Human maltase-glucoamylase (MGAM) and sucrase-isomaltase (SI) are the small-intestinal glucosidases responsible for catalyzing the last glucose-releasing step in starch digestion. MGAM and SI are each composed of duplicated catalytic domains, N- and C-terminal, which display complementary substrate specificities for the mixture of short linear and branch oligosaccharide substrates that typically make up terminal starch digestion products. As MGAM and SI are involved in post-prandial glucose production, regulating their activities with α-glucosidase inhibitors is an attractive approach to controlling blood glucose levels for the prevention and treatment of Type 2 diabetes. To better understand the complementary activities and mechanism of inhibition of these intestinal glucosidases, this thesis aims to characterize the individual N- and C-terminal MGAM and SI domains using a combination of X-ray crystallographic structural studies, enzyme kinetics, and inhibitor studies. First, the structure of the N-terminal domain of MGAM (ntMGAM) was determined in its apo form and in complex with the inhibitor acarbose. In addition to sequence alignments and kinetics studies, the structures provide insight into the preference of the N-terminal MGAM domain for short linear substrates and the C-terminal domain for longer substrates. Second, the structure of ntMGAM was determined in complex with various α-glucosidase inhibitors, including those currently on the market (acarbose and miglitol), a new class of inhibitors from natural extracts of Salacia reticulata (salacinol, kotalanol and de-O-sulfonated kotalanol) and chemically synthesized derivatives of salacinol. These studies reveal the features of the Salacia reticulata inhibitors that are essential for inhibitory activity and highlight their potential as future drug candidates. Third, the crystal structure of the N-terminal domain of SI (ntSI) was determined in apo-form and in complex with kotalanol. Structural comparison of ntSI and ntMGAM reveal key differences in active site architectures, which are proposed to confer differential substrate specificity.

L'objectif de ce travail est de présenter un modèle générique de la digestion dans l'intestin grêle. Dans la première partie de ce travail, un modèle mécaniste basé sur les équations différentielles ordinaires est utilisé pour présenter la digestion : les équations décrivent l'évolution de la position et de la composition du bolus provenant de l'estomac. Chaque bolus est représenté par un cylindre. Ce modèle prend en compte simultanément les différents aspects de la digestion à savoir le transport du bolus dans la lumière intestinale, la dégradation des aliments par des enzymes, et l'absorption des nutriments. Dans la deuxième partie de cette étude, nous utilisons les méthodes d'homogénéisation mathématique pour justifier le modèle de la digestion développé dans la première partie. Nous montrons que ce modèle peut être considéré comme une version macroscopique des modèles plus réalistes, qui contiennent des phénomènes biologiques à des échelles inférieures de l'intestin grêle : (i) les ondes péristaltiques à haute fréquence (échelle du temps microscopique) et leurs effets sur la vitesse du bolus, (ii) la présence des villosités intestinales (échelle microscopique de l'espace) et leur influence sur la digestion. Enfin, dans la troisième partie de ce travail, nous étudions l'influence du changement de la structure de bolus sur la digestion en intégrant les fibres alimentaires dans sa composition. Les deux principales caractéristiques des fibres alimentaires qui interagissent avec la fonction de l'intestin grêle, à savoir, la viscosité et la capacité de rétention d'eau ont été modélisées. Ce modèle nous a permis de considérer, en particulier, la relation entre l'évolution de la matière sèche et de l'eau au sein du bolus. Bien que ce modèle est générique et contient un grand nombre de paramètres, à notre connaissance, il est parmi les premiers modèles dynamiques qualitatives de l'influence des fibres sur la digestion intestinale.

