Chemical stability of prostacyclin (PGI2) in aqueous solutions

The rate constant for the hydrolysis of prostacyclin (PGI₂) to 6-keto-PGF_1α was measured by monitoring the UV spectral change, over a pH range 6 to 10 at 25°C and the total ionic strength of 0.5 M. The first-order rate constant (k_obs) extrapolated to zero buffer concentration follows an expression, k_obs = k_H+ (H+), where k_H+ is a second-order rate constant for the specific acid catalyzed hydrolysis. The value of k_H+ obtained (3.71 × 10⁴ sec⁻¹ M⁻¹) is estimated approximately 700-fold greater than a k_H+ value expected from the hydrolysis of other vinyl ethers. Such an unusually high reactivity of PGI₂ even for a vinyl ether is attributed to a possible ring strain release that would occur upon the rate controlling protonation of C₅. A Brønsted slope (α) of 0.71 was obtained for the acid (including H₃O+) catalytic constants, from which a pH independent first-order rate constant for the spontaneous hydrolysis (catalyzed by H₂O as a general acid) was estimated to be 1.3 × 10⁻⁶ sec⁻¹. An apparent activation energy (E_a) of 11.85 Kcal/mole was obtained for the hydrolysis at pH 7.48, from which a half-life of PGI₂ at 4°C was estimated to be approximately 14.5 min. when the total phosphate concentration is 0.165 M (cf. 3.5 min. at 25°C).     

Isotopic fractionation factors of intermolecular hydrogen bonds in water

Association constants for N---H⁺…⁻O hydrogen bond formation between substituted ammonium dications and phenolate ion were measured in water and deuterium oxide at 25°C and 2.0 ionic strength. In combination with isotopic fractionation factors for phenol and the conjugate diacid of 1,2-ethanediamine determined by ¹³C NMR spectroscopy, these yield isotopic fractionation factors for amine dication-phenolate ion hydrogen bonds in water: φ_AB = 0.69 for 1,2-propanediamine dication with a pK difference between donor and acceptor, ΔpK_a = −0.45, φ_AB = 0.88 for 1,2-ethanediamine dication (ΔpK_a = −2.1), and φ_AB = 1.1 for piperizine dication (ΔpK_a = −3.5). The hydrogen bond association constants follow Brønsted correlations α = 0.19 in water and α = 0.27 in deuterium oxide. The results are consistent with a double-minimum potential with a significant barrier for motion across the hydrogen bond.     

Reactivity with p-nitrophenyl acetate and interaction between the amino and imidazole groups of histidine and related compounds

The study of the reaction of p-nitrophenyl acetate (PNPA) with histidine and certain derivatives showed that the species in which the amino group is unprotonated (R(NH₂)Im) react with second-order rate constants ( ) that are higher than predicted by a Brønsted relation for a series of neutral amino acids. The reason for this behavior was investigated through an analysis of the kinetics of the reaction of PNPA with these compounds in order to assess the reactivities of the amino and imidazole groups in the two species . The rate constant for the reaction with the imidazole group ( ) of N^π-methyl histidine agrees with the value predicted by a Brønsted relation obtained from a series of model imidazole compounds. N^τ-Methyl histidine, however, is unreactive, indicating that N^τ is the reactive nitrogen in the imidazole ring of histidine. The values found for histidine, histidine methyl ester, and N^α-dimethyl histidine are lower than predicted by the Brønsted relation. This behavior was found to be due to low reactivity of the

Full-size image

. The evidence presented suggests that the lower reactivity of is due to an ion-dipole interaction between the protonated amino group and the unprotonated imidazole ring, which displaces the tautomeric equilibrium toward the unreactive N^τ-H form. The higher reactivity of the imidazole group in the species R(NH₂)Im, relative to that in , is responsible for the observed high values for histidine, for histidine methyl ester, for N^τ-methyl histidine, and for N^α-dimethyl histidine, in contrast with the normal value found for N^τ-methyl histidine. The conclusions from this study of histidine and its derivatives support the proposal of an interaction between the protonated N-terminal amino group and the imidazole ring of His⁶ in the octapeptide hormone angiotensin.     

