Organocatalysis in continuous flow
Berg, S.A. van den - \ 2016
Wageningen University. Promotor(en): Han Zuilhof, co-promotor(en): Tom Wennekes. - Wageningen : Wageningen University - ISBN 9789462576636 - 217
catalysis - organic chemistry - flow - microfluidics - alkenes - katalyse - organische scheikunde - stroming - microfluidics - alkenen
Continuous flow chemistry is an enabling technique in organic chemistry. Advantages include extremely fast mixing and heat transfer capabilities as well as rapid screening of reaction conditions. Combining continuous flow chemistry with solid-supported organocatalysis presents challenges that have been investigated in this thesis.
We have developed a technique to modify the surface of silicon carbide with alkenes using microwaves. Silicon carbide is an extremely tough bio-compatible material and is of interest as a solid support. By using monolayers of bifunctional alkenes we were able to perform follow-up reactions to attach other molecules of interest, thereby further modifying the surface chemistry of silicon carbide. The use of microwaves speeds up the attachment of monolayers by as much as sixteen times.
We have developed a new method for the surface functionalization of MCM-41, a type of mesoporous silica. Mesoporous silica is an interesting solid support for catalysts because of its large surface area. 1,7-octadiene can be covalently attached under mild conditions (24 h, 100 ºC). Follow-up thiol-ene click reactions can be performed using thiols. The use of 1,2-ethanedithiol leads to a material that can be reacted with the organocatalyst quinine, giving catalytically active mesoporous silica. This material was tested in flow and yielded full conversion of starting materials for the thio-Michael addition of 3-methoxythiophenol and 1-cyclohex-2-enone.
Sulfonic acid supported on mesoporous silica (SBA-15) made via co-condensation of a thiol-containing silica precursor was used in a packed-bed continuous flow setup. This solid-supported sulfonic acid efficiently catalyzes the protection and deprotection of various alcohols as the tetrahydropyranyl-, trimethylsilyl and dimethylphenylsilyl derivatives under continuous flow conditions. By passing the starting material (primary, secondary, benzylic and phenolic alcohols) together with the appropriate protecting group over the packed bed more than 100 mg of product was produced in flow.
Finally, the ring-opening polymerization of L-lactide using an organocatalyst was investigated. Using low catalyst loading and short residence times (0.25– 1.2% TBD catalyst, residence times as short as 2 s) well-defined poly(lactic acid) (PLA) is obtained with high (95–100%) monomer conversions and PDIs of typically 1.2. Use of the microreactor allows for a rapid screening of optimal reaction conditions, consistently yielding the optimal values for high conversion and low polydispersity. This quickly revealed that longer residence times will give rise to higher conversions and a concomitantly broader molecular weight distribution due to transesterification of the polymer backbone by TBD.
The organocatalytic, metal-free continuous flow method described is rapid and mild enough to allow the use of a PEG-5,000 macroinitiator (yielding block copolymers), exo-BCN alcohol and even a base-labile tetrazine-derived alcohol, the latter of which cannot be used under traditional batch conditions. The resulting BCN-PLA and tetrazine-PLA materials were readily functionalized with small molecules and large polymers bearing azides and norbornenes via SPAAC and inverse electron demand Diels-Alder click chemistry.
Despite the challenges that arise, the combination of continuous flow chemistry and (solid-supported) organocatalysis is a powerful concept that warrants further investigation.
Perinatal inhibition of NF-KappaB has long-term antihypertensive and renoprotective effects in fawn-hooded hypertensive rats
Koeners, Maarten P. ; Wesseling, Sebas ; Sánchez, Manuel ; Braam, Branko ; Joles, Jaap A. - \ 2016
American Journal of Hypertension 29 (2016)1. - ISSN 0895-7061 - p. 123 - 131.
blood pressure - developmental plasticity - glomerulosclerosis - hypertension - nuclear factor-kappa B - renal hemodynamics
BACKGROUND Inhibition of transcription factor nuclear factor-kappa B (NFκB) is beneficial in various models of hypertension and renal disease. We hypothesized first that NFκB inhibition during renal development ameliorates hereditary hypertensive renal disease and next whether this was mediated via suppression of peroxisome proliferator-activated receptor (PPAR)γ coactivator 1α (PGC-1α). METHODS AND RESULTS Prior to the development of renal injury in fawn-hooded hypertensive (FHH) rats, a model of hypertension, glomerular hyperfiltration, and progressive renal injury, NFkB activity, measured by nuclear protein expression of NFkB subunit p65, was enhanced twofold in 2-day-old male and female FHH kidneys as compared to normotensive Wistar-Kyoto (WKY) rats (P <0.05). Treating FHH dams with pyrrolidine di thio carbamate (PDTC), an NFκB inhibitor, from 2 weeks before birth to 4 weeks after birth diminished NFkB activity in 2-day-FHH offspring to 2-day-WKY levels (P <0.01). Perinatal PDTC reduced systolic blood pressure from 20 weeks onwards by on average 25mm Hg (P <0.001) and ameliorated proteinuria (P <0.05) and glomerulosclerosis (P <0.05). In kidneys of 2-day-, 2-week-, and adult offspring of PDTC-treated FHH dams, PGC-1α was induced on average by 67% (quantitative polymerase chain reaction (qPCR)) suggesting that suppression of this factor by NFkB could be involved in renal damage. Follow-up experiments with perinatal pioglitazone (Pio), a PPARγ agonist, failed to confer persistent antihypertensive or renoprotective effects. CONCLUSIONS Perinatal inhibition of enhanced active renal NFκB in 2-day FHH had persistent antihypertensive and renoprotective effects. However, this was not the case for PPARγ stimulation. NFkB stimulation is therefore involved in renal damage in the FHH model of proteinuric renal disease by pathways other than via PPARγ.
|Ideas to improve the efficacy and the robustness of bio-fumigation
Helder, J. ; Hallmann, J. ; Harkes, P. ; Quist, C.W. ; Vervoort, M.T.W. ; Vonk, A. - \ 2014
In: Proceedings of the 5th IS of Biofumigation. - - p. 114 - 116.
