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Pre-hydrolysis of the Phenyl Glycosidic Bond in a Model CompoundDeshpande, Sagar Nandkumar January 2008 (has links) (PDF)
No description available.
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Optimization of Hardwood Hemicellulose Hydrolysis by Xylanase from Trichoderma viride for the Development of an Integrated Forest BiorefineryGraham, LeRae Beth January 2008 (has links) (PDF)
No description available.
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Formadores de filmes híbridos orgânicos inorgânicos do tipo ureasil-poliéter para liberação controlada de fármacosMendes, Juliana Fernandes [UNESP] 27 June 2012 (has links) (PDF)
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mendes_jf_me_arafcf_parcial.pdf: 496742 bytes, checksum: 031c10b96f7070a5b508754d4d273375 (MD5) / Dentre os sistemas de liberação controlada de fármacos, os híbridos orgânico-inorgânicos destacam-se por apresentarem vantagens sobre os sistemas convencionais de liberação, tais como: flexibilidade, alta resistência mecânica e térmica, transparência e baixa solubilidade em água. Esses híbridos foram sintetizados através de uma rota química denominada sol-gel que usualmente utiliza como solvente o tetrahidrofurano (THF). O objetivo deste trabalho foi desenvolver formadores de filmes a partir de materiais híbridos com perfil de liberação controlada de fármacos e substituir o THF pelo etanol, por ser um solvente menos tóxico para aplicação na pele. Neste trabalho foram utilizados quatro polímeros, sendo dois à base de polióxido de etileno (POE) de massa molecular 500 e 1900 g mol-1 e dois à base de polióxido de propileno (POP) de massa molecular 400 e 2000 g mol-1. Os fármacos utilizados para o teste de liberação foram o cloridrato de pramoxina e o acetato de dexametasona. A caracterização desses materiais foi realizada por DSC, TG-DTA, DMA, infravermelho, RMN 1H, RMN 29Si, e SAXS. Os resultados das análises termo-mecânica revelaram que a substituição do solvente não influenciou na estabilidade térmica e na flexibilidade dos híbridos ureasil. Os resultados da caracterização estrutural por infravermelho, RMN e SAXS comprovaram a formação dos híbridos tanto em THF quanto em etanol, indicando que não houve mudanças estruturais significativas entre essas matrizes híbridas quando sintetizadas em ambos solventes. As curvas de SAXS coletadas in situ mostraram que o intumescimento na presença de água é acompanhado por um relaxamento da matriz polimérica, conduzindo a um afastamento dos grupos ureasil constituintes da molécula para a matriz híbrida do tipo POE 1900. O grau de... / Organic inorganic hybrids present several advantages as drug release systems such as: high flexibility, high mechanical and thermal resistance, transparency and low water solubility. These hybrids were synthesized by a chemical route called sol-gel usually used as the solvent tetrahydrofuran (THF). The aim of this study was to develop film formers from hybrid materials with drugs controlled release profile and to replace THF by ethanol, a less toxic solvent for skin application. In this work four polymers were used, two based on polyethylene oxide (PEO) of molecular weight 500 and 1900 g mol-1 and two based on polypropylene oxide (PPO) of molecular weight 400 and 2000 g mol1. The drugs used for the release experiment were pramoxine hydrochloride and dexamethasone acetate. The characterization of these materials was performed by DSC, TG-DTA, DMA, IR, NMR 1H, NMR 29Si, and SAXS. The results of thermo-mechanical analysis revealed that the solvent replacement has not affect the thermal stability and flexibility of the hybrid ureasil. The results of structural characterization by IR, NMR and SAXS confirmed the formation of hybrids both in THF and in ethanol, indicating no significant structural changes between these hybrid matrices when synthesized in both solvents. The SAXS measurements collected in situ showed that the swelling in the presence of water is followed by a relaxation of the polymer matrix, leading to an increase distance between siloxane nods present in the molecules of the hybrid matrix PEO-1900. The swelling degree of these matrices is related to the size of the polymer chain, to the ratio between the organic and inorganic domains and to the hydrophilic/hydrophobic ratio. The drugs incorporation was performed during the step of hydrolysis and condensation of hybrid matrices. The drug release experiments... (Complete abstract click electronic access below)
