Browsing by Subject "HYDROLYSIS"

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  • Collins, Scott; Hasan, Galib; Joshi, Anuj; McIndoe, J. Scott; Linnolahti, Mikko (2021)
    Density functional theory calculations on neutral sheet models for methylaluminoxane (MAO) indicate that these structures, containing 5-coordinate and 4-coordinate Al, are likely precursors to ion-pairs seen during the hydrolysis of trimethylaluminum (Me3Al) in the presence of donors such as octamethyltrisiloxane (OMTS). Ionization by both methide ([Me](-)) and [Me2Al](+) abstraction, involving this donor, were studied by polarizable continuum model calculations in fluorobenzene (PhF) and o-difluorobenzene (DFB) media. These studies suggest that low MW, 5-coordinate sheets ionize by [Me2Al](+) abstraction, while [Me](-) abstraction from Me3Al-OMTS is the likely process for higher MW 4-coordinate sheets. Further, comparison of anion stabilities per mole of aluminoxane repeat unit (MeAlO)(n), suggest that anions such as [(MeAlO)(7)(Me3Al)(4)Me](-)=[7,4](-) are especially stable compared to higher homologues, even though their neutral precursors are unstable.
  • Sugano, Junko; Linnakoski, Riikka; Huhtinen, Seppo; Pappinen, Ari; Niemela, Pekka; Asiegbu, Fred O. (2019)
    The initial stage of cellulose degradation has been studied via in vitro assays of fungi isolated from rotten wood in a boreal forest. Among the 37 isolates, Antrodia sinuosa appeared to be an effective cellulose degrader and was selected for studying the initial degradation process. In the liquid cultivation with carboxymethylcellulose (CMC), the increase of the mycelial dry weight coincided with the pH decrease of the culture medium from pH 5.7 to 3.9, between the 3rd and 6th cultivation day. At the same time, the cellulolytic activity increased; the CMCase activity increased sharply and the reducing sugars reached their maximum concentration in the culture medium. It seems that the decreasing pH enables the cellulose degradation by A. sinuosa at an early stage of the process. The results of this study may be useful for a more efficient industrial application of biomass by means of brown-rot fungi.
  • Ellilä, Simo; Bromann, Paul; Nyyssönen, Mari; Itävaara, Merja; Koivula, Anu; Paulin, Lars; Kruus, Kristiina (2019)
    Xylanases are in important class of industrial enzymes that are essential for the complete hydrolysis of lignocellulosic biomass into fermentable sugars. In the present study, we report the cloning of novel xylanases with interesting properties from compost metagenomics libraries. Controlled composting of lignocellulosic materials was used to enrich the microbial population in lignocellulolytic organisms. DNA extracted from the compost samples was used to construct metagenomics libraries, which were screened for xylanase activity. In total, 40 clones exhibiting xylanase activity were identified and the thermostability of the discovered xylanases was assayed directly from the library clones. Five genes, including one belonging to the more rare family GH8, were selected for subcloning and the enzymes were expressed in recombinant form in E. coli. Preliminary characterization of the metagenome-derived xylanases revealed interesting properties of the novel enzymes, such as high thermostability and specific activity, and differences in hydrolysis profiles. One enzyme was found to perform better than a standard Trichoderma reesei xylanase in the hydrolysis of lignocellulose at elevated temperatures.
  • Butler, Don H.; Koivisto, Satu Mirjami; Brumfeld, Vlad; Shahack-Gross, Ruth (2019)
    Salmonid resources currently foster socioeconomic prosperity in several nations, yet their importance to many ancient circumpolar societies is poorly understood due to insufficient fish bone preservation at archaeological sites. As a result, there are serious gaps in our knowledge concerning the antiquity of northern salmonid fisheries and their impacts on shaping biodiversity, hunter-gatherer adaptations, and human-ecological networks. The interdisciplinary study presented here demonstrates that calcium-magnesium phosphate minerals formed in burned salmonid bones can preserve at ancient northern sites, thus informing on the early utilization of these resources despite the absence of morphologically classifiable bones. The minerals whitlockite and beta magnesium tricalcium phosphate were identified in rare morphologically classifiable Atlantic salmonid bones from three Mid-Holocene sites in Finland. Large amounts of beta magnesium tricalcium phosphate were also experimentally formed by burning modern Atlantic salmonid and brown trout bones. Our results demonstrate the value of these minerals as proxies for ancient northern salmonid fishing. Specifically, the whitlockite mineral was discovered in hearth sediments from the 5,600 year old Yli-Ii Kierikinkangas site on the Iijoki River in northern Finland. Our fine sieving and mineralogical analyses of these sediments, along with zooarchaeological identification of recovered bone fragments, have confirmed for the first time that the people living at this village did incorporate salmonids into their economies, thus providing new evidence for early estuary/riverine fisheries in northern Finland.