Plants use at least two main strategies to protect themselves against pathogens and herbivores: by a physical barrier making the plant content inaccessible to the invader and/or by the production of appetite spoiling or even toxic components (or the precursors thereof). Apart from shaping the habitus of plants, cell walls, a rigid and tightly interwoven fabric consisting mainly of polysaccharides, constitute a formidable physical barrier. The primary cell wall is a relatively thin layer made up of carbohydrates and structural proteins. The secondary cell wall is a thicker layer rich in cellulose and lignin, polymers that greatly contribute to the physical strength of the plant cell wall. Combined the two cell wall elements constitute a compact protective layer. On top of this physical barrier, plants may produce secondary metabolites to reduce its attractiveness as a food source for pathogens and herbivores. Some plant families are well known by the production of specific categories of metabolites that reduce their attractiveness for herbivores. The plantain family (Plantaginaceae) harbours a wide diversity of genera including Plantago (about 200 species). Some Plantago species are known for their anti-toxic and antimicrobial characteristics. Iridoid glucosides such as aucubin and catalpol are terpenoid secondary metabolites produced by a range of plantain species, and the components make these plants unattractive for a wide spectrum of herbivores. Upon cleaving of the sugar residue by beta-glucosidases either from the plant or from the digestive tract of the invading organism, the resulting aglycon acts un-specifically by cross linking proteins and the inhibitions of enzymes. Iridoids are a class of compounds with as a core a cyclopentane fused to a 6-C oxygen heterocycle. All by all, iridoid glucosides are precursors of defence molecules against a broad range of pathogens and herbivores (Dobler et al., 2011). Members of the buttercup family (Ranunculaceae) produce another category of non-specific anti-feedants. The genus Ranunculus L. (buttercups) comprises about 600 herbaceous species, and fresh buttercup plant parts are poisonous for a wide range of organisms. This can be explained by a secondary metabolite named ranunculin. Upon de-compartimentation (chewing, maceration etc.), the enzyme beta-glucosidase, normally stored in the vacuole, comes into contact with ranunculin, and as a result this glucoside is broken down to form the volatile lactone protoanemonin (C5H4O2). This highly reactive component has anti-microbial properties, and ingestion by herbivorous mammals results in erythema and blistering (Sedivy et al., 2012; Martin et al., 1990). Yet another plant family that produces relatively harmless glycosides that are turned into toxic compounds upon the mechanical damaging of plant tissues is the mustard family (Brassicaceae). Over 100 types of glucosinolates are produced by members of this family. This class of secondary metabolites are the most important flavour compounds in edible representatives such as cabbage, 2 mustard and horseradish. The actual reason why these compounds are produced is (again) the defence against pathogens and herbivores. Just like we’ve seen for the Plantaginaceae and the Ranunculaceae, Brassicaceae produce precursors that are converted into toxic components upon de-compartimentation. The activating enzyme myrosinase (a thioglucoside glucohydrolase) is stored in the vacuole. Upon damaging or maceration of the plant tissue, myrosinase cleaves off the thio-linked glucose from its substrate, and the resulting chemically-unstable aglycone is converted into toxic isothiocyanate or related components (Textor & Gershenzon, 2009). The ban of certain categories of soil fumigantia and the severe restrictions with regard to the application of those fumigants that are still on the market to control soil-borne diseases including plant-parasitic fungi and nematodes have been the driving factors behind a search to find effective environmentally acceptable alternatives. One of the options was the use specific types of green manure. Incorporation in the topsoil of plant material harbouring compounds (or precursors thereof) that are toxic for a range of soil borne pathogens is called biofumigation. As compared to aglycones such as protoamonin from buttercups or iridoids from plantain species, the effective products released upon mechanically damaging Brassicaceous plant parts are relatively stable and volatile. Hence it will spread easily throughout the topsoil. In fact, the incorporation of mulched Brassicaceous material could be considered as a natural replacement of the soil fumigant metam sodium (Matthiessen & Kirkegaard, 2006). Recently we investigated in a field experiment in the western part of Germany the impact of Indian mustard (Brassica juncea) on plant parasitic and free-living nematode taxa (Vervoort et al., 2014). Just before the incorporation of plant material in the topsoil, aliquots of above and below plant parts were sampled and the glucosinolate content was determined both qualitatively and quantitatively. As a positive control 2-propenyl ITC was directed applied to the control plots. Although the impact of the incorporation of mulched Indian mustard material on the nematode community was clear and significant, this impact could not be attributed the effect of isothiocyanates on the nematode community. The severe physical disturbance of the soil in combination with addition of a large quantity of green manure alone sufficed to explain the observed changes. Even more remarkable: the positive control treatments by direct application of 2-propenyl ITC (two concentrations, the highest being two times the concentration predicted for the highest producing Indian mustard cultivar) had no significant effect on the nematode taxa under investigation. We cannot rule out the possibility that local circumstances such as soil type, organic matter type and content, moisture content and/or temperature of the soil at the day of biofumigation can be explanatory factors for the absence of a clear glucosinolate-related response. Our results suggest that a more widely acceptance of bio-fumigation as a tool to manage soil borne pathogens would require an increase in the efficacy and the robustness of this tool. In this paper we pay attention to some other plant families that a producing broad-spectrum toxic components (aglycones) upon maceration and/or physical damage. It could be worthwhile to investigate the impact of the simultaneous release of multiple plant-derived aglycones on plant pathogenic soil biota. The two ‘other’ plant families highlighted in this paper, the Plantaginaceae and the Ranunculaceae, should just be considered as examples (not necessarily the best ones) of families producing distinct broad-spectrum anti-microbial aglycones. We hypothesize that simultaneous exposure of soil pathogens to multiple plant defense-related aglycones could contribute to an improved efficacy and robustness of biofumigation.
Analysis of steady-state Förster resonance energy transfer data by avoiding pitfalls: Interaction of JAK2 tyrosine kinase with N-methylanthraniloyl nucleotides.
