Mass spectrometry and n-in-one analytics in early drug discovery: Combinatorial chemistry libraries, lipophilicity and absorption screening

This thesis describes current and past n-in-one methods and presents three early experimental studies using mass spectrometry and the triple quadrupole instrument on the application of n-in-one in drug discovery. N-in-one strategy pools and mix samples in drug discovery prior to measurement or analysis. This allows the most promising compounds to be rapidly identified and then analysed. Nowadays properties of drugs are characterised earlier and in parallel with pharmacological efficacy. Studies presented here use in vitro methods as caco-2 cells and immobilized artificial membrane chromatography for drug absorption and lipophilicity measurements. The high sensitivity and selectivity of liquid chromatography mass spectrometry are especially important for new analytical methods using n-in-one. In the first study, the fragmentation patterns of ten nitrophenoxy benzoate compounds, serial homology, were characterised and the presence of the compounds was determined in a combinatorial library. The influence of one or two nitro substituents and the alkyl chain length of methyl to pentyl on collision-induced fragmentation was studied, and interesting structurefragmentation relationships were detected. Two nitro group compounds increased fragmentation compared to one nitro group, whereas less fragmentation was noted in molecules with a longer alkyl chain. The most abundant product ions were nitrophenoxy ions, which were also tested in the precursor ion screening of the combinatorial library. In the second study, the immobilized artificial membrane chromatographic method was transferred from ultraviolet detection to mass spectrometric analysis and a new method was developed. Mass spectra were scanned and the chromatographic retention of compounds was analysed using extract ion chromatograms. When changing detectors and buffers and including n-in-one in the method, the results showed good correlation. Finally, the results demonstrated that mass spectrometric detection with gradient elution can provide a rapid and convenient n-in-one method for ranking the lipophilic properties of several structurally diverse compounds simultaneously. In the final study, a new method was developed for caco-2 samples. Compounds were separated by liquid chromatography and quantified by selected reaction monitoring using mass spectrometry. This method was used for caco-2 samples, where absorption of ten chemically and physiologically different compounds was screened using both single and nin- one approaches. These three studies used mass spectrometry for compound identification, method transfer and quantitation in the area of mixture analysis. Different mass spectrometric scanning modes for the triple quadrupole instrument were used in each method. Early drug discovery with n-in-one is area where mass spectrometric analysis, its possibilities and proper use, is especially important.

Suomen sahateollisuuden supistumisen ja tulevaisuuden kuvien aluetaloudelliset vaikutukset

Sahateollisuuden tuotantokapasiteetti on vähentynyt viime vuosina paljon. Syvä lama on leikannut vientiä ja myös rakentaminen on hiljentynyt. Tämä on pakottanut sopeuttamistoimiin samaan tapaan kuin massa- ja paperiteollisuuden kohdalla. Mikäli tällekin vuodelle suunnitellut toimet toteutuvat, yhteensä yksitoista sahaa ja seitsemän jalostetehdasta on suljettu. Kapasiteetin leikkaukset koskevat yhteensä kahtatoista maakuntaa ja viittätoista seutukuntaa. Puolet leikkauksista kohdistuu pieniin seutukuntiin, joka lisää aluetalouden sopeutumishaastetta. Tutkimuksessa on selvitetty sahojen ja jalostetehtaiden sulkemisten aluetaloudellisia vaikutuksia seutukuntien taloudelliseen kasvuun ja työllisyyteen. Sahateollisuuden menetyksiä verrattiin maakuntatasolla massa- ja paperiteollisuuden vastaaviin. Lisäksi tarkasteltiin alan kolmea tulevaisuuden kuvaa Suomen tasolla. Laskelmat suoritettiin alueellisella yleisen tasapainon RegFinDyn-simulointimallilla. Tarkasteluperiodi oli sulkemisten osalta vuodet 2004–2013 ja vuodet 2010–2014 tulevaisuuden kuvien kohdalla. Sulkemisten vaikutus seutukuntien taloudelliseen kasvuun näyttäisi jäävän suhteellisen pieneksi. Aluetalous sopeutuisi uuteen tilanteeseen muiden alojen kompensoidessa osan menetyksistä. Talouskasvun menetykset ovat yhteensä 173,8 miljoonaa euroa, jos Varkauden sahaa ei suljeta ja 184,7 miljoonaa, jos sulkeminen tapahtuu. Tilanne näyttäisi samalta työllisyyden suhteen. Työllisyyden menetykset ovat vastaavasti yhteensä 2 693 henkilötyövuotta tai 3 031. Menetysten osuus maakuntien yhteenlasketusta BKT:sta ja työllisyydestä on 0,3 %. Tämä on vähemmän kuin massa- ja paperiteollisuudessa. Ongelmalliseksi tilanteen tekee se, että molemmat alat ovat menettäneet talouskasvua ja työllisyyttä viidessä maakunnassa. ETLA:n sahateollisuuden ja rakentamisen tuotantoennusteiden samoin kuin PTT:n hintaennusteiden mukaan tulevaisuus näyttäisi valoisammalta. Laman pohjan saavuttamisen jälkeen sekä tuotanto että kv-hinnat alkaisivat vahvistua tästä vuodesta lähtien. Tulevaisuuden kuvat muodostettiin näiden suhdanne-ennusteiden pohjalta. Mikäli myönteinen kehitys toteutuisi, Suomen BKT voisi kasvaa yhteensä 300–900 miljoonalla eurolla vuoden 2014 loppuun mennessä. Työllisyys voisi vahvistua 1 000–5 000 henkilötyövuodella. Sahateollisuuden sopeuttaminen tulee jatkumaan, mikäli lama ei kohta hellitä. Alan kehitystä seurataan yleisellä tasolla, mutta myös sopeuttamisen aluetaloudellisten vaikutusten arviointia olisi syytä jatkaa.

