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  • Hakojärvi, Mikko (Helsingin yliopisto, 2015)
    Precision farming is a set of advanced technologies and cultivation practices that aim to enhance crop growing conditions in a site-specific manner. Current machinery offers many technical tools for such actions, but information about when and where it is necessary to use the developed machinery effectively is lacking. The use of crop models has been proposed to combine the effects of growing season conditions and field spatial properties. However, in a timely and spatially variable environment, which a field often is, much information about site-specific growing conditions should be available in order for an advanced crop model to reproduce the site-specific growth in a detailed manner. Unfortunately the information from fields has often been very limited, and insufficient for such purposes. Furthermore the set of precision farming tools and the number of growth factors that can be managed is limited. For describing maximal biomass accumulation, a simple crop model was introduced and evaluated in this thesis. The model is mechanistic, and it uses a minimal number of parameters that all are based on physics, chemistry or physiology. The model can be used for calculating the radiation or radiation and water limited biomass accumulation of a C3-crop. A field experiment equipped with continuous measurements was used for model establishment and after model establishment the model was evaluated with a field experiment with various radiation, nitrogen fertilization and precipitation conditions. In both the studies the crop model was found to produce the maximal biomass accumulation when parameter values measured in the experiment were used. The model was applied in a study evaluating the effects of selected site- and depth-specific soil properties on yield variation on three different clay soil fields located in southern Finland. In order to evaluate the effects of selected soil properties under various weather conditions a Monte Carlo method was used with the biomass accumulation model and generated precipitations. The yield variation was evaluated according to temporal mean biomass yield and temporal standard deviation of the biomass yield. For studying the use of the model in a spatial environment, the introduced biomass accumulation model was applied in a simulator built for simulating a fully automated crop farm. In addition, the use of continuous soil moisture measurements for measuring the crop water use and further for biomass accumulation was tested. According to the results, the crop model was capable of simulating the highest biomass accumulation of the crops used in the experiments. This was the case for all radiation-limited simulations and for most of the water-limited simulations. In a few cases the values of the observed soil properties were found to cause too low biomass accumulation in simulations but in such cases the problem was also present in the comparisons of observed soil properties and observed soil water content during the growing season. For future research with the model, the next phase will be to test the model use in precision farming-related decision making. The structure of the model enables its use with other C3-crops than small grain cereals. Therefore testing of the model with other C3-crops could be performed in future research.
  • Bauters, Merja (Helsingin yliopisto, 2007)
    The purpose of this study is to find a framework for a holistic approach to, and form a conceptual toolbox for, investigating changes in signs and in their interpretation. Charles S. Peirce s theory of signs in a communicative perspective is taken as a basis for the framework. The concern directing the study is the problem of a missing framework in analysing signs of visual artefacts from a holistic perspective as well as that of the missing conceptual tools. To discover the possibility of such a holistic approach to semiosic processes and to form a conceptual toolbox the following issues are discussed: i) how the many Objects with two aspects involved in Peirce s definition of sign-action, promote multiple semiosis arising from the same sign by the same Interpretant depending on the domination of the Objects; ii) in which way can the relation of the individual and society or group be made more apparent in the construction of the self since this construction is intertwined with the process of meaning-creation and interpretation; iii) how to account for the fundamental role of emotions in semiosis, and the relation of emotions with the often neglected topic of embodiment; iv) how to take into account the dynamic, mediating and processual nature of sign-action in analysing and understanding the changes in signs and in the interpretation of signs. An interdisciplinary approach is chosen for this dissertation. Concepts that developed within social psychology, developmental psychology, neurosciences and semiotics, are discussed. The common aspect of the approaches is that they in one way or another concentrate on mediation provided by signs in explaining human activity and cognition. The holistic approach and conceptual toolbox found are employed in a case study. This consists of an analysis of beer brands including a comparison of brands from two different cultures. It becomes clear that different theories and approaches have mutual affinities and do complement each other. In addition, the affinities in different disciplines somewhat provide credence to the various views. From the combined approach described, it becomes apparent that by the semiosic process, the emerging semiotic self intertwined with the Umwelt, including emotions, can be described. Seeing the interpretation and meaning-making through semiosis allows for the analysis of groups, taking into account the embodied and emotional component. It is concluded that emotions have a crucial role in all human activity, including so-called reflective thinking, and that emotions and embodiment should be consciously taken into account in analysing signs, the interpretation, and in changes of signs and interpretations from both the social and individual level. The analysis of the beer labels expresses well the intertwined nature of the relationship between signs, individual consumers and society. Many direct influences from society on the label design are found, and also some indirect attitude changes that become apparent from magazines, company reports, etc. In addition, the analysis brings up the issues of the unifying tendency of the visual artefacts of different cultures, but also demonstrates that the visual artefacts are able to hold the local signs and meanings, and sometimes are able to represent the local meanings although the signs have changed in the unifying process.
