Transport limitation of chlorine disinfection of Pseudomonas aeruginosa entrapped in alginate beads

An artificial biofilm system consisting of Pseudomonas aeruginosa entrapped in alginate and agarose beads was used to demonstrate transport limitation of the rate of disinfection of entrapped bacteria by chlorine. Alginate gel beads with or without entrapped bacteria consumed chlorine. The specific rate of chlorine consumption increased with increasing cell loading in the gel beads and decreased with increasing bead radius. The value of an observable modulus comparing the rates of reaction and diffusion ranged from less than 0.1 to 8 depending on the bead radius and cell density. The observable modulus was largest for large (3-mm-diameter) beads with high cell loading (1.8 x 10(9) cfu/cm(3)) and smallest for small beads (0.5 mm diameter) with no cells added. A chlorine microelectrode was used to measure chlorine concentration profiles in agarose beads (3.0 mm diameter). Chlorine fully penetrated cell-free agarose beads rapidly; the concentration of chlorine at the bead center reached 50% of the bulk concentration within approximately 10 min after immersion in chlorine solution. When alginate and bacteria were incorporated into an agarose bead, pronounced chlorine concentration gradients persisted within the gel bead. Chlorine did gradually penetrate the bead, but at a greatly retarded rate; the time to reach 50% of the bulk concentration at the bead center was approximately 46 h. The overall rate of disinfection of entrapped bacteria was strongly dependent on cell density and bead radius. Small beads with low initial cell loading (0.5 mm diameter, 1.1 x 10(7) cfu/cm(3)) experienced rapid killing; viable cells could not be detected (<1.6 x 10(5) cfu/cm(3)) after 15 min of treatment in 2.5 mg/L chlorine. In contrast, the number of viable cells in larger beads with a higher initial cell density (3.0 mm diameter, 2.2 x 10(9) cfu/cm(3)) decreased only about 20% after 6 h of treatment in the same solution. Spatially nonuniform killing of bacteria within the beads was demonstrated by measuring the transient release of viable cells during dissolution of the beads. Bacteria were killed preferentially near the bead surface. Experimental results were consistent with transport limitation of the penetration of chlorine into the artificial biofilm arising from a reaction-diffusion interaction. The methods reported here provide tools for diagnosing the mechanism of biofilm resistance to reactive antimicrobial agents in such applications as the treatment of drinking and cooling waters. (c) 1996 John Wiley & Sons, Inc.     

Chemical camouflage of nanospheres with a poorly reactive surface: towards development of stealth and target-specific nanocarriers

A two-step approach is described to chemically camouflage the inert surface of model polystyrene nanospheres of 60 nm in diameter against recognition by the body's defenses. The first step was based on the strong protein adsorbing potency of polystyrene, and the second step utilized the chemical reactivity of the adsorbed proteins for conjugation with cyanuric chloride-activated methoxypoly(ethyleneglycol)5000, mPEG5000. Bovine serum albumin (BSA) and rat IgG were used as models of non-immune and immune proteins, respectively. The maximum adsorbance values for both proteins were near expectation for a close-packed monolayer. Adsorption isotherms studies and analysis of the hydrodynamic thickness of the adsorbed protein layer confirmed the close-packed side-on mode of adsorption for BSA and the end-on mode of adsorption for IgG, respectively. Nucleophiles on the adsorbed proteins were then reacted with cyanuric chloride activated mPEG5000. The average poly(ethyleneglycol) (PEG) content for a 60-nm nanospheres was found to be 13.7+/-0.4 micromol PEG/micromol BSA and 3.6+/-0.3 micromol PEG/micromol IgG. The interaction of both PEG-bearing nanospheres with the hydrophobic column material octyl-agarose indicated surface heterogeneity among the nanospheres. Only nanospheres with the most hydrophilic phenotype (approximately 70% of the total population) exhibited stealth properties after intravenous injection to rats. In contrast to the described approach, incubation of uncoated nanospheres with preformed BSA-mPEG5000 conjugates failed to produce long circulating entities. For design of splenotropic particles cyanuric chloride-activated mPEG5000 was conjugated to BSA-coated polystyrene beads of 225 nm in diameter. Despite their stealth property to hepatic Kupffer cell recognition, these nanospheres were cleared by the splenic red pulp macrophages.     

