bluuebunny/crossref_metadata_2025 huggingface dataset:
Multipoint attachment to a support protects enzyme from inactivation by organic solvents: α‐Chymotrypsin in aqueous solutions of alcohols and diols
Authors: Mozhaev Vadim V. , Sergeeva Mariya V. , Belova Alla B. , Khmelnitsky Yuri L.
Published: May 2005

Inactivation of α‐chymotrypsin in aqueous solutions of alcohols and diols proceeds both reversibly and irreversibly. Reversible loss of the specific enzyme activity results from conformational changes (unfolding) of the enzyme detected by fluorescence spectroscopy. Multipoint covalent attachment to the matrix of polyacryl‐amide gel by copolymerization method stabilizes α‐chymotrypsin from denaturation by alcohols, the stabilizing effect increasing with the number of bonds between the protein and the support. Immobilization protects the enzyme also from irreversible inactivation by organic solvents resulting from bimolecular aggregation and autolysis.
A mathematical model for metal affinity protein partitioning
Authors: Suh Sung‐Sup , Arnold Frances H.
Published: May 2005

A mathematical model of metal affinity partitioning has been derived and used to describe protein partitioning in Cu (II)PEG/dextran systems. A working model has been extended to account for inhibition, which for metal affinity extraction is the inhibition of protein‐metal binding by hydrogen ion. PEG/dextran partitioning experiments were performed on four proteins, tuna heart cytochrome c,Candida kruseicytochrome c, horse myoglobin, and sperm whale myoglobin. The partition coefficients for these proteins are increased by the addition of Cu (II)PEG‐IDA, due to the affinity between the chelated copper atom and metal‐coordinating histidine residues on the protein surface. The results of experiments to determine the effects of the number of binding sites on the protein, the copper concentration, and pH on partitioning are all well‐described by the mathematical model. The pKavalue of the metal binding site was determined to be 6.5, which is in the range of pKavalues commonly observed for surface histidines. The average association constant for the binding of Cu (II)PEG‐IDA to accessible histidines was found to be 4.5 × 103. This value is comparable to stability constants measured by conventional potentiometry techniques for analogous small complexes.
Solvent selection for biocatalysis in mainly organic systems: Predictions of effects on equilibrium position
Authors: Halling Peter J.
Published: May 2005

Predictions may be made for the influence of solvent choice on the equilibrium position of biocatalyzed reactions, based on data for the liquid–liquid distribution of the reactants. The most reliable predictions are probably for dilute systems, based on partition coefficients or correlations derived from them. The effective equilibrium constant for esterification reactions is predicted to alter by more than four orders of magnitude on changing between different water‐immiscible solvents. The equilibrium constant correlates well with the solubility of water in the solvent, and is most favorable for synthesis in the least polar solvents (aliphatic hydrocarbons). Similar effects seem to apply for other reactions, including oxidation of alcohols and hydrolysis of chlorides. Predictions can be made for nondilute systems using the UNIFAC system of group contributions, but the reliability of these is more questionable.
Dynamic on‐line optimization of a bioreactor
Authors: Petersen James N. , Whyatt Greg A.
Published: May 2005

An algorithm was developed which uses recursive least squares to identify a dynamic, discrete time model of a poorly defined system and uses both the dynamic and static portions of the identified model for on‐line optimization. To test this new algorithm, a model of an continuous biochemical reactor was used as the “process.” The objective, here, was to maximize ethanol production from the reactor by manipulating the feed rate to the reactor. The new algorithm, which uses dynamic information, was found to be superior to previously published algorithms which use only the steady‐state portion of the identified model.
A macromodel for outdoor algal mass production
Authors: Guterman H. , Vonshak A. , Ben‐Yaakov S.
Published: May 2005

A model describing growth of an outdoor algal (Spir‐ulina platensis)culture was developed. The model can simulate biomass production, pH, growth rate, oxygen evolution, and CO2 fixation rate. It was calibrated and validated against experimental data obtained by a novel automatic data logger/controller instrumentation which can number most vital parameters of the culture including on line estimation of oxygen production rate (OPR). The importance of understanding light distribution through the pond and its effects on the photosynthesis and respiration processes are emphasized. A maximum yield of about 38g day−1 m−2 under optimal conditions is predicted. The present model can also be a useful tool for optimization of algal mass production sites.
Biosurfactant yields and nutrient consumption of Pseudomonas fluorescens 378 studied in a microcomputer controlled multifermentation system
Authors: Persson A. , Molin G. , Andersson N. , Sjöholm J.
Published: May 2005