  • Hämäläinen, Harri (Helsingin yliopisto, 2015)
    ABSTRACT The purpose of this study was to explore bone mineral content (BMC) and bone mineral density (BMD) development and related factors in patients with rheumatoid arthritis (RA) between 15 and 20 years from disease onset (I), in premenopausal women with RA (II), and in young adults with juvenile idiopathic arthritis (JIA) (III), and to ascertain osteoporosis (OP) drug use in patients with early RA (IV). BMD of the lumbar spine and the femoral neck were measured by dual-energy X-ray absorptiometry in patients with RA in two longitudinal studies and in young adults with JIA in a cross-sectional study. In assessing BMD at 15 years from disease onset in an inception cohort of RF-positive RA patients, it was found that eighteen out of 59 (31%) patients had OP. However, the decreases in central bone mineral in this patient group were of low degree and after the subsequent five years no essential change in central BMD was found. None of the explanatory variables: sex, age, ESR, HAQ, Larsen score, and cumulative prednisolone dose between 15-20 years from disease onset, proved to be a significant predictor of BMD change at the lumbar spine and femoral neck from 15- to 20-year check-ups (I). In assessment of BMC and BMD development in premenopausal, regularly cycling RA patients with and without GCs and in controls, it was found that RA patients with GCs had lower BMD values than those without GCs at commencement of follow-up. Furthermore, the mean BMD decreased significantly in both lumbar spine (P 0.002) and femoral neck (P smaller than 0.001) only in the RA patients with GCs during the 2-year follow-up. However, there was no statistically significant difference between the three groups in change in BMC or projectional area in the lumbar spine or femoral neck. Comparing results on bone mineral density change between the three groups it is relevant to report changes both in bone mineral content and in projectional area to clarify the basics of the bone mineral density change. BMD is expressed as BMC per projectional area. Only weight was found to be a significant predictor of BMD change (II). Assessment of BMC and BMD development in young adults with JIA and controls assumed to have reached their peak bone mass, showed that three (2.6%) out of 116 patients with JIA had OP. The male and female JIA patients had lower weight- and height-adjusted BMD values in the femoral neck than the controls. Dividing the patients into two groups, those with active and those with inactive JIA, both groups had lower BMC values in the femoral neck than the controls (P smaller than 0.001). Comparing BMC values in the femoral neck in both men and women with JIA a difference was found only among men (P 0.006). Among men, use of GCs and weight were significantly associated with BMC in the femoral neck. Among women, use of GCs, weight and also height were associated statistically significantly with BMC in the femoral neck, and among women GC use and height were also associated with BMC in the lumbar spine (III). A total of 14 878 incident cases of RA were studied when evaluating the implementation of pharmaceutical OP drug use. Out of this total, 1351 (9%) patients with RA had purchased OP drugs, mainly bisphosphonates, during the first year after commencement of antirheumatic treatment. Of GC users, 14% of women and 6% of men were prescribed OP drugs. In addition, 8% of females and 3% of males not taking GCs received OP medication. Women were more prone to use OP medication. Of the RA patients who took GCs, 38% of women and 24% of men received concomitant calcium and vitamin D preparations by prescription during the same year, whereas the corresponding percentages for patients without GCs were 21% and 13%. (IV). Study results suggest that bone loss takes place in earlier disease course in RA and JIA and bone loss is in the long-term disease course in RA of low degree. Further studies are needed to elucidate bone loss and OP development in early rheumatoid arthritis and to better focus the timing and means of OP prevention.
  • Arponen, Heidi (Helsingin yliopisto, 2012)
    Cranial base and craniocervical junction anatomy can be evaluated from CT and MR scans, and lateral skull radiographs. Cranial base anatomy changes during growth, as the form of the anatomic structures and their relative positions alter. In disorders of compromised bone quality, abnormal changes in the craniocervical junction can lead to pathological conditions with possibly life-threatening neurological complications. In this investigation these issues have first been addressed by making skull base measurements of healthy growing individuals to determine the age-specific normative values used to evaluate cranial base structures. These normative values were then employed in the following analysis of growing patients suffering from osteogenesis imperfecta (OI), a disorder of bone fragility due to abnormal collagen composition. In healthy individuals cranial base measurements, young children demonstrated significantly different results from those of older growing individuals and adults. Thus studies on patients younger than 9 years should include age-appropriate controls. Significant individual variation in skull base measurements during growth may appear, as revealed by longitudinal observation. The observed wide ranges, and the fact that the position of the odontoid process in relation to skull base structures may individually display alternating up- and downward movements, are important to consider when conducting follow-up examinations. Nevertheless, a notable deviation from the documented normal values is suggestive of pathological development in the craniocervical junction. When comparing the age-specific normal values, skull base abnormalities were found to be present in approximately one fifth of OI patients; platybasia was the most frequent finding. Patients with haploinsufficiency mutations (which lead to a relatively mild phenotype) were less likely to have skull base abnormalities than patients with helical glycine substitutions. However, the strongest predictor of skull base abnormality was not the type of collagen mutation underlying OI but the clinical severity of the disorder, as expressed by the height Z-score. No evidence was found for a protective effect of bisphosphonate treatment, nor for the progression of craniocervical junction pathology with age. Based on this normative data and the observations made in patients with OI, age-appropriate screening limits for skull base pathology are proposed here, in addition to a recommendation for timing of radiological screening in patients with OI.
