An Investigation into the Effect of Fine Lactose Particles on the Fluidization Behaviour and Aerosolization Performance of Carrier-Based Dry Powder Inhaler Formulations

The effect of milled and micronized lactose fines on the fluidization and in vitro aerosolization properties of dry powder inhaler (DPI) formulations was investigated, and the suitability of static and dynamic methods for characterizing general powder flow properties of these blends was assessed. Lactose carrier pre-blends were prepared by adding different lactose fines (Lactohale® (LH) 300, 230 and 210) with coarse carrier lactose (Lactohale100) at 2.5, 5, 10 and 20 wt% concentrations. Powder flow properties of lactose pre-blends were characterized using the Freeman Technology FT4 and Schulze RST-XS ring shear tester. A strong correlation was found between the basic flow energy (BFE_Norm) measured using the Freeman FT4 Rheometer and the flowability number (ff_c) measured on Schulze RST-XS. These data indicate that both static and dynamic methods are suitable for characterizing general powder flow properties of lactose carriers. Increasing concentration of fines corresponded with an increase in the normalized fluidization energy (FE_Norm). The inclusion of fine particles of lactose resulted in a significant (p < 0.05) increase in fine particle delivery of budesonide and correlated with FE_Norm. This trend was strongest for lactose containing up to 10 wt% LH300. A similar trend was found for the milled lactose grades LH230 and LH210. However, the increase in FE_Norm upon addition of milled fines only corresponded to a very slight improvement in the performance. These data suggest that whilst the fluidization energy correlated with fine particle delivery, this relationship is specific to lactose grades of similar particle size.KEY WORDS: dry powder inhaler, fluidisation, lactose, powder flow     

Cumulative geoecological effects of 62 years of infrastructure and climate change in ice‐rich permafrost landscapes,Prudhoe Bay Oilfield,Alaska

Many areas of the Arctic are simultaneously affected by rapid climate change and rapid industrial development. These areas are likely to increase in number and size as sea ice melts and abundant Arctic natural resources become more accessible. Documenting the changes that have already occurred is essential to inform management approaches to minimize the impacts of future activities. Here, we determine the cumulative geoecological effects of 62 years (1949–2011) of infrastructure‐ and climate‐related changes in the Prudhoe Bay Oilfield, the oldest and most extensive industrial complex in the Arctic, and an area with extensive ice‐rich permafrost that is extraordinarily sensitive to climate change. We demonstrate that thermokarst has recently affected broad areas of the entire region, and that a sudden increase in the area affected began shortly after 1990 corresponding to a rapid rise in regional summer air temperatures and related permafrost temperatures. We also present a conceptual model that describes how infrastructure‐related factors, including road dust and roadside flooding are contributing to more extensive thermokarst in areas adjacent to roads and gravel pads. We mapped the historical infrastructure changes for the Alaska North Slope oilfields for 10 dates from the initial oil discovery in 1968–2011. By 2010, over 34% of the intensively mapped area was affected by oil development. In addition, between 1990 and 2001, coincident with strong atmospheric warming during the 1990s, 19% of the remaining natural landscapes (excluding areas covered by infrastructure, lakes and river floodplains) exhibited expansion of thermokarst features resulting in more abundant small ponds, greater microrelief, more active lakeshore erosion and increased landscape and habitat heterogeneity. This transition to a new geoecological regime will have impacts to wildlife habitat, local residents and industry.     

Interaction of phosphorylase kinase with thin filament proteins of rabbit skeletal muscles

The binding of phosphorylase kinase to thin filaments and their effects on the enzyme activity as well as the contribution of the enzyme to contractile protein phosphorylation have been studied. The data obtained suggest that the kinase binding to thin filaments is controlled by the regulatory proteins, troponin and tropomyosin. The bulk of the enzyme is bound to the F-actin-tropomyosin-troponin complex which activates the enzyme in a far greater degree than each of its constituent components. Ca2+ and ATP control the kinase binding to F-actin. ATP increases the enzyme binding 6-fold; Ca2+ decrease the S0.5 value for F-actin 5-fold. In acetone powder extracts phosphorylase kinase phosphorylates thin filament-bound phosphorylase b, troponin T and troponin I as well as 51-58 kDa and 114 kDa proteins. These results suggest that phosphorylase kinase plays a role in the mechanism of synchronization of glycogenolysis and muscle contraction rates.     

