Non-enzymatic Degradation of (1→3)(1→4)-β-D-glucan in Aqueous Processing of Oats

Cereal water-soluble β-glucan [(1→3)(1→4)-β-D-glucan] has well-evidenced health benefits and it contributes to the texture properties of foods. These functions are characteristically dependent on the excellent viscosity forming ability of this cell wall polysaccharide. The viscosity is affected by the molar mass, solubility and conformation of β-glucan molecule, which are further known to be altered during food processing. This study focused on demonstrating the degradation of β-glucan in water solutions following the addition of ascorbic acid, during heat treatments or high pressure homogenisation. Furthermore, the motivation of this study was in the non-enzymatic degradation mechanisms, particularly in oxidative cleavage via hydroxyl radicals. The addition of ascorbic acid at food-related concentrations (2-50 mM), autoclaving (120°C) treatments, and high pressure homogenisation (300-1000 bar) considerably cleaved the β-glucan chains, determined as a steep decrease in the viscosity of β-glucan solutions and decrease in the molar mass of β-glucan. The cleavage was more intense in a solution of native β-glucan with co-extracted compounds than in a solution of highly purified β-glucan. Despite the clear and immediate process-related degradation, β-glucan was less sensitive to these treatments compared to other water-soluble polysaccharides previously reported in the literature. In particular, the highly purified β-glucan was relatively resistant to the autoclaving treatments without the addition of ferrous ions. The formation of highly oxidative free radicals was detected at the elevated temperatures, and the formation was considerably accelerated by added ferrous ions. Also ascorbic acid pronounced the formation of these oxidative radicals, and oxygen was simultaneously consumed by ascorbic acid addition and by heating the β-glucan solutions. These results demonstrated the occurrence of oxidative reactions, most likely the metal catalysed Fenton-like reactions, in the β-glucan solutions during these processes. Furthermore, oxidized functional groups (carbonyls) were formed along the β-glucan chain by the treatments, including high pressure homogenisation, evidencing the oxidation of β-glucan by these treatments. The degradative forces acting on the particles in the high pressure homogenisation are generally considered to be the mechanical shear, but as shown here, carbohydrates are also easily degraded during the process, and oxidation may have a role in the modification of polysaccharides by this technique. In the present study, oat β-glucan was demonstrated to be susceptible to degradation during aqueous processing by non-enzymatic degradation mechanisms. Oxidation was for the first time shown to be a highly relevant degradation mechanism of β-glucan in food processing.

New chelating resins based on polybenzimidazole. Selective sorption of copper(II) from ammoniacal media on resin with anchored l-cysteine

Microporous polybenzimidazole (PBI) of 250–500 μm bead size has been epoxidized and subsequently reacted with l-cysteine in the presence of a phase-transfer catalyst at room temperature to obtain a sorbent having anchored l-cysteine, EPBI(Cyst). The sorption of Cu(II), Ni(II), Co(II), and Zn(II) in mildly acidic and ammoniacal solutions has been measured under comparable conditions on EPBI(Cyst) and Dowex 50W-X8(H+) resins. While the latter shows no appreciable difference in sorption of the four metals in acidic and ammoniacal media and has 40–60 % selectivity for copper(II) over the other three, EPBI(Cyst) shows a threefold increase in copper sorption and more than 90% copper selectivity over the other metals in ammoniacal media, compared to mildly acidic media. The copper binding constant and saturation capacity of EPBI(Cyst) in ammoniacal media decrease only slowly beyond pH 11.6 with the result that the resin shows significant sorption of Cu(II) even in strongly ammoniacal solutions. The sorbed copper is stripped with HCl relatively easily. The copper sorption kinetics on EPBI(Cyst) is unusually fast in ammoniacal media with more than 90 % of equilibrium sorption being attained in one minute.