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Formadores de filmes híbridos orgânicos inorgânicos do tipo ureasil-poliéter para liberação controlada de fármacos /Mendes, Juliana Fernandes. January 2013 (has links)
Orientador: Leila Aparecida Chiavacci / Banca: Marlus Chorilli / Banca: Renata Fonseca Vianna Lopez / Resumo: Dentre os sistemas de liberação controlada de fármacos, os híbridos orgânico-inorgânicos destacam-se por apresentarem vantagens sobre os sistemas convencionais de liberação, tais como: flexibilidade, alta resistência mecânica e térmica, transparência e baixa solubilidade em água. Esses híbridos foram sintetizados através de uma rota química denominada sol-gel que usualmente utiliza como solvente o tetrahidrofurano (THF). O objetivo deste trabalho foi desenvolver formadores de filmes a partir de materiais híbridos com perfil de liberação controlada de fármacos e substituir o THF pelo etanol, por ser um solvente menos tóxico para aplicação na pele. Neste trabalho foram utilizados quatro polímeros, sendo dois à base de polióxido de etileno (POE) de massa molecular 500 e 1900 g mol-1 e dois à base de polióxido de propileno (POP) de massa molecular 400 e 2000 g mol-1. Os fármacos utilizados para o teste de liberação foram o cloridrato de pramoxina e o acetato de dexametasona. A caracterização desses materiais foi realizada por DSC, TG-DTA, DMA, infravermelho, RMN 1H, RMN 29Si, e SAXS. Os resultados das análises termo-mecânica revelaram que a substituição do solvente não influenciou na estabilidade térmica e na flexibilidade dos híbridos ureasil. Os resultados da caracterização estrutural por infravermelho, RMN e SAXS comprovaram a formação dos híbridos tanto em THF quanto em etanol, indicando que não houve mudanças estruturais significativas entre essas matrizes híbridas quando sintetizadas em ambos solventes. As curvas de SAXS coletadas in situ mostraram que o intumescimento na presença de água é acompanhado por um relaxamento da matriz polimérica, conduzindo a um afastamento dos grupos ureasil constituintes da molécula para a matriz híbrida do tipo POE 1900. O grau de... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Organic inorganic hybrids present several advantages as drug release systems such as: high flexibility, high mechanical and thermal resistance, transparency and low water solubility. These hybrids were synthesized by a chemical route called sol-gel usually used as the solvent tetrahydrofuran (THF). The aim of this study was to develop film formers from hybrid materials with drugs controlled release profile and to replace THF by ethanol, a less toxic solvent for skin application. In this work four polymers were used, two based on polyethylene oxide (PEO) of molecular weight 500 and 1900 g mol-1 and two based on polypropylene oxide (PPO) of molecular weight 400 and 2000 g mol1. The drugs used for the release experiment were pramoxine hydrochloride and dexamethasone acetate. The characterization of these materials was performed by DSC, TG-DTA, DMA, IR, NMR 1H, NMR 29Si, and SAXS. The results of thermo-mechanical analysis revealed that the solvent replacement has not affect the thermal stability and flexibility of the hybrid ureasil. The results of structural characterization by IR, NMR and SAXS confirmed the formation of hybrids both in THF and in ethanol, indicating no significant structural changes between these hybrid matrices when synthesized in both solvents. The SAXS measurements collected in situ showed that the swelling in the presence of water is followed by a relaxation of the polymer matrix, leading to an increase distance between siloxane nods present in the molecules of the hybrid matrix PEO-1900. The swelling degree of these matrices is related to the size of the polymer chain, to the ratio between the organic and inorganic domains and to the hydrophilic/hydrophobic ratio. The drugs incorporation was performed during the step of hydrolysis and condensation of hybrid matrices. The drug release experiments... (Complete abstract click electronic access below) / Mestre