  • Dong, Yujiao; Paukkonen, Heli; Fang, Wenwen; Kontturi, Eero; Laaksonen, Timo; Laaksonen, Paivi (2018)
    Drug release from a new type of matrix material consisting of partially fibrillated microcrystalline cellulose was investigated. A mechanical treatment of novel AaltoCell T cellulose microcrystals caused partial opening of the nanofibrillary structure of the cellulose particles and entanglement of individual particles led into formation of an elastic network of microcrystalline cellulose. The rheological properties of the stable hydrogel-like materials were characterised by shear rheometry. Model compounds metronidazole and lysozyme were successfully employed in drug release experiments carried out by delignified (bleached) and lignin-containing matrices. The viscosity as well as the lignin-content played a role in the release dynamics of the drugs. Microcrystalline AaltoCell T was proven as high-performing material for diffusion controlled release of the chosen model compounds and can be seen as a safe and economical alternative for novel matrix materials such as nanocellulose or cellulose derivatives.
  • Heikkinen, Emma M.; del Amo, Eva M.; Ranta, Veli-Pekka; Urtti, Arto; Vellonen, Kati-Sisko; Ruponen, Marika (2018)
    Corneal esterases are utilized in the activation of topically applied ester prodrugs. Esterases may also be involved in the metabolism of drugs in posterior eye tissues, but their physiological activity is unknown. Furthermore, extrapolation of the esterase activity from protein level to the tissues is missing. The aims of the current study were to determine esterase activities in porcine and albino rabbit ocular tissues, calculate the activities for whole tissues and compare esterase activity between the species. We conducted a hydrolysis study with ocular tissue homogenates using an esterase probe substrate 4-nitrophenyl acetate. The hydrolysis rates were first normalized to protein content and then scaled to whole tissues. The hydrolytic rate normalized to protein content was high in the cornea and iris-ciliary body and low in the lens and aqueous humor, and in general, the rabbit tissues had higher hydrolytic rates than the porcine ones. Esterase activity scaled to whole tissue was high in cornea and iris-ciliary body and low in aqueous humor and retinal pigment epithelium in both species. The current study revealed differences in esterase activities among the ocular tissues and the species. This basic knowledge on ocular esterases provides background information particularly for posterior segment drug development.
  • Kouzi, Afamia; Puranen, Matti; Kontro, Merja H. (2020)
    Biogas production from sewage sludge volatile solids (VS) by anaerobic digestion slows down towards the end of the process, among inhibitory factors being pH increase upon ammonia accumulation, poorly digestible biomaterials, and high fixed solid (FS) content. The possibility of concentrating the digested sludge VS (41.7-56.6% on a dry weight basis) by surface and bottom layer separation with biogas post-production was studied. Furthermore, the potential to recycle concentrated VS and digested sludge back to the process after adjusting pH 7.0 to optimal for biogas-producing microbes and after acid, alkali, thermal, and sonolytic treatments was examined. In general, pH 7.0 control alone improved biogas production from the recycled digested sludge the most. An equally good improvement in biogas production was achieved by recycling the digested sludge, which had been heated until ammonia had evaporated and the pH dropped to 7.0 (1-2 h, 75 degrees C), and at the same time, VS was degraded. The biogas production from the sonicated and recycled sludge was almost as good as from the pH-adjusted, or heat-treated recycled sludge. After the acid and base treatments of the digested sludge, the recycled sludge yielded often the lowest biogas volume, as the added chemicals increased the FS concentration, which proved to be a more important inhibitory factor than poorly degradable VS. The high FS content significantly reduced the benefits of the treatments. By separating the surface and bottom layers with biogas post-production, the surface layer of VS was concentrated to 51.6-61.8%, while different compositions of the layers affected the biogas production.
  • Monschein, Mareike; Jurak, Edita; Paasela, Tanja; Koitto, Taru; Lambauer, Vera; Pavicic, Mirko; Enjalbert, Thomas; Dumon, Claire; Master, Emma R. (2022)
    Background Substrate accessibility remains a key limitation to the efficient enzymatic deconstruction of lignocellulosic biomass. Limited substrate accessibility is often addressed by increasing enzyme loading, which increases process and product costs. Alternatively, considerable efforts are underway world-wide to identify amorphogenesis-inducing proteins and protein domains that increase the accessibility of carbohydrate-active enzymes to targeted lignocellulose components. Results We established a three-dimensional assay, PACER (plant cell wall model for the analysis of non-catalytic and enzymatic responses), that enables analysis of enzyme migration through defined lignocellulose composites. A cellulose/azo-xylan composite was made to demonstrate the PACER concept and then used to test the migration and activity of multiple xylanolytic enzymes. In addition to non-catalytic domains of xylanases, the potential of loosenin-like proteins to boost xylanase migration through cellulose/azo-xylan composites was observed. Conclusions The PACER assay is inexpensive and parallelizable, suitable for screening proteins for ability to increase enzyme accessibility to lignocellulose substrates. Using the PACER assay, we visualized the impact of xylan-binding modules and loosenin-like proteins on xylanase mobility and access to targeted substrates. Given the flexibility to use different composite materials, the PACER assay presents a versatile platform to study impacts of lignocellulose components on enzyme access to targeted substrates.