Cloning, expression, purification and preliminary X-ray crystallographic studies of 2-methylisocitrate lyase from Salmonella typhimurium. Acta Crystallogr

In Salmonella typhimurium, propionate is oxidized to pyruvate via the 2-methylcitric acid cycle. The last step of this cycle, the cleavage of 2-methylisocitrate to succinate and pyruvate, is catalysed by 2-methylisocitrate lyase (EC 4.1.3.30). Methylisocitrate lyase (molecular weight 32 kDa) with a C-terminal polyhistidine affinity tag has been cloned and overexpressed in Escherichia coli and purified and crystallized under different conditions using the hanging-drop vapour-diffusion technique. Crystals belong to the orthogonal space group P2(1)2(1)2(1), with unit-cell parameters a = 63.600, b = 100.670, c = 204.745 Angstrom. A complete data set to 2.5 Angstrom resolution has been collected using an image-plate detector system mounted on a rotating-anode X-ray generator.

Mesoporous silica- and silicon-based materials as carriers for poorly water soluble drugs

New chemical entities with unfavorable water solubility properties are continuously emerging in drug discovery. Without pharmaceutical manipulations inefficient concentrations of these drugs in the systemic circulation are probable. Typically, in order to be absorbed from the gastrointestinal tract, the drug has to be dissolved. Several methods have been developed to improve the dissolution of poorly soluble drugs. In this study, the applicability of different types of mesoporous (pore diameters between 2 and 50 nm) silicon- and silica-based materials as pharmaceutical carriers for poorly water soluble drugs was evaluated. Thermally oxidized and carbonized mesoporous silicon materials, ordered mesoporous silicas MCM-41 and SBA-15, and non-treated mesoporous silicon and silica gel were assessed in the experiments. The characteristic properties of these materials are the narrow pore diameters and the large surface areas up to over 900 m²/g. Loading of poorly water soluble drugs into these pores restricts their crystallization, and thus, improves drug dissolution from the materials as compared to the bulk drug molecules. In addition, the wide surface area provides possibilities for interactions between the loaded substance and the carrier particle, allowing the stabilization of the system. Ibuprofen, indomethacin and furosemide were selected as poorly soluble model drugs in this study. Their solubilities are strongly pH-dependent and the poorest (< 100 µg/ml) at low pH values. The pharmaceutical performance of the studied materials was evaluated by several methods. In this work, drug loading was performed successfully using rotavapor and fluid bed equipment in a larger scale and in a more efficient manner than with the commonly used immersion methods. It was shown that several carrier particle properties, in particular the pore diameter, affect the loading efficiency (typically ~25-40 w-%) and the release rate of the drug from the mesoporous carriers. A wide pore diameter provided easier loading and faster release of the drug. The ordering and length of the pores also affected the efficiency of the drug diffusion. However, these properties can also compensate the effects of each other. The surface treatment of porous silicon was important in stabilizing the system, as the non-treated mesoporous silicon was easily oxidized at room temperature. Different surface chemical treatments changed the hydrophilicity of the porous silicon materials and also the potential interactions between the loaded drug and the particle, which further affected the drug release properties. In all of the studies, it was demonstrated that loading into mesoporous silicon and silica materials improved the dissolution of the poorly soluble drugs as compared to the corresponding bulk compounds (e.g. after 30 min ~2-7 times more drug was dissolved depending on the materials). The release profile of the loaded substances remained similar also after 3 months of storage at 30°C/56% RH. The thermally carbonized mesoporous silicon did not compromise the Caco-2 monolayer integrity in the permeation studies and improved drug permeability was observed. The loaded mesoporous silica materials were also successfully compressed into tablets without compromising their characteristic structural and drug releasing properties. The results of this research indicated that mesoporous silicon/silica-based materials are promising materials to improve the dissolution of poorly water soluble drugs. Their feasibility in pharmaceutical laboratory scale processes was also confirmed in this thesis.

