Molecular mass and chain conformations of Rhizoma Panacis Japonici polysaccharides

The molecular mass and size of five water-soluble polysaccharides isolated from Rhizoma Panacis Japonici (RPJ) were determined with laser light scattering (LLS), size-exclusion chromatography (SEC) combined with LLS (SEC–LLS), dynamic light scattering (DLS), as well as transmission electron microscope (TEM). Their weight-average molecular masses (M_w) were 3.5 × 10⁴, 1.47 × 10⁵, 1.24 × 10⁶, 9.26 × 10⁵ and 1.36 × 10⁶, radii of gyration (<s²>_z^1/2) were 14.7, 31.7, 50.8, 41.8 and 40.4 nm, and hydrodynamic radii (R_h) were 19.9, 37.5, 66.2, 52.1, and 55.2 nm, respectively. The results showed that molecular masses and sizes of the polysaccharides were influenced by the pH and temperature of the extraction mediums. The conformation parameters were calculated from the above data according to polymer solution theory. The values of ρ (= <s²>_z^1/2/R_h) were from 0.7 to 0.8, exponents (ν) of <s²>_z^1/2 = k M_w^ν were from 0.31 to 0.43, and fractal dimension (d_f) were from 2.3 to 3.2, respectively. The results revealed that all of the polysaccharides existed as spheres in 0.15 M NaCl aqueous solution. TEM and atomic force microscope (AFM) further confirmed the spherical morphologies of these molecules. The spherical conformations of the polysaccharides were a result of their highly branched structures.     

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.     

Conformation of (2→1)-β-d-fructan in aqueous solution

The conformation and dilute solution properties of (2→1)-β-d-fructan in aqueous solution were studied by gel permeation chromatography, low-angle laser light-scattering photometry, viscometry, small-angle X-ray scattering and electron microscopy. Fractions covering a broad range of weight-average molecular weights (M_w) from 1.49 × 10⁴ to 5.29 × 10⁶ were obtained from a native sample by ultrasonic degradation and fractional precipitation. For M_w < 4 × 10⁴, the intrinsic viscosity [η] varies with M_w^0.71, indicating that the fructan chain behaves as a random coil expanded by an excluded-volume effect in this molecular weight region. For M_w > 10⁵, [η] exhibits an unusually weak dependence on M_w and finally becomes almost independent of molecular weight. This behaviour is interpreted in terms of a globular conformation of the high-molecular-weight fructan molecules. Small-angle X-ray-scattering measurements and electron microscopic observations support this interpretation of the values of [η] observed.     

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.     

A Conjugative Transfer System for the Rumen Bacterium, Butyrivibrio fibrisolvens, Based on Tn916-Mediated Transfer of the Staphylococcus aureus Plasmid pUB110

A limitation of genetic studies of the rumen bacterium, Butyrivibrio fibrisolvens, has been the availability of suitable vectors and transfer systems. Using the conjugative tetracycline resistant transposon, Tn916, the Staphylococcus aureus plasmid, pUB110, and the pUB110-based shuttle vector, pUBLRS, a conjugative transfer system was developed for B. fibrisolvens. B. fibrisolvens donor strains H17c2 and H17c12, containing Tn916 and pUB110 or pUBLRS, respectively, were used in mating experiments with selected B. fibrisolvens strains. Kanamycin resistant transconjugants, containing pUB110, of strains 193, 194, and 195 were detected at a combined average frequency of 7.78 × 10^-7 per donor and 1.11 × 10^-5 per recipient. Transconjugants of strains 193 and 194, containing pUBLRS, were detected at an average frequency of 1.22 × 10^-6 per donor and 4.70 × 10^-8 per recipient. Southern hybridization analysis confirmed the presence of pUB110 and pUBLRS in transconjugants. Results indicated that Tn916 was necessary for mobilization of pUB110 as transconjugants were not detected when the transposon was absent from the donor strains. The ability to mobilize pUB110 and pUBLRS between B. fibrisolvens strains provides a conjugative transfer system that circumvents problems encountered with electroporation.     

