Phosphorus-Containing Fluoro-Sulfonated Polytriazole Membranes with High Proton Conductivity: Understanding Microstructural and Thermomechanical Behaviors as a Function of Degree of Sulfonation

The current article describes the design and synthesis of a new series of phosphorus-containing fluoro-sulfonated polytriazoles through click polymerization. The synthesized copolytriazoles (PTPFDSH-70 to 90) with different degrees of sulfonation (DS) from 70% to 90% are structurally interpreted by various spectroscopic techniques (¹H, ¹³C, and FTIR). The high molecular weight (weight average molecular weight as high as 77 500 g mol⁻¹ with polydispersity index of 2.29) polymers exhibits excellent mechanical (elongation at break up to 95%), thermal (10% decomposition temperature: 266–317 °C), and oxidative (>14.5 h) stability. The PTPFDSH-70 to 90 possess outstanding water-holding ability in hydrated conditions (swelling ratio [in-plane]: 6.2–7.3% at 80 °C). The microstructural alterations by their thermal relaxations and transitions with increasing DS in the polymers have been thoroughly investigated by dynamic mechanical analysis. The atomic force microscopy and transmission electron microscope images of the PTPFDSH-70 to 90 polymer membranes demonstrated the phase segregated interconnected ionic cluster-like morphology between hydrophilic and hydrophobic domains. The PTPFDSH-90 (DEB:PFAZ:DSAZ = 100:10:90) polymer membrane displays the proton conductivity (176 and 190 ms cm⁻¹ at 80 and 90 °C, respectively) higher than Nafion117 under similar test conditions.     

Evaluation of Genetic Stability of Recombinant Human Factor VIII by Peptide Mapping and On-line Mass Spectrometric Analysis

Purpose. The genetic stability of a recombinant human factor VIII (rhFVIII) product expressed in Chinese hamster ovary cells (Recombinate) has been evaluated through comparisons of the protein produced at the beginning, middle and end of a typical production campaign. Methods. Recombinant human factor VIII was incubated with thrombin, the resulting four polypeptides were isolated by RP-HPLC, subjected to proteolysis with trypsin, and the peptide mixtures were resolved by RP-HPLC. Tryptic peptide mixtures were subjected to online mass spectrometric analysis using an electrospray ionization source interfaced to a quadrupole mass analyzer scanning from 1950–200 amu, and the peptide ion data were compared for three lots produced from the beginning, middle and end of a production campaign. Results. The UV elution profiles for each of the rhFVIIIa polypeptides were highly similar for factor VIII isolated from the beginning, middle and end of production. Total ion data from the peptide maps derived from three lots of rhFVIII were compared by MH¹⁺ values as a function of scan range. A total of 918 ions were analyzed for the four polypeptides of rhFVIII produced at the beginning, middle and end of a production campaign. The ions were detected at the same relative retention times, as indicated by the similar scan numbers for the three lots. Conclusions. These observations support that rhFVIII preparations produced from the beginning, middle and end of a production campaign were highly similar, and demonstrate genetic stability in the manufacturing process of Recombinate.     

A new long shelf life formulation of modified Ham's F-10 medium: Biochemical and clinical evaluation

Purpose To evaluate biochemically and clinically a new formulation of modified Ham's F-10 medium made without the inclusion of hypoxanthine. The medium was formulated for long-term storage and use by separately preparing a stable liquid (basal) portion and a freeze-dried supplement containing the labile medium components.Results Following 18 months of storage the basal medium was biochemically analyzed for its amino acid (aa's) and vitamin content. Cysteine and tryptophan were decreased to less than 30% of their starting theoretical concentrations (STCs). Asparagine, serine, tyrosine, histidine and lysine were present at 50% to 70% of their STC. The remaining aa's were all within 90% of their STCs except arginine which was at 77%. All of the vitamins were present at 90% or more of their STCs except inositol, riboflavin and'thiamine which were present at 70% of their STCs. IVF with the new formulation resulted in 13 deliveries from 51 aspirations (25%) as compared with 10/39 (26%) in 1991, when standard medium preparation was used. Oocyte donation resulted in 30 deliveries from 84 cycles (36%) with the new formulation as compared with 21/65 (32%) in 1991.Conclusions (1) The new basal with lyophilized supplement formulation produces similar clinical results in the IVF laboratory as medium prepared in the standard fashion, (2) certain amino acids and vitamins are not stable in the liquid basal medium, and (3) the separate formulation of a liquid basal medium with lyophilized supplement is convenient, viable alternative to modified Ham's F-10 medium prepared in the standard manner (i.e., from powder) and may decrease the need for frequent medium preparation.Modified Ham's F-10 Medium, Irvine Scientific, Santa Ana, California.Presented at the 42nd Annual Meeting of the Pacific Coast Fertility Society, Indian Wells, California, April 20–24, 1994.     