Specific growth rate as a determinant of the carbon isotope composition of the temperate seagrass Zostera marina

The seasonal variability of specific growth rate and the carbon stable isotope ratio (δ¹³C) of leaf blades (δ¹³C_leaf) of a temperate seagrass, Zostera marina (within 10 days old) were measured simultaneously, together with the δ¹³C of dissolved inorganic carbon (δ¹³C_DIC) at three sites in the semi-closed Akkeshi estuary system, northeastern Japan, in June, September, and November 2004. The δ¹³C_leaf ranged from −16.2 to −6.3‰ and decreased from summer to winter. The simultaneous measurement of the δ¹³C_leaf, growth rate, and morphological parameters (mean leaf length and width, mean number of leaves per shoot, and sheath length) of the seagrass and δ¹³C_DIC in the surrounding water allowed us to compare directly the δ¹³C_leaf and specific growth rate of seagrass. The difference in the δ¹³C of seagrass leaves relative to the source DIC (Δδ¹³C_{leaf − DIC}) was the least negative (−11 to −7‰) in June at all three sites and became more negative (−17 to −8‰) as the specific growth rate decreased. This positive correlation between Δδ¹³C_{leaf − DIC} and specific growth rate can be used to diagnose the growth of seagrasses. Δδ¹³C_{leaf − DIC} changed by −1.7 ± 0.2‰ when the leaf specific growth rate decreased by 1% d⁻¹.     

o-Phthalaldehyde–N-acetylcysteine polyamine derivatives: formation and stability in solution and in C18 supports

A comparative study of different derivatization procedures has been performed in order to improve the stability of the reaction products o-phthalaldehyde–N-acetylcysteine (OPA–NAC) polyamines. Procedures such as solution derivatization, solution derivatization followed by retention on a packing support, derivatization on different packing supports and on-column derivatization, have been optimized and compared. The degradation rate constant (k) of the derivative was dependent on the procedure used and on the analyte. For the spermine (the most unstable isoindol tested) k was 8±2×10⁻² min⁻¹ in solution versus 7.7±1.1×10⁻⁴ min⁻¹ on the (C₁₈) solid support. The results obtained showed that forming the derivative on the packing support (C₁₈) gave the best results following this procedure: conditioning the cartridges with borate buffer (1 ml, 0.5 M, pH 8), retention of the analyte, addition of 0.8 ml of OPA–NAC reagent, 0.2 ml borate buffer 0.8 M (pH 8) and elution of the isoindol with 3 ml of MeOH–borate buffer (9:1). The different derivatization procedures have been used to study the stability of the reaction products OPA–NAC polyamines formed in urine matrix using spermine as model compound. Similar results were obtained for standard solutions and urine samples.     

Leaf traits and photosynthetic parameters of saplings and adult trees of co-existing species in a temperate broad-leaved forest