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Characterization of the hydantoin-hydrolysing system of Pseudomonas putida RU-KM3sMatcher, Gwynneth Felicity January 2005 (has links)
The biocatalytic conversion of 5-monosubstituted hydantoin derivatives to optically pure amino acids involves two reaction steps: the hydrolysis of hydantoin to N-carbamylamino acid by an hydantoinase or dihydropyrimidinase enzyme, followed by conversion of the Ncarbamylamino acid to the corresponding amino acid by an N-carbamoylase enzyme. This biocatalytic process has been successfully applied in several industrial processes for the production of enantiomerically pure amino acids used in the synthesis of pharmaceuticals, insecticides, hormones, and food additives. P. putida RU-KM3S was selected for study based on inherent high levels of hydantoinase and N-carbamoylase activity. Subsequent biocatalytic analysis of the enzyme activity within this strain revealed unique properties thus prompting further characterization. The main focus of this research was the isolation of the genes encoding the hydantoin-hydrolysing pathway in RU-KM3S. A genomic library was constructed and screened for heterologous expression of the hydantoin-hydrolysing enzymes. However, this approach was unsuccessful prompting the use of transposon mutagenesis in order to circumvent the drawbacks associated with complementation studies. The enzymes responsible for hydantoin-hydrolysis were identified by insertional inactivation as a dihydropyrimidinase and b-ureidopropionase encoded by dhp and bup respectively. A third open reading frame, encoding a putative transport protein, was identified between the dhp and bup genes and appeared to share a promoter with bup. Analysis of the amino acid sequence deduced from bup and dhp substantiated the distinctive properties and potential industrial application of the L-enantioselective b-ureidopropionase and provided targets for potential optimisation of the substrate-selectivity and activity of the dihydropyrimidinase by site directed mutagenesis. Several transposon-generated mutants with an altered phenotype for growth on minimal medium with hydantoin as the sole source of nitrogen were also isolated. Analysis of the insertion events in these mutants revealed disruptions of genes encoding key elements of the Ntr global regulatory pathway. However, inactivation of these genes had no effect on the dihydropyrimidinase and b-ureidopropionase activity levels. An additional mutant in which the gene coding for the dihydrolipoamide succinyltransferase, which is involved in the TCA cycle, was isolated with reduced levels of both dihydropyrimidinase and b-ureidopropionase activities. These results indicated that the hydantoin-hydrolysis pathway in RU-KM3S is regulated by carbon rather than nitrogen catabolite repression. This was confirmed by the reduction of hydantoin-hydrolysis in cells grown in excess carbon as opposed to nitrogen. Identification of a putative CRP-binding site within the promoter region of these enzymes further supported the regulatory role of carbon catabolite repression (CCR). As CCR in Pseudomonads is poorly understood, elucidation of the mechanism by which the hydantoinhydrolysing pathway in RU-KM3S is regulated would provide valuable insight into this complex process.
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Studies on the mechanism of action of proteolytic enzymesHawkins, M. J. January 1965 (has links)
No description available.
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Studies on the enzymic hydrolysis of adenosine triphosphateWheeler, Kenneth Peter January 1964 (has links)
No description available.