Incorporating fluorescent dyes and quantum dots into magnetic microbeads for immunoassays

Microbeads that are both paramagnetic and fluorescently labeled are commercially available in colors spanning the visible spectrum. Although these commercial beads can be bright, polydispersity in both size and fluorescent intensity limit their use in quantitative assays. Very recently, more monodisperse beads have become available, but their large size and surface properties make them less than ideal for some bioassay applications. Here we describe methods to customize commercial nonfluorescent magnetic microparticles with fluorescent dyes and quantum dots (QDs) without affecting their magnetic or surface chemical properties. Fluorescent dyes and 3.3-nm diameter CdSe/ZnS QDs were sequestered within 0.8-micron diameter magnetic beads by swelling the polystyrene matrix of the bead in organic solvent, letting the chromophores partition, and then collapsing the matrix in polar solvents. Chromophore incorporation has been characterized using both UV-visible absorption spectroscopy and fluorescence microscopy, with an average of 3 x 10(8) rhodamine 6G molecules/bead and 6 x 10(4) QDs/bead. The modified beads are uniform in size and intensity, with optical properties comparable to currently available commercial beads. Immunoassay results obtained with our custom fluorescent magnetic microbeads are consistent with those obtained using conventional magnetic microbeads.     

Role of integrins in regulation of collagen phagocytosis by human fibroblasts

Phagocytosis of collagen fibrils by fibroblasts is an important pathway for degradation of extracellular matrix in mature connective tissues. To study regulatory mechanisms in phagocytosis, 2-μm fluorescent beads coated with either collagen (COL) or bovine serum albumin (BSA) were incubated with human gingival fibroblasts in vitro. For these studies single cell suspensions were prepared by trypsinization, and bead internalization and collagen receptor expression were assessed by flow cytometry. After 3-h incubations, up to 8-fold more cells internalized COL beads than BSA-coated beads. Increased collagen coating concentration was associated with elevated proportions of cells that internalized COL beads, and was observed also in the presence of competing fibronectin-coated beads. The number of beads per cell and the percent of phagocytic cells increased proportionally with higher bead loadings. At > 4 beads per cell a maximum of ∼︁80% of cells were phagocytic. Cells reacted with mAbs against the α1, α2, and α3 integrin subunits were, respectively, 5%, 98% and 93% positively stained above background controls. All cells that internalized COL beads exhibited α2 staining but there were large proportions of phagocytic cells that were not stained for α1. In unfixed cells, bead internalization caused an immediate reduction of surface staining of membrane-bound α2 by ∼︁55% which returned to control levels within 3 h, indicating that cell-surface α2 was internalized by phagocytosis. Preincubation of cells with up to 8 COL beads per cell reduced the proportion of phagocytic cells and the number of internalized beads after a second COL bead incubation 4 h later. To assess the relationship between the percent of phagocytic cells and α2 integrin levels, serum starvation and cycloheximide experiments were conducted. Compared to controls, serum starvation for 24 h induced a 3.2-fold increase of cells internalizing COL beads but did not alter α2 staining levels. In contrast, 3 h cycloheximide treatment reduced α2 staining to 60% of control levels and this treatment also inhibited COL bead internalization. GRGDTP peptide as well as mAbs against the α1 and α2 subunits significantly reduced internalization of COL beads by 1.8 to 2.6-fold, whereas GRGESP peptide and α3 mAb exerted no effect. Internalization of BSA beads was not affected by any of these treatments. Collectively, these data indicate that the α2 integrin, along with other, as yet unidentified components, is likely involved in COL bead internalization. The α2 integrin subunit is rapidly recycled or synthesized following a phagocytic load. In contrast, the α1 integrin is not directly required for phagocytosis but may regulate the internalization step. © 1996 Wiley-Liss, Inc.     