Production of biosurfactant AP‐6 and consumption of carbon (succinic acid) and nitrogen (ammonium ions) by Pseudomonas fluorescens 378 were studied under different growth conditions. The study was performed in a microcomputer controlled multibatch fermentation system which enabled simultaneous running of 10 fermentors. The fermentors were mantled glass vessels, temperature controlled by circulated water, and mixing was arranged by magnetic stirrers. They were connected to the computer system (pH measurement and control) via signal conditioning cards. The microcomputer had a 128 kbytes RAM, two 800‐kbyte floppy disc drives, a graphic terminal, and expansion cards. Biosurfactant production was independent of the carbon‐to‐nitrogen ratio and the phosphorus content in the medium. Omitting the Fe(III) supplement to the medium increased the product yield by 120%. Changes in oxygen transfer rate and pH in the iron deficient cultures did not have any effect on the product yield. Iron deficiency increased the cell consumption of carbon source. Consumption of carbon source in relation to nitrogen uptake (carbon/nitrogen quotient) increased with increasing quotient in the growth medium. The uptake of carbon and nitrogen changed in the intervals of 1.2–1.5 g/g biomass and 0.09–0.16 g/g biomass, respectively. The consumption of carbon increased from 1.5 g/g biomass to 2.0 g/g biomass when the medium concentration of phosphorus was decreased from 0.18 to 0.027 g/L.
Enzymatic synthesis of alkyds
Authors: Geresh Shimona , Giiboa Yigal
Published: May 2005

Lipases were used as catalysts in the synthesis of “all‐trans” polyester oligomers in organic solvents. Esters of fumaric acid and 1,4‐butane diol served as the substrates in the enzyme‐catalyzed polytransesterification. No isomerization of the double bond was found under the mild conditions of enzymatic catalysis used by us, as opposed to the extensive isomerization found during chemical polycondensation. The alkoxy leaving group of the ester fumarate was found to be responsible for the rate of transesterification. Low (Mw ≃ 600–800) and high (Mw = 1250) molecular weight alkyds were synthesized depending on whether tetrahedrofuran or acetonitrile, respectively, was used as the solvent.
Dynamic model development for a continuous culture of Saccharomyces cerevisiae
Authors: O'Neil D. G. , Lyberatos G.
Published: May 2005

The problem of dynamically modeling a chemostat is addressed. Using the results of continuous culture experiments for the growth of a strain of Saccharomyces cerevisiae on a glucose‐limited medium, a general approach to developing dynamic models is discussed. The approach to develop and verify the model involves three different types of experiments: steady‐state, dynamic step response, and feedback identification.
Adsorption of cellulase from Trichoderma reesei on cellulose and lignacious residue in wood pretreated by dilute sulfuric acid with explosive decompression
Authors: Ooshima Hiroshi , Burns Douglas S. , Converse Alvin O.
Published: May 2005

The adsorption of cellulase on cellulose and a lignacious residue was examined by using cellulase from Trichoderma reesei, hardwood pretreated by dilute sulfuric acid under high pressure, and a lignacious residue prepared by a complete enzymatic hydrolysis of the pretreated wood. A significant amount of cellulase was found to adsorb on the lignacious residue during the hydrolysis of the pretreated wood. Hence, the adsorption of enzyme on the lignacious residue as well as cellulose must be taken into account in the development of the hydrolysis kinetics. It was found that the adsorption of enzyme on cellulose and on the lignacious residue could be represented by Langmuir type isotherms. The data show that the pretreatment at a higher temperature results in more enzyme adsorption on the cellulose fraction and less on the lignacious residue fraction. The relationship between the hydrolysis rate and the amount of enzyme adsorbed is discussed.
A stochastic model to simulate the growth of anchorage dependent cells on flat surfaces
Authors: Lim J. H. R. , Davies G. A.
Published: May 2005