Multihost, Multiparasite systems: An application of bifurcation theory

Author to whom all correspondence should be addressed (email: j.v.greenman{at}stir.ac.uk). The local analysis of multihost multiparasite models has beenhampered by algebraic intractability. There have been two responsesto this difficulty: extensive numerical investigation, and simplificationto a level where analytical techniques work. In this paper wedescribe another approach, based on bifurcation theory, in whichthe qualitative properties of the model equilibrium structureare realized on an array of maps drawn in parameter space. Thisapproach is described in the context of two models: the basictwo-host shared microparasite S–I model and the single-hosttwo-microparasite S–I (susceptible-infective) model. Theprocedure involved does not require model simplification througha reduction in dimensionality. It can handle intraspecific aswell as parasite-mediated competition and, in the second model,single-host parasite coexistence. The map arrays provide a concisecatalogue of the possible modes of behaviour of a system andan explanation for changes in that behaviour. In particular,the reasons why the conjectures made about the behaviour ofthe first of these models do not hold throughout parameter spaceare immediately clear from the map structure, as are the conditionsfor collusive and competitive behaviour between the two typesof parasite in the second model.     

Purification and Partial Characterization of an Indomethacin Hydrolyzing Enzyme from Pig Liver

Purpose. Indomethacin is well known to be metabolized via O-demethylation and N-deacylation. In this paper we found an enzyme involved in the hydrolysis of amide-linkage of indomethacin and partially characterized it as well as its substrate specificity. Methods. An indomethacin hydrolyzing enzyme was purified to homogeneity from pig liver microsomes using columns of Q-Sepharose, Red-Sepharose and Blue-Sepharose. The enzyme activity was assayed by measuring of -chlorobenzoic acid liberated from indomethacin by HPLC. Results. The purified enzyme effectively hydrolyzed the amide linkage in indomethacin but not those in -naphthylacetate and -nitrophenylacetate, which are typical substrates for carboxylesterase. The subunit molecular mass of the enzyme was 65 kDa according SDS-polyacrylamide gel electrophoresis. The Michaelis constant (Km) and maximum velocity (Vmax) values for indomethacin were 67.8 µM and 9.02 nmol/min/mg protein, respectively. The amino acid sequence analysis of the enzyme after cyanogen bromide cleavage showed high homology with a mouse carboxylesterase isozyme designated as ES-male. The activity of indomethacin hydrolysis was relatively high in the pig, rabbit and human liver homogenate, but not in those from rat and mouse. On the other hand, purified human liver carboxylesterases pl 5.3 and 4.5, and pig liver carboxylesterases have no catalytic activity for indomethacin. Conclusions. These results indicate that the hydrolysis of amide-linkage of indomethacin in humans would be associated with an enzyme similar to the indomethacin hydrolyzing enzyme from pig liver microsomes described here.     

Overexpression and purification of enzymatically active recombinant integrase protein of rous sarcoma virus

The carboxy-terminal domain of polymerase gene of Rous sarcoma virus was cloned into an expression vector under the control oflac regulatory elements, resulting in the plasmid pMF1413. Upon isopropyl--D-thiogalactopyranoside induction, viral integration (IN) protein was expressed in large quantity inEscherichia coli. The expressed recombinant protein was prepurified by successive washing of the bacterial pellet with 0.1 M NaCl and detergents. Further purification was performed in high yield by standard chromatography methods. The purified enzyme revealed selective DNA cleaving activity on supercoiled plasmid with the LTR-LTR junction fragment. The reaction was metal ion dependent, with a preference for Mn²⁺ over Mg²⁺, and showed substrate specificity at 1 mM MnCl₂.     