Direct Growth of Gold Nanorods on Gold and Indium Tin Oxide Surfaces: Spectral, Electrochemical, and Electrocatalytic Studies

The surfactant-assisted seed-mediated growth method was used for the formation of gold nanorods (GNRs) directly on gold (Au) and indium tin oxide (ITO) surfaces. Citrate-stabilized similar to 2.6 nm spherical gold nanoparticles (AuNPs) were first self-assembled on ITO or Au surfaces modified with (3-mercaptopropyl)-trimethoxysilane (MPTS) sol-gel film and then immersed in a cationic surfactant growth solution to form GNRs. The growth of GNRs on the MPTS sol gel film modified ITO surface was monitored by UV-visible spectroscopy. The ITO surface with the attached spherical AuNPs shows a surface plasmon resonance band at 550 nm. The intensity of this absorption band increases while increasing the immersion time of the AuNP-modified ITO surface into the growth solution, and after 5 h, an additional shoulder band around 680 nm was observed. The intensity of this shoulder band increases, and it was shifted to longer wavelength as the immersion time of the AuNP-modified ITO surface into the growth solution increases. After 20 h, a predominant wave at 720 nm was observed along with a band at 550 nm. Further immersion of the modified ITO surface into the growth solution did not change the absorption characteristics. The bands observed at 550 and 720 nm were characteristics of GNRs, corresponding to transverse and longitudinal waves, respectively. The AFM images showed the presence of GNRs on the surface of the MPTS sol gel modified ITO surface with a typical length of similar to 100-120 nm and a width of similar to 20-22 nm in addition to a few spherical AuNPs, indicating that seeded spherical AuNPs were not completely involved in the GNRs' formation. Finally, the electrocatalytic activity of the surface-grown GNRs on the MPTS sol gel film modified Au electrode toward the oxidation of ascorbic acid (AA) was studied. Unlike a polycrystalline Au electrode, the surface-grown GNR-modified electrode shows two well-defined voltammetric peaks for AA at 0.01 and 0.35 V in alkaline, neutral, and acidic pHs. The cause for the observed two oxidation peaks for AA was due to the presence of both nanorods and spherical nanoparticles on the electrode surface. The presence of spherical AuNPs on the MPTS sol gel film oxidized AA at more positive potential, whereas the GNRs oxidized AA at less positive potential. The observed 340 mV less positive potential shift in the oxidation of AA suggested that GNRs are better electrocatalysts for the oxidation of AA than the spherical AuNPs.

A study of symbolic processing and computational aspects in helicopter dynamics

Even research models of helicopter dynamics often lead to a large number of equations of motion with periodic coefficients; and Floquet theory is a widely used mathematical tool for dynamic analysis. Presently, three approaches are used in generating the equations of motion. These are (1) general-purpose symbolic processors such as REDUCE and MACSYMA, (2) a special-purpose symbolic processor, DEHIM (Dynamic Equations for Helicopter Interpretive Models), and (3) completely numerical approaches. In this paper, comparative aspects of the first two purely algebraic approaches are studied by applying REDUCE and DEHIM to the same set of problems. These problems range from a linear model with one degree of freedom to a mildly non-linear multi-bladed rotor model with several degrees of freedom. Further, computational issues in applying Floquet theory are also studied, which refer to (1) the equilibrium solution for periodic forced response together with the transition matrix for perturbations about that response and (2) a small number of eigenvalues and eigenvectors of the unsymmetric transition matrix. The study showed the following: (1) compared to REDUCE, DEHIM is far more portable and economical, but it is also less user-friendly, particularly during learning phases; (2) the problems of finding the periodic response and eigenvalues are well conditioned.

Diruthenium(III) compounds, Ru2O(O2CAr)2(MeCN)4(PPh3)2(ClO4)2.cntdot.H2O and Ru2O(O2CAr)4(PPh3)2, with an (Ru2(.mu.-O)(.mu.-O2CAr)22+) core as model systems for the diiron centers in hemerythrin

