A preparative method for the enzymatic 5'-monophosphorylation of nucleosides

A simple procedure to prepare a stable preparation of bovine liver adenosine kinase was developed. The adenosine kinase was used to phosphorylate a variety of purine nucleosides using ATP or [gamma-32P]ATP as the phosphoryl donor. A convenient scheme to purify the nucleotides was also developed.     

Development of a model solid-state fermentation system

Summary A model solid substrate was developed, consisting of cassava starch and other nutrients embedded in kappa-carrageenan. A suitable fermentation system and analytical techniques were also developed to cope with the constraints imposed by solid-state fermentation.     

Framework for online optimization of recombinant protein expression in high-cell-density Escherichia coli cultures using GFP-fusion monitoring

A framework for the online optimization of protein induction using green fluorescent protein (GFP)-monitoring technology was developed for high-cell-density cultivation of Escherichia coli. A simple and unstructured mathematical model was developed that described well the dynamics of cloned chloramphenicol acetyltransferase (CAT) production in E. coli JM105 was developed. A sequential quadratic programming (SQP) optimization algorithm was used to estimate model parameter values and to solve optimal open-loop control problems for piecewise control of inducer feed rates that maximize productivity. The optimal inducer feeding profile for an arabinose induction system was different from that of an isopropyl-beta-D-thiogalactopyranoside (IPTG) induction system. Also, model-based online parameter estimation and online optimization algorithms were developed to determine optimal inducer feeding rates for eventual use of a feedback signal from a GFP fluorescence probe (direct product monitoring with 95-minute time delay). Because the numerical algorithms required minimal processing time, the potential for product-based and model-based online optimal control methodology can be realized.     

Refractory heart failure and age-related differences in adriamycin-induced myocardial changes in rats

Clinical use of adriamycin, an effective chemotherapeutic agent, has been restricted because of a demonstrated dose-limiting cardiotoxicity. To study age-related differences in adriamycin-induced cardiotoxicity, clinical status, developed force, ultrastructure, and lipid peroxide changes in the myocardium were investigated in two age groups of rats termed younger (Cy) and older (Co). Experimental animals (Cy + A; Co + A) received a cumulative dose of 15 mg/kg of adriamycin over 2 weeks. Animals in the Co + A group showed hydroperitoneum, higher mortality, and a greater decline in weight and feed consumption. Decline in base-line developed force in papillary muscles from Cy + A and Co + A group was not significant, but responsiveness to different interventions was attenuated. Papillary muscles from the Co + A group showed a significantly lesser increase over its control (Co) group in peak developed force in response to higher Ca2+. The decline in the peak developed force due to low Ca2+ and frequency increase was also significantly less in the Co + A group. Qualitatively similar but quantitatively less or even statistically insignificant changes were seen in the younger treated (Cy + A) group compared with its controls. A greater cell damage indicated by the loss of myofibrils, swelling of the mitochondria as well as the tubular system and accumulation of lipofuscin granules was seen in the Co + A group. While there was no significant change in the malondialdehyde content in the lipofuscin granules was seen in the Co + A group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynamic analysis of a microbial process: A systems engineering approach

A general mathematical model of the chemostat system is developed in order to define an experimental program of dynamic testing. A glucose-limited culture ofSaccharomyces cerevisiae was grown in a chemostat using chemically defined medium. The chemostat was perturbed from an initial steady state by changes in input glucose concentration, dilution rate, pH, and temperature. Dynamic responses of cell mass, glucose, cell number, RNA, and protein concentrations were measured. A number of simulation techniques were used in developing a dynamic mathematical model and in comparing the developed model with experimental data as well as the Monod model. The resulting model was found to be quantitatively accurate and superior to the Monod model. The developed model was interpreted in the light of cell physiology. Adjustment of intracellular RNA fraction was found to be rate limiting in acceleration of cell specific growth rate.     