Effect of prostaglandin F2α on propulsive activity of the isolated segmental colon of the guinea-pig

The effects of prostaglandin F_2α (PGF_2α) on propulsive activity in segments of isolated colon and on isolated strips of guinea-pig colon were investigated.Using experimental conditions under which spontaneous propulsive activity was negligible, PGF_2α (5×10⁻⁸×1×10⁻⁶M), added to the bathing medium, increased propulsive activity in a concentration dependent manner. This increase of propulsive activity was abolished in the presence of atropine or tetrodotoxin (1×10⁻⁷g/ml).The contractions produced by PGF_2α(5×10⁻⁷ − 1×10⁻⁵M) in isolated longitudinal and circular smooth muscle strips of guinea-pig colon were unaffected in the presence of atropine or tetrodotoxin (1×10⁻⁷ g/ml).From these results it is concluded that under the conditions employed in this study propulsive activity stimulated by PGF_2α may depend on the contractions of both muscle layers and stimulation of the peristalic reflex.     

Membrane Permeability Characteristics of Metaphase II Mouse Oocytes at Various Temperatures in the Presence of Me2SO

In this study, the hydraulic conductivity (L_p), Me₂SO permeability ( _Me2SO), and the reflection coefficients (ς) and their activation energies were determined for Metaphase II (MII) mouse oocytes by exposing them to 1.5 M Me₂SO at temperatures of 30, 20, 10, 3, 0, and −3°C. These data were then used to calculate the intracellular concentration of Me₂SO at given temperatures. Individual oocytes were immobilized using a holding pipette in 5 μl of an isosmotic PBS solution and perfused with precooled or prewarmed 1.5 M Me₂SO solutions. Oocyte images were video recorded. The cell volume changes were calculated from the measurement of the diameter of the oocytes, assuming a spherical shape. The initial volume of the oocytes in the isoosmotic solution was considered 100%, and relative changes in the volume of the oocytes after exposure to the Me₂SO were plotted against time. Mean (means ± SEM) L_pvalues in the presence of Me₂SO ( ^Me₂SO_p) at 30, 20, 10, 3, 0, and −3°C were determined to be 1.07 ± 0.03, 0.40 ± 0.02, 0.18 ± 0.01, 7.60 × 10⁻²± 0.60 × 10⁻², 5.29 × 10⁻²± 0.40 × 10⁻², and 3.69 × 10⁻²± 0.30 × 10⁻²μm/min/atm, respectively. The _Me2SOvalues were 3.69 × 10⁻³± 0.3 × 10⁻³, 1.07 × 10⁻³± 0.1 × 10⁻³, 2.75 × 10⁻⁴± 0.15 × 10⁻⁴, 7.83 × 10⁻⁵± 0.50 × 10⁻⁵, 5.24 × 10⁻⁵± 0.50 × 10⁻⁵, and 3.69 × 10⁻⁵± 0.40 × 10⁻⁵cm/min, respectively. The ς values were 0.70 ± 0.03, 0.77 ± 0.04, 0.81 ± 0.06, 0.91 ± 0.05, 0.97 ± 0.03, and 1 ± 0.04, respectively. The estimated activation energies (E_a) for ^Me₂SO_p, _Me2SO, and ς were 16.39, 23.24, and −1.75 Kcal/mol, respectively. These data may provide the fundamental basis for the development of more optimal cryopreservation protocols for MII mouse oocytes.     

Biodegradation of lindane by a native bacterial consortium isolated from contaminated river sediment

The aerobic biodegradation of lindane (γ-hexachlorocyclohexane) by a consortium of acclimated bacteria from sediment at a polluted site on the Suquia River, Cordoba, Argentina, is reported. The bacteria were acclimated for 30 days under aerobic conditions, using a minimal culture medium containing lindane (0.034 mM) as sole carbon source. Growth of the bacterial consortium decreased at a lindane concentration of 1.03 mM and was totally inhibited at 2.41 mM. The consortium showed initial lindane degradation rates of 4.92×10⁻³, 11.0×10⁻³ and 34.8×10⁻³ mM h⁻¹ when exposed to lindane concentrations of 0.069, 0.137 and 0.412 mM, respectively. Chloride concentration increased during aerobic biodegradation, indicating lindane mineralization. A metabolite identified as γ-2,3,4,5,6-pentachlorocyclohexene appeared during the first 24 h of biodegradation. Four different bacteria, identified as Sphingobacterium spiritivorum, Ochrobactrum anthropi, Bosea thiooxidans and Sphingomonas paucimobilis, were isolated. Pure strains of B. thiooxidans and S. paucimobilis degraded lindane after 3 days of aerobic incubation. This is the first report of lindane biodegradation by B. thiooxidans.     