Assessment of antral grinding of a model solid meal with echo-planar imaging

Mathematical modeling of how physical factors alter gastric emptying is limited by lack of precise measures of the forces exerted on gastric contents. We have produced agar gel beads (diameter 1.27 cm) with a range of fracture strengths (0.15-0.90 N) and assessed their breakdown by measuring their half-residence time (RT(1/2)) using magnetic resonance imaging. Beads were ingested either with a high (HV)- or low (LV)-viscosity liquid nutrient meal. With the LV meal, RT(1/2) was similar for bead strengths ranging from 0.15 to 0.65 N but increased from 22 +/- 2 min (bead strength <0.65 N) to 65 +/- 12 min for bead strengths >0.65 N. With the HV meal, emptying of the harder beads was accelerated. The sense of fullness after ingesting the LV meal correlated linearly (correlation coefficient = 0.99) with gastric volume and was independently increased by the harder beads, which were associated with an increased antral diameter. We conclude that the maximum force exerted by the gastric antrum is close to 0.65 N and that gastric sieving is impaired by HV meals.     

Determination of the affinity of vitamin D metabolites to serum vitamin D binding protein using assay employing lipid-coated polystyrene beads.

We have developed an assay to measure the affinity of serum vitamin D binding protein for 25-hydroxyvitamin D3, 1,25-dihydroxyvitamin D3, and vitamin D3, using uniform diameter (6.4 microns) polystyrene beads coated with phosphatidylcholine and vitamin D metabolites as the vitamin D donor. The lipid metabolite coated beads have a solid core, and thus all of the vitamin D metabolites are on the bead surface from which transfer to protein occurs. After incubating these beads in neutral buffer for 3 h, essentially no 3H-labeled vitamin D metabolites desorb from this surface. Phosphatidylcholine/vitamin D metabolite-coated beads (1 microM vitamin D metabolite) were incubated with varying concentrations of serum vitamin D binding protein under conditions in which the bead surfaces were saturated with protein, but most of the protein was free in solution. After incubation, beads were rapidly centrifuged without disturbing the equilibrium of binding and vitamin D metabolite bound to sDBP in solution was assayed in the supernatant. All three vitamin D metabolites became bound to serum vitamin D binding protein, and after 10 min of incubation the transfer of the metabolites to serum vitamin D binding protein was time independent. The transfer followed a Langmuir isotherm, and the Kd for each metabolite binding to serum vitamin D binding protein was derived by nonlinear least-squares fit analysis. From this analysis the following values for the Kd were obtained: 5.59 x 10(-6) M, 25-hydroxyvitamin D; 9.45 x 10(-6) M, 1,25-dihydroxyvitamin D; and 9.17 x 10(-5) M, vitamin D. In conclusion, we have developed a method which avoids problems encountered in previous assays and allows the precise and convenient determination of binding affinities of vitamin D metabolites and serum vitamin D binding protein.     

Mobility of integrin alpha5beta1 measured on the isolated ventral membranes of human skin fibroblasts

We have measured the lateral mobility of individual alpha5 integrin molecules in ventral plasma membranes of fibroblasts, which were prepared by removal of apical surfaces and nuclei followed by elimination of actin filaments with gelsolin, an actin-severing protein. The cytoplasmic domain of individual integrin molecules was tagged with 100 nm fluorescent polystyrene bead, and motion of the bead was observed and video-recorded. Position of the bead in each frame was determined from the centroid of the fluorescence image, from which plots of the mean-square displacement against time intervals were derived. Within short intervals of time (<100 ms) the mean-square displacement was proportional to the time interval, and the averaged translational diffusion coefficient of (5.3+/-4.4) x 10(-10) cm2/s was obtained with a broad distribution of (1.3-20) x 10(-10) cm2/s. The broad distribution might reflect the oligomerized state of integrin. The largest diffusion coefficient was comparable to that of lipid molecules previously measured in cells and probably represented the diffusion of a single integrin molecule in the presence of little interference of actin cytoskeleton or extracellular matrix. In longer time intervals (>100 ms) the motion of the bead was confined in an area, the average diameter of which was 410+/-160 nm. This was similar to the values described in previous reports, in which the motion of other membrane receptors labeled on their extracellular domain was measured in living cells.     