In Central European forestry the establishment of broad-leaved mixed forests is attaining increasing importance, but little information exists about gas exchange characteristics of some of the tree species involved, which are less abundant today. In an old-growth forest in Central Germany (Hainich, Thuringia), (i) I compared morphological and chemical leaf traits that are indicative of leaf gas exchange characteristics among eight co-existing species, and (ii) analysed photosynthetic parameters of saplings and adult trees (lower and upper canopy level) in four of these species (Acer pseudoplatanus L., Carpinus betulus L., Fraxinus excelsior L. and Tilia platyphyllos Scop.).Leaves from the upper canopy in the eight species studied varied significantly in their specific leaf area (12.9–19.4 m² kg⁻¹), stomatal density (125–313 stomata mm⁻²), leaf nitrogen concentration (95–157 mmol N m⁻²) and δ¹³C content (–27.81 to –25.85‰). F. excelsior and C. betulus were largely contrasting species, which suggests that the species, which were studied in more detail, include the widest difference in leaf gas exchange among the co-existing species. The saplings of the four selected species exhibited shade acclimated leaves with net photosynthesis rates at saturating irradiance (A_max) between 5.0 and 6.4 μmol m⁻² s⁻¹. In adult trees A_max of fully sunlit leaves was more variable and ranged from 10.5 (C. betulus) to 16.3 μmol m⁻² s⁻¹ (F. excelsior). However, less negative δ¹³C values in F. excelsior sun leaves point to a strong limitation in gas exchange. In the lower canopy of adult trees A_max of F. excelsior (12.0 μmol m⁻² s⁻¹) was also greater than that of A. pseudoplatanus, C. betulus and T. platyphyllos (5.0–5.6 μmol m⁻² s⁻¹). This can be explained by the small leaf area and the absence of shade leaves in mature F. excelsior trees. Thus, a considerable variation in leaf traits and gas exchange was found among the co-existing tree species. The results suggest that species-specific characteristics increase the spatial heterogeneity of canopy gas exchange and should be taken into account in the interpretation and prediction of gas flux from mixed stands.In der Forstwirtschaft Mitteleuropas gewinnt die Begründung von Laubmischwäldern zunehmend an Bedeutung, aber über Eigenschaften im Gasaustausch einiger beteiligter Baumarten, die heute nicht so häufig sind, ist wenig bekannt. In einem Altbestand in Mitteldeutschland (Hainich, Thüringen) habe ich (i) morphologische und chemische Eigenschaften von Sonnenblättern, die Hinweise auf Charakteristika im Blattgaswechsel geben, an acht koexistierenden Baumarten untersucht, und (ii) Photosyntheseparameter von juvenilen und adulten Bäumen (unteres und oberes Kronenniveau) von vier dieser Arten (Acer pseudoplatanus L., Carpinus betulus L., Fraxinus excelsior L. and Tilia platyphyllos Scop.) erhoben.Blätter aus dem oberen Kronenraum der acht untersuchten Arten variierten signifikant in der spezifischen Blattfläche (12.9–19.4 m² kg⁻¹), der Stomatadichte (125–313 Stomata mm⁻²), dem Blattstickstoffgehalt (95–157 mmol N m⁻²) und den δ¹³C-Werten (–27.81 bis –25.85‰). In diesem Kollektiv zeigten F. excelsior und C. betulus groβe Unterschiede, was darauf hindeutet, dass die Arten, die genauer untersucht wurden, die Spannweite an Gaswechseleigenschaften unter den koexistierenden Baumarten umfassen. Die Jungpflanzen der vier ausgewählten Arten besaßen Schattenblätter, deren Netto-Photosyntheserate bei hoher Lichtintensität (A_max) zwischen 5.0 and 6.4 μmol m⁻² s⁻¹ variierte. An Sonnenblättern von Altbäumen war A_max variabler und lag zwischen 10.5 (C. betulus) und 16.3 μmol m⁻² s⁻¹ (F. excelsior). Allerdings weisen hohe δ¹³C-Werte in Sonnenblättern von F. excelsior auf eine starke Limitierung des Gasaustauschs hin. Auch in der unteren Krone der Altbäume war A_max von F. excelsior (12.0 μmol m⁻² s⁻¹) höher als A_max von A. pseudoplatanus, C. betulus und T. platyphyllos (5.0–5.6 μmol m⁻² s⁻¹). Dies kann durch die geringe Blattfläche und die Abwesenheit von Schattenblättern in der Krone adulter Bäume von F. excelsior erklärt werden. Zwischen den koexistierenden Baumarten wurde somit in Bezug auf Blatteigenschaften und Photosyntheseparameter eine erhebliche Variation festgestellt. Die Ergebnisse legen nahe, dass artspezifische Eigenschaften die räumliche Heterogenität des Gaswechsels im Kronenraum erhöhen und bei der Interpretation und Vorhersage von Gasflüssen über Mischbeständen berücksichtigt werden sollten.     