Niranjan, Y. ; Ungureanu, D. ; Hammarén, H. ; Sanz-Sanz, A. ; Westphal, A.H. ; Borst, J.W. ; Silvennoinen, O. ; Hilhorst, M.H. - \ 2013
Analytical Biochemistry 442 (2013)2. - ISSN 0003-2697 - p. 213 - 222.
pseudokinase domain - protein-kinase - fluorescence - atp - binding - receptor - analogs - site - autophosphorylation - mechanism
Förster resonance energy transfer (FRET) between the fluorescent ATP analogue 2'/3'-(N-methyl-anthraniloyl)-adenosine-5'-triphosphate (MANT–ATP) and enzymes is widely used to determine affinities for ATP–protein binding. However, in analysis of FRET fluorescence data, several important parameters are often ignored, resulting in poor accuracy of the calculated dissociation constant (Kd). In this study, we systematically analyze factors that interfere with Kd determination and describe methods for correction of primary and secondary inner filter effects that extend the use of the FRET method to higher MANT nucleotide concentrations. The interactions of the fluorescent nucleotide analogues MANT–ATP, MANT–ADP [2'/3'-O-(N-methylanthraniloyl) adenosine diphosphate], and MANT–AMP [2'/3'-O-(N-methylanthraniloyl) adenosine monophosphate] with the JAK2 tyrosine kinase domain are characterized. Taking all interfering factors into consideration, we found that JAK2 binds MANT–ATP tightly with a Kd of 15 to 25 nM and excluded the presence of a second binding site. The affinity for MANT–ADP is also tight with a Kd of 50 to 80 nM, whereas MANT–AMP does not bind. Titrations of JAK2 JH1 with nonhydrolyzable ATP analogue MANT–ATP-¿-S [2'/3'-O-(N-methylanthraniloyl) adenosine-5'-(thio)- triphosphate] yielded a Kd of 30 to 50 nM. The methods demonstrated here are applicable to other enzyme–fluorophore combinations and are expected to help improve the analysis of steady-state FRET data in MANT nucleotide binding studies and to obtain more accurate results for the affinities of nucleotide binding proteins.
Efficient Functionalization of Oxide-Free Silicon(111) Surfaces: Thiol-yne versus Thiol-ene Click Chemistry
Bhairamadgi, N.S. ; Gangarapu, S. ; Caipa Campos, M.A. ; Paulusse, J.M.J. ; Rijn, C.J.M. van; Zuilhof, H. - \ 2013
Langmuir 29 (2013)14. - ISSN 0743-7463 - p. 4535 - 4542.
copper(i)-catalyzed azide-alkyne - self-assembled monolayers - free-radical addition - cycloaddition - nanoparticles - polymers
Thiol-yne click (TYC) chemistry was utilized as a copper-free click reaction for the modification of alkyne-terminated monolayers on oxide-free Si(111) surfaces, and the results were compared with the analogous thiol–ene click (TEC) chemistry. A wide range of thiols such as 9-fluorenylmethoxy-carbonyl cysteine, thio-ß-d-glucose tetraacetate, thioacetic acid, thioglycerol, thioglycolic acid, and 1H,1H,2H,2H-perfluorodecanethiol was immobilized using TYC under photochemical conditions, and all modified surfaces were characterized by static water contact angle measurements, X-ray photoelectron spectroscopy (including a simulation thereof by density functional calculations), and infrared absorption reflection spectroscopy. Surface-bound TYC proceeds with an efficiency of up to 1.5 thiols per alkyne group. This high surface coverage proceeds without oxidizing the Si surface. TYC yielded consistently higher surface coverages than TEC, due to double addition of thiols to alkyne-terminated monolayers. This also allows for the sequential and highly efficient attachment of two different thiols onto an alkyne-terminated monolayer.
Hydrogen Bonding in Phosphine Oxide/Phosphate-Phenol Complexes
Cuypers, R. ; Sudhölter, E.J.R. ; Zuilhof, H. - \ 2010
ChemPhysChem 11 (2010)10. - ISSN 1439-4235 - p. 2230 - 2240.
solvent-impregnated resins - isothermal titration calorimetry - plesset perturbation-theory - log k-values - density functionals - bonded complexes - noncovalent interactions - interaction energies - binding-energies - water clusters
To develop a new solvent-impregnated resin (SIR) system for the removal of phenols and thiophenols from water, complex formation by hydrogen bonding of phosphine oxides and phosphates is studied using isothermal titration calorimetry (ITC) and quantum chemical modeling. Six different computational methods are used: B3LYP, M06-2X, MP2, spin component-scaled (SCS) MP2 [all four with 6-311+G(d,p) basis set], a complete basis set extrapolation at the MP2 level (MP2/CBS), and the composite CBS-Q model. This reveals a range of binding enthalpies (¿H) for phenol–phosphine oxide and phenol–phosphate complexes and their thio analogues. Both structural (bond lengths/angles) and electronic elements (charges, bond orders) are studied. Furthermore, solvent effects are investigated theoretically by the PCM solvent model and experimentally via ITC. From our calculations, a trialkylphosphine oxide is found to be the most promising extractant for phenol in SIRs, yielding ¿H=-14.5 and -9.8 kcal¿mol-1 with phenol and thiophenol, respectively (MP2/CBS), without dimer formation that would hamper the phenol complexation. In ITC measurements, the ¿H of this complex was most negative in the noncoordinating solvent cyclohexane, and slightly less so in p–p interacting solvents such as benzene. The strongest binding is found for the dimethyl phosphate–phenol complex [-15.1 kcal¿mol-1 (MP2/CBS)], due to the formation of two H-bonds (PO···H-O- and P-O-H···O-H); however, dimer formation of these phosphates competes with complexation of phenol, and would thus hamper their use in industrial extractions. CBS-Q calculations display erroneous trends for sulfur compounds, and are found to be unsuitable. Computationally relatively cheap SCS-MP2 and M06-2X calculations did accurately agree with the much more elaborate MP2/CBS method, with an average deviation of less than 1 kcal¿mol-1
Hydrogen bonding in the recovery of phenols and methyl-t-butyl ether : molecular modeling and calorimetry
Cuypers, R. - \ 2010
Wageningen University. Promotor(en): Han Zuilhof; Ernst Sudhölter. - [S.l. : S.n. - ISBN 9789085857754 - 189
afvalwaterbehandeling - verwijdering - fenol - ethers - derivaten - harsen - extractie - waste water treatment - removal - phenol - ethers - derivatives - resins - extraction
The purification of waste water is very important, for clean potable water is a common good and a necessity. Surface water purification is nowadays carried out on a massive industrial scale, and clean water is at our disposal virtually everywhere and always. However, cleaning industrial waste water can be a difficult task. Although apolar and slightly polar compounds can be removed from water relatively easily e.g. by extraction to an apolar phase, more polar pollutants like phenol and methyl-tert-butyl ether (MTBE), the two main compounds that this thesis deals with, cannot be removed as easily. A more effective method is therefore needed to clean water that is contaminated with either phenol or MTBE.