Suomen metsäteollisuuden uusien mahdollisuuksien aluetaloudelliset vaikutukset

Biojalostamo lisäisi talouskasvua ja työllisyyttä Tutkimuksessa selvitettiin biojalostamotoiminnan aluetaloudellisia vaikutuksia Kymenlaakson ja Satakunnan maakuntiin. Biodieselin valmistus kohentaisi molemmilla alueilla talouskasvua yhteensä 3,0–3,5 prosenttiyksiköllä eli noin 200 miljoonalla eurolla vuoteen 2020 mennessä. Työllisyys paranisi Kymenlaaksossa 437 ja Satakunnassa 420 henkilötyövuodella. Biojalostamotoiminta näyttäisi tukevan työllisyyttä suhteellisesti vähemmän kuin talouskasvua johtuen alan pääomavaltaisuudesta. Kymenlaakso hyötyisi biojalostamosta hiukan enemmän kuin Satakunta johtuen aluetalouksien rakenteellisista eroista. Metsäteollisuuden menetyksiä voitaisiin kompensoida Kymenlaakso on kärsinyt metsäteollisuuden supistumisesta tähän asti suurimmat menetykset. Biojalostamon perustaminen Kymenlaaksoon voisi merkittävästi korvata paperin tuotannon laskusta aiheutuneita menetyksiä. Talouskasvun suhteen biojalostamo voisi korvata puolet menetyksistä. Työllisyysmenetyksistä biojalostamotoiminta pystyisi kompensoimaan noin neljäsosan. Tukien merkitystä biojalostamon aluetaloudellisiin vaikutuksiin selvitettiin tukityypeittäin ja -tasoittain. Sekä raaka-aine- että tuotantotukivaikutukset jäivät simuloinneissa vaatimattomiksi. Tukien tehottomuutta selittää niiden pienuus. Muut toimialat kärsivät biojalostamoalan tukemisesta, mikä vähentää tukien aluetaloudellista vaikuttavuutta. Toisaalta reaalimaailmassa tuet voivat olla ratkaisevassa asemassa bioenergia-alan käynnistyessä ja sen kehityksen alkumetreillä. Bioenergia-strategian mukainen kehitys tuottaisi tulosta Bioenergian käytön lisäämisen aluetaloudellista vaikuttavuutta tutkittiin Keski-Suomen maakuntaa koskevan tapaustutkimuksen avulla. Vaikutusten laskenta pohjautui maakunnan bioenergian käytön tavoitteiden mukaisiin määriin. Mikäli bioenergian käyttö kasvaisi Keski-Suomessa neljällä terawattitunnilla vuoteen 2015 mennessä, vaikutus talouskasvuun olisi yhteensä 0,5 prosenttiyksikköä eli 35 miljoonaa euroa. Maakunnan työllisyys paranisi yli 200 henkilötyövuodella. Todennäköisesti myönteisiä aluetalousvaikutuksia vahvistaisi vielä alueen energiaostoista johtuvien vuotojen väheneminen energiaomavaraisuuden kasvaessa. Keski-Suomelle voisi kertyä hyötyjä myös bioenergiateknologian alan laitevalmistuksen osaamisen lisääntymisen kautta. Biojalostamotoiminta on täysin uudenlaista toimintaa, joten tilastollista seurantatietoa tältä alalta ei ole käytettävissä. Julkisia teknis-taloudellisia selvityksiä biodieselin tuotannosta on saatavilla suhteellisen niukasti. Koska simuloinneista saadut tulokset perustuvat biojalostamolle oletettuun kustannusrakenteeseen, tuloksia on tästä syystä käsiteltävä vain suuntaa-antavina. Ruralia-instituutti suosittaa biojalostamotoiminnan aluetaloudellisten vaikutusten seuraamista ja laskelmien päivittämistä, kun tarkempaa tietoa biojalostamon tuotannon kustannuksista ja tuotteiden kysynnän rakenteesta on saatavilla. Biojalostamon kustannusrakenne. Biojalostamotoiminta kompensoisi noin puolet massa- ja paperiteollisuuden supistumisesta aiheutuneista talouskasvun menetyksistä Kymenlaaksossa.