Kinetic studies on dextransucrase from the cariogenic oral bacterium Streptococcus mutans

The kinetic mechanism of dextransucrase was studied using the Streptococcus mutans enzyme purified by affinity chromatography to a specific activity of 36.9 mumol/min/mg of enzyme. In addition to dextran synthesis, the enzyme catalyzed sucrose hydrolysis and isotope exchange between fructose and sucrose. The rates of sucrose hydrolysis and dextran synthesis were partitioned as a function of dextran concentration such that exclusive sucrose hydrolysis was observed in the absence of dextran and exclusive dextran synthesis at high dextran concentrations. An analogous situation was observed with fructose-dependent partitioning of sucrose hydrolysis and fructose exchange. Steady state dextran synthesis and fructose isotope exchange kinetics were simplified by assay at dextran or fructose concentrations high enough to eliminate significant contributions from sucrose hydrolysis. This limited dextran synthesis assays to dextran concentrations above apparent saturation. The limitation was diminished by establishing conditions in which the enzyme does not distinguish between dextran as a substrate and product which allowed initial discrimination among mechanisms on the basis of the presence or absence of dextran substrate inhibition. No inhibition was observed, which excluded ping-pong and all but three common sequential mechanisms. Patterns of initial velocity fructose production inhibition and fructose isotope exchange at equilibrium were consistent with dextran synthesis proceeding by a rapid equilibrium random mechanism. A nonsequential segment was apparent in the exchange reaction between fructose and sucrose assayed in the absence of dextran. However, the absence of detectable glucosyl exchange between dextrans and the lack of steady state dextran substrate inhibition indicate that glucosyl transfer to dextran must occur almost exclusively through the sequential route. A review of the kinetic constants from steady state dextran synthesis, fructose product inhibition, and fructose isotope exchange showed a consistency in constants derived from each reaction and revealed that dextran binding increases the affinity of sucrose and fructose for dextransucrase.     

低温对植物叶片中超氧物歧化酶、过氧化氢酶和过氧化氢水平的影响

番茄和鸡蛋果叶片中可提取的SOD活性不受低温的影响。在电泳谱带上SOD主同工酶带被氰化物而不被低温抑制,次同工酶带在低温下不稳定,且活性很低,它的变化不影响总的SOD活性。一些冷敏感植物叶片中CAT活性被低温抑制,而H_2O_3水平在低温下稳定或有增加,这可能使毒性更强的羟基离子(OH·)易于形成。     

Protein-carbohydrate complementarity in mammalian gamete recognition

Recent studies suggest that gamete recognition in a number of species is mediated by complementary proteins and carbohydrates on opposing gamete surfaces. Studies in invertebrates and vertebrates have shown that carbohydrate-binding proteins on the sperm surface recognize and bind to complementary glycoconjugates on the egg's extracellular coat. This chapter reviews our current knowledge of gamete recognition in the mouse. The complementary receptors for both mouse sperm and egg have been identified, purified, and characterized. Their synthesis during gametogenesis has been defined, as have the effects of sperm capacitation and of the acrosome reaction on their expression and distribution. Their relationship to gamete receptors that function in other species is discussed. Finally, evidence is presented that suggests that one of the receptors that mediate mouse gamete recognition belongs to a family of cell surface receptors that function during multiple cellular interactions in development.     