Diruthenium(II1) compounds, Ru20(02CAr)2(MeCN)4(PPh3)2(C104)(z1~) Hazn0d R U ~ O ( O ~ C A ~ ) ~(2() P(PA~r ~= )P~h,C6H4-p-OMe), were prepared by reacting R U ~ C I ( O ~ CaAnd~ P)P~h 3 in MeCN and characterized by analytical and spectral data. The molecular structures of 1 with Ar = Ph and of 2 with Ar = C&p-OMe were determined by X-ray crystallography. Crystal data for Ru~~(~~CP~)~(M~CN),(PP~(~la)):~ m(oCnIoc~lin,ic), n~/~cH, ~a O= 27.722 (3) A, b = 10.793 (2) A, c = 23.445 ( 2 )A , fi = 124.18 (l)', V = 5803 A3, and 2 = 4. Cr stal data for Ru~O(O~CC~H~-~-O(M2b~): )o~rth(orPhoPm~bi~c, )Pn~n a, a = 22.767 (5) A, b = 22.084 (7) A, c = 12.904 (3) 1, V = 6488 AS; and 2 = 4. Both 1 and 2 have an (Ruz0(02CAr)z2t1 core that is analogous to the diiron core present in the oxidized form of the nonheme respiratory protein hemerythrin. The Ru-Ru distances of 3.237 (1) and 3.199 ( I ) A observed in 1 and 2, respectively, are similar to the M-M distances known in other model systems. The essentially diamagnetic nature of 1 and 2 is due to the presence of two strongly interacting t22 Ru"' centers. The intense colors of 1 (blue) and 2 (purple) are due to the charge-transfer transition involving an ( R ~ ~ ( f i - 0m)o~ie~ty.) The presence of labile MeCN and carboxylato ancillary ligands in I and 2, respectively, makes these systems reactive toward amine and heterocyclic bases.

A New Pathway for the Degradation of a Sesquiterpene Alcohol, Nerolidol by Alcaligenes eutrophus

An oxidative pathway hitherto unknown for tile degradation of a sesquiterpene alcohol, nerolidol (I) by Alcaligenes eutrophus is presented. Fermentation of nerolidol (I) by this organism in a mineral salts medium resulted in the formation of geranylacetone (II) and an optically active alcohol (S)-(+)-geranylacetol (III), as major metabolites. Nerolidol (I) induced cells readily transformed 1,2-epoxynerolidol (IV) and 1,2-dihydroxynerolidol (V) into geranylacetone (II). These cells also exhibited their ability to carry out stereospecific reduction of II into (S)-(+)-geranylacetol (III). Oxygen uptake studies clearly indicated that nerolidol induced cells oxidized compounds II, III, IV, V and ethyleneglycol. Based on these observations a new oxidative pathway for the degradation of I is suggested which envisages the epoxidation of the terminal double bond, opening of the epoxide and cleavage between C-2 and C-3 in a manner similar to the periodate oxidation of diol.

Rihmamatolääkkeiden vaikutuskohteita. Betuliinin johdannaisia mahdollisiksi uusiksi yhdisteiksi

Jokisokeus eli onkosersiaasi on ihmisen loismatotauti, jota aiheuttaa Onchocerca volvulus -rihmamato. Tautia esiintyy trooppisilla alueilla Afrikassa ja Latinalaisessa Amerikassa. Tartunnan saaneita on noin 37 miljoonaa. Jokisokeus ilmenee iho- ja silmäoireina. Oireet johtuvat loisen nuorimmista muodoista eli mikrofilarioista. Jokisokeutta vastaan on käyty jakamalla lähinnä mikrofilarioihin tehoavaa ivermektiiniä. Tarvetta olisi lääkkeelle, joka tappaisi aikuiset madot tai steriloisi naaraat. Rokote olisi vielä parempi vaihtoehto. Antibiootit on uusi hoitokeino, sillä O. volvuluksella on elintärkeänä symbionttina Wolbachia-bakteeri. Doksisykliini tappaa vähintään 60 prosenttia aikuisista madoista ja steriloi naaraita, mutta kuuri kestää viikkoja. Yksi lupaava yhdiste on emodepsidi, jolla on loismatolääkkeille uusi vaikutusmekanismi. Rihmamatolääkkeiksi on testattu lukuisia yhdisteitä. Jotkut niistä inhiboivat entsyymejä, joilla madot kiertävät ihmisen immuunipuolustusta. Toiset häiritsevät neljä kertaa tapahtuvaa nahanvaihtoa. Hyvä lääkkeiden vaikutuskohde on loiselle välttämätön mutta puuttuu nisäkkäiltä. Betuliini on triterpeeni, jota on runsaasti koivun tuohessa. Betuliini ja monet sen johdannaiset ovat farmakologisesti aktiivisia yhdisteitä, joita tutkitaan etenkin syöpä- ja HIV-lääkkeiksi. Helsingin yliopiston lääkekemian ryhmä on syntetisoinut ja tutkinut lukuisia johdoksia. Jotkin niistä ovat lupaavia esimerkiksi Leishmania-alkueläimiin, Chlamydia pneumoniae -bakteeriin ja alfaviruksiin. Siksi yhdisteitä kannattaisi tutkia muihinkin taudinaiheuttajiin, kuten rihmamatoihin. Sekä Wolbachialla että C. pneumoniaella on sama lipidisynteesireitti, joka on kummallekin elintärkeä. Betuliinista syntetisoitiin johdoksia, joissa betuliinin alkoholeja on hapetettu karbonyyleiksi ja joihin on liitetty typpiheterorengas. Sekä Leishmania donovani että L. braziliensis -tutkimuksissa tehokkain oli formyylibetuliinin heterosykli. Vaikka valmistettuja yhdisteitä ei ole tutkittu rihmamatotesteissä, jatkossa voisi syntetisoida johdannaisia, joissa karbonyylien tilalla on typpirakenteita, koska C. pneumoniaehen hyvin tehonneessa yhdisteessä betuliinin OH-ryhmien tilalla on oksiimi.