Quantitative determination of anhydroleandomycin and glycol in oleandomycin phosphate]

A spectrophotometric method for determination of inactive admixtures in oleandomycin phosphate-anhydrooleandomycin was developed. The method was based on measuring optical density of methanol solutions of oleandomycin and its admixtures at a wave length of 235 nm of UV-spectrum. The minimun detectable amount was 0.5 per cent. A method of thin-layer chromatography in a buffered fixed layer was developed for determination of the 2nd inactive admixture, i.e. glycole. The chromatogrammes were developed with xanthydrol. The minimum detectable amount was 3 mug.     

Computerized Chronothermodynamic Breast Examinations Under Ambulatory Conditions

A new approach to the assessment of breast health was developed based upon continuous ambulatory recording of breast temperature for 24 hrs. A new data acquisition system consisting of a portable, multi-channel, microprocessor-based recorder worn on a belt was developed to record temperatures. Dramatic thermal fluctuations were noted during the recordings in response to multiple stimuli (physiological, behavioral, environmental). Specific algorithms and software programs were developed to process the thermobiologic and chronobiologic information. Data processing was fully automatic and included an examination report indicating a score and a chronothermodynamic class for each of the five areas examined on each breast. The preliminary clinical studies have concerned more than 450 patients with healthy or diseased breasts.     

Granulation and immobilisation of methanogenic and sulfate-reducing bacteria in high-rate anaerobic reactors

The formation of anaerobic granular sludge on a sulfate-containing waste-water was studied in up-flow anaerobic sludge blanket reactors. Three systems were examined: a sulfidogenic system, a methanogenic system and a mixed sulfidogenic/methanogenic system. No significant granulation was observed in the sulfidogenic system. For the methanogenic and the mixed methanogenic/sulfidogenic system granulation proceeded well, and no significant difference in the granule diameter could be detected. In the three systems studied, different types of sludge developed. A (mainly) methanogenic granular sludge was developed in the methanogenic system, a (more) sulfate-reducing granular sludge was developed in the mixed methanogenic/sulfidogenic system, and a flocculant sulfate-reducing sludge was developed in the sulfidogenic system. Correspondence to: A Visser     

Computerized Chronothermodynamic Breast Examinations Under Ambulatory Conditions

A new approach to the assessment of breast health was developed based upon continuous ambulatory recording of breast temperature for 24 hrs. A new data acquisition system consisting of a portable, multi-channel, microprocessor-based recorder worn on a belt was developed to record temperatures. Dramatic thermal fluctuations were noted during the recordings in response to multiple stimuli (physiological, behavioral, environmental). Specific algorithms and software programs were developed to process the thermobiologic and chronobiologic information. Data processing was fully automatic and included an examination report indicating a score and a chronothermodynamic class for each of the five areas examined on each breast. The preliminary clinical studies have concerned more than 450 patients with healthy or diseased breasts.     

Numerical simulation of flow field around a cow using 3-D body-fitted coordinate system

A 3-D theoretical model that characterizes the flow field around a single cow or multiple cows was developed. A general computer code that simulates the flow field that takes into account the shape of a cow and the boundary conditions of the enclosure was developed. A system of visualization of flow field that provides theoretical basis for the study of heat and mass transfer between a cow and its environment was developed. Qualitative and quantitative analyses were performed to determine the effect of cow orientation with respect to direction of airflow on convective heat exchange between the cow and its environment. The direction of air movement is not an important factor in influencing convective heat transfer between a cow and its environment.     

A simple technique for nonsurgical embryo transfer in mice

A simple method was developed for nonsurgical transfer of mouse embryos which enabled transfer to both uterine horns. Embryos were picked up in a modified capillary tube and transferred through the cervix into each uterine horn of an unanesthetized mouse. A glass speculum was used to facilitate location of the cervix. The technique was found to be as successful (up to 60% of embryos transferred developed to term) as surgical methods yet was simpler and eliminated surgical trauma to the recipient mouse.     