  • Pulkkinen, Marjo; Coda, Rossana; Lampi, Anna-Maija; Varis, Jutta; Katina, Kati; Piironen, Vieno (2019)
    Vicine and convicine may be removed from faba bean by hydrolysis to the corresponding aglycones, divicine and isouramil. For total elimination of their toxicity, further degradation of the aglycones should be shown. The aim of the study was to investigate hydrolysis of vicine and convicine using the enzymatic activity in faba bean in flour suspensions and selected lactic acid bacteria used as starters for faba bean fermentation. In addition, the effect of acidity on the stability of vicine and convicine was investigated. Sourdoughs were used in a baking process to obtain breads as final products. Vicine, convicine, and their aglycones were analyzed using reversed phase high pressure liquid chromatography with UV detection (RP-HPLC-UV). Incubation of the suspensions showed rather small vicine and convicine losses. Acidity itself did not cause losses under the conditions studied, apart from that of convicine at low pH. In sourdough fermentation with strains of Lactobacillus plantarum and Pediococcus pentosaceus, losses of vicine and convicine were dependent on the fermentation temperature and the β-glucosidase activity of the starter. Compared to fermentation at 20 °C, more intense acidification at 25 °C resulted in decrease of vicine up to 85% and convicine up to 47%. Levels of vicine and convicine in breads were comparable to levels in sourdoughs. Furthermore, the aglycones were not detected from breads.
  • Mäkelä, Noora; Sontag-Strohm, Tuula; Schiehser, Sonja; Potthast, Antje; Maaheimo, Hannu; Maina, Ndegwa H. (2017)
    Oxidation of cereal beta-glucans may affect their stability in food products. Generally, polysaccharides oxidise via different pathways leading to chain cleavage or formation of oxidised groups within the polymer chain. In this study, oxidation pathways of oat and barley beta-glucans were assessed with different concentrations of hydrogen peroxide (H2O2) or ascorbic acid (Asc) with ferrous iron (Fe2+) as a catalyst. Degradation of beta-glucans was evaluated using high performance size exclusion chromatography and formation of carbonyl groups using carbazole-9-carbonyloxyamine labelling. Furthermore, oxidative degradation of glucosyl residues was studied. Based on the results, the oxidation with Asc mainly resulted in glycosidic bond cleavage. With H2O2, both glycosidic bond cleavage and formation of carbonyl groups within the beta-glucan chain was found. Moreover, H2O2 oxidation led to production of formic acid, which was proposed to result from Ruff degradation where oxidised glucose (gluconic acid) is decarboxylated to form arabinose. (C) 2016 Elsevier Ltd. All rights reserved.
  • Pitkänen, Leena M.; Heinonen, Marina; Mikkonen, Kirsi S. (2018)
    A growing population and concern over the sufficiency of natural resources for feeding this population has motivated researchers and industries to search for alternative and complementary sources of food ingredients and additives. Numerous plant species and parts of plants are explored as raw materials for food production. An interesting example is wood; to date, few wood-based additives or ingredients are authorized for food use. Wood hemicelluloses, such as softwood galactoglucomannans (GGM), constitute an abundant bioresource that shows a highly potential functionality in edible materials. Spruce GGM—“spruce gum”—acts as a multi-functional emulsion stabilizer, and it could be used in various processed food products, replacing less effective, conventional emulsifiers. Before new materials can be released onto the food market, their safety must be evaluated, according to the Novel Food regulation. This review focuses on the safety aspects that must be considered before polysaccharide- and phenolic-rich plant extracts can be awarded the status of authorized food ingredients. In this review, GGM is presented as a case study and examples are given of plant-based polysaccharides that are already authorized for food purposes. The legislation regarding Novel Food ingredients in Europe is also briefly reviewed.