Intracellular singlet oxygen generation by phagocytosing neutrophils in response to particles coated with a chemical trap.

To determine if singlet oxygen (O2(1 delta g)) is produced by neutrophils (PMNs) during the process of phagocytosis, glass beads were coated with a specific chemical trap for O2(1 delta g), 9,10-diphenylanthracene (DPA). Singlet oxygen, but not other reactive oxygen species, reacts rapidly with DPA at a rate of kr = 1.3 x 10(6) M-1 s-1 to form a stable product, DPA-endoperoxide (Corey, E. J., and Taylor, W. C. (1964) J. Am. Chem. Soc. 86, 3881-3882; Wasserman, H. H., Scheffer, J. R., and Cooper, J. L. (1972) J. Am. Chem. Soc. 94, 4991-4996; Turro, N. J., Chow, M.-F., and Rigaudy, J. (1981) J. Am. Chem. Soc. 103, 7218-7224). The production of DPA-endoperoxide was determined by ultraviolet spectroscopy as a decrease in DPA absorbance at 355 nm. The absorbance of DPA was normalized to the absorbance of perylene, which was included in the coating on the beads as a nonreactive, internal standard. In the present study, DPA- and perylene-coated beads were initially allowed to adhere to fibronectin-coated coverslips. PMNs were then added to the bead-coated coverslips and allowed to adhere and phagocytose the beads for 1 h at 37 degrees C. In some experiments, 4B-phorbol-12-myristate-13-acetate (PMA) (1 ng/2.5 x 10(7) cells/ml), a known activator of the PMN NADPH-oxidase, was added as a co-stimulant. The amount of O2(1 delta g) produced by phagocytically stimulated PMNs was calculated to be 11.3 +/- 4.9 nmol of O2(1 delta g)/1.25 x 10(6) cells. Low dose PMA co-stimulation increased the production of O2(1 delta g) to 14.1 +/- 4.1 nmol/1.25 x 10(6) cells. Averaged together these amounts represent approximately 19 +/- 5.0% of the total oxygen consumed by PMNs in response to DPA- and perylene-coated beads. The specificity of the DPA reaction with O2(1 delta g) was confirmed by warming to 120 degrees C, which releases O2(1 delta g) from the DPA-endoperoxide, regenerating the parent DPA compound (Wasserman et al., 1972; Turro et al., 1981) and the absorbance at 355 nm. In addition, beta-carotene, an avid quencher of O2(1 delta g), was included in the coating of some bead preparations; assays in which these beads were used showed no change in the absorbance at 355 nm. Singlet oxygen production by myeloperoxidase was also measured using the coated bead assay and the results suggest that this is a major pathway by which singlet oxygen is generated in phagocytically stimulated PMNs.(ABSTRACT TRUNCATED AT 400 WORDS)     

Influence of size of emboli on extravascular lung water

We examined the influence of the size of emboli on the vascular volume (QL) and extravascular volume (Qev) accessible to 3HOH during a single pass through an isolated dog lung lobe using the double indicator-dilution method with 125I-human serum albumin as the vascular indicator. As successively more beads of a given diameter (58, 548, or 3,175 microns) were introduced into a lung lobe, a linear relationship between QL and Qev was obtained as they both decreased. The slope of the graph of QL vs. Qev with progressive embolism was directly proportional to the bead diameter. This suggested an approach for estimating the total vascular volume in vessels smaller than the diameter of the beads before embolization, referred to as Qm. If it is assumed that most of the transvascular diffusional exchange of 3HOH occurs in vessels smaller than the smallest beads (mainly capillaries) and that vessel obstruction does not change the ratio of Qev to the perfused capillary volume, the slope of the plot of QL vs. Qev is an estimate of the fraction, Qm/QL, of the total vascular volume in vessels smaller than the bead diameter. In the dog lung lobes studied, Qm/QL was approximately 0.64 for 58-microns vessels, 0.75 for 548-microns vessels, and 0.82 for 3,175-microns vessels. The results suggest that, with occlusion of vessels greater than or equal to 58 microns, 3HOH does not diffuse significantly into unperfused regions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intestinal NaCl transport in NHE2 and NHE3 knockout mice