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Sulphide-enhanced hydrolysis of primary sewage sludge : implications for the bioremediation of sulphate-enriched wastewatersWhittington-Jones, Kevin John January 2000 (has links)
The potential application of sulphate reducing bacteria for the bioremediation of acid mine drainage has already been recognised, and offers significant financial advantages over conventional chemical treatment approaches. Although the technology has been demonstrated successfully on both small- and large-scale, it’s extensive implementation has been constrained by the provision of suitable and cost effective electron donor and carbon sources. Primary sewage sludge is readily available in large quantities, but the slow rate of solubilization and low yield of soluble products do not apparently favour its use for this application. A number of pre-treatment steps have been introduced in an attempt to improve the yield and rates under methanogenic conditions. However, although early work suggested that degradation of lignocellulose and proteins may be more rapid under sulphate reducing conditions, the fate of primary sewage sludge under these conditions has been ignored. It was proposed that by combining the hydrolysis of primary sewage sludge and biological sulphate reduction, in a settling sludge bed, both processes would be enhanced. The aim of this study was to test this hypothesis on laboratory- and pilot-scale, and attempt to elucidate the underlying mechanism involved. The solubilization of primary sewage sludge was enhanced in the presence of sulphate reduction in continuous laboratory-scale reactors. Particulate matter accumulated in the bed of non-sulphidogenic systems, but not in sulphidogenic ones. This was attributed to increased solubilization and the smaller average floc size in the latter. Solubilization occurred within the settling sludge bed of the reactors, and offered a possible explanation for the better performance of the multiple- over single-stage reactor. A pilot-scale Falling Sludge Bed Reactor was constructed at Grootvlei Gold Mine, Springs, South Africa, and resulted in the solubilization of more than 70% of the influent primary sewage sludge. The system was also found to be highly resilient to severe perturbations, and returned rapidly to steady-state. Flask studies revealed that the hydrolysis of both proteins and complex carbohydrates was accelerated in the presence of biological sulphate reduction or sulphide. A study of the enzymology of sludge digestion revealed that sulphate reduction had little direct effect on the activity of the hydrolytic enzymes, but that reactor design was critical in the prevention of washout of these enzymes. Finally, a descriptive model was developed to explain the enhanced hydrolysis of primary sewage sludge. The model incorporated the effect of sulphidogenesis on floc fracture and reflocculation, and likely implications for mass transfer limitations.
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An investigation of the isolation, characterisation and application of hydantoinases for the industrial production of amino acidsKirchmann, Shaun January 2003 (has links)
This thesis describes a series of investigations into the hydantoin-hydrolysing activity of bacterial strains RU-KM1 and RU-OR, which were previously isolated for their ability to hydrolyse hydantoins to amino acids. The main aim of the study was to develop biotransformations with potential application in the production of enantiomerically pure amino acids using a bioreactor based system utilising the hydantoin hydrolysing enzymes of the two isolated microorganisms. Different substituted hydantoins may be used as substrates by these enzymes for the production of a variety of amino acids. These are not only important for amino acid production, but they may be used for production of other industrially important compounds, such as semisynthetic penicillin/ampicillin, L-aspartame (sweetener), Fluvalinate (insecticide), Enalapril (ACE inhibitor). Thus, the ability of the above-mentioned strains to hydrolyse these substrates was investigated, with the view to utilizing the maximum potential of these biocatalysts. Hydantoin conversion involves a two-step hydrolysis reaction which yields, initially, an N-carbamylamino acid intermediate, and subsequently, an amino acid. The hydantoin-hydrolysing enzymes of a Pseudomonas sp. RU-KM1, and an Agrobacterium sp. RU-OR were characterised as whole cells and in a crude extract preparation, and reaction conditions for its biocatalytic application were optimised. The optimum conditions for conversion of hydantoin to glycine were found to be 1 hour at 40 °C, with conversion yields greater than 30 % achieved. The enzymes of RU-KM1 demonstrated considerable stability, retaining 80 % of their activity after storage for 2 weeks at 4 °C. The activities of the enzymes were increased by the addition of a detergent to the extraction medium, suggesting that the enzymes might be membrane-bound. The results of the determination of the metal-dependence of the hydantoinase and N-carbamoylase of RU-KM1 suggested that these enzymes required metal