Solvent-Impregnated Resins (SIRs) are porous polymer beads containing apolar organic extractant liquids. They are used as three-phase separation systems. When brought into contact with a SIR, a solute will preferentially partition from the aqueous phase into the impregnated solvent phase. A drawback to the use of solely the organic extraction liquid in SIRs is the limited solubility of more polar compounds like phenols and ethers in such a medium. In order to enhance extraction, complex-forming extractants can be added to the organic solvent. By means of complex formation inside the organic solvent, the overall equilibrium distribution can be shifted towards the SIR, with a concomitant enhancement of the extraction efficiency. A tight binding of the pollutant molecules to the extractant will eventually ensure high distribution coefficients. However, a moderate binding strength would enable a relatively easy regeneration of the complexing agent after a binding event, enabling multiple uses of the same compound.
In this thesis hydrogen-bond (H-bond) complexation, a specific and strong yet reversible way of binding is investigated for phenol recovery and MTBE recovery from aqueous environments, involving the use of organic complexing agents that can be used inside the SIR to enhance the extraction. Potentially interesting compounds were investigated on a molecular scale by quantum chemical modeling methods and subsequent synthesis and physical characterization by primarily calorimetric means. H-bond complex formation has been evaluated and the important parameters determining the binding process have been described.
After a general introduction in Chapter 1, Chapters 2 through 4 describe phenol complexation by several different classes of complexing agents. In Chapter 2, the binding of phenol and thiophenol by phosphine oxides, phosphates, and their thio-analogs was investigated. Modeling experiments, isothermal titration calorimetry (ITC) measurements, and liquid-liquid extraction experiments showed that, in principle, the binding affinity of the oxide compounds for phenol is high, whereas the sulfide compounds show only low affinity. In particular, the binding behavior of tri-n-octylphosphine oxide towards phenol and a series of electron-withdrawing group (EWG)-substituted phenols was studied, both in the presence and absence of water in the system. It was found that the presence of water in the system – as can be expected in industrial applications – yields lower binding affinities by as much as 60 %, but the binding stoichiometry remained specific and 1 : 1 complexes were still found. Electronic and steric effects were shown to play an important role in phenol binding in the investigated environment.
In Chapter 3, these investigations were extended to the modeling of the full homologous series of mono-, di- and tri-substituted phosphine oxides and phosphates and their thio-analogs. Different modeling methods were used to investigate both structural and electronic elements. Dimethylphosphate was found to form the strongest complexes to the investigated phenols, but because this compound forms very strong homo-dimers in solution it cannot be used as an effective extractant. The SCS-MP2 method, that was relatively unexplored for H-bonding until now, was found to yield very accurate energy predictions, whereas the CBS-Q method was found to predict false binding affinities. Solvent effects are shown to immensely influence the binding behavior.
Another, even stronger group of H-bond acceptors, amine-N-oxides, was investigated as described in Chapter 4. The binding properties for phenol and thiophenol with three different amine-N-oxides yielded very high binding affinities (up to 30 times higher than for the phosphine oxide compounds). Introduction of EWGs in the amine-N-oxides was shown to yield markedly lower binding affinities towards phenols, providing a handle to fine-tune the interaction and facilitating easier regeneration of the complexing agent in future SIR applications. Solvent effects and the influence of water in the system were investigated, and it was shown that they both influence the phenol binding strength. The results in Chapters 2 through 4 show that phosphates, phosphine oxides, and amine-N-oxides could all be used in future SIR extraction systems, and the choice between these classes of compounds can be made based on more detailed considerations.
MTBE binding by several complexing agents was described in Chapter 5. A detailed modeling study of a number of different substituted phenols for MTBE binding was carried out, and the influence of solvents on the binding behavior was investigated, using a.o. the recently developed M06-2X functional and SMD solvent model. The investigated complexing agents were found to show moderate binding affinities to MTBE with binding strengths being closely linked to the acidity of the extractant. Steric effects and a proper consideration of entropic effects are also found to be important to yield successful binding of MTBE. In combination with the existing MTBE distribution coefficient for apolar phases, these moderate binding affinities were found to be able to enhance extraction, in principle, up to the point where it becomes industrially relevant to use such extractants in SIR-based extractions.
Finally, in Chapter 6 the performed research is reviewed, and conclusions, recommendations and a wider perspective for future scientific challenges are given.