Characterization of galactomannans isolated from legume endosperms of Caesalpinioideae and Faboideae subfamilies by multidetection aqueous SEC

Galactomannans isolated from legume seed endosperms, including those of commercial interest, have been characterized by multidetection aqueous SEC. Galactomannans derived from seeds of the Faboideae subfamily had substantially higher M_w than those from Caesalpinioideae seeds (M_w,Fab = 2.4–3.1 × 10⁶ g/mol, M_w,Caes. = 0.86–2.1 × 10⁶ g/mol) and within the latter botanical subfamily, an apparent correlation between M_w and the degree of galactose substitution DG was found. The molar mass distributions were unimodal and differed primarily by a scale factor, with distributional widths narrower than a true Flory ‘most-probable distribution’; good fits to Schulz–Zimm model were obtained. Across subfamilies no differences were found in the exponents of [η]–M and R_v–M relationships (0.61 ± 0.02, 0.54 ± 0.01, respectively), the Flory chain stiffness ratio (C_∞ = 20 ± 1 (BSF analysis)), or the persistence length (L_p = 5.5 ± 0.2 nm) obtained from SEC fraction data. However, it was found that prefactors in the [η]–M and R_v–M relationships as well as the unperturbed parameter K_Θ decrease in proportion to DG and therefore chain density. Generalized relationships incorporating galactose-dependent prefactors were therefore developed to model SEC fraction data of native galactomannans ([η]_GM = (1800 ± 200) × M_o^−1.61 × M^0.61±0.02, R_v,GM = 0.63 ± 0.05 × M_o^−0.54 × M^0.54±0.01) as well as lower-M fractions obtained by ultrasonication ([η]_GM = (730 ± 100) × M_o^−1.71 × M_w^0.71±0.02, R_v,GM = 0.49 ± 0.05 × M_o^−0.57 × M_w^0.57±0.01, M ≈ 1 × 10⁵-native). As a consequence of this dependence and the observed patterns in molar mass variation, [η] varies within a narrow range for galactomannans as a whole despite substantial M_w differences.     

Seasonal and spatial variability of coccolithophore export production at the South-Western margin of Crete (Eastern Mediterranean)

Six moorings were deployed at different locations in the deep submarine canyons along the south–west margin of Crete, providing a total of eight sediment-trap time series from June 2005 to May 2006. Within this dataset, we analyzed the record from intact coccospheres, which represent the signal of export production from the coccolithophore community. The most abundant species at all stations during the whole investigated period were E. huxleyi and A. robusta, followed by S. pulchra HET, G. flabellatus, H. carteri, F. profunda, S. pulchra HOL oblonga, while the rest of the species represented ≤ 1% of the assemblage. Overall the assemblage composition was comparable at all stations, with slight variations mostly related to the different preservation of coccosphere integrity at the different collection depths. The consistent pattern of seasonal variation in species distribution and total coccolithophore export allowed us to define the occurrence of three main periods: a) March to June, with high overall coccosphere flux (up to 4.3 × 10⁵–3.4 × 10⁶ coccospheres m^− 2 day^− 1), increased abundance of E. huxleyi and subordinate H. carteri s.s., Umbilicosphaera spp. and S. pulchra; b) June to November, with high but gradually decreasing total coccosphere flux (up to 7 × 10⁵–1.4 × 10⁶ coccospheres m^− 2 day^− 1) and high relative abundance of the deep photic zone species A. robusta, F. profunda, G. flabellatus as well as S. pulchra and Coronosphaera spp., R. clavigera, U. tenuis, D. tubifera and holococcolithophores; c) November to February, with low overall export fluxes (3.5–9 × 10⁴ coccospheres m^− 2 day^− 1) and high relative abundance of A. robusta, S. pulchra and Syracosphaera spp. These three periods correspond to the seasonal changes in sea surface temperature, surface mixed layer depth and rainfall and are associated with varying total surface primary production, as detected through remote sensing in the surface waters.     