头部位移检测法动物悬尾实验仪的研制

采用红外传感、激光定位、51系列单片微机控制和记忆等技术,研制一种检测动物悬尾特性的仪器,用于药物研究中的动物悬尾实验。经30例小鼠实验证明,效果良好,该仪器为动物悬尾实验提供了一种全新的先进方法。     

耕作的数值模型及其应用

种子发芽与出苗主要受土壤水分与温度的影响。在研究非均质土壤起伏不平的地表与大气间水分能量交换过程的数值分析方法的基础上,发展了耕作对土壤水分与温度影响的基于有限元分析方法的数值模型。作为应用研究,讨论了不同的耕作方式如镇压,少耕与沟种垄作对土壤水分、温度的影响,并介绍了模型用于陕北的沟种设计。本项研究为农业生态学提供了一种基于模型－模拟－优化的定量分析方法。     

A novel technology coupling extraction and foam fractionation for separating the total saponins from Achyranthes bidentata

A novel technology coupling extraction and foam fractionation was developed for separating the total saponins from Achyranthes bidentata. In the developed technology, the powder of A. bidentata was loaded in a nylon filter cloth pocket with bore diameter of 180?µm. The pocket was fixed in the bulk liquid phase for continuously releasing saponins. Under the optimal conditions, the concentration and the extraction rate of the total saponins in the foamate by the developed technology were 73.5% and 416.2% higher than those by the traditional technology, respectively. The foamates obtained by the traditional technology and the developed technology were analyzed by ultraperformance liquid chromatography–mass spectrometry to determine their ingredients, and the results appeared that the developed technology exhibited a better performance for separating saponins than the traditional technology. The study is expected to develop a novel technology for cost effectively separating plant-derived materials with surface activity.     

A general high-performance liquid chromatography-based assay for the hydrolysis of N-acyl glutamates.

A rapid and simple HPLC assay has been developed to separate and quantify N-acyl glutamates and the corresponding carboxylic acids of the acyl moiety. This method was specifically developed to assay hydrolytic activity for glutamate carboxypeptidases. Although established assays for specific substrates of such enzymes exist, they may not be amenable for examining the hydrolytic activity of new substrate probes. This assay was developed to accommodate such probes.     

The protoplasts of Acremonium chrysogenum. Biochemical and morphological studies

A procedure for preparing stable A. chrysogenum protoplasts capable of 60 per cent regeneration was developed. Two morphogenetic types of the regeneration were detected. The variants isolated after the protoplast regeneration were characterized by wide ranges of morphological variation. Capacity for the antibiotic production varied from 60 to 160 per cent of the activity of the starting strain. A procedure for isolating functionally active mitochondria from protoplasts of A. chrysogenum was also developed. Their main bioenergetic parameters were studied. In the respiratory chain of the A. chrysogenum mitochondria there were detected three conjugation sites of oxidative phosphorylation.     

Production of asymmetric hybrids between Arabidopsis thaliana and Brassica napus utilizing an efficient protoplast culture system

Application of the protoplast culture method developed for Brassica protoplasts to protoplasts of Arabidopsis thaliana has increased the opportunities for interspecific hybridizations involving Arabidopsis. A more-efficient and much-simpler method was established compared to the earlier-reported protocol developed for A. thaliana protoplasts in which alginate beads were utilized. Mesophyll protoplasts of A. thaliana (ecotypes 'Landsberg erecta' and 'Wassilewskija') were cultured in the modified 8p liquid medium, which had been developed for Brassica protoplasts. For comparison, protoplasts were cultured in sodium alginate beads supplied with B5 medium according to the protocol for A. thaliana. The protoplasts divided with high frequencies in the 8p medium, and calli proliferated more rapidly than in the sodium alginate beads. High frequencies of shoot differentiation and regeneration were observed in calli of both ecotypes, from about 30% in the ecotype 'Wassilewskija' to about 60% for 'Landsberg erecta'. The more-rapidly the calli developed, the higher the regeneration frequencies were. Asymmetric hybrids between A. thaliana and Brassica napus were obtained by treating the protoplasts of A. thaliana with iodoacetamide (IOA) and B. napus protoplasts with UV-irradiation before fusion with polyethylene glycol (PEG). By using the culture procedure developed for Brassica protoplasts, calli developed and plants were regenerated. Although most of the plants regenerated after cell fusion were A. thaliana-like and were judged to be escapes from IOA treatment, more than ten plants showed hybrid features of both morphological and molecular characters. Among the hybrids that have flowered so far, both male-fertile and male-sterile plants have been obtained. Back-crossings to A. thaliana are now in progress as is morphological and molecular characterization of the plants.     