Quantification of hydrogen peroxide and glucose using 3-methyl-2-benzothiazolinonehydrazone hydrochloride with 10,11-dihydro-5H-benz(b,f)azepine as chromogenic probe

A sensitive, selective, and rapid enzymatic method is proposed for the quantification of hydrogen peroxide (H₂O₂) using 3-methyl-2-benzothiazolinonehydrazone hydrochloride (MBTH) and 10,11-dihydro-5H-benz(b,f)azepine (DBZ) as chromogenic cosubstrates catalyzed by horseradish peroxidase (HRP) enzyme. MBTH traps free radical released during oxidation of H₂O₂ by HRP and gets oxidized to electrophilic cation, which couples with DBZ to give an intense blue-colored product with maximum absorbance at 620 nm. The linear response for H₂O₂ is found between 5 × 10⁻⁶ and 45 × 10⁻⁶ mol L⁻¹ at pH 4.0 and a temperature of 25 °C. Catalytic efficiency and catalytic power of the commercial peroxidase were found to be 0.415 × 10⁶ M⁻¹ min⁻¹ and 9.81 × 10⁻⁴ min⁻¹, respectively. The catalytic constant (k_cat) and specificity constant (k_cat/K_m) at saturated concentration of the cosubstrates were 163.2 min⁻¹ and 4.156 × 10⁶ L mol⁻¹ min⁻¹, respectively. This method can be incorporated into biochemical analysis where H₂O₂ undergoes catalytic oxidation by oxidase. Its applicability in the biological samples was tested for glucose quantification in human serum.     

Variation in δC values for the seagrass Thalassia testudinum and its relations to mangrove carbon

Carbon isotope ratios (¹³C/¹²C) were measured for the leaves of the seagrass Thalassia testudinum Banks ex König and carbonates of shells collected at the seagrass beds from seven sites along the coast of southern Florida, U.S.A. The δ¹³C values of seagrass leaves ranged from −7.3 to −16.3‰ among different study sites, with a significantly lower mean value for seagrass leaves from those sites near mangrove forests (−12.8 ± 1.1‰) than those far from mangrove forests (−8.3 ± 0.9‰; P < 0.05). Furthermore, seagrass leaves from a shallow water area had significantly lower δ¹³C values than those found in a deep water area (P < 0.01). There was no significant variation in δ¹³C values between young and mature leaves (P = 0.59) or between the tip and base of a leaf blade (P = 0.46). Carbonates of shells also showed a significantly lower mean δ¹³C value in the mangrove areas (−2.3 ± 0.6‰) than in the non-mangrove areas (0.6 ± 0.3‰; P <0.025). In addition, the δ¹³C values of seagrass leaves were significantly correlated with those of shell carbonates (δ¹³C seagrass leaf = −9.1 + 1.3δ¹³C shell carbonate (R² = 0.83, P < 0.01)). These results indicated that the input of carbon dioxide from the mineralization of mangrove detritus caused the variation in carbon isotope ratios of seagrass leaves among different sites in this study.     

Inter and intra-species-related differences in the regulation of the cardiac autonomic system

The heart rate response to isoproterenol (HR-Iso), density and affinity (k_d) of β-adrenergic (β-AR) and muscarinic (M₂) receptors were compared among three rodents with different generation-life histories of confinement and of high altitude exposure. The European guinea pig (Cavia porcellus) (EGp), a laboratory animal that arrived in Europe after the Spanish Conquest of South America and the Peruvian guinea pig (C. porcellus) (PGp), a semi-wild animal that came from the altiplano to sea level at least 25 generations ago, were used for intra-species comparison. Wistar rats (WR) were used for inter-species comparison as representative of a typical sea level laboratory animal. The HR-Iso was lower in EGp than in the PGp. The PGp showed the highest β-AR density (P<0.0005) and the highest β-AR k_d values (P<0.0005) when compared to both EGp and WR groups (β-AR B_max (fmol mg⁻¹ prot), WR, 19±4; Egp, 34±10; PGp, 74±15. β-AR k_d (pM), WR, 24±10; Egp, 17±7; PGp, 39±14). In contrast, PGp showed lower M₂ receptor density values than the EGp (P<0.0005). The WR had the highest M₂ receptor densities (M₂ B_max (fmol mg⁻¹ prot), WR, 188±15; Egp, 147±9; PGp, 118±6 and M₂ k_d (pM), WR, 65±12; Egp, 67±6; PGp, 92±2). The inter and intra-species differences found may be related to their respective history of confinement rather than to their history of exposure to high altitude.     