Porphyrazines with oligo(ethylene oxide) thio alkoxy chains: Synthesis, aggregation, photophysics, and complexation with redox-active ions
Piet, D.P. ; Veldhuis, H. ; Zuilhof, H. ; Sudhölter, E.J.R. - \ 2004
Journal of Porphyrins and Phthalocyanines 8 (2004)8. - ISSN 1088-4246 - p. 1055 - 1061.
transition-metal ions - crystal-structure - solar-cells - titanium-dioxide - phthalocyanines - sensitization - porphyrin - electrode - phase - rings
A series of new porphyrazines with oligo(ethylene oxide) thioalkoxy side chains has been synthesized. Study of the phase transitions reveals the absence of liquid crystallinity. Due to the ethylene oxide moieties, the solubility in a variety of solvents is dramatically increased. This allowed the first study of solvent effects on the photophysical properties (absorption and emission) in water and chloroform. The complexation of the porphyrazines with the redox couple LiI/I2 has been studied. No effect on the aggregate formation has been observed in solution, while in a film incorporation of an increased amount of LiI/I2 leads to a disturbance of the aggregates. At temperatures up to 100 °C the porphyrazine - LiI/I2 complexes are stable in the liquid phase, but from 115 °C onwards, small crystallites of LiI are formed in the isotropic liquid.
Bicyclooctane derivatives as plasticisers
Jansen, J.C. ; Luitjes, H. - \ 1999
Octrooinummer: WO9945060, gepubliceerd: 1999-09-10.
The use of isosorbide esters, isosorbide polyesters, isosorbide ethers, isosorbide carbonates, isosorbide thioethers, isosorbide thioesters, isosorbide amides, isosorbide (thio)urethanes, isosorbide urea, isosorbide phosphates and isosorbide phosphonates as a plasticizer is described. Instead of isosorbide it is also possible for corresponding diols such as isomannide and similar compounds, in which the ring oxygen atoms of the isosorbide have been replaced by carbon or by other heteroatoms, to serve as a base. In particular, this relates to C3-C11 alkanoates of isosorbide, isomannide and the like. In particular, the compounds are used as a plasticizer inpoly(vinylchloride), optionally in conjunction with usual plasticizers such as dioctyl phthalates.
Direct coating of poly(lys) or acetyl-thio-acetyl peptides to polystyrene: the effects in an enzyme-linked immunosorbent assay
Loomans, E.E.M.G. ; Petersen-van Ettekoven, A. ; Bloemers, H.P.J. ; Schielen, W.J.G. - \ 1997
Analytical Biochemistry 248 (1997). - ISSN 0003-2697 - p. 117 - 129.
Cloning and expression of DNA encoding a ripening form a polypeptide having sulfhydryl oxidase activity
Maat, J. ; Musters, W. ; Stam, H. ; Schaap, P.J. ; Vondervoort, P.J.J. van de; Visser, J. ; Verbakel, J.M.A. - \ 1993
Octrooinummer: EP0565172, gepubliceerd: 1993.
The invention relates to recombinant DNA technology for the production of an enzyme having sulfhydryl oxidase ("SOX") activity. This SOX-enzyme can be used where the oxidation of free sulfhydryl groups (thio compounds) to the corresponding disulfides is desirable. SOX enzyme may be used for treatment of bakery products or for removal of off-flavour from milk or beer.
|Health-based recommended occupational exposure limits for methyl acrylate.
Maclaine Pont, M.A. - \ 1990
Unknown Publisher - 27 p.
Me-acr can be absorbed totally after dermal, and oral exposure. Me-acr is metabolized fairly rapidly and elimination occurs for a large part via exhalation of CO2 and to a lesser extent via urinary excretion of thio-ethers. Me-acr is irritant to corrosive to the skin. Skin sensitization and cross sensitization to some of the related compounds occurs. In a two-year inhalatory study in rats (highest dose tested: 483 mg/m3) no signs of systemic toxicity or carcinogenicity are observed. Dose-related changes are observed in the nasal mucosas. The lowest dose tested (54 mg/m3 = 15 ppm) induces a slight effect on the respiratory and olfactory nasal mucose. Me-acr induces mutations and chromosomal aberrations in mammalian cells in vitro. However, in view of the abovementioned local effects these results are considered irrelevant for the MAC evaluation. Since rats ar obligatory nasal breathers this region has a higher sensitivity and this fact has little significance for the extrapolation to humans. To take into account intraspecies variation a safety factor of 3 is introduced. An occupational exposure limit of 18 mg/m3 (5 ppm) TWA 8 hr is recommended. Since dermal absorption can add to the inhalatory exposure a "H" (skin notation ) is advised.
Bepaling van thiouracyl en methylthiouracyl in toedieningsplaatsen
Hooijerink, H. ; Korbee, H.J. ; Ruig, W.G. de - \ 1985
Wageningen : RIKILT (Rapport / RIKILT 85.82) - 13
vlees - vleeswaren - thio-uracil - vloeistofchromatografie - hplc - methylthio-uracil - meat - meat products - thiouracil - liquid chromatography - methylthiouracil
|Matara dictrict integrated rural development project: A critical analysis of project design
Thio, K.S. - \ 1985
Unknown Publisher - 83 p.
|Programme de formation en matiere d'analyse de project de developpement rural au Niger
Moens, N. ; Keita, M. ; Thio, K.S. - \ 1985
Unknown Publisher - 53 p.
|Maatschappelijke aspecten van landbouwontwikkeling in ontwikkelingslanden
Thio, K.S. - \ 1985
L&O, kwartaaltijdschrift van de NRLO (1985)september. - 6 p.
|Methodologie d'une etude pour l'elaboration d'un programme de formation et de recyclage des cadres en matiere de planification pour le developp. rural au Niger
Thio, K.S. ; Moens, N.M. - \ 1984
|Evaluation du projet hydro-agricole 'Riziculture irrigue du Logome et Chari', finance par le Fonds Europaen de Developpement en Republique Unie du Cameroun
Thio, K.S. ; Zijlstra, G. - \ 1984
Reduced planning efforts:Identification of rural potentials in selected key regions of Ratnapura
Schipper, R.A. ; Thio, K.S. ; Black-Michaud, J. - \ 1982
|Kurunegala District Integrated Rural Development Project: Viability and replicability of project: management
Thio, K.S. ; Samad, M. ; Black-Michaud, J. - \ 1982
Unknown Publisher - 87 p.
|Matara District Integrated Rural Development Project: A critical analysis of project design
Thio, K.S. - \ 1982
Unknown Publisher - 83 p.
|A demonstration of resource based socio-economic planning in Matara District Report No. 1 in 'Regional planning for agricultural development in Sri Lanka'
Thio, K.S. ; Polman, F.J. ; Samad, M. - \ 1982
|Integrated rural development in Kurunegala: Viability and replicability of project management
Black-Michaud, J. ; Samad, M. ; Thio, K.S. - \ 1982
Unknown Publisher - 87 p.
|Bacterial Bioluminescence: Spectral Study of the Emitters in the In Vivo Reaction
Matheson, I.B.C. ; Lee, J. ; Muller, F. - \ 1981
Proceedings of the National Academy of Sciences of the United States of America 78 (1981)2. - ISSN 0027-8424 - p. 948 - 952.