  • Sheehy, Jatta (Helsingin yliopisto, 2015)
    Soils comprise more carbon (C) than any other terrestrial source and hence even a small change in the C content can be significant in regards to atmospheric CO2 concentration. Cultivated soils have lost soil organic carbon (SOC) during the latest decades in Finland. New cereal crop management practices, like no-till (NT) and reduced tillage (RT), can affect not only soil organic carbon (SOC) stocks and stabilization, but also nitrous oxide (N2O) emissions. The aim of this study was to gain better understanding about the changes in soil C dynamics and N2O emissions as a result of management practice changes in the boreal region, and the implications of these changes to climate change mitigation. Changes in SOC stocks and stabilization rates under different tillage (NT, RT, CT (conventional tillage with a moldboard plow)) and straw management (straw retention, straw removal, straw burning) practices, were studied at different sites with clayey and coarse textured soil across southern Finland. This was done by soil fractionation method (wet sieving and microaggregate isolation) to elucidate the composition of different soil fractions, namely large and small macroaggregates, microaggregates, silt and clay and macroaggregate-occluded soil fractions, and where the C is stored within them. The effects of Lumbricus terrestris on SOC were studied using the same method. Nitrous oxide fluxes were monitored biweekly for 2 years under CT, NT and RT practices using closed chambers. Measurements of several environmental and soil parameters were taken to study the underlying factors controlling the observed changes in soil C stocks and N2O emissions under the different management practices. Climate change mitigation potential through the studied cereal crop management practices seems small in the humid boreal region based on the results of this study. The minimum tillage treatments did not sequester SOC at any of the study sites which had been under NT or RT for a decade and the total C stocks were lower in the 0 15 cm topsoil layer at one clayey site under RT compared to CT after implementing RT for 30 years. All clayey sites had a fairly high SOC content originally and all sites had higher decomposition rate of crop residues in NT compared to CT, possibly hindering C sequestration. However, the aggregate stability was enhanced in NT cropping systems compared to CT, and NT increased the amount of SOC in large macroaggregates at several sites and in microaggregates within macroaggregates in the coarse textured site. L. terrestris mediated the formation of soil aggregates and the increase of SOC in the topsoil but possibly enhanced the decomposition rate in the soils. Cumulative N2O emissions were higher under NT compared to both CT and RT at the clayey sites and lower at the coarse textured site. However, the coarse textured site under NT received slightly less N fertilizer compared to CT. Increased N2O emissions under NT on clayey soils were likely due to denser soil structure with consistently higher soil moisture content and poor aeration. Therefore, mitigating N2O emissions requires special attention to soil structure and drainage. This study suggests that RT is a notable option to control N2O emissions. In the future, climate change could increase precipitation and freeze-thaw cycles in boreal agroecosystems possibly enhancing N2O fluxes and C losses of cultivated soils which puts pressure on finding new mitigation measures.