A stochastic model is proposed to simulate the growth of anchorage dependent cells on a flat surface. The model, based on representing the cell shapes on the surface as external irregular polygons with the nuclei distributed as a set of Poisson points (producing a modified Voronoi tessellation of 2 space) and incorporating a distribution function to describe cell division of the perimeter cells of the colony, provides data not only on population dynamics but also on the patterns produced by clusters of cells in the colony. These patterns produced by the model are qualitatively similar to observations reported for some cell cultures. The periods of induction, rapid growth, and decreasing growth asymptoting to zero as confluence is reached are predicted by the model. Quantitative comparison with published experimental data for this is good. The specific growth rate computed for the period of rapid growth predicted by the model is dependent on the distribution function describing the cell division time. As the standard deviation of this increases, the specific growth rate decreases as with a consequent increase in time to achieve confluence. The removal of cells from the colony by shear forces or death is considered in the model. As the probability for removal increases, the cell density at confluence and specific growth rate decrease. The clusters of cells, patterns, in the colony are very sensitive to cell removal. By analyzing these patterns in experiments, an estimate of cell removal can be made. The areas covered by cells on a substrate are fractal patterns. The fractal dimension is always greater than 1 and is a function of the removal probability.
Continuous production of prourokinase in feed harvest and perfusion cultures
Authors: Wagner A. , Marc A. , Engasser J. M. , Villermaux S. , Einsele A.
Published: May 2005

The production of prourokinase (PUK) by a human kidney tumor cell line is studied in long term cultures. Cells are grown on microcarriers which are retained inside the reactor by sedimentation or with a spin filter. Two modes of operation are compared: feed harvest at an average medium exchange rate of 0.3 d−1 and continuous perfusion at a higher dilution rate of 1.5 d−1. In the two systems a stable production of PUK has been maintained for more than 400 h. Kinetics of cellular growth, nutrient consumption, and metabolite and PUK excretion are similar. After an initial rapid growth period, one observes a 10‐fold reduction in all the cellular metabolic activities during the stationary phase. Continuous perfusion yields a higher cell density (7 × 106 cells·mL−1) than feed harvest (3 × 106 cells·mL−1), which results in a twofold increase in the reactor productivity. But higher final enzyme activities, about 250 ru·mL−1, are obtained in the feed harvest recovered medium than in the perfusion medium, 100–150 ru·mL−1. The cumulative medium consumption per mass of product is the same in the repeated batch and in the continuous operation mode.
Effects of cell density and glucose and glutamine levels on the respiration rates of hybridoma cells
Authors: Wohlpart Dave , Kirwan Donald , Gainer John
Published: May 2005

The effects of cell density as well as the concentration levels of glucose and glutamine on the specific respiration rate of a hybridoma cell line were investigated. The experimental oxygen consumption rate was found to be constant over a wide range of dissolved oxygen levels if the suspension medium contained glutamine. In glutamine‐free medium, however, the rate of oxygen consumption decreased slowly with time.

In a stationary flask batch culture, the specific respiration rate decreased from about 7 to 2.9 μmol/min per 109 cells as the cell density increased exponentially from 1 × 105 to 1.2 × 106/mL. To isolate the effect of cell density, cells were re suspended in fresh culture medium so that nutrient concentrations were the same for all experiments. The specific respiration rate decreased with increasing cell density in the same manner as in the stationary flask culture, falling from 8 to 4 μmol/min per 109 cells as the cell density increased from 105 to 106 cells/mL, then declining to 2 μmol/min per 109 cells when the cell density reached 107 cells/mL.

Cells suspended in Hanks balanced sale solution (HBSS) were used to elucidate the effect of glucose and glutamine levels on respiration. The addition of glucose in concentrations of 0.25, 0.50, and 0.75 g/L had no observable effect on the specific oxygen uptake rate; however, a glucose concentration of 1 g/L reduced the uptake rate by 22%. Glutamine in a concentration of 0.30 g/L increased the specific respiration rate in HBSS containing 0 and 1 g/L glucose by approximately 13%.
Enzymatic production of hydrogen peroxide and acetaldehyde in a pressure reactor
Authors: Nelles L. P. , Arnold J. A. , Willman D. S.
Published: May 2005