A specific complement-fixation test for human hepatitis a employing CR326 virus antigen. Diagnosis and epidemiology.

A specific diagnostic complement-fixation test for hepatitis A antibody in human serum was described employing livers of marmosets infected with CR326 strain human hepatitis A virus. Persons with hepatitis A, but not hepatitis B, developed hepatitis A CF antibody shortly after the onset of illness and this persisted thereafter. Good agreement was noted in the development of CF and neutralizing antibodies in hepatitis A cases. Hepatitis A was shown to occur in a person with hepatitis B antigenemia and hepatitis B occurred in persons with hepatitis A antibody. Most persons with hepatitis A who were tested, but none of those with hepatitis B, developed increased anticomplementary activity in their sera at the time of onset of illness. At least one patient with hepatitis A developed antibody against normal liver that persisted. The possible inplications of this in relation to pathogenesis and to non-specific diagnostic tests in hepatitis were discussed. A limited epidemiologic study of a family outbreak of hepatitis in Costa Rica and of a group of young adults in our epidemic country acquire their infections at an early age and are immune thereafter; persons in areas of relatively low incidence may proceed into adulthood without experience with hepatitis A. The CF test should provide an excellent tool for diagnosis and for epidemiologic investigation of hepatitis A and should be of considerable value to detect hepatitis A virus in attempts to propagate the virus in cell culture.     

A New Method for Quantitative Determination of Steroid Metabolites of Cytochrome P450-Dependent Reactions Using Fluorescent Spectroscopy

A method for determination of hydroxylase activity of cytochrome P450 3A4 (CYP3A4) towards its substrate hydrocortisone using fluorescent analysis of the product was developed. 6β-hydroxycortisol, formed during CYP3A4-dependent electrocatalysis, has a characteristic fluorescent peak at λ = 427 ± 2 nm after treating with the sulfuric acid : ethanol (3 : 1) mixture and excitation at λ = 365 nm, which is different from the substrate (hydrocortisone) fluorescence (λ = 525 ± 2 nm). The limit of detection of 6β-hydroxycortisol was 0.32 μM. The developed analytical approach was used to determine the kinetic parameters of CYP3A4-dependent hydrocortisone hydroxylation.     

Using situs for flexible and rigid-body fitting of multiresolution single-molecule data

We describe here a set of multiresolution visualization and docking procedures that we refer to as the Situs package. The package was developed to provide an efficient and robust method for the fitting of atomic structures into low-resolution data. The current release was optimized specifically for the visualization and docking of single molecules. A novel 3D graphics viewer, volslice3d, was developed for the package to provide an immersive virtual reality environment for measuring and rendering volumetric data sets. The precision of single-molecule, rigid-body docking was tested on simulated (noise-free) low-resolution density maps. For spatial resolutions near 20 A typically arising in electron microscopy image reconstructions, a docking precision on the order of 1 A can be achieved. The shape-matching score captured the correct solutions in all 10 trial cases and was sufficiently stringent to yield unique matches in 8 systems. Novel routines were developed for the flexible docking of atomic structures whose shape deviates from the corresponding low-resolution shape. Test calculations on isoforms of actin and lactoferrin demonstrate that the flexible docking faithfully reproduces conformational differences with a precision < 2 A if atomic structures are locally conserved.