This paper reports on environmental impacts and health hazards as a result from inadequate management of organic household waste in developing countries. It gives details of water and soil contamination, air pollution and spread of diseases through expanding breeding grounds for pathogens, vectors and rodents. To manage this waste flow sustainably, decentralised composting and anaerobic digestions technologies have been studied to give an overall picture of existing appropriate technologies, including: windrow, box/bin/barrel, THM, aerated static pile, in-vessel, vermi, ARTI compact biogas digester and BARC’S NISARG-RUNA. Comparing different technologies showed that it is crucial to consider local conditions and markets when choosing which method to implement. However, the manual composting methods: windrow/box/bin/barrel, THM and vermi, are more likely to be appropriate in regard to current conditions in developing countries. A comparison between the environmental impacts of anaerobic digestion and composting did not result in a clear indication of which technology is most favourable. However, in the literature studied, biogas production showed an overall better energy balance, and composting a better result regarding nutrient recycling and xenobiotic compounds. In terms of the mitigation effect on global warming, the results varied essentially depending on the technology used and its loss of methane during the biogas production process. Finally, this paper investigates common constraints for implementation of the above-mentioned technologies, as well as recommendations for future projects. The study of general constraints revealed the need for directing attention to education, key consequences and benefits, co-operation, exchange of knowledge and bottom-up driving forces, for sustainable and successful implementation of organic household waste management practices in developing countries.

The finite amount of energy carriers affects all of us. It is important to utilize all available sources and to find new sources of energy. The confectionery industry generates both solid and liquid waste during the production of confectioneries, which could be utilized as a substrate for biogas production. However, problems might arise during the biogas process since these kinds of waste are very rich in carbohydrates. The initial breakdown of the substrate would probably cause an accumulation of fermentation products such as volatile fatty acids (VFA) and a low pH. A solution to this might be to use a two-stage process. The first stage would be a pre-fermentation that should be optimized for production of fermentation products such as ethanol and VFA. The aim of this master thesis is to evaluate the biogas potential of confectionery waste products. The confectionery waste products are evaluated using a continuous two-stage process, batch experiments and theoretical calculations of the methane potential. The potential from process wastewater was examined. Depending on COD reduction for a reactor and COD content of process wastewater, an annual amount of 75 000 m3 or of 857 000 m3 of process wastewater is necessary to produce enough biogas for a gas engine to continuously convert the biogas to electricity. A batch experiment evaluating the methane production potential of nine different confectionery waste products from a large confectionery industry gave a range of 430 - 690 NmL/g VS, which is relatively high. A continuous experiment in two lab-scale reactors with a HRT of two days worked satisfactory. The gas production was stable periodically with a carbon dioxide content above 60%. The pH was low (3.4 - 3.6) throughout the experiment for one of the reactors. However, addition of digester sludge from a methane-producing reactor towards the end of the experiment resulted in a higher pH and more VFA available for utilization in the second stage. The main fermentation products were: acetic acid, lactic acid, ethanol and carbon dioxide. A second batch experiment showed that the methane potential was not affected by pre-fermentation. A carbon balance calculation of the process indicates that 57% of the ingoing organic matter is fermented within only two days and ends up in the known fermentation products. The study shows that confectionery waste products are well suited for two-stage anaerobic digestion. / Den ändliga mängden av energibärare påverkar oss alla. Det är därför viktigt att utnyttja alla tillgängliga men även att finna nya energibärare. Konfektyrindustrin generar restprodukter (avfall) både i fast och flytande form, vilka båda kan utnyttjas för produktion av biogas. Det kan dock uppstå problem i biogasprocessen eftersom dessa innehåller en stor mängd kolhydrater. Den initiala nedbrytningen kan ge upphov till en ackumulering av VFA och ett lågt pH. En lösning på detta problem kan vara att använda en två-stegs process. Första