Brain oscillatory responses, working memory and cognitive developmental stages in 11- and 14-year-old children: an ERD/ERS study

Aim: So far, most of the cognitive neuroscience studies investigating the development of brain activity in childhood have made comparisons between different age groups and ignored the individual stage of cognitive development. Given the wide variation in the rate of cognitive development, this study argues that chronological age alone cannot explain the developmental changes in brain activity. This study demonstrates how Piaget s theory and information on child s individual stage of development can complement the age-related evaluations of brain oscillatory activity. In addition, the relationship between cognitive development and working memory is investigated. Method: A total of 33 children (17 11-year-olds, 16 14-year-olds) participated in this study. The study consisted of behavioural tests and an EEG experiment. Behavioral tests included two Piagetian tasks (the Volume and Density task, the Pendulum task) and Raven s Standard Progressive Matrices task. During EEG experiment, subjects performed a modified version of the Sternberg s memory search paradigm which consisted of an auditorily presented memory set of 4 words and a probe word following these. The EEG data was analyzed using the event-related desynchronization / synchronization (ERD/ERS) method. The Pendulum task was used to assess the cognitive developmental stage of each subject and to form four groups based on age (11- or 14-year-olds) and cognitive developmental stage (concrete or formal operational stage). Group comparisons between these four groups were performed for the EEG data. Results and conclusions: Both age- and cognitive stage-related differences in brain oscillatory activity were found between the four groups. Importantly, age-related changes similar to those reported by previous studies were found also in this study, but these changes were modified by developmental stage. In addition, the results support a strong link between working memory and cognitive development by demonstrating differences in memory task related brain activity and cognitive developmental stages. Based on these findings it is suggested that in the future, comparisons of development of brain activity should not be based only on age but also on the individual cognitive developmental stage.

Wood and fiber properties of Norway spruce and its suitability for thermomechanical pulping.