Transient fluorescent species are observed in the bioluminescent reactions of three reduced flavin mononucleotides with aliphatic aldehydes and oxygen, catalyzed by bacterial luciferase. In each case the fluorescence spectral distribution is similar to that of the bioluminescence but is readily distinguishable from it on the basis of a significantly greater signal strength. The corrected bioluminescence maxima using Beneckea harveyi luciferase are 479 nm (iso-FMNH2), 490 nm (FMNH2), and 560 nm (2-thio-FMNH2). In an ethanol glass at 77 K, 2-thioriboflavin is fluorescent (ϕF = 0.03, λmax = 562 nm). These results are interpreted by a sensitized chemiluminescence mechanism in which the flavins bound to luciferase act as acceptors of excitation energy. For 2-thio-FMNH2, this acceptor species appears to be the oxidized 2-thio-FMN on the basis of the spectral evidence, whereas for the other flavins, some form of reduced species is a more likely candidate.
L' efficacite de la planification agricole en Tunisie
Thio, K.S. - \ 1979
Landbouwhogeschool Wageningen. Promotor(en): F.P. Jansen; A.A. Kampfraath. - Wageningen : Veenman - 344
plattelandsplanning - plattelandsontwikkeling - sociale economie - landbouw - economische sectoren - ontwikkeling - ontwikkelingsplanning - economisch beleid - industrie - tunesië - economische situatie - economische planning - landbouw als bedrijfstak - internationale economie - organisatievormen - economische productie - rural planning - rural development - socioeconomics - agriculture - economic sectors - development - development planning - economic policy - industry - tunisia - economic situation - economic planning - agriculture as branch of economy - international economy - forms of organization - economic production
Dit boek richt zich tot allen die op een of andere wijze betrokken zijn bij de problematiek van de planning in ontwikkelingslanden: internationale organisaties, instituten voor onderzoek en onderwijs, hulpverlenende instanties en de planners in de ontwikkelingslanden zelf.
Het doel van de studie is bij te dragen tot de verbetering van methoden van planning die in praktijk in ontwikkelingslanden worden of kunnen worden toegepast. Tot dat doel wordt het begrip efficiency van de planning in zijn componenten ontleed en toegepast op een speciaal geval, nl. de landbouwplanning in Tunesië. Onidat de Franse samenvatting de inhoud van het boek per hoofdstuk weergeeft zullen wij ons hier beperken tot de algemene kenmerken van deze studie.
In de litteratuur over ontwikkelingsplanning wordt voornamelijk aandacht besteed aan de inhoudelijke kant van de ontwikkelingsproblematiek. Te weinig wordt benadrukt dat de planning zelf een instrument voor ontwikkeling is dat gepland kan en moet worden. Immers, in een ontwikkelingsstrategie is de strateeg, d.w.z. de overheid, zelf betrokken. Planning van de sociaal-economische ontwikkeling vereist ontwikkeling van de planning, inhoudelijk en bestuurlijk. Een belangrijk uitgangspunt van het onderzoek is dan ook dat verbetering van methoden van planning niet alleen betrekking dient te hebben op verbetering van de methoden van informatieverzameling en van sociaal-economische analyse. Zulke pogingen kunnen nutteloos zijn indien men niet ook rekening houdt met de bestuurlijke en politieke context waarin de planning
De efficiency van de planning kan alleen begrepen worden in het totale cybernetische planproces, dat globaal in de volgende cycli verloopt: informatieverzameling ->planopstelling ->planuitvoering ->evaluatie en bijsturing. De evaluatie van de resultaten van het planproces dient op drie niveaux te geschieden:
De efficiency van de planning dient echter niet alleen afgemeten te worden aan de uitkomst van zijn voorspellingen. In ieder geval dient de werkelijke invloed van het overheidsingrijpen op de eindresultaten te worden onderscheiden van toevallige faktoren die in gunstige of ongunstige zin aan de eindresultaten hebben bijgedragen. Bovendien kan de efficiency van de planning alleen geappreciterd worden in het licht van de reikwijdte van de planning, waaronder wij verstaan de omvang van de te plannen probleemgebieden, het detail waarin deze gepland worden en de in de beschouwing betrokken samenhangen. Het onderscheid tussen trefzekerheid en reikwijdte komt voort uit twee mogelijke benaderingen van de evaluatie van de efficiency van de planning, te weten respectievelijk: de immanente kritiek, die uitgaat van de veronderstellingen van het planningsysteem zelf en de transcendente kritiek, die uitgaat van veronderstellingen die aan criteria van buiten het planningsysteem ontleend zijn.
Ieder efficiency-onderzoek komt neer op een doeleinden-middelen analyse. Terwijl trefzekerheid en reikwijdte tot de doeleinden van een planningsysteem gerekend kunnen worden, behoren tot de middelen: de tijd, de mankracht en het geld nodig voor het opstellen en evalueren van plannen en de overheadkosten van het uitvoerende apparaat. Wij hebben niet getracht tot een nauwkeurige kwantitatieve berekening van deze middelen te kornen, hoewel zij in ontwikkelingslanden vaak een reële beperking op de planning en dus op de ontwikkeling in het algemeen vormen. Wij menen echter dat in de beginfase, waarin dit type onderzoek zich nog bevindt, een kwantitatieve benadering niet relevant is. Een kwalitatieve benadering ter besparing van de kosten van de planning is vooralstiog vruchtbaarder. Deze houdt o.a. in: een goede definiëring van de ontwilekelingsproblernatiek, waarvoor de planning gesteld is, het toepassen van arbeids- en tijdsbeparende methoden van informatieverzameling en analyse (b.v. het toezien op de benodigde precisie van gegevens), het bewust kiezen van de beslissingsprocedures, een adekwate verdeling van de expertise over de planningsorganen, een juiste keuze van de expertise in functie van de op te lossen problemen, het zo goed mogelijk verdelen van de planningactiviteiten in de tijd, enz... .