steget är en för-fermentering, som ska optimeras för att producera fermentationsprodukter så som etanol och VFA. Syftet med detta arbete är att utvärdera biogaspotentialen från konfektyrrestprodukter genom att använda en kontinuerlig två-stegs process, batchförsök och teoretiska beräkningar av metanpotentialen. Potentialen undersöktes från processvatten. Beroende på COD reduktion i en reaktor och COD innehåll i processvatten, är en årlig mängd av 75 000 m3 eller av 857 000 m3 processvatten nödvändig för att producera en tillräcklig mängd biogas så att en gasmotor kontinuerligt kan omvandla biogasen till elektricitet. Ett batchförsök med nio olika restprodukter från en storskalig konfektyrproducent visade en relativt hög metanpotential (430 - 690 NmL/g VS). Ett kontinuerligt reaktorexperiment genomfördes i laboratorieskala med två reaktorer, där uppehållstiden var två dagar. De två för-fermenteringsreaktorerna presterade tillfredsställande. Gasproduktionen var periodvis stabil med en koldioxidhalt över 60%. pH var lågt (3,4 och 3,6) genom hela experimentet för en av reaktorerna. För den andra reaktorn gjordes tillsatser av reaktormaterial från en metanproducerande reaktor i slutet av experimentet. Dessa tillsatser ökade pH och totalmängden av VFA, som kan utnyttjas i det andra steget. Huvudfermentationsprodukterna är acetat, laktat, etanol och koldioxid. Ytterligare batchförsök visade att för-fermentation inte verkar påverka metanpotentialen för konfektyrrestprodukter. En kolbalans av processen indikerar att 57% av ingående kol återfinns i de kända fermentationsprodukterna inom två dagar. Studiens resultat visar att avfallsprodukter från konfektyrindustrin lämpar sig väl för två-stegs anaerob rötning.

Med ett växande energibehov i världen, sinande energikällor i form av fossila bränslen och en miljö som vi under en längre tid har förorenat behövs det nya energiformer som är mer långsiktiga och framförallt miljövänliga. En sådan energiform är biogas. Biogasprocessen är dock inte helt optimerad. Flera av de substrat som används idag tar lång tid att röta och bryts bara ner till viss del i processen eller innehåller onödigt mycket vatten, vilket ger höga transportkostnader. Med syfte att göra biogasprocessen mer ekonomisk lönsam utvärderas i denna rapport på uppdrag från E.ON några olika förbehandlingstekniker: Ångexplosion, extrusion, avvattning och kalkbehandling. Förhoppningen är att dessa ska kunna öka lönsamheten för storskalig biogasproduktion och kanske möjliggöra biogasproduktion från tidigare obrukbara substrat som fjädrar och halm.  För att jämföra och utvärdera förbehandlingsteknikerna utfördes batchrötningsförsök i 330 ml flaskor med obehandlade och förbehandlade substrat. De flesta förbehandle substraten gav snabbare nedbrytning och några gav även högre metanutbyte än de obehandlade. Fjädrar och halm, som från början hade ett lågt utbyte, påverkades mest av förbehandlingen. Ångexploderade fjädrar gav efter 44 dagars rötning 141% högre metanutbyte och extruderad halm gav 22% högre metanutbyte än obehandlad.  För ekonomiska beräkningarna användes en referensanläggning med en förutbestämd substratmix: 12500 ton majs och 11500 ton hästgödsel. Att tillgå för referensanläggningen finns dessutom fjädrar. Cambis THP-anläggning för ångexplosion visade sig vara alldeles för dyr för referensanläggningen. En THP-anläggning kräver en större biogasanläggning där en större mängd svårnedbrytbara substrat rötas för att bli lönsam. En extruder skulle kunna vara lönsam för för refernsanläggningen om hästgödseln som de har tillgång till innehåller halm som strömaterial. En investering i en extruder bara för att förbehandla majsensilage visade sig inte lönsam.  Avvattning av gödsel gav signifikant lägre utbyte av biogas per torrvikt men signifikant högre utbyte per våtvikt. Avvattningsutrustningen från Splitvision, som testades, var för dyr för att bli lönsam. Först när gården låg 4 mil från biogasanläggningen blev det billigare att avvattna gödsel och transportera den jämfört med att transportera den obehandlad. Andra avvattningsutrustningar i studien var billigare i drift så det finns möjligheter att tekniken kan bli lönsam med någon av dessa. / The world needs new energy sources that are durable for long time and which not affect the environment negatively. Biogas fulfills those demands. The biogas process is however not completely optimized. Several of the substrates used today for biogas production are slowly degraded and only partly digested in the process. Other substrates consist of unnecessarily much water which makes transportation costly. To optimize the process and make the biogas process more profitable, several pretreatment techniques are evaluated by direction of E.ON in this report: steam explosion, extrusion, lime treatment and dewatering. The hope is that one of those could increase the profitability and hopefully