Sodium/proton exchangers [Na(+)/H(+) (NHEs)] play an important role in salt and water absorption from the intestinal tract. To investigate the contribution of the apical membrane NHEs, NHE2 and NHE3, to electroneutral NaCl absorption, we measured radioisotopic Na(+) and Cl(-) flux across isolated jejuna from wild-type [NHE(+)], NHE2 knockout [NHE2(-)], and NHE3 knockout [NHE3(-)] mice. Under basal conditions, NHE(+) and NHE2(-) jejuna had similar rates of net Na(+) (approximately 6 microeq/cm(2) x h) and Cl(-) (approximately 3 microeq/cm(2) x h) absorption. In contrast, NHE3(-) jejuna had reduced net Na(+) absorption (approximately 2 microeq/cm(2) x h) but absorbed Cl(-) at rates similar to NHE(+) and NHE2(-) jejuna. Treatment with 100 microM 5-(N-ethyl-N-isopropyl) amiloride (EIPA) completely inhibited net Na(+) and Cl(-) absorption in all genotypes. Studies of the Na(+) absorptive flux (J) indicated that J in NHE(+) jejunum was not sensitive to 1 microM EIPA, whereas J in NHE3(-) jejunum was equally sensitive to 1 and 100 microM EIPA. Treatment with forskolin/IBMX to increase intracellular cAMP (cAMP(i)) abolished net NaCl absorption and stimulated electrogenic Cl(-) secretion in all three genotypes. Quantitative RT-PCR of epithelia from NHE2(-) and NHE3(-) jejuna did not reveal differences in mRNA expression of NHE3 and NHE2, respectively, when compared with jejunal epithelia from NHE(+) siblings. We conclude that 1) NHE3 is the dominant NHE involved in small intestinal Na(+) absorption; 2) an amiloride-sensitive Na(+) transporter partially compensates for Na(+) absorption in NHE3(-) jejunum; 3) cAMP(i) stimulation abolishes net Na(+) absorption in NHE(+), NHE2(-), and NHE3(-) jejunum; and 4) electroneutral Cl(-) absorption is not directly dependent on either NHE2 or NHE3.     

Measurement of phagocytosis using fluorescent latex beads

Fluorescent monodisperse latex beads and a computer-centered spectrofluorimeter were used to devise a sensitive new assay for phagocytosis. LM fibroblasts, a transformed cell line with a high endocytic rate, were exposed to fluoresbrite beads and the following parameters were investigated: incubation time, incubation temperature and bead/cell ratio. The bead uptake was linear for 60 min over a wide range of bead/cell ratios up to 130 beads/cell. Phagocytosis was inhibited at 4 degrees C, by incubation in the presence of colchicine, and by glucose deprivation. Scanning and transmission electron microscopy were used to confirm that at 37 degrees C both bead adsorption and internalization occurred while at 4 degrees C only bead adsorption but not endocytosis occurred. Large bead sizes (0.86 and 1.72 micrometer diameter) were most useful due to higher fluorescence and higher signal to noise ratios than smaller beads (0.25 and 0.57 micrometer diameter). Beads (0.86 micrometer diameter) were taken up at a rate of 4.4 beads/cell/h at 37 degrees C when a bead/cell ratio of 70 was used. The uptake was zero when assayed at zero time. These criteria establish that fluoresbrite beads provide a useful new fluorimetric assay for phagocytosis.     