In the first part of the study, the selected wood and fiber properties were investigated in terms of their occurrence and variation in wood, as well as their relevance from the perspective of thermomechanical pulping process and related end-products. It was concluded that the most important factors were the fiber dimensions, juvenile wood content, and in some cases, the content of heartwood being associated with extremely dry wood with low permeability in spruce. With respect to the above properties, the following three pulpwood assortments of which pulping potential was assumed to vary were formed: wood from regeneration cuttings, first-thinnings wood, and sawmill chips. In the experimental part of the study the average wood and fiber characteristics and their variation were determined for each raw material group prior to pulping. Subsequently, each assortment - equaling about 1500 m3 roundwood - was pulped separately for a 24 h period, at constant process conditions. The properties of obtained newsgrade thermomechanical pulps were then determined. Thermomechanical pulping (TMP) from sawmill chips had the highest proportion of long fibers, smallest proportion of fines, and had generally the coarsest and longest fibers. TMP from first-thinnings wood was just the opposite, whereas that from regeneration cuttings fell in between the above two extremes. High proportion of dry heartwood in wood originating from regeneration cuttings produced a slightly elevated shives content. However, no differences were found in pulp specific energy consumption. The obtained pulp tear index was clearly best in TMP made from sawmill chips and poorest in pulp from first-thinnings wood, which had generally inferior strength properties. No dramatical differences in any of the strength properties were found between pulp from sawmill residual wood and regeneration cuttings. Pulp optical properties were superior in TMP from first-thinnings. Unexpectedly, no noticeable differences, which could be explained with fiber morphology, were found in sheet density, bulk, air permeance or roughness between the three pulps. The most important wood quality factors in this study were the fiber length, fiber cross-sectional dimensions and percentage juvenile wood. Differences found in the quality of TMP manufactured from the above spruce assortments suggest that they could be segregated and pulped separately to obtain specific product characteristics, i.e., for instance tailor-made end-products, and to minimize unnecessary variation in the raw material quality, and hence, pulp quality.

Accurate mass-based analysis in human sport doping control : Application of liquid chromatography - time-of-flight mass spectrometry

Human sport doping control analysis is a complex and challenging task for anti-doping laboratories. The List of Prohibited Substances and Methods, updated annually by World Anti-Doping Agency (WADA), consists of hundreds of chemically and pharmacologically different low and high molecular weight compounds. This poses a considerable challenge for laboratories to analyze for them all in a limited amount of time from a limited sample aliquot. The continuous expansion of the Prohibited List obliges laboratories to keep their analytical methods updated and to research new available methodologies. In this thesis, an accurate mass-based analysis employing liquid chromatography - time-of-flight mass spectrometry (LC-TOFMS) was developed and validated to improve the power of doping control analysis. New analytical methods were developed utilizing the high mass accuracy and high information content obtained by TOFMS to generate comprehensive and generic screening procedures. The suitability of LC-TOFMS for comprehensive screening was demonstrated for the first time in the field with mass accuracies better than 1 mDa. Further attention was given to generic sample preparation, an essential part of screening analysis, to rationalize the whole work flow and minimize the need for several separate sample preparation methods. Utilizing both positive and negative ionization allowed the detection of almost 200 prohibited substances. Automatic data processing produced a Microsoft Excel based report highlighting the entries fulfilling the criteria of the reverse data base search (retention time (RT), mass accuracy, isotope match). The quantitative performance of LC-TOFMS was demonstrated with morphine, codeine and their intact glucuronide conjugates. After a straightforward sample preparation the compounds were analyzed directly without the need for hydrolysis, solvent transfer, evaporation or reconstitution. The hydrophilic interaction technique (HILIC) provided good chromatographic separation, which was critical for the morphine glucuronide isomers. A wide linear range (50-5000 ng/ml) with good precision (RSD<10%) and accuracy (±10%) was obtained, showing comparable or better performance to other methods used. In-source collision-induced dissociation (ISCID) allowed confirmation analysis with three diagnostic ions with a median mass accuracy of 1.08 mDa and repeatable ion ratios fulfilling WADA s identification criteria. The suitability of LC-TOFMS for screening of high molecular weight doping agents was demonstrated with plasma volume expanders (PVE), namely dextran and hydroxyethylstarch (HES). Specificity of the assay was improved, since interfering matrix compounds were removed by size exclusion chromatography (SEC). ISCID produced three characteristic ions with an excellent mean mass accuracy of 0.82 mDa at physiological concentration levels. In summary, by combining TOFMS with a proper sample preparation and chromatographic separation, the technique can be utilized extensively in doping control laboratories for comprehensive screening of chemically different low and high molecular weight compounds, for quantification of threshold substances and even for confirmation. LC-TOFMS rationalized the work flow in doping control laboratories by simplifying the screening scheme, expediting reporting and minimizing the analysis costs. Therefore LC-TOFMS can be exploited widely in doping control, and the need for several separate analysis techniques is reduced.