Comparison of oxygen mass transfer coefficient in simple and extractive fermentation systems

Aeration and agitation are important variables to ensure effective oxygen transfer rate during aerobic bioprocesses; therefore, the knowledge of the volumetric mass transfer coefficient (k_La) is required. In view of selecting the optimum oxygen requirements for extractive fermentation in aqueous two-phase system (ATPS), the k_La values in a typical ATPS medium were compared in this work with those in distilled water and in a simple fermentation medium, in the absence of biomass. Aeration and agitation were selected as the independent variables using a 2² full factorial design. Both variables showed statistically significant effects on k_La, and the highest values of this parameter in both media for simple fermentation (241 s⁻¹) and extractive fermentation with ATPS (70.3 s⁻¹) were observed at the highest levels of aeration (5 vvm) and agitation (1200 rpm). The k_La values were then used to establish mathematical correlations of this response as a function of the process variables. The exponents of the power number (N³D²) and superficial gas velocity (V_s) determined in distilled water (α = 0.39 and β = 0.47, respectively) were in reasonable agreement with the ones reported in the literature for several aqueous systems and close to those determined for a simple fermentation medium (α = 0.38 and β = 0.41). On the other hand, as expected by the increased viscosity in the presence of polyethylene glycol, their values were remarkably higher in a typical medium for extractive fermentation (α = 0.50 and β = 1.0). A reasonable agreement was found between the experimental data of k_La for the three selected systems and the values predicted by the theoretical models, under a wide range of operational conditions.     

Inhibition of escherichia coli glucosamine synthetase by novel electrophilic analogues of glutamine—comparison with 6-diazo-5-oxo-norleucine

A series of electrophilic glutamine analogues based on 6-diazo-5-oxo-norleucine has been prepared, using novel synthetic routes, and evaluated as inhibitors of Escherichia coli. glucosamine synthetase. The γ-dimethylsulphonium salt analogue of glutamine was found to be one of the most potent inactivators of this enzyme yet reported, with an apparent second order rate constant (k₂/K_i) of 3.5×10⁵ M⁻¹ min⁻¹.     

The stereochemical course of the reaction mechanism of trehalose phosphorylase from Schizophyllum commune

Phosphorolysis of α,α-trehalose catalyzed by trehalose phosphorylase from the basidiomycete Schizophyllum commune proceeds via net retention of anomeric configuration and yields α- -glucose 1-phosphate and α- -glucose as the products. In reverse reaction, only the α-anomers of -glucose 1-phosphate and -glucose are utilized as glucosyl donor and acceptor, respectively, and give exclusively the α,α-product. Trehalose phosphorylase converts α- -glucose 1-fluoride and phosphate into α- -glucose 1-phosphate, a reaction requiring the stereospecific protonation of the glucosyl fluoride by a Brønsted acid. The results are discussed with regard to a plausible reaction mechanism of fungal trehalose phosphorylase.     