also enable substrates that not are working today like feathers and straw. To compare and evaluate the different pretreatment batch digester, experiments were carried out during 31-44 days for untreated and pretreated substrates. Most pretreated substrates were faster degraded than untreated and some also gave a higher methane yield. Chicken waste feathers and wheat straw, which had low methane yields untreated, were affected most by pretreatment. Steam exploded feathers gave after 44 days of digestion 141% higher methane yield and extruded straw gave 22% higher methane yield than untreated samples of the same substrate. A reference plant with a substrate mixture of 12500 tonnes of maize silage and 11500 tons of horsemanure annually was used to make economical calculations. Additionally, chicken waste feathers waste could be included. Obtainable for the reference plant were also chicken waste feathers. Steam explosion appeared to be too expensive for a plant in the size of the reference plant. Its large capacity could probably make it profitable for a much larger biogas plant running on a lot of hard digestible substrates. An extruder could be a profitable investment for the reference plant if the plant gets horse manure with straw as bedding material. To just use the extruder to pretreat maize silage could not make the investment profitable. Dewatering of manure gave significantly lower methane yield per dry weight but significantly higher methane yield per wet weight. The increase in methane yield per wet weight makes the substrate better for transportation. The dewatering equipment from Splitvision tried in this study had too high operational costs and was too expensive to make dewatering particularly profitable. Only when the farm was situated farther away than 40km from the biogas plant it was cheaper to dewater the manure before transport than to transport the manure without any pretreatment. Other dewatering equipments evaluated in this study had much lower operational costs and among those an equipment that makes dewatering profitable might therefore be found. The world needs new energy sources that are durable for long time and which not affect the environment negatively. Biogas fulfills those demands. The biogas process is however not completely optimized. Several of the substrates used today for biogas production are slowly degraded and only partly digested in the process. Other substrates consist of unnecessarily much water which makes transportation costly. To optimize the process and make the biogas process more profitable, several pretreatment techniques are evaluated by direction of E.ON in this report: steam explosion, extrusion, lime treatment and dewatering. The hope is that one of those could increase the profitability and hopefully also enable substrates that not are working today like feathers and straw. To compare and evaluate the different pretreatment batch digester, experiments were carried out during 31-44 days for untreated and pretreated substrates. Most pretreated substrates were faster degraded than untreated and some also gave a higher methane yield. Chicken waste feathers and wheat straw, which had low methane yields untreated, were affected most by pretreatment. Steam exploded feathers gave after 44 days of digestion 141% higher methane yield and extruded straw gave 22% higher methane yield than untreated samples of the same substrate. A reference plant with a substrate mixture of 12500 tonnes of maize silage and 11500 tons of horsemanure annually was used to make economical calculations. Additionally, chicken waste feathers waste could be included. Obtainable for the reference plant were also chicken waste feathers. Steam explosion appeared to be too expensive for a plant in the size of the reference plant. Its large capacity could probably make it profitable for a much larger biogas plant running on a lot of hard digestible substrates. An extruder could be a profitable investment for the reference plant if the plant gets horse manure with straw as bedding material. To just use the extruder to pretreat maize silage could not make the investment profitable. Dewatering of manure gave significantly lower methane yield per dry weight but significantly higher methane yield per wet weight. The increase in methane yield per wet weight makes the substrate better for transportation. The dewatering equipment from Splitvision tried in this study had too high operational costs and was too expensive to make dewatering particularly profitable. Only when the farm was situated farther away than 40km from the biogas plant it was cheaper to dewater the manure before transport than to transport the manure without any pretreatment. Other dewatering equipments evaluated in this study had much lower operational costs and among those an equipment that makes dewatering profitable might therefore be found.