Stimulation by prostaglandin E2 of glucagon and insulin release from isolated rat pancreas

To ascertain whether prostaglandins (PG) may play a role in the secretion of glucagon and in an attempt to elucidate the conflicting observations on the effects of PG on insulin release, the isolated intact rat pancreas was perfused with solutions containing 1.1 × 10⁻⁹ to 1.8 × 10⁻⁵M PGE₂. In the presence of 5.6 mM glucose significant increments in portal venous effluent levels of glucagon and insulin were observed in response to minimal concentrations of 2.8 × 10⁻⁸ and 1.4 × 10⁻⁷M PGE₂, respectively; a dose-response relationship was evident for both hormones at higher concentrations of PGE₂. When administered over 60 seconds, 1.4 × 10⁻⁶M PGE₂ resulted in a significant increase in glucagon levels within 24 seconds and in insulin within 48 seconds. Ten-minute perfusions of 1.4 × 10⁻⁶M PGE₂ elicited biphasic release of both islet hormones; Phase I glucagon release preceded that of insulin. Both phases of the biphasic glucagon and insulin release which occurred in response to 15-minute perfusions of 10 mM arginine were augmented by PGE₂. These observations indicate that PGE₂ can evoke glucagon and insulin release at concentrations close to those observed by others in the extracts of rat pancreas. We conclude that PG may be involved in the regulation of secretion of glucagon and insulin and may mediate and/or modify the pancreatic islet hormone response to other secretagogues.     

Laccase-catalyzed oxidation of phenolic compounds in organic media

Rhus vernificera laccase-catalyzed oxidation of phenolic compounds, i.e., (+)-catechin, (−)-epicatechin and catechol, was carried out in selected organic solvents to search for the favorable reaction medium. The investigation on reaction parameters showed that optimal laccase activity was obtained in hexane at 30 °C, pH 7.75 for the oxidation of (+)-catechin as well as for (−)-epicatechin, and in toluene at 35 °C, pH 7.25 for the oxidation of catechol. E_a and Q₁₀ values of the biocatalysis in the reaction media of the larger log p solvents like isooctane and hexane were relatively higher than those in the reaction media of lower log p solvents like toluene and dichloromethane. Maximum laccase activity in the organic media was found with 6.5% of buffer as co-solvent. A wider range of 0–28 μg protein/ml in hexane than that of 0–16.7 μg protein/ml in aqueous medium was observed for the linear increasing conversion of (+)-catechin. The kinetic studies revealed that in the presence of isooctane, hexane, toluene and dichloromethane, the K_m values were 0.77, 0.97, 0.53 and 2.9 mmol/L for the substrate of (+)-catechin; 0.43, 0.34, 0.14 and 3.4 mmol/L for (−)-epicatechin; 2.9, 1.8, 0.61 and 1.1 mmol/L for catechol, respectively, while the corresponding V_max values were 2.1 × 10⁻², 2.3 × 10⁻², 0.65 × 10⁻² and 0.71 × 10⁻² δA/μg protein min); 1.8 × 10⁻², 0.88 × 10⁻², 0.19 × 10⁻² and 1.0 × 10⁻² δA/μg protein min); 0.48 × 10⁻², 0.59 × 10⁻², 0.67 × 10⁻² and 0.54 × 10⁻² δA/μg protein min), respectively. FT-IR indicated the formation of probable dimer from (+)-catechin in organic solvent. These results suggest that this laccase has higher catalytic oxidation capacity of phenolic compounds in suitable organic media and favorite oligomers could be obtained.     

Inhibition of gibberellin biosynthesis by paclobutrazol in cell-free homogenates ofCucurbita maxima endosperm andMalus pumila embryos

The plant growth retardant paclobutrazol, (PP333) (2RS, 3RS)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)pentan-3-ol, inhibits specifically the three steps in the oxidation of the gibberellin-precursorent-kaurene toent-kaurenoic acid in a cell-free system fromCucurbita maxima endosperm. The KI₅₀ for this inhibition is 2×10^–8 M. The KI₅₀ values for the separated2S, 3S, and2R, 3R enantiomers of paclobutrazol in this system are 2×10^–8 M and 7×10^–7 M, respectively. A cell-free preparation from immatureMalus pumila embryos convertsent-kaurene to gibberellin A₉, whereas no conversion occurs in a similar preparation fromMalus endosperm. The conversion ofent-kaurene by the embryo preparation is inhibited by paclobutrazol with KI₅₀ values for the2S,3S and2R,3R enantiomers of 2×10^–8 M and 6×10^–8 M, respectively.     