Emerging bioenergy business at Finnish independent industrial sawmills

Finnish forest industry is in the middle of a radical change. Deepening recession and the falling demand of woodworking industry´s traditional products have forced also sawmilling industry to find new and more fertile solutions to improve their operational preconditions. In recent years, the role of bioenergy production has often been highlighted as a part of sawmills´ business repertoire. Sawmilling produces naturally a lot of by-products (e.g. bark, sawdust, chips) which could be exploited more effectively in energy production, and this would bring more incomes or maybe even create new business opportunities for sawmills. Production of bioenergy is also supported by government´s climate and energy policies favouring renewable energy sources, public financial subsidies, and soaring prices of fossil fuels. Also the decreasing production of domestic pulp and paper industry releases a fair amount of sawmills´ by-products for other uses. However, bioenergy production as a part of sawmills´ by-product utilization has been so far researched very little from a managerial point of view. The purpose of this study was to explore the relative significance of the main bioenergy-related processes, resources and factors at Finnish independent industrial sawmills including partnerships, cooperation, customers relationships and investments, and also the future perspectives of bioenergy business at these sawmills with the help of two resource-based approaches (resource-based view, natural-resource-based view). Data of the study comprised of secondary data (e.g. literature), and primary data which was attracted from interviews directed to sawmill managers (or equivalent persons in charge of decisions regarding bioenergy production at sawmill). While a literature review and the Delphi method with two questionnaires were utilized as the methods of the study. According to the results of the study, the most significant processes related to the value chain of bioenergy business are connected to raw material availability and procurement, and customer relationships management. In addition to raw material and services, the most significant resources included factory and machinery, personnel, collaboration, and geographic location. Long-term cooperation deals were clearly valued as the most significant form of collaboration, and especially in processes connected to raw material procurement. Study results also revealed that factors related to demand, subsidies and prices had highest importance in connection with sawmills´ future bioenergy business. However, majority of the respondents required that certain preconditions connected to the above-mentioned factors should be fulfilled before they will continue their bioenergy-related investments. Generally, the answers showed a wide divergence of opinions among the respondents which may refer to sawmills´ different emphases and expectations concerning bioenergy. In other words, bioenergy is still perceived as a quite novel and risky area of business at Finnish independent industrial sawmills. These results indicate that the massive expansion of bioenergy business at private sawmills in Finland is not a self-evident truth. The blocking barriers seem to be connected mainly to demand of bioenergy and money. Respondents´ answers disseminated a growing dissatisfaction towards the policies of authorities, which don´t treat equally sawmill-based bioenergy compared to other forms of bioenergy. This proposition was boiled down in a sawmill manager´s comment: “There is a lot of bioenergy available, if they just want to make use of it.” It seems that the positive effects of government´s policies favouring the renewables are not taking effect at private sawmills. However, as there anyway seems to be a lot of potential connected to emerging bioenergy business at Finnish independent industrial sawmills, there is also a clear need for more profound future studies over this topic.