  • Ahlgrén-Rimpiläinen, Aulikki (Helsingin yliopisto, 2014)
    Aims and background: The purpose of this study was to investigate movement disturbances in schizophrenia and Restless Legs Syndrome (RLS) with the help of Transcranial magnetic stimulation (TMS) and Computerized force platform posturography (CFPP). Motor and postural controls are supported by central regulation of dopamine transmission. Dysregulation of dopamine is considered to be one of the background factors in schizophrenia and RLS. Methods: TMS and intramuscular recording electrodes were applied to study the function of descending motor pathways and the inhibitionary motor control. After stimulation of motor cortex motor evoked potentials, motor conduction time and central silent periods (CSP) were measured in the Abductor digiti minimi (ADM) and Tibialis anterior (TA) muscles. CFPP was used to investigate postural control. Center point of pressure forces (CPPF) and the center point of force velocity (CPFV) were measured during the subject`s stance eyes open and eyes closed on a stable computerized platform. Volunteers with schizophrenia having a long-term antipsychotic medication and volunteers with RLS were compared to healthy controls with these two methods. The CFPP in RLS study was repeated after a single day intervention with a dopaminergic agent. Findings: The function of the corticospinal motor pathways was normal in all subjects. Central inhibition was found to be disrupted into multiple separate CSPs. The number of CSPs was significantly higher in the dominant ADM in subjects with schizophrenia and in subjects with RLS compared to the controls. In subjects with schizophrenia the closing of the eyes had less impact on the CPFV than in controls. Subjects with RLS demonstrated lower sway velocity eyes open compared to controls. Pramipexole intervention balanced the CPFV differences. Conclusions: Motor inhibitory control was different in subjects with RLS and with schizophrenia compared to controls, appearing as repeated suppressions of muscle activity preferably in the dominant ADM. The ability of controlling the upright stance was not impaired per se in schizophrenia or in RLS. It was discovered a defect of visual compensation in schizophrenia, and in turn, a hyper compensatory effect of vision on postural control in RLS. The findings may refer to different feedback mechanisms in the CNS and to a shared subcortical CNS origin involved in the changed inhibitionary motor control and related dopamine dysregulation.
  • Virtanen, Pirjo Kristiina (Helsingin yliopisto, 2007)
    This thesis discusses the contemporary construction of the lived worlds of indigenous Amazonian youths. Today’s native peoples are considerably affected by the processes of globalization and urbanization, which have led to new ways of relating to their cultural traditions. This work presents a case study of Manchineri youngsters aged between 14 and 24 years old living in Acre state in Brazilian Amazonia. The Arawak-speaking Manchineri number some 1,000 people; their legally demarcated reserve is situated next to the River Yaco. The research is based on ethnographic material collected in the Mamoadate reserve and in the state capital, Rio Branco. By comparing the youth in different physical and social environments (the reserve and the city), my attempt has been to search for the most typical elements maintained, altered and created in the current lived worlds of Manchineri youths. Fieldwork methods included interviews, participant observation, photographs, video recordings, and drawings. The material was analyzed within the multidisciplinary framework of the social and cultural construction of knowledge. The study applies the concepts of social field, symbolic capital, and habitus as they have been used by Pierre Bourdieu; perspective as developed recently in Amazonian ethnology; the sacred as a cultural category as understood in the study of religion; and individual and person as concepts central to anthropology and sociology. Additionally, the study can be contextualized within youth studies, Latin American studies, and urban studies. The results of the study show that the everyday lives of young Amazonian native people are formed by a complex mixture of ‘modernity’ and ‘tradition’, fragmentation, and transitions between different conceptual frameworks. Part II discusses the ethnographic material in depth and shows that indigenous adolescents act from a variety of social perspectives: the native youth’s own ethnic group, divided into sub-groups, especially into urban residents and those living in the reserve; ancestors, super-human agents and spirits; other indigenous groups and non-natives. Consequently, besides the traditional initiation ritual, we find various contemporary rites of passage to adulthood: state-education, learning traditional practices, shamanism, matrimony, and transitions between the reserve and urban areas. According to these results, new social roles, political organization, responsibilities, and in general the desire to be respected, require both ‘modern’ and ‘traditional’ abilities. In Part III, the study shows that the current power relations constituted by new social contacts, ethnic recognition, and cooperation with different institutions have resulted in the formation of new social fields: youth cultures, the ethnic group, shamanic practices, the ethnopolitical movement, and indigenous students. The capacity of young Amazonian Indians to act in contemporary social fields produces them as full persons. The study also argues that the elements of the lived worlds can be divided into these social fields. When focusing on these fields, it became evident that these comprise the strategies adopted by young Indians to break through social and cultural barriers.
  • Koskinen, Inkeri (Helsingin yliopisto, 2015)
    This dissertation examines the effects of a change in the roles that extra-academic agents have in academic research: they are participating in the production of academic knowledge more actively than used to be the case. The focus is mainly on disciplines that fall under the umbrella of cultural research. Former informants are nowadays often becoming collaborators, co-researchers or co-authors in collaborative or participatory projects, or conducting activist research on their own cultures. Cultural research is in a unique position when facing the contemporary urge towards more "democratic" knowledge production. In disciplines such as anthropology, folkloristics and ethnology, researchers have always interacted with their informants' knowledge systems and developed research practices for approaching these systems. These practices are presently colliding with new demands that arise from the now common call for collaborative and participatory research. I focus on normative epistemic questions related to relativistic research practices and to objectivity. When the roles of the extra-academic agents change, the composition of research communities is also changed. An interactive notion of objectivity has recently been defended in social epistemology. It takes a research community as the unit whose objectivity is to be assessed. In the articles I identify shortcomings in the interactive objectivity of the emerging research communities, and develop analytical tools that can hopefully be of use in improving the situation. As the composition of research communities is changed, the established ways of approaching extra-academic knowledge systems also have to change. The moderately relativistic practice of avoiding the appraisal of alien knowledge systems is no longer as practicable as it used to be. When former informants join research teams in participatory projects, or indigenous activists become activist researchers, they become part of communities whose interactive objectivity can and should be assessed. This is the case even if some or all of the members of the communities are taken to represent, or see themselves as representing, extra-academic knowledge systems. The contributions of everyone belonging to a research community must be met with the same critical attitude, or the objectivity of the community will suffer.