  • van den Bosch, Tijs J. M.; Niemi, Outi; Welte, Cornelia U. (2020)
    Plants of the Brassicales order, including Arabidopsis and many common vegetables, produce toxic isothiocyanates to defend themselves against pathogens. Despite this defence, plant pathogenic microorganisms like Pectobacterium cause large yield losses in fields and during storage of crops. The bacterial gene saxA was previously found to encode isothiocyanate hydrolase that degrades isothiocyanates in vitro. Here we demonstrate in planta that saxA is a virulence factor that can overcome the chemical defence system of Brassicales plants. Analysis of the distribution of saxA genes in Pectobacterium suggests that saxA from three different phylogenetic origins are present within this genus. Deletion of saxA genes representing two of the most common classes from P. odoriferum and P. versatile resulted in significantly reduced virulence on Arabidopsis thaliana and Brassica oleracea. Furthermore, expressing saxA from a plasmid in a potato-specific P. parmentieri strain that does not naturally harbour this gene significantly increased the ability of the strain to macerate Arabidopsis. These findings suggest that a single gene may have a significant role in defining the host range of a plant pathogen.
  • Linares, Nancy Coconi; Di Falco, Marcos; Benoit-Gelber, Isabelle; Gruben, Birgit S.; Peng, Mao; Tsang, Adrian; Mäkelä, Miia R.; de Vries, Ronald P. (2019)
    Guar gum consists mainly of galactomannan and constitutes the endosperm of guar seeds that acts as a reserve polysaccharide for germination. Due to its molecular structure and physical properties, this biopolymer has been considered as one of the most important and widely used gums in industry. However, for many of these applications this (hemi-) cellulosic structure needs to be modified or (partially) depolymerized in order to customize and improve its physicochemical properties. In this study, transcriptome, exoproteome and enzyme activity analyses were employed to decipher the complete enzymatic arsenal for guar gum depolymerization by Aspergillus niger. This multi-omic analysis revealed a set of 46 genes encoding carbohydrate-active enzymes (CAZymes) responding to the presence of guar gum, including CAZymes not only with preferred activity towards galactomannan, but also towards (arabino-) xylan, cellulose, starch and pectin, likely due to trace components in guar gum. This demonstrates that the purity of substrates has a strong effect on the resulting enzyme mixture produced by A. niger and probably by other fungi as well, which has significant implications for the commercial production of fungal enzyme cocktails.
  • Mäkelä, Noora; Brinck, Outi; Sontag-Strohm, Tuula (2020)
    The physiological functionality of cereal beta-glucan (beta-glucan) has been mainly attributed to its ability to form viscous solutions in the gastrointestinal (GI) tract. The viscosity is dependent on the concentration, extractability and molecular weight of beta-glucan, and to enable maximal functionality, these factors should therefore be acknowledged and their role in the physiological functionality of cereal beta-glucan further studied. An in vitro GI simulation with separate oral, gastric and small intestine phases was used to model the state of beta-glucan from various oat products in the GI tract. A rather large variation (from 26% to 99%) was observed in the extractabilities between product categories, with the highest extractabilities observed in spoonable products. The viscosities also varied highly within categories. When the comparison was done at similar concentration levels, the highest viscosities were observed in the products produced through dry processes, and moisture content during processing was suggested to be essential to the extent of beta-glucan degradation. The viscosity in samples that were likely to exhibit enzymatic activity was shown to be rather low, and thus the physiological functionality of beta-glucan may be threatened if the product also contains grain ingredients other than kiln-dried oat. Clear differences were observed in the functionality of beta-glucan in the GI tract model depending on a product type, and these were explained by differences in ingredients and processes. However, further studies are needed to specify the influence of each factor and to clarify the factors determining the physiological functionality of beta-glucan in food products.
  • Lopes Graça, Carla Alexandra; Edelmann, Minnamari; Raymundo, Anabela; Sousa, Isabel; Coda, Rossana; Sontag-Strohm, Tuula; Huang, Xin (2022)
    The incorporation of yoghurt as a starter in sourdough wheat bread, to improve technological and nutritional properties, was investigated. Two bread dough matrices were considered: endosperm wheat flour and blended with whole-grain flour. Two fermentation's types were performed, two-stage sourdough bread and yeast bread fermentation. Compared with yeast dough, yoghurt-sourdough fermentation promoted considerable changes in chemical composition, particularly when whole-grain flour was conjoined with wheat flour: higher protein proteolysis degree, increase of peptide and free amino nitrogen content, solubilization of phenolic compounds (46-53%), increase of DPPH radical scavenging (54-65%) and ferric-reducing power (85-88%), were observed. As a baking ingredient, yoghurt-sourdough improved the bread crumb softness (15-12%) and delayed the staling (40-35%). Nutritionally, the glycemic index was reduced (18-32%) while protein digestibility (6-12%) and free amino acids bioavailability (50-100%) were enhanced. The addition of yoghurt and sourdough fermentation techniques offered a promising tool to improve wheat bread's technological and functional properties.