Batch fermentation with entrapped growing cells ofLactobacillus casei

Summary Growing cells ofLactobacillus casei were entrapped in-carrageenan/locust bean gum (LBG) (2:1 or 2.75%:0.25% w/w respectively) mixed gel beads (two ranges of diameter: 0.5–1.0 and 1.0–2.0 mm) to fermentLactobacillus Selection (LBS) medium and produce biomass. The results showed significant influence of initial cell loading of the beads and bead size on the fermentation rate. The highest cell release rates were obtained with 2.75%:0.25%-carrageenan/LBG small diameter gel beads. However, 17 h fermentation of LBS medium with immobilized cells resulted in substantial softening of the gel matrix, prohibiting reuse of immobilized biocatalysts as inoculum in subsequent batch fermentation. A dynamic shear rheological study showed that the gel weakness was related to chemical interactions with the medium. Results indicated that part of the matrix-stabilizing K⁺ ions diffused back to the medium. Stabilization of the gel was obtained by adding potassium ions to the LBS medium;L. casei growth was not altered by this supplementation. Fermentation of LBS medium supplemented with KCl byL. casei showed higher cell counts in the broth medium with immobilized cells than with free cells, reaching 10¹⁰ cells/ml after about 10 h with entrapped cells in 0.5–1.0 mm diameter beads and 17 h with free cells. Counts in the gel beads after fermentation were higher than 10¹¹ cells/ml and bead integrity was maintained throughout fermentation.     

稻草覆盖对冬闲稻田二氧化碳通量的影响

通过研究稻草覆盖对土壤含水量、土壤温度及田间杂草生长的影响,探讨了亚热带红壤性稻田生态系统冬闲期稻草覆盖对CO2通量的影响及其机理.结果表明:稻草覆盖主要通过两个途径影响冬闲稻田的CO2通量:一是稻草覆盖对土壤温度有正效应,使稻田生态系统CO2排放量显著增加,试验期间覆盖处理平均净排放1.99 g CO2·m-2·d-1,而对照处理平均净吸收2.68 g CO2·m-2·d-1,两者差异达极显著水平(P《0.01);二是稻草覆盖使田间杂草的生物量及其吸收的光合有效辐射显著减少, 导致覆盖处理白天CO2排放量提高.稻草覆盖处理土壤表层(0～15 cm)相对含水量比无覆盖处理提高了9%以上,但未对CO2通量的变化产生显著影响.     

Fibronectin-mediated binding and phagocytosis of polystyrene latex beads by baby hamster kidney cells

The binding and phagocytosis of fibronectin (pFN)-coated latex beads by baby hamster kidney (BHK) cells was studied as a function of fibronectin concentration and bead diameter. Cells were incubated with radioactive pFN-coated beads, and total bead binding (cell surface or ingested) was measured as total radioactivity associated with the cells. Of the bound beads, those that also were phagocytosed were distinguished by their insensitivity to release from the cells by trypsin treatment. In continuous incubations, binding of pFN-coated beads to cells occurred at 4 degrees C or 37 degrees C, but phagocytosis was observed only at 37 degrees C. In addition, degradation of 3H-pFN from ingested beads occurred at 37 degrees C, as shown by the release of trichloroacetic acid-soluble radioactivity into the incubation medium. When the fibronectin density on the beads was varied, binding at 4 degrees C and ingestion at 37 degrees C were found to have the same dose-response dependencies, which indicated that pFN densities that permitted bead binding were sufficient for phagocytosis to occur. The fibronectin density for maximal binding of ingestion was approximately 250 ng pFN/cm2. When various sized beads (0.085-1.091 micron), coated with similar densities of pFN, were incubated with cells at 4 degrees C, no variation in binding as a function of bead size was observed. Under these conditions, the absolute amount of pFN ranged from less than 100 molecules on the 0.085-micron beads to greater than 15,000 molecules on the 1.091-micron beads. Based upon these results it can be concluded that the critical parameter controlling fibronectin-mediated binding of latex beads by BHK cells is the spacing of the pFN molecules on the beads. Correspondingly, it can be suggested that the spacing between pFN receptors on the cell surface that is optimal for multivalent interactions to occur is approximately 18 nM. When phagocytosis of various sized beads was compared, it was found that the largest beads were phagocytosed slightly better (two fold) than the smallest beads. This occurred both in continuous incubations of cells with beads and when the beads were prebound to the cells. Finally, the kinetic constants for the binding of 0.085 microM pFN-coated beads to the cells were analyzed. There appeared to be approximately 62,000 binding sites and the KD was 4.03 X 10(-9) M. Assuming a bivalent interaction, it was calculated that BHK cells have approximately 120,000 pFN receptors/cell and the binding affinity between pFN and its receptor is approximately 6 X 10(-5) M.     