Alcohol oxidase, an enzyme which exhibits relatively weak substrate specificity among short chain alcohols, forms the corresponding aldehyde and hydrogen peroxide as coproduct. The ability of alcohol oxidase from Pichia pastoris yeast to convert ethanol to acetaldehyde and hydrogen peroxide was examined in an oxygen pressure reactor under conditions, such that oxygen availability was sufficient to permit rapid catalysis. Hydrogen peroxide levels of ∼1.8/M (6% w/w) were attained in 2–3 h with 2.8 μM enzyme, corresponding to a productivity of ∼30 g peroxide/g enzyme. Optimal conditions (within equipment limitations) were 900 psi oxygen, 2.6M ethanol, at 4 °C. Similar levels of products were reached in the reactor using enzyme immobilized covalently on controlled pore glass and noncovalently on an anion exchange support. Recycle of covalently immobilized enzyme was not possible as a result of enzyme inactivation after a single run. Limited recycle of noncovalently immobilized enzyme was accomplished with substantial decreases in levels of product attainable on each cycle.
Hydrophobic interaction chromatography of autoproteolysis products of human seminal plasma
Authors: Mazzini M. N. , Di Giacomo M. A. , Cerezo A. S.
Published: May 2005

Hydrophobic interaction chromatography was employed for the fractionation of the glycopeptide mixture obtained after autoproteolysis and delipidization treatment of normal human seminal plasma samples. This sequence of procedures led to the elimination of highly hydrophobic components and to the concentration of the carbohydrate‐rich fractions. A partial separation between the O‐ and N‐glycosidic‐linked oligopeptides was also achieved after the chromatographic step.
HPLC determination of polyamines in the femtomole range
Authors: Yun Zhong , Zhang Ren‐en
Published: May 2005

A new method for the chromatographic determination of polyamines is presented. The procedure consists of the reaction of 9‐fluorenylmethyl chloroformate (FMOC‐Cl) with polyamines at 50 ± 1°C for 10 min, followed by stepwise elution chromatography. The reaction is simple and the derivatives are stable. All the polyamine‐FMOC derivatives can be separated within 13 min. The linearity of this method is satisfactory in the range of 0.5–100 pmol. The detection limits for putrescine, cadaverine, spermidine and spermine are 83.3 fmol, 109 fmol, 25.5 fmol and 22.7 fmol respectively.
A rapid method for the determination of benzylpenicillin in serum by reversed‐phase HPLC
Authors: Soto‐Otero R. , Mendez‐Alvarez E. , Sierra‐Paredes G. , Galan‐Valiente J. , Aguilar‐Veiga E. , Sierra‐Marcuño G.
Published: May 2005

A simple procedure for the determination of benzylpenicillin in serum is described. The assay involves the extraction of the drug and the internal standard (phenoxymethylpenicillinic acid) from the sample into dichloromethane, using tetrabutylammonium hydrogen sulfate neutralized with NaOH and buffered with citrate as an ion‐pairing reagent. RP–HPLC was performed on a Spherisorb 5 ODS column, eluting the drugs isocratically with 14% acetonitrile in 10 mM ammonium acetate buffer. Monitoring was by UV detection at 208 nm. Our results show that the method is accurate and reproducible, permitting quantification of serum levels of benzylpenicillin without interference from other drugs commonly used in therapy. Analytical recovery was greater than 79.2%.
A high performance liquid chromatographic post‐column fluorescent ion pair extraction system: Application to physostigmine and its metabolite eseroline in human serum
Authors: Quinn Karen D. , Stewart James T.
Published: May 2005

A high performance liquid chromatographic post‐column fluorescent ion pair extraction system was developed for the analysis of quaternary ammonium and amine drugs in serum. A new fluorescent ion pair reagent, sodium α‐(3,4‐dimethoxyphenyl) cinnamonitrile‐2′‐sulfonate (DPS), was synthesized and characterized. The post‐column extraction system consisted of a three‐dimensional knitted teflon mixing coil and a membrane phase separator which was modified from an original literature design. Physostigmine and its metabolite eseroline were used as model cations. A solid phase extraction procedure using octadecylsilane columns was developed to extract the compounds and neostigmine bromide (internal standard) from human serum. The compounds were chromatographed on a diol column using a 80:20 aqueous phosphate buffer pH 4 absolute methanol mobile phase at a flow rate of 1 mL/min. Methylene chloride was used as the on‐line extraction solvent for the DPS ion pairs formed. Fluorescence of the extracted ion pairs was measured using an excitation of 243 nm and an emission cut‐off filter at 418 nm. Linearity was in the 2–100 ng/mL and 5–100 ng/mL ranges for physostigmine and eseroline, respectively. Detection limits based on a signal‐to‐noise ratio of 2, were 2 and 5 ng/mL, respectively. Precision of the method was found to be in the 1.5–3% range and percentage error in the 1.5–7% range for both compounds.
Liquid chromatographic analysis of mexiletine in serum, with alternate application to tocainide, procainamide, and N‐acetylprocainamide
Authors: vasBinder Eric , Annesley Thomas
Published: May 2005