Differential effects of prazosin and rauwolsin on the release of prostaglandins I2 and E2 from the perfused mesenteric arterial bed of the rabbit following nerve stimulation

Sympathetic nerve stimulation of the perfused mesenteric arterial bed of the rabbit, , increase the secretion of prostaglandin (PG)I₂ and PGE₂. Prazosin (4.8 × 10⁻⁶), and α₁ adrenergic receptor antagonist, inhibited this inrease in release of PGI₂ but not of PGE₂ whereas rauwolsin (10⁻⁷ M), an α₂ adrenergic receptor antagonist, inhibited the increase in release of PGE₂ but not of PGI₂. Prazosin (10⁻⁶ M) completely blocked the vasoconstrictor response to nerve stimulation, and to norepinephrine and phenylephrine administration, suggesting there to be little of an α₂ adrenergic receptor component in this response. It is concluded that the increase in PGI₂ release follows the activation of α₁ adrenergic receptors and is therefore post-junctional in origin, whereas the increase in PGE₂ release follows the activation of α₂ adrenergic receptors and may be pre- and/or post-junctional in origin.Indomethacin (2.8 × 10⁻⁷, 5.6 × 10⁻⁷ and 1.12 × 10^{−6 M} did not affect the vasoconstrictor responses to nerve stimulation at 10 Hz, whereas rauwolsin (10⁻⁷ M) in the presence of indomethacin substantially increased them. These results indicate that PGE₂ does not regulate norepinephrine release following nerve stimulation at 10 Hz to rabbit mesenteric arteries, and that the inhibition of norepinephrine release following stimulation of α₂ pre-junctional receptors is independent of PG involvement.     

Avian oviduct motility induced by prostaglandin E1

Oviduct segments from infundibulum, magnum, uterus, uterovaginal junction and vagina of actively laying hens at preoviposition time were tested for their contractile reaction to prostaglandin E₁ by or methods. Maximum stimulatory response was observed from the muscular strips of the proximal oviduct segment (infundibulum) and a complete relaxation was recorded from the distal part (vagina) at molar concentrations of 1.4 × 10⁻⁷, 3.4 × 10⁻⁷ and 7.0 × 10⁻⁷. The uterine strips reacted with a stimulatory response at higher concentrations (1.4 × 10⁻⁶ and 2.8 × 10⁻⁶ moles), but lacked any significant change at lower concentrations. The uterovaginal muscular strips showed a mild but prolonged inhibitory response, while the magnum responded with a significant increase in the luminal pressure when tested . It is concluded that PGE₁ exerts a stimulatory effect on the uterus to initiate transport of the egg to subsequent segments (uterovaginal junction and vagina), which relax under PGE₁ influence and allow passage of the egg by pressure differences.     

Prostaglandins and the rat seminal vesicle: Effects of mating and adrenergic stimulation

The concentrations of PGE, PGF, and 6-keto-PGF_1α were increased in rat seminal vesicle tissue following mating activity. Likewise, synthesis of PGE and PGF was stimulated by epinephrine (3 × 10⁻⁷to 3 × 10⁻⁶ M) in tissues and media from incubations of intact rat seminal vesicles. The stimulation was inhibited by phentolamine, an α-adrenoreceptor blocking agent. Carbamylcholine (2 × 10⁻⁶ M) and bradykinin (1 × 10⁻⁶ M) had no effect on PGE or PGF synthesis, even though both compounds stimulated contractility of the rat seminal vesicle at these concentrations. These data suggest that mating and adrenergic stimulation increase prostaglandin synthesis in] the rat seminal vesicle, probably through an α-adrenergically mediated mechanism.     

Seasonal occurrence of coccoliths in sediment traps from West Caroline Basin, equatorial West Pacific Ocean

Coccolith fluxes were investigated by sediment trap studies in the West Caroline Basin, which is located in the equatorial western Pacific. The investigation was conducted from June 1991 to March 1992 at two water depths, 1592 and 3902 m, as part of the Northwest Pacific Carbon Cycle Study (NOPACCS) program. Two seasonal maxima of coccolith fluxes were observed during September–early October and late December–January. The average coccolith and coccosphere fluxes at the depth of the shallow trap were 1800×10⁶ coccoliths m⁻² day⁻¹ and 1.9×10⁶ coccospheres m⁻² day⁻¹, respectively. The flux of coccoliths followed the same trend as the total flux, and was closely correlated with the flux of organic matter flux. Florisphaera profunda, Gladiolithus flabellatus, Gephyrocapsa oceanica, Umbilicosphaera sibogae var. sibogae, Emiliania huxleyi, and Oolithotus fragilis were the most abundant species together comprising more than 85% of the total flora. Observed seasonal changes of the species composition of coccolith flora, as well as analysis of the R-mode cluster, revealed that during the summer, the assemblage was marked by the dominance of G. oceanica and U. sibogae. However, during the winter, the assemblage was dominated by E. huxleyi and O. fragilis. These assemblage changes were influenced by monsoonal events, which were observed off the New Guinea coast. F. profunda dominated the community in the shallow trap throughout most of the year; peak values of this species were recorded during the winter. The coccosphere assemblage was dominated by G. oceanica at both water depths. In the deep trap, the sedimentation pattern was similar to that observed at the shallow depth. Mean coccolith and coccosphere fluxes at the deep trap were 2000×10⁶ coccolith m⁻² day⁻¹ and 0.08×10⁶ coccospheres m⁻² day⁻¹, respectively. The increase in coccolith flux with water depth suggests a lateral influx. The estimated average daily mass of CaCO₃ flux in coccoliths and coccospheres was 16.6 mg m⁻² day⁻¹ at the 1592 m trap and 17.9 mg m⁻² day⁻¹ at the 3902 m trap, respectively. These calculated values contributed only 23.3% to the total CaCO₃ flux at the shallow trap and 27.9% at the deep trap.     