Long-Term Temperature Acclimation of Photosynthesis in Steady-State Cultures of the Polar Diatom <Emphasis Type="Italic">Fragilariopsis cylindrus</Emphasis>

Cultures of the obligate psychrophilic diatom Fragilariopsis cylindrus (Grunow) were grown for 4 months under steady-state conditions at −1 °C and +7 °C (50 μmol photons m⁻² s⁻¹) prior to measurements in order to investigate long-term acclimation of photosynthesis to both temperatures. No differences in maximum intrinsic quantum yield of PS II (F_V/F_M) and relative electron transport rates could be detected at either temperature after 4 months of acclimation. Measurements of photosynthesis (relative electron transport rates) vs. irradiance (P vs. E curves) revealed similar values for relative light utilization efficiency (α = 0.57 at −1 °C, α = 0.60 at +7 °C) but higher values for irradiance levels at which photosynthesis saturates (E_K) at −1 °C and, therefore, higher maximum photosynthesis (P_MAX = 54 (relative units) at −1 °C, P_MAX = 49 at +7 °C). Nonphotochemical quenching (NPQ) measurements at 385 μmol photons m⁻² s⁻¹ indicated higher (37%) NPQ for diatoms grown at −1 °C compared to +7 °C, which was possibly related to a 2-fold increase in the concentration of the pigment diatoxanthin and a 9-fold up-regulation of a gene encoding a fucoxanthin chlorophyll a,c-binding protein. Expression of the D1 protein encoding gene psbA was ca. 1.5-fold up-regulated at −1 °C, whereas expression levels of other genes from Photosystem II (psbC, psbU, psbO), as well as rbcL, the gene encoding the Rubisco large subunit were similar at both temperatures. However, a 2-fold up-regulation of a plastid glyceraldehyde-P dehydrogenase at −1 °C indicated enhanced Calvin cycle activity. This study revealed for the first time that a polar diatom could efficiently acclimate photosynthesis over a wide range of polar temperatures given enough time. Acclimation of photosynthesis at −1 °C was probably regulated similarly to high light acclimation.     

Direct oxidation of boronates by peroxynitrite: Mechanism and implications in fluorescence imaging of peroxynitrite

In this study, we show that boronates, a class of synthetic organic compounds, react rapidly and stoichiometrically with peroxynitrite (ONOO⁻) to form stable hydroxy derivatives as major products. Using a stopped-flow kinetic technique, we measured the second-order rate constants for the reaction with ONOO⁻, hypochlorous acid (HOCl), and hydrogen peroxide (H₂O₂) and found that ONOO⁻ reacts with 4-acetylphenylboronic acid nearly a million times (k = 1.6 × 10⁶ M^− 1 s^− 1) faster than does H₂O₂ (k = 2.2 M^− 1 s^− 1) and over 200 times faster than does HOCl (k = 6.2 × 10³ M^− 1 s^− 1). Nitric oxide and superoxide together, but not alone, oxidized boronates to the same phenolic products. Similar reaction profiles were obtained with other boronates. Results from this study may be helpful in developing a novel class of fluorescent probes for the detection and imaging of ONOO⁻ formed in cellular and cell-free systems.     

Kinetic analyses for bindings of concanavalin A to dispersed and condensed mannose surfaces on a quartz crystal microbalance

A biotinylated mannotriose (Man3-bio) was dispersively immobilized in the matrix of biotinylated lactose (Gal-Glc-bio) on a streptavidin-covered, 27-MHz quartz crystal microbalance (QCM), and binding kinetics of concanavalin A (Con A) to Man3-bio in the Gal-Glc-bio matrix could be obtained from frequency decreases (mass increases) of the QCM. Association constants (K_a) and binding and dissociation rate constants (k_on and k_off) could be determined separately as the 1:1 and 1:2 bindings of Con A to Man3-bio on the surface. When Man3-bio was immobilized with content of 1 to 5 mol% in the matrix, the 1:1 binding of Con A to Man3-bio was obtained as K_a = (4 ± 1) × 10⁶ M⁻¹, k_on = (4 ± 1) × 10⁴ M⁻¹ s⁻¹, and k_off = (12 ± 2) × 10^–3 s⁻¹. On the contrary, when Man3-bio was immobilized with content of 20 to 100 mol% in the matrix, the 1:2 binding of Con A to Man3-bio was obtained as K_a = (14 ± 2) × 10⁶ M⁻¹, k_on = (14 ± 2) × 10⁴ M⁻¹ s⁻¹, and k_off = (7 ± 2) × 10^–3 s⁻¹. Thus, K_a for the 1:2 binding was 10 times larger than that for the 1:1 binding, with a three times larger binding rate constant (k_on) and a three times smaller dissociation rate constant (k_off). This is the first example to obtain separate kinetic parameters for the 1:1 and 1:2 bindings of lectins to carbohydrates on the surface.     