Intercalation of n-alkylamines in iron thiohypophosphate (FePS3)

The intercalation of linear alkylamines (C1-C4) in the two-dimensional (2D) Ising antiferromagnet, FePS3, has been investigated. Intercalation proceeds with a dilation of the interlayer distance. The expansion (approximately 3.8 angstrom) is the same for all four amine molecules, suggesting that they are oriented flat with respect to the layers. From an analysis of the products of deintercalation, it is concluded that the intercalated species are the alkylammonium cations and neutral amine molecules. The intercalated compounds are highly moisture sensitive, as reflected in the chemical nature of the intercalated species. Charge neutrality of the lattice after intercalation is preserved by the loss of Fe2+ ions from the lattice. These Fe2+ ions are further oxidized to form superparamagnetic Fe2O3 clusters, as confirmed by Mossbauer spectra and magnetic measurements. This was further corroborated by in situ EPR studies. The Fe-57 Mossbauer spectra of the intercalated compounds showed evidence for two species other than Fe2O3. On the basis of the observed isomer shifts and quadrupole splittings, they have been assigned to Fe2+ in an environment similar to that in FePS3 and in a distorted FePS3 environment. The temperature and field dependence of the magnetic susceptibility of single crystals of the amine-intercalated FePS3 have been measured. Their magnetic behavior shows many of the features expected of a 2D Ising antiferromagnet with random defects, Fe1-xPS3, in agreement with the mechanism of intercalation.

Conformational analysis and design of cross-strand disulfides in antiparallel beta-sheets

Cross-strand disulfides bridge two cysteines in a registered pair of antiparallel beta-strands. A nonredundant data set comprising 5025 polypeptides containing 2311 disulfides was used to study cross-strand disulfides. Seventy-six cross-strand disulfides were found of which 75 and 1 occurred at non-hydrogen-bonded (NHB) and hydrogen-bonded (HB) registered pairs, respectively. Conformational analysis and modeling studies demonstrated that disulfide formation at HB pairs necessarily requires an extremely rare and positive chi(1) value for at least one of the cysteine residues. Disulfides at HB positions also have more unfavorable steric repulsion with the main chain. Thirteen pairs of disulfides were introduced in NHB and HB pairs in four model proteins: leucine binding protein (LBP), leucine, isoleucine, valine binding protein (LIVBP), maltose binding protein (MBP), and Top7. All mutants LIVBP T247C V331C showed disulfide formation either on purification, or on treatment with oxidants. Protein stability in both oxidized and reduced states of all mutants was measured. Relative to wild type, LBP and MBP mutants were destabilized with respect to chemical denaturation, although the sole exposed NHB LBP mutant showed an increase of 3.1 degrees C in T-m. All Top7 mutants were characterized for stability through guanidinium thiocyanate chemical denaturation. Both exposed and two of the three buried NHB mutants were appreciably stabilized. All four HB Top7 mutants were destabilized (Delta Delta G(0) = -3.3 to -6.7 kcal/mol). The data demonstrate that introduction of cross-strand disulfides at exposed NHB pairs is a robust method of improving protein stability. All four exposed Top7 disulfide mutants showed mild redox activity. Proteins 2011; 79: 244-260. (C) 2010 Wiley-Liss, Inc.