Molecular mass and antitumor activities of sulfated derivatives of α-glucan from Poria cocos mycelia

Two kinds of water-insoluble (1 → 3)-α-d-glucan samples, ab-PCM3-I and ac-PCM3-I, isolated from different Poria cocos mycelia were sulfated, to produce two series of water-soluble derivatives ab-PCM3-I-S1–S5 and ac-PCM3-I-S1–S5, respectively. The derivatives having different weight-average molecular mass (M_w) were produced by changing reaction temperature and time as well as molar ratios between chlorosulfonic acid and number of hydroxyl groups in the glucan. The degrees of substitution (DS) of the sulfated derivatives were analyzed by elemental analysis (EA) to be 0.39–0.67 for ab-PCM3-I-S and 0.73–0.96 for ac-PCM3-I-S, respectively. The M_w and the intrinsic viscosity ([η]) of the samples ab-PCM3-I-S and the ac-PCM3-I-S were measured by size exclusion chromatography combined with laser light scattering (SEC–LLS) and viscometry in phosphate buffer solution (PBS) at 37 °C. The results indicated that their M_w ranged from 2.0 to 11.3 × 10⁴ for the samples ab-PCM3-I-S, and 4.7 to 40.0 × 10⁴ for the samples ac-PCM3-I-S. Moreover, the antitumor activities of the sulfated derivatives ab-PCM3-I-S and ac-PCM3-I-S against Sarcoma 180 tumor cell tested both in vitro and in vivo are significantly higher than those of the native α-d-glucans. Therefore, a moderate range of molecular mass from 2.0 × 10⁴ to 40.0 × 10⁴, relatively high chain stiffness and good water solubility of the sulfated derivatives are beneficial to the enhancement of their antitumor activities.     

Electrotransformation ofClostridium thermosaccharolyticum

Transformation of the thermophileClostridium thermosaccharolyticum ATCC 31960 was achieved using plasmid pCTC1 and electroporation. Evidence supporting transformation was provided by Southern blots, detection of the plasmid in 10 out of 10 erythromycin-resistant clones, retransformation ofE. coli andC. thermosaccharolyticum with plasmid DNA isolated fromC. thermosaccharolyticum, and a proportional relationship between the number of transformants and the amount of DNA added. Transformation efficiencies were very low for plasmid DNA prepared fromE. coli (0.6 transformants mg^–1 DNA), although somewhat higher for plasmid DNA prepared fromC. thermosaccharolyticum (52 transformants mg^–1 DNA). Transformation-dependent erythromycin resistance indicates that an adenosine methylase gene originating fromEnterococcus faecalis, a mesophile, is expressed inC. thermosaccharolyticum. The plasmid pCTC1 appears to be replicated independently of the chromosome, as indicated by visualization of recovered plasmid on gels, and retransformation using recovered plasmid. pCTC1 is maintained inC. thermosaccharolyticum at both 45 and 60°C. Restriction analysis showed little or no rearrangement occurred upon passage through the thermophile.     

Transformation of Zymomonas mobilis by a hybrid plasmid

The transformation of Zymomonas mobilis by plasmid DNA was achieved using a modification of the CaCl₂ method for Escherichia coli. The highest frequency of transformation obtained was 5 × 10³ transformants/μg DNA. The success of the method depended upon the use of a plasmid which is a cointegrate between a Z. mobilis cryptic plasmid and an E. coli plasmid carrying two selectable drug resistance markers.