  • Hoppu, Pekka (Helsingin yliopisto, 2008)
    The number of drug substances in formulation development in the pharmaceutical industry is increasing. Some of these are amorphous drugs and have glass transition below ambient temperature, and thus they are usually difficult to formulate and handle. One reason for this is the reduced viscosity, related to the stickiness of the drug, that makes them complicated to handle in unit operations. Thus, the aim in this thesis was to develop a new processing method for a sticky amorphous model material. Furthermore, model materials were characterised before and after formulation, using several characterisation methods, to understand more precisely the prerequisites for physical stability of amorphous state against crystallisation. The model materials used were monoclinic paracetamol and citric acid anhydrate. Amorphous materials were prepared by melt quenching or by ethanol evaporation methods. The melt blends were found to have slightly higher viscosity than the ethanol evaporated materials. However, melt produced materials crystallised more easily upon consecutive shearing than ethanol evaporated materials. The only material that did not crystallise during shearing was a 50/50 (w/w, %) blend regardless of the preparation method and it was physically stable at least two years in dry conditions. Shearing at varying temperatures was established to measure the physical stability of amorphous materials in processing and storage conditions. The actual physical stability of the blends was better than the pure amorphous materials at ambient temperature. Molecular mobility was not related to the physical stability of the amorphous blends, observed as crystallisation. Molecular mobility of the 50/50 blend derived from a spectral linewidth as a function of temperature using solid state NMR correlated better with the molecular mobility derived from a rheometer than that of differential scanning calorimetry data. Based on the results obtained, the effect of molecular interactions, thermodynamic driving force and miscibility of the blends are discussed as the key factors to stabilise the blends. The stickiness was found to be affected glass transition and viscosity. Ultrasound extrusion and cutting were successfully tested to increase the processability of sticky material. Furthermore, it was found to be possible to process the physically stable 50/50 blend in a supercooled liquid state instead of a glassy state. The method was not found to accelerate the crystallisation. This may open up new possibilities to process amorphous materials that are otherwise impossible to manufacture into solid dosage forms.
  • Niemi, Jarkko (Helsingin yliopisto, 2007)
    Aerosol particles can cause detrimental environmental and health effects. The particles and their precursor gases are emitted from various anthropogenic and natural sources. It is important to know the origin and properties of aerosols to efficiently reduce their harmful effects. The diameter of aerosol particles (Dp) varies between ~0.001 and ~100 μm. Fine particles (PM2.5: Dp < 2.5 μm) are especially interesting because they are the most harmful and can be transported over long distances. The aim of this thesis is to study the impact on air quality by pollution episodes of long-range transported aerosols affecting the composition of the boundary-layer atmosphere in remote and relatively unpolluted regions of the world. The sources and physicochemical properties of aerosols were investigated in detail, based on various measurements (1) in southern Finland during selected long-range transport (LRT) pollution episodes and unpolluted periods and (2) over the Atlantic Ocean between Europe and Antarctica during a voyage. Furthermore, the frequency of LRT pollution episodes of fine particles in southern Finland was investigated over a period of 8 years, using long-term air quality monitoring data. In southern Finland, the annual mean PM2.5 mass concentrations were low but LRT caused high peaks of daily mean concentrations every year. At an urban background site in Helsinki, the updated WHO guideline value (24-h PM2.5 mean 25 μg/m3) was exceeded during 1-7 LRT episodes each year during 1999-2006. The daily mean concentrations varied between 25 and 49 μg/m3 during the episodes, which was 3-6 times higher than the mean concentration in the long term. The in-depth studies of selected LRT episodes in southern Finland revealed that biomass burning in agricultural fields and wildfires, occurring mainly in Eastern Europe, deteriorated air quality on a continental scale. The strongest LRT episodes of fine particles resulted from open biomass-burning fires but the emissions from other anthropogenic sources in Eastern Europe also caused significant LRT episodes. Particle mass and number concentrations increased strongly in the accumulation mode (Dp ~ 0.09-1 μm) during the LRT episodes. However, the concentrations of smaller particles (Dp < 0.09 μm) remained low or even decreased