Cell surface galactosyltransferase as a recognition molecule during development

Recent results from our laboratory suggest that a variety of cellular interactions during development are mediated, in part, by the binding of a cell surface enzyme, galactosyltransferase (GalTase), to its specific lactosaminoglycan (LAG) substrate on adjacent cell surfaces and in the extracellular matrix. Our present interest in surface GalTase developed from earlier biochemical studies of a series of morphogenetic mutations in the mouse which map to the T/t-complex. These studies identified a specific defect in the regulation of surface GalTase activity on morphogenetically abnormal cells, while eight other enzymes showed normal activity. This led us to consider the unique function of surface GalTase in those cell interactions that are influenced by mutations of the T/t-complex. By using a multidisciplinary approach, which included genetic, biochemical and immunological probes, we have found that GalTase functions as a surface receptor during fertilization, early embryonic cell adhesions, and embryonic cell migration on basal lamina matrices. Recently, we have examined the expression of surface GalTase during spermatogenesis, as well as the fate of sperm GalTase following the acrosome reaction. This paper summarizes the results of these studies, as well as others, which suggest that GalTase functions as a surface receptor during those cell interactions regulated by the T/t-complex alleles.     

Galactosyltransferase activity is restricted to the plasma membranes of equine and bovine sperm

beta 1, 4-Galactosyltransferase (GalTase) is localized to the plasma membrane of mouse sperm, in which it mediates the binding of sperm to glycoconjugate residues in the egg zona pellucida. In this study, the presence of subcellular distribution of sperm GalTase were determined in two other mammalian species that yield sufficient sperm for subcellular fractionation. Equine and bovine semen were collected, and the plasma membranes (PM), outer acrosomal membranes (OAM), and inner acrosomal membranes (IAM) were sequentially removed. The purities of the isolated membrane preparations were determined by transmission electron microscopy and found to be greater than or equal to 90%, 96%, and 98% for equine PM, OAM, and IAM, respectively, and greater than or equal to 80%, 94%, and 97% for bovine PM, OAM, and IAM, respectively. GalTase activity was assayed under optimal conditions in all membrane preparations and was preferentially localized to the isolated PM both in equine and in bovine spermatozoa. The selective localization of GalTase to the sperm PM in two other species suggest that it may serve as a generalized gamete receptor during initial sperm-egg binding in mammals.     

Cell surface beta 1,4-galactosyltransferase functions during neural crest cell migration and neurulation in vivo

Mesenchymal cell migration and neurite outgrowth are mediated in part by binding of cell surface beta 1,4-galactosyltransferase (GalTase) to N-linked oligosaccharides within the E8 domain of laminin. In this study, we determined whether cell surface GalTase functions during neural crest cell migration and neural development in vivo using antibodies raised against affinity-purified chicken serum GalTase. The antibodies specifically recognized two embryonic proteins of 77 and 67 kD, both of which express GalTase activity. The antibodies also immunoprecipitated and inhibited chick embryo GalTase activity, and inhibited neural crest cell migration on laminin matrices in vitro. Anti-GalTase antibodies were microinjected into the head mesenchyme of stage 7-9 chick embryos or cranial to Henson's node of stage 6 embryos. Anti-avian GalTase IgG decreased cranial neural crest cell migration on the injected side but did not cross the embryonic midline and did not affect neural crest cell migration on the uninjected side. Anti-avian GalTase Fab crossed the embryonic midline and perturbed cranial neural crest cell migration throughout the head. Neural fold elevation and neural tube closure were also disrupted by Fab fragments. Cell surface GalTase was localized to migrating neural crest cells and to the basal surfaces of neural epithelia by indirect immunofluorescence, whereas GalTase was undetectable on neural crest cells prior to migration. These results suggest that, during early embryogenesis, cell surface GalTase participates during neural crest cell migration, perhaps by interacting with laminin, a major component of the basal lamina. Cell surface GalTase also appears to play a role in neural tube formation, possibly by mediating neural epithelial adhesion to the underlying basal lamina.