Competing peroxidase and oxidase reactions in scopoletin-dependent H2O2-initiated oxidation of NADH by horseradish peroxidase

Addition of NADH inhibited the peroxidative loss of scopoletin in presence of horseradish peroxidase and H2O2 and decreased the ratio of scopoletin (consumed):H2O2 (added). Concomitantly NADH was oxidized and oxygen was consumed with a stoichiometry of NADH: O-2 of 2:1. On step-wise addition of a small concentration of H2O2 a high rate of NADH oxidation was obtained for a progressively decreasing time period followed by termination of the reaction with NADH:H2O2 ratio decreasing from about 40 to 10. The rate of NADH oxidation increased linearly with increase in scopoletin concentration. Other phenolic compounds including p-coumarate also supported this reaction to a variable degree. A 418-nm absorbing compound;d accumulated during oxidation of NADH. The effectiveness of a small concentration of H2O2 in supporting NADH oxidation increased in presence of SOD and decreased in presence of cytochrome c, but the reaction terminated even in their presence. The results indicate that the peroxidase is not continuously generating H2O2 during scopolerin-mediated NADH oxidation and that both peroxidase and oxidase reactions occur simultaneously competing for an active form of the enzyme.

Evidence for the involvement of multiple pathways in the biodegradation of 1- and 2-methylnaphthalene by pseudomonas putida CSV86

Pseudomonas putida CSV86, a soil bacterium, grows on 1- and 2-methylnaphthalene as the sole source of carbon and energy. In order to deduce the pathways for the biodegradation of 1- and 2-methylnaphthalene, metabolites were isolated from the spent medium and purified by thin layer chromatography. Emphasis has been placed on the structural characterisation of isolated intermediates by CC-MS, demonstration of enzyme activities in the cell free extracts and measurement of oxygen uptake by whole cells in the presence of various probable metabolic intermediates. The data obtained from such a study suggest the possibility of occurrence of multiple pathways in the degradation of 1- and 2-methylnaphthalene. We propose that, in one of the pathways, the aromatic ring adjacent to the one bearing the methyl moiety is oxidized leading to the formation of methylsalicylates and methylcatechols. In another pathway the methyl side chain is hydroxylated to -CH2-OH which is further converted to -CHO and -COOH resulting in the formation of naphthoic acid as the end product. In addition to this, 2-hydroxymethylnaphthalene formed by the hydroxylation of the methyl group of 2-methylnaphthalene undergoes aromatic ring hydroxylation. The resultant dihydrodiol is further oxidised by a series of enzyme catalysed reactions to form 4-hydroxymethyl catechol as the end product of the pathway.

Novel synthesis of alpha-SI3N4 fibers

Alpha-Si3N4 fibres have been synthesized by carbothermal reduction and nitridation of pre-oxidized SiO1.7. The fibres were characterized using X-ray diffraction, infrared spectroscopy and electron microscopic techniques. The likely mechanism of reaction has been outlined

A pathway for biodegradation of 1-naphthoic acid by Pseudomonas maltophilia CSV89

Pseudomonas maltophilia CSV89, a bacterium isolated from soil in our laboratory, grows on 1-naphthoic acid as the sole source of carbon and energy. To elucidate the pathway for degradation of 1-naphthoic acid, the metabolites were isolated from spent medium, purified by TLC, and characterized by gas chromatography-mass spectrometry. The involvement of various metabolites as intermediates in the pathway was established by demonstrating relevant enzyme activities in cell-free extracts, oxygen uptake and transformation of metabolites by the whole cells. The results obtained from such studies suggest that the degradation of 1-naphthoic acid is initiated by double hydroxylation of the aromatic ring adjacent to the one bearing the carboxyl group, resulting in the formation of 1,2-dihydroxy-8-carboxynaphthalene. The resultant diol was oxidized via 3-formyl salicylate, 2-hydroxyisophthalate, salicylate and catechol to TCA cycle intermediates.