Purification and properties of a novel raw starch degrading-cyclodextrin glycosyltransferase from Klebsiella pneumoniae AS- 22

A novel raw starch degrading α-cyclodextrin glycosyltransferase (CGTase; E.C. 2.4.1.19), produced by Klebsiella pneumoniae AS-22, was purified to homogeneity by ultrafiltration, affinity and gel filtration chromatography. The specific cyclization activity of the pure enzyme preparation was 523 U/mg of protein. No hydrolysis activity was detected when soluble starch was used as the substrate. The molecular weight of the pure protein was estimated to be 75 kDa with SDS-PAGE and gel filtration. The isoelectric point of the pure enzyme was 7.3. The enzyme was most active in the pH range 5.5–9.0 whereas it was most stable in the pH range 6–9. The CGTase was most active in the temperature range 35–50°C. This CGTase is inherently temperature labile and rapidly loses activity above 30°C. However, presence of soluble starch and calcium chloride improved the temperature stability of the enzyme up to 40°C. In presence of 30% (v/v) glycerol, this enzyme was almost 100% stable at 30°C for a month. The K_m and k_cat values for the pure enzyme were 1.35 mg ml⁻¹ and 249 μM mg⁻¹ min⁻¹, respectively, with soluble starch as the substrate. The enzyme predominantly produced α-cyclodextrin without addition of any complexing agents. The conditions employed for maximum α-cyclodextrin production were 100 g l⁻¹ gelatinized soluble starch or 125 g l⁻¹ raw wheat starch at an enzyme concentration of 10 U g⁻¹ of starch. The α:β:γ-cyclodextrins were produced in the ratios of 81:12:7 and 89:9:2 from gelatinized soluble starch and raw wheat starch, respectively.     

Decarboxylation kinetics for the carbonatotris(pyridine)cobalt(III) ion in perchloric acid solutions

The kinetics of the decomposition reactions of the CO(py)₃(CO₃)(H₂O)⁺ ion have been investigated in aqueous perchloric acid solutions over a range of hydrogen ion concentrations (0.10 to 5.0 M) and at two ionic strengths (I = 1.0 and 5.0 M). At the lower ionic strength, plots of ln (A_t–A_∞ versus time show a nonlinearity that is consistent with that expected for consecutive first-order reactions. The rates of the faster reaction are similar to those reported for the spontaneous reduction of aquopyridine-cobalt(III) cations. At the higher ionic strength, the above noted curvature is not apparent and the decarboxylation kinetics of the title complex may be described by a pseudo-first-order rate constant: k_obs = k[H₃O⁺]. At 20°C, k = (1.75−+0.09) s⁻¹ M⁻¹ with activation parameters ofΔH^‡ = (97 −+ 4) kJ mol⁻¹ and ΔS^‡ = −(54 −+ 32) J deg⁻¹ mol⁻¹. These kinetic parameters are compared with those previously reported for the similar complexes, Co(py)₄CO₃⁺ and Co(py)₂(CO₃)(H₂O)₂⁺.     