The effects of urinastatin on the plasma levels of granulocyte elastase during open heart surgery under simple deep hypothermia

Changes in granulocyte elastase (GLE) and -gluculonidase (-gl) were observed during open heart surgeries which were performed under deep hypothermia with surface cooling. In addition, the effect of urinary trypsin inhibitor, urinastatin, on the activities of these enzymes was studied. The patients were divided into three groups, namely group U-I with intravenous injection of 6000u·kg^–1 of urinastatin before cooling, group U-II administered with an additional 6000u·kg^–1 after warming to 30°C, and an untreated group (Group C). The plasma level of GLE increased significantly in the three groups compared with the level before cooling respectively. In the group U-II, the GLE level after the warming was lower than that in the control group. The serum level of -gl increased significantly in the three groups at the end of rewarming (36°C). The release of GLE from lysosomes in granulocytes was inhibited in the group U-II. The insufficient inhibition of GLE release in the group U-I is probably due to relatively short half-life of urinastatin. Therefore double administration of 6000u·kg^–1, before and after the cooling, may be required to achieve the therapeutic effect. Consequently, urinastatin appears to be useful in open heart surgery under deep hypothermia with surface cooling.(Kawamura T, Shimoda Y, Wakusawa R: The effects of urinastatin on the plasma levels of granulocyte elastase during open heart surgery under simple deep hypothermia. J Anesth 6: 269–276, 1992)     

Time-dependent kinetics. V: Time course of drug levels during enzyme induction (one-compartment model)

Equations were derived to describe the time course of drug levels during auto- and heteroinduction under a variety of input conditions. These equations were based on a pharmacokinetic theory of induction which assumes that metabolic clearance increases exponentially to a maximum value and that the rate of this increase is governed by the degradation rate constant of the induced enzyme (k). Closed form solutions could be obtained only for intravenous single-dose (case I) and multiple-dose (case IV) administration. For each of the other cases, constant-rate intravenous infusion (case III), oral single-dose administration (case II), and multiple-dose administration (case V), an exact solution (not closed form) and an approximation (closed form) were derived. Two sets of equations were derived for each of the five cases to take into consideration the possibility of a latency term ().Plots of drug amount X(or concentration C) vs. time (t) were constructed. In case I, a log Xvs. tplot was convex, the slope increasing with time. In case II, Xincreased,reached a peak, and decayed as in case I. In case III ( > 5In 2V/Q) Creached a preinduction steady state before decreasing to a lower (induced) steady state. When =0, Creached a maximum before decreasing to the same induced steady state. The behavior of Cvs. tfor cases IV and V was similar to that for case III. Determination of parameters was attempted in case III. Nonlinear least-square fitting of generated data with 3–9% error yielded reasonable estimates of k.This work was supported by NIH Research Contracts N0l-NS-1-2282 and N01-NS-6-2341.Parts I, II, III, IV, and VI of this series can be found in theJournal of Pharmaceutical Sciences.     

降纤酶低分子肝素治疗短暂性脑缺血发作的研究

目的 观察降纤酶与低分子肝素治疗短暂性脑缺血发作的效果及副作用。方法 选择本院神经内科住院患者36例应用降纤酶10U加入加入250ml生理盐水中静脉滴注，隔日1次，共3次；低分子肝素0．5ml脐旁皮下注射，12h 1次，连用7—10d,同时常规给予复方丹参滴注，口服尼莫地平，维生素E，维生素C，停用低分了肝素后给予肠溶阿斯匹林75mg，每日1次口服。结果 治疗开始后TLA发作相继减少，停止发作时间分别为1d内9例，3d内15例，5d内12例。随访6个月—1年，1例2个月后复发，重新应用上药治愈。结论 降纤酶与低分子肝素治疗TLA安全有效、无明显副作用、不易复发。     

亚硝基谷胱甘肽对大鼠肝微粒体谷胱甘肽转移酶的激活及机制

目的　探索一氧化氮供体亚硝基谷胱甘肽(GSNO)能否在体外通过S 亚硝酰化机制激活大鼠肝微粒体谷胱甘肽转移酶 (mGST)。方法　微粒体粗提物与GSNO体外共孵育 ,测定mGST催化动力学改变 ,结合N 乙基马来酰亚胺 (NEM )再激活实验和二巯基苏醇 (DTT)逆转实验 ,以及酶蛋白游离巯基和酶S 亚硝酰化蛋白的改变 ,研究酶的激活机制。结果　GSNO在 0 .12 5～ 2mmol·L- 1浓度范围内呈浓度和时间 (3～ 15min)依赖性地激活mGST ,NEM对酶的再激活效应消失 ,DTT可以逆转上述激活作用 ,同时酶蛋白游离巯基浓度依赖性减少 ,而S 亚硝酰化蛋白浓度依赖性增多。结论　GSNO体外可激活大鼠肝mGST ,激活机制可能与mGST第 4 9位半胱氨酸 (Cys4 9)的巯基被亚硝酰化形成S 亚硝基硫醇结构有关。