Grote nadruk valt in dit onderzoek op de case-study van de landbouwplanning in Tunesië. Daarmee is aangegeven dat dit soort onderzoek zich nog in een empirisch stadium bevindt. Na een reeks van dergelijke deelstudies kan later tot systematisering van de kennis worden gekomen. Wij menen, dat in de litteratuur nog een groot gebrek is aan nauwkeurige, concrete beschrijvingen van totale planprocessen. Uiteraard zijn met de case-study ook de beperkingen van dit onderzoek gegeven. In feite is de problematiek van ieder land specifiek en vraagt daarom om aarigepaste methoden van planning. De kenmerken van Tunesië zijn: het is een klein, al relatief ontwikkeld land, dat sterk in de economische en politieke invloedssfeer van Europa ligt. Naar Europese standaarden gemeten zijn de problemen van armoede en werkloosheid er echter nog zeer groot en zullen dat op lange termijn nog blijven. De planning in Tunesië kan bogen op een vrij lange traditie en staat relatief op een hoog peil. (Dit maakt van de Tunesische planning overigens een geschikt objekt van evaluatie). Een bijzondere omstandigheid in Tunesië is de overgang van de politiek van collectivisatie van de jaren 60 naar een meer liberale gemengde economie. Dit schept de gelegenheid twee totaal verschillende planningsystemen binnen de context van één land te vergelijken. De behandeling van de landbouwplanning in het bijzonder geeft ook een specifiek karakter aan deze case-study. De landbouwsector kenmerkt zich immers door zijn heterogeniteit als een zeer omvangrijk gebied van planning, waarvan de trefzekerheid echter gering is. Men kan dan ook constateren dat de landbouwplanning zich vaak eenzijdig richt op de makkelijk grijpbare aspekten van de landbouwontwikkeling: infrastrukturele werken, de technische aspecten van de produktie, de grote landbouwbedrijven, de regio's met grote potenties. De sociale problematiek van de landbouw in ontwikkelingslanden (de lage inkomens en de werkloosheid) wordt relatief verwaarloosd.
Tenslotte moet worden opgemerkt dat de koers, die een klein land als Tunesië in de toekomst gaat varen, in hoge mate bepaald wordt door de internationale politieke situatie (relatie tot de EEG en tot Frankrijk in het bijzonder, relatie tot de andere Arabische landen, invloed van de V.S. en de Sowjet-Unie in het Middellandse zeegebied). Wij hebben deze randvoorwaarden niet in de beschouwing betrokken, in de eerste plaats orndat wij niet competent zijn hierover een wetenschappelijk oordeel te vellen, in de tweede plaats orndat wij menen dat de interne ontwikkeling, ook van kleine landen, niet uitsluitend door externe faktoren wordt bepaald. Er blijft een ruime marge voor een eigen ontwikkelingspolitiek, waarin het probleem van de efficiency van de planning zeer relevant is.
|Plattelandsontwikkeling in de derde wereld: een techniek van landbouwplanning
Luning, H.A. ; Thio, K.S. - \ 1977
Intermediair 13 (1977)43. - ISSN 0020-5605 - p. 47 - 55.
|Project Landbouwsectorplanning Tunesie: Verslag en evaluatie van een methodologisch onderzoek
Polman, F.J. ; Thio, K.S. ; Woersem, L.M. van - \ 1976
Pyridine nucleotide transhydrogenase
Broek, H.W.J. van den - \ 1971
Wageningen University. Promotor(en): C. Veeger. - Wageningen : Veenman - 117
oxidoreductasen - hydrogenase - pyridinenucleotiden - oxidoreductases - hydrogenase - pyridine nucleotides
A method for the isolation and purification of a reversible transhydrogenase from Azotobacter vinelandii is described (CHAPTER 3). The purification of the enzyme is hampered by association-dissociation phenomena, resulting in large losses of transhydrogenase activity. The relation between these losses and removal of lipoamide dehydrogenase, pyruvate dehydrogenase complex and a NADPH-ferricyanide diaphorase activity suggests the existence of a multi-enzyme complex, dissociation of this complex resulting in association of the transhydrogenase molecules.The purified transhydrogenase is a flavoprotein with FAD as prosthetic group. The absorption spectrum of the NADP +-free enzyme is characterized by a maximum at 442 nm. A minimum molecular weight of 60,000 daltons can be calculated from the experimentally determined extinction coefficient at 442 nm. The oxidized enzyme is stable at elevated temperatures and at high dilution; storage at -20° results in aggregation of the enzyme. The reduced enzyme is thermolabile; inactivation at elevated temperatures can be prevented by addition of FAD. It is difficult to prepare the apoenzyme and incubation with FAD only results in partial restoration of transhydrogenase activity.Ultracentrifugation and light-scattering studies demonstrate that the transhydrogenase preparations are inhomogeneous. In two types of enzyme preparations three sedimentation coefficients (s 20,w ) are determined: 24, 48 and 88 S, respectively. By addition of NADPH and NADP +upon sedimentation of the enzyme a relation is demonstrated between the 48 and 88 S components. The sedimentation coefficient of the main component is concentration independent beyond 1 mg/ml. From light-scattering experiments a molecular weight (M z ) of 30-50 million daltons is determined; the calculated length for a rod-like structure is 10,000-15,000 Å .Electron microscopic investigations (CHAPTER 4) confirm the inhomogeneity of the transhydrogenase preparations. Long helical-like structures up to 10,000 Å are present apart from large amounts of rod-like structures and separate fragments. The diameter of both the threads and rods is 116-120 Å . No clearcut substructures are observable; possibly they represent projections of sphericle particles; the threads may represent spiral-like structures. Addition of NADP +to the enzyme solution can result in fragmentation of the thread-like structures, but the dissociation is never complete. Extensive removal of NADP +results in more homogeneous preparations with structures up to 15,000-18,000 Å in length. The ratio NADPH/NADP +seems to be regulatory for the length of the structure. Other nucleotides have no clear-cut effect on the structure of the enzyme.In some preparations a totally unknown, ladder-like structure is observed. The