Production of alginate beads by a rotative atomizer

Summary The use of a rotative flat disc atomizer for the production of biocatalysts immobilized in alginate gel was investigated. The influence of viscosity (0.28 to 1.53 Pa · s), disc diameter (5 to 20 cm), flow rate (0.5 to 2 L/min) and rotational speed (1.6 to 5 rev/s) on drop formation mechanisms, bead diameter and their distributions were investigated. Bead diameters ranging from 1 to 3 mm were been obtained. Flow rate and viscosity increases resulted in higher bead diameters but increasing disc diameter and rotational speed had a decreasing effect. A viscosity of 0.65 Pa · s would prove best for production of beads with good size distribution and mechanical strength. Predictive equations for the mean volumetric bead diameter were determined.     

Preparation of magnetically susceptible polyacrylamide/magnetite beads for use in magnetically stabilized fluidized bed chromatography

Spherical polyacrylamide/magnetite (PAM) composite beads, suitable for use in a magnetically stabilized fluidized bed (MSFB), were manufactured by a suspension polymerization method. Yield of beads depended on the type and concentration of buffer used during polymerization as well as the pH. More stabilizer was needed to prevent bead agglomeration as magnetite concentration increased. Bead diameter ranged from less than 60 to 600 mum, depending on reaction conditions, and the bead mean diameter and size distribution decreased with increasing impeller speed. The density and roundness factor of the beads were 1.19 +/- 0.02 g cm(-3) and 1.08 +/- 0.03, respectively. The beads had high magnetization at a low applied magnetic field strength (60 mT at 75 kA m(-1)) and retained little residual magnetization (<2 mT) after the field was removed. Incorporation of magnetite did not significantly affect the physical strength of the beads: the beads' average elastic modulus was 14 +/- 4 kPa, similar to reported values for polyacrylamide gels (15.8 kPa). The beads were stable in a range of buffers from pH 1 to 10 and were resistant to microbial degradation. The fluidization and stabilization behavior of the beads was examined in a bench-scale MSFB. The minimum fluidization velocity (U(mf)) of the beads (0.035 mm s(-1)) allowed the MSFB to be operated at superficial velocities close to those used in HPLC systems. Against expectations, at high superficial velocities, the stabilized bed of the MSFB had a greater expansion than the unstabilized bed. The PAM beads could be derivatized and activated for soybean trypsin inhibitor immobilization by a standard carbodiimide method, and the affinity separation of trypsin from chymotrypsin was demonstrated. The PAM beads show excellent potential for use in MSFB chromatography. (c) 1997 John Wiley & Sons, Inc.     

Influence of pulmonary embolism on absorption of inhaled iodide-125

To evaluate the influence of embolus size on the absorption of 125I- deposited on the bronchoalveolar surface, we exposed isolated perfused rabbit lungs to an aerosol containing 125I- for 5 min. We monitored the blood radioactivity for the subsequent 2 h. Several groups of lungs were studied, including those in which blood flow was varied and those in which enough glass beads ranging in size from 58 to 548 micron were injected into the pulmonary artery to approximately double the vascular resistance. The results indicated that under control conditions approximately 94% of the 125I- deposited on the intrapulmonary bronchoalveolar surface was able to reach the pulmonary circulation during the 2-h perfusion period, and the bronchoalveolar surface was sufficiently perfused so that absorption was limited by the rate of diffusion into the blood rather than the rate of blood flow. In the absence of embolization, the initial absorption rate was approximately 10.4%/min regardless of the total flow rate. The 58-micron beads reduced the rate to approximately 7.5%/min, whereas the beads greater than or equal to 194 micron in diameter reduced the rate to approximately 4.5%/min. Thus the effect of the embolization on the absorption rate was directly related to the bead diameter, even though the number of beads injected was adjusted to produce about the same increase in vascular resistance.     

