME-GC-MS, thus we can assume that kamut is more aromatic than wheat, so using it in sourdough process can be a successful approach to improve the bread taste and flavor. The determination of whole grain biormakers such as alkylresorcinols and others using FIE-MS AND GC-tof-MS is a valuable alternative for further metabolic investigations. The decrease of N-acetyl-glucosamine and 3-methyl-hexanedioic acid in kamut faecal samples suggests that kamut can have a role in modulating mucus production/degradation or even gut inflammation. This work gives a new approach to the innovation strategies in bakery functional foods, that can help to choose the right or best combination between stages of kernel maturation-fermentation process and baking temperature.