due to the uptake of vapours and molecular clusters by LRT particles. The chemical analysis of individual particles showed that the proportions of several anthropogenic particle types increased (e.g. tar balls, metal oxides/hydroxides, spherical silicate fly ash particles and various calcium-rich particles) in southern Finland during an LRT episode, when aerosols originated from the polluted regions of Eastern Europe and some open biomass-burning smoke was also brought in by LRT. During unpolluted periods when air masses arrived from the north, the proportions of marine aerosols increased. In unpolluted rural regions of southern Finland, both accumulation mode particles and small-sized (Dp ~ 1-3 μm) coarse mode particles originated mostly from LRT. However, the composition of particles was totally different in these size fractions. In both size fractions, strong internal mixing of chemical components was typical for LRT particles. Thus, the aging of particles has significant impacts on their chemical, hygroscopic and optical properties, which can largely alter the environmental and health effects of LRT aerosols. Over the Atlantic Ocean, the individual particle composition of small-sized (Dp ~ 1-3 μm) coarse mode particles was affected by continental aerosol plumes to distances of at least 100-1000 km from the coast (e.g. pollutants from industrialized Europe, desert dust from the Sahara and biomass-burning aerosols near the Gulf of Guinea). The rate of chloride depletion from sea-salt particles was high near the coasts of Europe and Africa when air masses arrived from polluted continental regions. Thus, the LRT of continental aerosols had significant impacts on the composition of the marine boundary-layer atmosphere and seawater. In conclusion, integration of the results obtained using different measurement techniques captured the large spatial and temporal variability of aerosols as observed at terrestrial and marine sites, and assisted in establishing the causal link between land-bound emissions, LRT and air quality.
  • Salusjärvi, Tuomas (Helsingin yliopisto, 2006)
    The work covered in this thesis is focused on the development of technology for bioconversion of glucose into D-erythorbic acid (D-EA) and 5-ketogluconic acid (5-KGA). The task was to show on proof-of-concept level the functionality of the enzymatic conversion or one-step bioconversion of glucose to these acids. The feasibility of both studies to be further developed for production processes was also evaluated. The glucose - D-EA bioconversion study was based on the use of a cloned gene encoding a D-EA forming soluble flavoprotein, D-gluconolactone oxidase (GLO). GLO was purified from Penicillium cyaneo-fulvum and partially sequenced. The peptide sequences obtained were used to isolate a cDNA clone encoding the enzyme. The cloned gene (GenBank accession no. AY576053) is homologous to the other known eukaryotic lactone oxidases and also to some putative prokaryotic lactone oxidases. Analysis of the deduced protein sequence of GLO indicated the presence of a typical secretion signal sequence at the N-terminus of the enzyme. No other targeting/anchoring signals were found, suggesting that GLO is the first known lactone oxidase that is secreted rather than targeted to the membranes of the endoplasmic reticulum or mitochondria. Experimental evidence supports this analysis, as near complete secretion of GLO was observed in two different yeast expression systems. Highest expression levels of GLO were obtained using Pichia pastoris as an expression host. Recombinant GLO was characterised and the suitability of purified GLO for the production of D-EA was studied. Immobilised GLO was found to be rapidly inactivated during D-EA production. The feasibility of in vivo glucose - D-EA conversion using a P. pastoris strain co-expressing the genes of GLO and glucose oxidase (GOD, E.C. of A. niger was demonstrated. The glucose - 5-KGA bioconversion study followed a similar strategy to that used in the D-EA production research. The rationale was based on the use of a cloned gene encoding a membrane-bound pyrroloquinoline quinone (PQQ)-dependent gluconate 5-dehydrogenase (GA 5-DH). GA 5-DH was purified to homogeneity from the only source of this enzyme known in literature, Gluconobacter suboxydans, and partially sequenced. Using the amino acid sequence information, the GA 5-DH gene was cloned from a genomic library of G. suboxydans. The cloned gene was sequenced (GenBank accession no. AJ577472) and found to be an operon of two adjacent genes encoding two subunits of GA 5-DH. It turned out that GA 5-DH is a rather close homologue of a sorbitol dehydrogenase from another G. suboxydans strain. It was also found that GA 5-DH has significant polyol dehydrogenase activity. The G. suboxydans GA 5-DH gene was poorly expressed in E. coli. Under optimised conditions maximum expression levels of GA 5-DH did not exceed the levels found in wild-type G. suboxydans. Attempts to increase expression