Renewable Energy and Climate Policies: Studies in the Forest and Energy Sector

‪This dissertation examines the impacts of energy and climate policies on the energy and forest sectors, focusing on the case of Finland. The thesis consists of an introduction article and four separate studies. The dissertation was motivated by the climate concern and the increasing demand of renewable energy. In particular, the renewable energy consumption and greenhouse gas emission reduction targets of the European Union were driving this work. In Finland, both forest and energy sectors are in key roles in achieving these targets. In fact, the separation between forest and energy sector is diminishing as the energy sector is utilizing increasing amounts of wood in energy production and as the forest sector is becoming more and more important energy producer.‬ ‪The objective of this dissertation is to find out and measure the impacts of climate and energy policies on the forest and energy sectors. In climate policy, the focus is on emissions trading, and in energy policy the dissertation focuses on the promotion of renewable forest-based energy use. The dissertation relies on empirical numerical models that are based on microeconomic theory. Numerical partial equilibrium mixed complementarity problem models were constructed to study the markets under scrutiny. The separate studies focus on co-firing of wood biomass and fossil fuels, liquid biofuel production in the pulp and paper industry, and the impacts of climate policy on the pulp and paper sector.‬ ‪The dissertation shows that the policies promoting wood-based energy may have have unexpected negative impacts. When feed-in tariff is imposed together with emissions trading, in some plants the production of renewable electricity might decrease as the emissions price increases. The dissertation also shows that in liquid biofuel production, investment subsidy may cause high direct policy costs and other negative impacts when compared to other policy instruments. The results of the dissertation also indicate that from the climate mitigation perspective, perfect competition is the favored wood market competition structure, at least if the emissions trading system is not global.‬ ‪In conclusion, this dissertation suggests that when promoting the use of wood biomass in energy production, the favored policy instruments are subsidies that promote directly the renewable energy production (i.e. production subsidy, renewables subsidy or feed-in premium). Also, the policy instrument should be designed to be dependent on the emissions price or on the substitute price. In addition, this dissertation shows that when planning policies to promote wood-based renewable energy, the goals of the policy scheme should be clear before decisions are made on the choice of the policy instruments.‬

Cruzando el umbral de la postmodernidad

Resumen: El presente escrito intenta ser un acercamiento al momento histórico actual conocido bajo el nombre de postmodernidad. El objetivo del artículo es alcanzar la esencia de esta época enunciando sus características, sus implicancias en el arte y su relación con el nihilismo y el fracaso del humanismo. También se busca explicar el vínculo existente entre ésta y la modernidad, el fracaso de la misma evidenciado en el Holocausto judío, la caída de la concepción unitaria de la historia, la desconfianza en los grandes relatos y la consiguiente superación de la modernidad

La leyenda artúrica en la literatura alemana : el Parzival de Wolfram von Eschenbach

Integran este número de la revista ponencias presentadas en Studia Hispanica Medievalia VIII: Actas de las IX Jornadas Internacionales de Literatura Española Medieval, 2008, y de Homenaje al Quinto Centenario de Amadis de Gaula.

Hegel, relações internacionais e globalismo jurídico

Resumen: Las Relaciones Internacionales son uno de los temas predominantes en la agenda política actual. ¿Hasta qué punto la teoría hegeliana legitima y, al mismo tiempo, explicita la contradicción-mediación en el escenario de la globalización? Teniendo en cuenta esta cuestión, el objetivo de este artículo es, en primer lugar, presentar la teoría hegeliana del Estado en el contexto de las Relaciones Internacionales y, a continuación, actualizar este debate tratándolo al respecto de la soberanía brasileña. Por último, se presentan los desafíos de la articulación de Derecho Interno e Internacional, y se reconstruyen las posiciones de algunas teorías políticas actuales en vistas a la posición hegeliana

Nietzsche y la filosofía de la consciencia en Más allá del bien y el mal

Resumen: Aunque de innegable utilidad, el énfasis cientificista en el estudio de la consciencia tal como lo ha desarrollado la filosofía de la mente contemporánea ha obnubilado otras perspectivas. Hoy resulta imperativo retrotraernos a ellas en el estudio de tesis sobre la consciencia. En aras de contribuir al debate actual, el presente trabajo se propone una interpretación de las tesis de Friedrich Nietzsche sobre la consciencia. Se analizarán, además, los presupuestos epistemológicos nietzscheanos involucrados en su crítica de la consciencia. Finalmente, se estudiará la interpretación de Paul Katsafanas sobre cómo debemos entender el concepto “consciencia” en Nietzsche

La filosofía de la liberación en la Argentina

Resumen: El artículo trata primero del surgimiento de la filosofía de la liberación en Argentina (1971); luego, de su caracterización, en cuanto su punto de partida y lugar hermenéutico es la opción preferencial por los pobres, en cuyo horizonte se replantean todos los grandes problemas filosóficos, desde América Latina, con vigencia universal. En tercer lugar se describen su historia y corrientes, dando especial relevancia a la histórico-cultural. Finalmente se habla de su vigencia actual, debida a la situación de los excluidos, la crisis del neoliberalismo excluyente, el nuevo mestizaje cultural de imaginarios en la sabiduría popular latinoamericana y la pertinencia del método interdisciplinar y anadialéctico