Synthesis of [Tl4Se16]: a novel tetranuclear Tl polyselenide

(Ph₄P)₄[Tl₄Se₁₆] was prepared hydrothermally in a sealed pyrex tube by the reaction of TlCl, K₂Se₄ and Ph₄PCl in a 1:1:1 molar ratio at 110 °C for one day. The red crystals were obtained in 50% yield. Crystals of (Ph₄P)₄[Tl₄Se₁₆]: triclinic P (No. 2), Z=1, a=12.054(9), b=19.450(10), c=11.799(6) Å, α=104.63(4), β=98.86(6), γ=101.99(6)° and V=2555(3) ų at 23 °C, 2θ_max=40.0°, μ=120.7 cm⁻¹, D_calc=2.23. The structure was solved by direct methods. Number of data collected: 5206. Number of unique data having F_o²>3σ(F_o²): 1723. Final R=0.075 and R_w=0.089. [Tl₄Se₁₆]⁴⁻ consists of four, almost already linearly arranged, tetrahedral thallium centers which are coordinated by two chelating Se₄²⁻, two bridging Se₂²⁻ and four bridging Se²⁻ ligands. [Tl₄Se₁₆]⁴⁻ sits on an inversion center and possesses a central {Tl₂Se₂}²⁺ planar core. The Tl(1)–Tl(1)′ distance in this core is 3.583(6) Å. These two thallium atoms are then each linked to two cyclic Tl(Se₄) fragments via bridging Se₂²⁻ and Se²⁻ ligands forming Tl₂Se(Se₂) five-membered rings.     

Glycan-binding profile of a D-galactose binding lectin purified from the annelid, Perinereis nuntia ver. vallata

A lectin recognizing D-galactose was purified from the pacific annelid Perinereis nuntia ver. vallata (Polychaeta) by affinity chromatography. Hemagglutinating activity, with a very low titer suggesting the presence of lectin appeared in the supernatant from the homogenization of body with Tris-buffered saline. However, dialyzed supernatant from the precipitate homogenized by galactose in the buffer revealed strong hemagglutinating activity against human erythrocytes. The crude supernatant was applied onto lactosyl–agarose column, and only the supernatant eluted from precipitate with galactose was obtained a galactose-binding lectin with 32 kDa polypeptide was obtained from the supernatant of the precipitate, extracted in presence of galactose. It suggests that the lectin tightly binds with glycoconjugate as endogenous ligand(s) in the tissue. Hemagglutinating activity against trypsinized and glutaraldehyde-fixed human erythrocytes was specifically inhibited by D-galactose, N-acetyl-D-galactosamine, lactose, melibiose, and asialofetuin. Glycan-binding profile of the lectin analyzed by frontal affinity chromatography shows that the lectin recognizes branched complex type N-linked oligosaccharides and both type 1 (Galβ1-3GlcNAc) and type 2 (Galβ1-4GlcNAc) lactosamine. The surface plasmon resonance study of the lectin against asialofetuin showed the k_ass and k_diss values are 5.14 × 10⁴ M^− 1 s^− 1 and 2.9 × 10⁻³ s^− 1, respectively. The partial primary structure of the lectin reveals 182 amino acids with novel sequence.     

Purification and substrate characterization of α-ketobutyrate decarboxylase from Pseudomonas putida

α-Ketobutyrate decarboxylase encoded in the -methionine catabolism operon of Pseudomonas putida is homologous with the E1 component of pyruvate dehydrogenase complex from gram-negative bacteria. The enzyme was purified to homogeneity from the cell extract of an Escherichia coli transformant. The purified enzyme was homodimeric with a subunit of M_r 93,000 on SDS-PAGE. The enzyme activity was activated by the addition of both thiamine pyrophosphate (TPP) and a divalent cation, such as Mg²⁺, Mn²⁺, and Co²⁺. The enzyme showed high activity for α-ketobutyrate and α-keto-n-valerate rather than pyruvate, but the α-keto acids with increasing length of the side chain as well as branching, such as α-keto-n-caproate and α-keto-3-methylvalerate, were not used by the enzyme. The K_m values for α-ketobutyrate and pyruvate were 0.016 and 0.147 mM, respectively, and the k_cat/K_m value (10.69 s⁻¹ mM⁻¹) for α-ketobutyrate was 29-fold greater than that for pyruvate. Thus, α-ketobutyrate decarboxylase is distinguished from the pyruvate dehydrogenase E1 component with respect to the substrate specificity, although their structural and enzymological properties were similar. These results suggest that the unique substrate specificity of α-ketobutyrate decarboxylase is due to a slight difference in the highly conserved active sites of both enzymes.