levels resulted in repression of growth and extensive cell lysis. However, the expression levels were sufficient to demonstrate the possibility of bioconversion of glucose and gluconate into 5-KGA using recombinant strains of E. coli. An uncharacterised homologue of GA 5-DH was identified in Xanthomonas campestris using in silico screening. This enzyme encoded by chromosomal locus NP_636946 was found by a sequencing project of X. campestris and named as a hypothetical glucose dehydrogenase. The gene encoding this uncharacterised enzyme was cloned, expressed in E. coli and found to encode a gluconate/polyol dehydrogenase without glucose dehydrogenase activity. Moreover, the X. campestris GA 5-DH gene was expressed in E. coli at nearly 30 times higher levels than the G. suboxydans GA 5-DH gene. Good expressability of the X. campestris GA-5DH gene makes it a valuable tool not only for 5-KGA production in the tartaric acid (TA) bioprocess, but possibly also for other bioprocesses (e.g. oxidation of sorbitol into L-sorbose). In addition to glucose - 5-KGA bioconversion, a preliminary study of the feasibility of enzymatic conversion of 5-KGA into TA was carried out. Here, the efficacy of the first step of a prospective two-step conversion route including a transketolase and a dehydrogenase was confirmed. It was found that transketolase convert 5-KGA into TA semialdehyde. A candidate for the second step was suggested to be succinic dehydrogenase, but this was not tested. The analysis of the two subprojects indicated that bioconversion of glucose to TA using X. campestris GA 5-DH should be prioritised first and the process development efforts in future should be focused on development of more efficient GA 5-DH production strains by screening a more suitable production host and by protein engineering.
  • Blom, Titta S. (Helsingin yliopisto, 2003)
  • Vainio, Ulla (Helsingin yliopisto, 2007)
    Wood is an important material for the construction and pulping industries. Using x-ray diffraction the microfibril angle of Sitka spruce wood was studied in the first part of this thesis. Sitka spruce (Picea sitchensis [Bong.] Carr.) is native to the west coast of North America, but due to its fast growth rate, it has also been imported to Europe. So far, its nanometre scale properties have not been systematically characterised. In this thesis the microfibril angle of Sitka spruce was shown to depend significantly on the origin of the tree in the first annual rings near the pith. Wood can be further processed to separate lignin from cellulose and hemicelluloses. Solid cellulose can act as a reducer for metal ions and it is also a porous support for nanoparticles. By chemically reducing nickel or copper in the solid cellulose support it is possible to get small nanoparticles on the surfaces of the cellulose fibres. Cellulose supported metal nanoparticles can potentially be used as environmentally friendly catalysts in organic chemistry reactions. In this thesis the size of the nickel and copper containing nanoparticles were studied using anomalous small-angle x-ray scattering and wide-angle x-ray scattering. The anomalous small-angle x-ray scattering experiments showed that the crystallite size of the copper oxide nanoparticles was the same as the size of the nanoparticles, so the nanoparticles were single crystals. The nickel containing nanoparticles were amorphous, but crystallised upon heating. The size of the nanoparticles was observed to be smaller when the reduction of nickel was done in aqueous ammonium hydrate medium compared to reduction made in aqueous solution. Lignin is typically seen as the side-product of wood industries. Lignin is the second most abundant natural polymer on Earth, and it possesses potential to be a useful material for many purposes in addition to being an energy source for the pulp mills. In this thesis, the morphology of several lignins, which were produced by different separation methods from wood, was studied using small-angle and ultra small-angle x-ray scattering. It was shown that the fractal model previously proposed for the lignin structure does not apply to most of the extracted lignin types. The only lignin to which the fractal model could be applied was kraft lignin. In aqueous solutions the average shape of the low molar mass kraft lignin particles was observed to be elongated and flat. The average shape does not necessarily correspond to the shape of the individual particles because of the polydispersity of the fraction and due to selfassociation of the particles. Lignins, and especially lignosulfonate, have many uses as dispersants, binders and emulsion stabilisers. In this thesis work the selfassociation of low molar mass lignosulfonate macromolecules was observed using small-angle x-ray scattering. By taking into account the polydispersity of the studied lignosulfonate fraction, the shape of the lignosulfonate particles was determined to be flat by fitting an oblate ellipsoidal model to the scattering intensity.