Effective Bead Preparation of Coimmobilized Methanogenic and Methanotrophic Bacteria for Tetrachloroethene Degradation

Three types of coimmobilized methanogenic and methanotrophic bacterial beads – Ca-alginate, Ba-alginate, and Ca-alginate chitosan – were used for tetrachloroethene (PCE) degradation. For the purpose of effective preparation of coimmobilized bacterial beads, the diameter and broken-loading of beads were measured. The activity tests to find the optimal bacteria concentration in the bead were performed. It was found that Ba-alginate beads had superiority in bacterial growth and the degree of strength of beads from the diameter and broken-loading tests. Also, it was shown that it is most effective to add 200 mL of methanogens into 500 mL of 2% alginate solution and 20 mL of methanotrophs into 500 mL to 2% alginate solution. When methanogens and methanotrophs were applied with the Ba-alginate bead in the actual dechlorination of PCE, the biological PCE dechlorination rate was 92%, and there was highly effective degradation of PCE based on the coimmobilized bead. Additionally, relation to the diameter (X) and broken-loading (Y) of the Ba-alginate bead was derived following equation, Y = 438.02 exp(–1.4815 X).     

三江平原春小麦农田生态系统氧化亚氮通量特征

利用静态暗箱-气相色谱法对三江平原春小麦农田生态系统N2O排放通量进行连续2.5年的田间原位观测.结果表明:三江平原春小麦农田生态系统N2O排放通量具有较明显的季节变化和年际变化,并主要与年际间降水及田间水分管理差异有关;春小麦农田生态系统N2O排放日变化与气温及地下5 cm温度变化有关.生长期N2O的排放较强,休耕期N2O排放量显著下降,冰冻期N2O的排放较微弱,融冻时N2O排放缓慢增强.生长期N2O平均排放通量为0.190 mg.m-2.h-1,收割后到冰冻期间为0.077 mg.m-2.h-1,冻融期间为0.017 mg.m-2.h-1.     

Biotinylated photocleavable polyethylenimine: capture and triggered release of nucleic acids from solid supports

A biotinylated photocleavable polyethylenimine (B-PC-PEI) was designed and synthesized for the capture and controlled release of nucleic acids from solid supports. B-PC-PEI was synthesized via a three-step reaction process and verified by 1H NMR and mass spectrometry. In aqueous solution, the o-nitrobenzyl group within B-PC-PEI was efficiently cleaved by 5 min of 365 nm light exposure from a distance of 20 cm (9 mW/cm2). When coupled to streptavidin-coated beads, the PEI domain of Cy5-labeled B-PC-PEI was released by 365 nm light exposure. In contrast, a Cy5-labeled biotinylated PEI (B-PEI) was used as a control and negligible fluorescence loss was observed. Cy5-labeled siRNA was electrostatically captured to streptavidin-coated beads preabsorbed with B-PC-PEI or B-PEI, and flow cytometry demonstrated significant loss of fluorescence from the bead surface after 5 min of light exposure only for B-PC-PEI, demonstrating controlled release of siRNA from the bead surface. Finally, the release of the Cy5-labeled siRNA into the supernatant was quantified. The release of Cy5-siRNA into the supernatant was significantly greater after 5 min of light exposure for B-PC-PEI/streptavidin beads compared to 0 min exposure and remained unchanged for B-PEI/streptavidin beads. B-PC-PEI facilitates capture and triggered release of surface-tethered nucleic acids with light exposure and is fully compatible with streptavidin-based applications.