Produktions- och optimeringspotentialen vid anaerob rötning av flytgödsel från nötkreatur granskades. Detta genom att, i laboratoriemiljö, undersöka förändringar i processtabilitet, gasproduktion och metanhalt vid tillsats av kycklinggödsel och pH-buffert med flytgödsel från nötkreatur som grundsubstrat. Tillsatsen av pH-buffert underlättade tillväxtfasen för mikroorganismerna, snabbare ökning i metanhalt och den totala metangasproduktionen ökade. Skillnaden var som störst under tillväxtfasen och avtog från stationärfas och framåt. Tillsats av kycklinggödsel har i detta fall inhiberat processen, möjligtvis på grund av ammoniakinhibering vid tillsats av det kväverika substratet som kycklinggödsel är. / Production potential and basis for optimization of anaerobic digestion with cattle manure was reviewed by examining process stability, gas production and methane content in a laboratory environment via addition of pH buffer and chicken manure with slurry from cattle manure as basic feedstock. Addition of pH buffer facilitated the growth of microorganisms by more rapid increase in methane content and greater quantities of produced methane gas. The effects were high during the growth stage but decreased, during and after, the stationary phase. Addition of chicken manure caused inhibition of the process, probably caused by ammonia inhibition from high nitrogen content in chicken manure.

Ultrasonic sludge disintegration &ndash / the most commonly used mechanical pretreatment method- enables the occurrence of cavitation bubbles to extract intracellular material from the cell into aqueous phase. However, there is a lack of information on the volatile solids loading on the anaerobic digestion process performance of ultrasonically treated sludge. In this thesis work, the effect of sonication on disintegration of waste activated sludge (WAS) and an important parameter digester F/M (food to microorganism) ratio on ultrasonically treated WAS were investigated. First, preliminary studies were conducted. It was obtained that when the sonication power and time increased, soluble COD in the supernatant increased as well. Then, batch anaerobic digestion tests were conducted. Effect of F/M ratio in the digesters by using sonicated sludges at different powers was analyzed. For the sludge sonicated at high power, the methane content increased up to 55.1 % at F/M ratio of 10 compared to untreated sludge. On the other hand, methane generation rate slowed down with the increase in F/M ratio. Moreover, 10 % and 15 % increase in the destruction of MLVSS and total COD content was observed for sonicated sludges compared to the untreated sludges, respectively. In summary, both the sonication as a pretreatment method and the increase in digester F/M ratio increased the biogas production and the solids reduction during anaerobic digestion prosess. These results may have important implications for the operation of full scale systems in terms of system efficiency and operation.

In the last few decades, numerous studies have been conducted on xenobiotic compounds due to their vast use in industries, households, etc. and consequently high exposure of these compounds. The main focus of this study is nonylphenol compounds such as nonylphenol monoethoxylate (NP1EO), nonylphenol diethoxylate (NP2EO), nonylphenoxy acetic acid (NP1EC) and nonylphenol (NP), which are among the harmful xenobiotic compounds that can cause endocrine disruption, cancer and other health problems and which are used widely in the production of surfactants and personal care products. In this study, laboratory scale aerobic semi continuous reactors containing Waste Activated Sludge (WAS) spiked with NP2EO were operated for a period of 91 days, to inspect the decomposition of NP2EO in solid and liquid phases. The results obtained on the final day of operation (91st day) showed that NP2EO degraded into product compounds among which NP1EC contributed to 90% of molar mass. In general, NP2EO showed a sharp degradation while NP1EC was produced rapidly. NP1EO also showed a steady degradation. However, NP was accumulated in the reactor. During the study, TS, VS, TSS and VSS degradation was also monitored and the percentage removals were found to be between 40-60%. COD removal on the other hand was between 64-66%.