constituents are very similar to the thread-like structures of the transhydrogenase and the tetramer-like structures, present in pyruvate dehydrogenase complex preparations. Only transhydrogenase and transacetylase activities are detectable in these preparations. This structure supports the idea of a complex between the transhydrogenase and the pyruvate dehydrogenase. The ladder-like structures resist the influence of the different nucleotides.Ammonium sulfate crystalline suspensions of the enzyme show regular structures, some kind of 'stacked disc' structures; solubilisation results in the appearance of thread-like structures, fragments and rosettes, but no close relation between the different species can be established.From an examination of the different purification stages it can be concluded that the thread-like transhydrogenase structure is not a purification artefact due to the procedure followed; however, a relation seems to exist between the presence of the thread-like structures, the concentration of the transhydrogenase and the-removal of the pyruvate dehydrogenase complex.Spectral studies (CHAPTER 5) have shown that 1 mole NADP +(per mole of flavin) can be easily bound to the oxidized enzyme. This results in a spectral shift in the region 300-500 nm. of the absorption spectrum, a quenching of flavin fluorescence and a shift of the emission maximum; moreover it affects the ORD and CD characteristics.. The dissociation constant (K D ) of this NADP +complex is ~ 3-4 μM. Changes in the 300-350 nm region of the absorption spectrum at higher NADP +concentrations point to the possible existence of a second binding site for NADP +. These shifts are possibly due to either changes in polarity around the flavin, or to interaction between NADP +and the isoalloxazine ring of the flavin, or to weakening of ribityl side chain interactions; protein conformational changes, however, cannot be excluded.Anaerobic titrations of the transhydrogenase with NAD(P)H in the presence of NADase show that two moles of reductant are required to reduce one mole of enzyme-bound flavin; this observation is confirmed by titrations with sodium dithionite. The existence of an unknown reducible group X next to the flavin is proposed. On the basis of the differences found in the titrations with dithionite, with NADPH in the absence and presence of NADP +and with NADH in the absence and presence of NADP +and NAD +a reduction scheme is proposed. The titration experiments show that NAD +and NADP +interact in a different manner with the different forms of the enzyme, e.g. NADP +has a higher affinity for the oxidized enzeym, while NAD +has a higher affinity for the reduced enzyme. Furthermore the level of reduction of the enzyme-bound flavin is determined by the ratio of reduced and oxidized pyridine nucleotides. Titration experiments of the reduced enzyme with NADP +and photoreduction experiments support the proposals. On the basis of these findings a model is proposed giving an explanation for a specific hydrogen transfer from the 4B site of NADH to the 4B site of NADP +.Kinetic studies (CHAPTER 6) performed with naturally occurring coenzymes and their thio-analogues point to the existence of a ping-pong bi-bi mechanism for the NADH-TNAD +and NADPH-TNAD +reactions. The inhibition of NAD +with respect to NADH in the reaction NADH-TNAD +and the dependence of the reaction velocity on [NADPH] 2in the reaction NADPH-TNAD +in the presence of NADP +, however, exclude this mechanism. A ternary complex mechanism is proposed, in which the enzyme is first reduced to the 4-equivalent reduced state by 2 moles of donor before 2 moles of acceptor are bound to the reduced enzyme; the possibility of complex formation of the oxidised enzyme with NADP +is included in the scheme. The kinetic patterns of the NADPH-TNAD +reaction in the absence and presence of NADP +can be explained by this mechanism. The explanation of the kinetic patterns of the NADH-TNAD +reaction can be given by slightly modifying the mechanism in such a way that the sequence of addition of the acceptor molecule and the second donor molecule is changed and assuming irreversible binding of the first acceptor molecule to the reduced enzyme; this latter assumption is justified by the high affinity of NAD +for the reduced enzyme. The reactions of NADP +and TNADP +with NAD(P)H proceed due to the reversible binding of both substrates according to a rapid equilibrium random bi-bi mechanism. The data of the reaction NADH-NADP +and the NADP +inhibition patterns of the reaction NADPH-TNADP +are consistent with this mechanism. The proposed ternary complex or random bi-bi mechanisms are more reasonable for a 4B-4B hydrogen transfer than a ping-pong bi-bi mechanism.In CHAPTER 7 results are presented from studies on Azotobacter lipoamide dehydrogenase. The results show that the Azotobacter enzyme has several properties in common with the pig heart lipoamide dehydrogenase. Gel filtration and ultracentrifugation studies show that the holoenzyme and the apoenzyme have a molecular weight of 102,000 and 52,000 daltons, respectively, supporting the idea of the existence of a monomer-dimer system. The apoenzyme-FAD complex, however, does not show a pronounced DCIP activity; the absorption spectrum points to a more polar environment of the flavin.Kinetic studies with reduced lipoamide and NAD +support, as proposed for the pig heart enzyme, an ordered bi-bi mechanism. Assuming this mechanism, kinetic parameters are calculated which are in the same order of magnitude as for the pig heart enzyme. The affinity of the Azotobacter enzyme for both substrates, however, is much higher.The apoenzyme prepared according to the acid ammonium sulfate procedure is very stable and can be easily reconstituted with FAD; the recombination process is temperature and FAD dependent. Preparation of the apoenzyme by dialysis against guanidine-HCl, however, results in a totally different structure, as concluded from recombination experiments and from the different characteristics with respect to the holoenzyme and the acid ammonium sulfate treated enzyme.