ions for activity, with metal ions such as Cu[superscript (2+)], Fe[superscript (2+)], and Co[superscript (2+)] resulting in no significant change in enzyme activity, however there was an activation of the enzymes when Mn[superscript (2+)] was added to the enzymes. The stereoselectivity of the enzymes was investigated, and the results suggested that the hydantoinase was D-selective, whereas the N-carbamoylase was shown to be L-selective by other researchers. The hydantoin substrate selectivity of RU-KM1 and RU-OR was investigated, and the organisms were shown to be able to hydrolyse all of the seven substrates tested. However, there was a difference in activity levels between crude extract preparations and whole cells, with crude extracts generally showing slightly lower activity than whole cells in RU-KM1, and the whole cells or RU-OR showing the lower activity than its crude extract. Some difference was also observed in the order of preference of substrates between whole cells and crude extracts. The preferred substrate for RU-KM1 whole cells was isopropylhydantoin, whereas the crude extract preparation preferentially hydrolysed p-hydroxyphenylhydantoin, achieving 57 % and 52 % conversions respectively. RU-OR whole cells preferred methylhydantoin where as the crude extract preferred isopropylhydantoin, and showed 49 % and 51 % conversions respectively. The enzymes were characterised in terms of their temperature and pH optima, inducer requirements, and product inhibition studies. The hydantoinase of RU-KM1 was shown to be inducible with low levels of hydantoin, and thermostable upto 75 °C with its optima between 60 and 70 °C. The N-carbamoylase was shown to have its optima at 50 °C. The addition of ATP (0.5 mM), DTT (1 mM) and a protease inhibitor (2 mg.mL[superscript (-1)]) all increased the hydantoinase activity of RU-KM1 crude extract, however they had very little effect on the N-carbamoylase activity. The hydantoinase enzyme from extracts of RU-KM1 was partially purified by development of cell disruption methods using mechanical and lysing enzymes, followed by precipitation and chromatographic resolution. The results obtained showed a hydantoinase enzyme of between 48 and 66 kDa. RU-KM1 was grown under fermentation conditions using different minimal media. The activity and yields under these conditions were low. Previous attempt to grow the organism in a rich medium had resulted in an increase in biomass but no hydantoinase activity. A rich medium was developed by carbon and nitrogen optimisation and yielded biomass up to 30 g.L[superscript (-1)] dry cell weight. The hydantoinase activity was restored by nitrogen starvation in stationary phase. This resulted in high biomass with increased activity. This data is currently in press. Crude extract and whole cells were immobilised on flat sheet membranes, hollow fibre membranes and in alginate beads. Low hydantoinase activity was measured in bioreactors using membranes in different configurations. A significant increase in hydantoinase activity was measured when the crude extract was immobilised in sodium alginate, as a result of stabilisation of the N-carbamoylase. Temperature and pH optima were unaffected by the immobilisation procedure, however the durability of the enzymes increased 2-fold. Different configurations of the bioreactor were investigated, as well as a hydroxyphenylhydantoin as an alternative substrate in this study. The bioreactors showed a near 95 % conversion of the hydantoin to glycine, and a 99 % conversion using HPG. In conclusion, the hydantoin-hydrolysing enzymes of RU-KM1 have been shown to be possibly membrane associated, which is a novel finding. This study has shown that the hydantoinase of RU-KM1 is D-stereoselective, with high temperature stability. A growth medium was developed for the rapid production of active biomass. A bioreactor was developed using a single and a dual biocatalyst configuration, which was capable of hydrolysing hydantoin and monosubstituted hydantoins to produce amino acids. To our knowledge this system is the first such dual biocatalyst system reported for the production of amino acids.
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Characterization of amide bond hydrolysis in novel hydantoinase-producing bacteriaSkepu, Zoleka G January 2000 (has links)
This thesis describes a series of investigations into the amide bond-hydrolyzing activity of bacterial strains RU-KM1, RU-KM3L, RU-KM3S, and RU-OR, which were previously isolated for their ability to hydrolyze hydantoins to amino acids. The main aim of the study was to develop biotransformations with potential application in the production of enantiomerically pure amino acids and related compounds. Several compounds may be used as substrates by biocatalysts for the production of amino acids, such as hydantoins, amino nitriles, and amides. These compounds are not only important for amino acid production, but they may be used for production of other industrially important compounds, such as 2- arylpropionic acids, which are non-steroidal anti-inflammatory drugs. Thus, the ability of the above-mentioned strains to hydrolyze these substrates was investigated, with