Despite tremendous advances in mass spectrometry instrumentation and mass spectrometry-based methodologies, global protein profiling of organellar, cellular, tissue and body fluid proteomes in different organisms remains a challenging task due to the complexity of the samples and the wide dynamic range of protein concentrations. In addition, large amounts of produced data make result exploitation difficult. To overcome these issues, further advances in sample preparation, mass spectrometry instrumentation as well as data processing and data analysis are required. The presented study focuses as first on the improvement of the proteolytic digestion of proteins in in-gel based proteomic approach (Gel-LCMS). To this end commonly used bovine trypsin (BT) was modified with oligosaccharides in order to overcome its main disadvantages, such as weak thermostability and fast autolysis at basic pH. Glycosylated trypsin derivates maintained their cleavage specifity and showed better thermostability, autolysis resistance and less autolytic background than unmodified BT. In line with the “accelerated digestion protocol” (ADP) previously established in our laboratory modified enzymes were tested in in-gel digestion of proteins. Kinetics of in-gel digestion was studied by MALDI TOF mass spectrometry using 18O-labeled peptides as internal standards as well as by label-free quantification approach, which utilizes intensities of peptide ions detected by nanoLC-MS/MS. In the performed kinetic study the effect of temperature, enzyme concentration and digestion time on the yield of digestion products was characterized. The obtained results showed that in-gel digestion of proteins by glycosylated trypsin conjugates was less efficient compared to the conventional digestion (CD) and achieved maximal 50 to 70% of CD yield, suggesting that the attached sugar molecules limit free diffusion of the modified trypsins into the polyacrylamide gel pores. Nevertheless, these thermostable and autolysis resistant enzymes can be regarded as promising candidates for gel-free shotgun approach. To address the reliability issue of proteomic data I further focused on protein identifications with borderline statistical confidence produced by database searching. These hits are typically produced by matching a few marginal quality MS/MS spectra to database peptide sequences and represent a significant bottleneck in proteomics. A method was developed for rapid validation of borderline hits, which takes advantage of the independent interpretation of the acquired tandem mass spectra by de novo sequencing software PepNovo followed by mass-spectrometry driven BLAST (MS BLAST) sequence similarity searching that utilize all partially accurate, degenerate and redundant proposed peptide sequences. It was demonstrated that a combination of MASCOT software, de novo sequencing software PepNovo and MS BLAST, bundled by a simple scripted interface, enabled rapid and efficient validation of a large number of borderline hits, produced by matching of one or two MS/MS spectra with marginal statistical significance.

<p>Produktions- och optimeringspotentialen vid anaerob rötning av flytgödsel från nötkreatur granskades. Detta genom att, i laboratoriemiljö, undersöka förändringar i processtabilitet, gasproduktion och metanhalt vid tillsats av kycklinggödsel och pH-buffert med flytgödsel från nötkreatur som grundsubstrat. Tillsatsen av pH-buffert underlättade tillväxtfasen för mikroorganismerna, snabbare ökning i metanhalt och den totala metangasproduktionen ökade. Skillnaden var som störst under tillväxtfasen och avtog från stationärfas och framåt. Tillsats av kycklinggödsel har i detta fall inhiberat processen, möjligtvis på grund av ammoniakinhibering vid tillsats av det kväverika substratet som kycklinggödsel är.</p> / <p>Production potential and basis for optimization of anaerobic digestion with cattle manure was reviewed by examining process stability, gas production and methane content in a laboratory environment via addition of pH buffer and chicken manure with slurry from cattle manure as basic feedstock. Addition of pH buffer facilitated the growth of microorganisms by more rapid increase in methane content and greater quantities of produced methane gas. The effects were high during the growth stage but decreased, during and after, the stationary phase. Addition of chicken manure caused inhibition of the process, probably caused by ammonia inhibition from high nitrogen content in chicken manure.</p>