  • Koskinen, Kaisa (Helsingin yliopisto, 2013)
    Microbes are essential for all life on Earth. They are found in all viable habitats from deep sea sediments and bedrock to high up in the atmosphere with a variety that exceeds by far the eukaryotic diversity. Ecosystem services provided by microorganisms, such as degradation of organic material and mediation of biogeochemical cycles are fundamentally important for the whole biosphere and its inhabitants. Microbes also form symbiotic relationships with multicellular organisms, and play important roles in nutrition and disease. Recent developments in molecular techniques, especially the next generation sequencing technologies and microarray applications, have opened new possibilities in studying diverse microbial communities. In this thesis, the aim was to determine the diversity and community structure of environmental samples collected from the northern Baltic Sea water column and anaerobic digestion reactor, and to assess how the prevailing abiotic factors affect the microbial community structure. We applied 16S rRNA and ITS gene amplicon sequencing method with 454 sequencing technology to form a detailed taxonomic description of studied communities. The produced sequence data was further utilised in designing probes for a new padlock probe based ligation detection reaction (LDR) microarray that could be employed for specific and sensitive taxonomic identification of microbial groups in diverse communities. The functionality, specificity and sensitivity of the microarray were assessed using artificial and real environmental samples. Additionally, selected amplicon sequencing data analysis methods were compared in order to discover which algorithms work most reliably. In this subproject, we aimed to clarify how significantly the selected analysis methods, specifically denoising and clustering algorithms, affect the results and how comparable the results derived from different analysis pipelines are. Amplicon sequencing revealed diverse microbial communities in the northern Baltic Sea water column and anaerobic digestion reactor. The pelagic bacterial communities in the northern Baltic Sea were strongly stratified, with aerobic Bacteria such as Pseudomonas and Flavobacterium dominating in the surface layer and Oleispira and sulfate-reducing bacteria in the anoxic deep waters. Based on the sequence data the diversity was assessed one order of magnitude less diverse compared to Atlantic and Pacific ocean bacterial communities. The anaerobic digestion reactor communities were dominated by Bacteria belonging to phyla Bacteroidetes, Firmicutes and Thermotogae and methanogenic Archaea, all essential and typical degraders in anaerobic digestion. The process also supported a diverse fungal community of phyla Ascomycota and Basidiomycota, including several taxa capable of degrading organic material in anaerobic conditions. The LDR microarray technology proved sensitive, specific and semiquantitative method for identifying microbes in diverse communities. The proof of principle tests and experiments with real environmental samples showed that if the probes are designed carefully, the detection is comparable to qPCR and amplicon sequencing. The detection limit was 0.01 fmol/µl/template. Data analysis method comparisons revealed prominent differences in observed operational taxonomic units and relative abundance of identified taxa. The majority of tested methods assessed the species richness too high. Using a functioning denoising method evened out the differences in the number of observed OTUs caused by various clustering algorithms. The ability to filter out the spurious taxa produced by amplification and sequencing, but still retain all the real diversity varied between methods. This study shows both the potential and the challenges in the use of amplicon sequencing and microarray technologies in studying diverse microbial communities. The results indicate that the padlock based LDR microarray can be designed for very accurate and sensitive identification of microbial groups of interest. The data suggest that amplicon sequencing is a powerful tool in identifying microbes and assessing the diversity but distinguishing between spurious and true community members remain a challenge. There is still work to be done in the development and application of data analysis tools.
  • Sarén, Matti-Paavo (Helsingin yliopisto, 2006)
    In recent years there has been growing interest in selecting suitable wood raw material to increase end product quality and to increase the efficiency of industrial processes. Genetic background and growing conditions are known to affect properties of growing trees, but only a few parameters reflecting wood quality, such as volume and density can be measured on an industrial scale. Therefore research on cellular level structures of trees grown in different conditions is needed to increase understanding of the growth process of trees leading to desired wood properties. In this work the cellular and cell wall structures of wood were studied. Parameters, such as the mean microfibril angle (MFA), the spiral grain angles, the fibre length, the tracheid cell wall thickness and the cross-sectional shape of the tracheid, were determined as a function of distance from the pith towards the bark and mutual dependencies of these parameters were discussed. Samples from fast-grown trees, which belong to a same clone, grown in fertile soil and also from fertilised trees were measured. It was found that in fast-grown trees the mean MFA decreased more gradually from the pith to the bark than in reference stems. In fast-grown samples cells were shorter, more thin-walled and their cross-sections were rounder than in slower-grown reference trees. Increased growth rate was found to cause an increase in spiral grain variation both within and between annual rings. Furthermore, methods for determination of the mean MFA using x-ray diffraction were evaluated. Several experimental arrangements including the synchrotron radiation based microdiffraction were compared. For evaluation of the data analysis procedures a general form for diffraction conditions in terms of angles describing the fibre orientation and the shape of the cell was derived. The effects of these parameters on the obtained microfibril angles were discussed. The use of symmetrical transmission geometry and tangentially cut samples gave the most reliable MFA values.
  • Rantakari, Anssi (Helsingin yliopisto, 2005)