the view to utilizing the maximum potential of these biocatalysts. The compounds used as substrates in the investigation are all essentially amides. Thus, the ability of the strains to hydrolyze imides, hydantoins, and amides, was investigated. In particular, imides have a structure which is very similar to that of hydantoins, and thus it was an objective of the study to determine whether these strains could hydrolyze imides. Imidehydrolyzing activity has only recently been discovered in microorganisms. Hydantoin conversion involves a two-step hydrolysis reaction which yields, initially, an Ncarbamylamino acid intermediate, and subsequently, an "-amino acid. The hydantoinhydrolyzing enzymes of a Pseudomonas putida strain, RU-KM3S, were characterized in a crude extract preparation and reaction conditions for its biocatalytic application were optimized. The optimum conditions for conversion of 5-methylhydantoin were found to be 3 hours at 40°C, with conversion yields greater than 50% achieved. The enzymes of RU-KM3S demonstrated considerable stability, retaining 80% of their activity after incubation at 40°C for 3 hours. The activities of the enzymes were increased by the addition of a detergent to the extraction medium, suggesting that the enzymes might be membrane-bound. The results of the determination of the metal-dependence of the hydantoinase and N-carbamylase of RUKM3S suggested that these enzymes required metal ions for activity, with metal ions such as Mg²⁺, Mn²⁺, Zn²⁺, and Co²⁺ resulting in activation of the enzymes. However, Cu²⁺ and Fe²⁺ caused inactivation of these enzymes. The stereoselectivity of the enzymes was investigated, and the results suggested that the hydantoinase was non-selective, whereas the N-carbamylase was L-selective. The hydantoin substrate selectivity of RU-KM3S was compared to that of three other hydantoinase-producing bacteria, RU-KM1, RU-KM3L, and RU-OR. The four strains were able to hydrolyze all of the seven substrates tested. However, there was a difference in activity levels between crude extract preparations and whole cells, with crude extracts generally showing higher activity than whole cells, except in the case of RU-KM1. Some difference was also observed in the order of preference of substrates between whole cells and crude extracts. The preferred substrate for RU-KM1 whole cells was isopropylhydantoin, whereas the crude extract preparation preferentially hydrolyzed p-hydroxyphenylhydantoin. RU-KM3L whole cells achieved a higher conversion yield with isobutylhydantoin, whereas the crude extract achieved a higher yield with 5-t-butylhydantoin. RU-KM3S whole cells and crude extract preferentially hydrolyzed 5-n-butylhydantoin, although the yield was greater with the crude extract. The highest conversion yields were observed with RU-KM3S crude extract, with conversion yields of 71.6% and 100% for n-butylhydantoin and phydroxyphenylhydantoin, respectively.The ability of RU-KM1, RU-KM3L, and RU-KM3S to hydrolyze nitriles, initially to amides and subsequently to carboxylic acids, was investigated. These strains were demonstrated to be unable to utilize acrylonitrile, propionitrile and benzonitrile as nitrogen sources, but were able to hydrolyze acrylonitrile, propionitrile and acetonitrile, in resting cell reactions. Nitrile hydrolysis was demonstrated to be inducible in all three strains, and the enzyme system responsible for nitrile hydrolysis was proposed to be a nitrile hydratase-amidase system. Amidase activity in the four bacterial strains was investigated. The ability of RU-KM1, RUKM3L, RU-KM3S, and RU-OR to utilize amides as a nitrogen source was investigated, and the results showed that propionamide was a good nitrogen source for all four of the strains. Amide-hydrolyzing activity, by resting cells, was shown to be inducible by propionamide in all four strains. RU-KM3S demonstrated superior amide-hydrolyzing ability in that it hydrolyzed propionamide, acetamide, and acrylamide to a greater extent than the other strains. Resting cells of RU-KM1 and RU-OR were demonstrated to have the ability to hydrolyze the imide substrate, succinimide, and this imidase activity was found to be inducible. These strains were also able to utilize this imide as the sole source of nitrogen for growth, which is a novel finding, as to date, bacteria have only be reported to utilize imides as a carbon source. The identity of the enzyme system responsible for succinimide hydrolysis is not yet clear. In conclusion, the hydantoin-hydrolyzing enzymes of RU-KM3S have been shown to be possibly membrane associated, which is a novel finding that has also been proposed in three other hydantoinase-producing strains in our laboratory. This study has shown that the Ncarbamylase of RU-KM3S is L-stereoselective, which, to our knowledge, is the first report of an L-stereospecific N-carbamylase in a Pseudomonas putida. Publication of these findings is already in progress. This is the first report on the study of imide hydrolysis in either an Agrobacterium tumefaciens or a Pseudomonas sp., and publications reporting these results are in preparation.
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