A simple and precise high performance liquid chromatographic method for the determination of mexiletine in human serum or plasma is described. Following addition of N‐propionylprocainamide as internal standard the specimens are extracted, under basic conditions, into methylene chloride. After removal of the aqueous layer the drug is back‐extracted into dilute acid, which is then injected directly for analysis. The extraction efficiency is 79% for both mexiletine and internal standard, and the assay is linear to 4 mg/L (twice upper therapeutic concentration). Inter‐run coefficients of variation are 3.0% or less. The relative retention time of mexiletine to internal standard averages 1.3. An adaptation of this method is described for an alternate application to the analysis of tocainide, procainamide and N‐acetylprocainamide.
A high performance liquid chromatographic procedure for the simultaneous determination of norfloxacin and furprofen in rat plasma
Authors: Carlucci Giuseppe , Mazzeo Pietro , Palumbo Giancarlo
Published: May 2005

A rapid and simple high performance liquid chromatographic analytical method is described for the simultaneous determination of norfloxacin and furprofen in rat plasma. Following dichloromethane extraction, the solution was chromatographed in a Vydac anion exchange column using a mobile phase of 0.05 M phosphate buffer (pH=7.0):acetonitrile (80:20, v/v) at a flow‐rate of 1.8 mL/min. The drugs were detected by UV absorption at 278 nm. The total chromatographic analysis time was 10 min. The response was linear, 0.1‐5.0 μg/mL for norfloxacin and 0.1‐3.0 μg/mL for furprofen, respectively. This method is useful for pharmacokinetic studies of these compounds and will facilitate detailed investigations into the interactions between quinolones and furprofen.
Development of a chemiluminescence detection system using bis[4‐nitro‐2‐(3,6,9‐trioxadecyloxycarbonyl)phenyl] oxalate for the sensitive determination of the fluorescent compounds separated with an acidic mobile phase
Authors: Sugiura Masaki , Kanda Susumu , Imai Kazuhiro
Published: May 2005

A rotating flow mixing device having three directional inlets was developed for a high performance liquid chromatographic peroxyoxalate chemiluminescence detection system for the sensitive detection of fluorescent compounds separated with acidic mobile phases, such as those containing trifluoroacetic acid. The diameters of the three inlets were 0.3, 0.3 and 0.5 mm i.d., for an acidic column eluate, chemiluminogenic reagents (bis[4‐nitro‐2‐(3,6,9‐trioxadecyloxycarbonyl)phenyl] oxalate and hydrogen peroxide) and imidazole solution (for neutralization and as a catalyst), respectively. The three mixed solutions flowed out through an outlet (0.5 mm i.d.) at the centre of the top of the vessel. The reaction conditions to give the appropriate signal‐to‐noise ratio (S/N) were investigated using dipyridamole (an antiplatelet aggregation drug) as a typical fluorescent compound. The system was applied to the detection of dipyridamole and dansylated amino acids with a mobile phase containing trifluoroacetic acid. The detection limits for dipyridamole and each dansylated amino acid were 10 amol and sub‐fmol (S/N=2), respectively.
Sensitive determination of enantiomers of amino acids derivatized with the fluorogenic reagent, 4‐fluoro‐7‐nitro‐2,1,3‐benzoxadiazole, separated on a pirkle‐type column, sumichiral OA 2500(S)
Authors: Imai Kazuhiro , Fukushima Takeshi , Uzu Sonoko
Published: May 2005

Some L‐ and D‐amino acids (Phe or Leu) were derivatized with a fluorogenic reagent, 4‐fluoro‐7‐nitro‐2,1,3‐benzoxadiazole (NBD‐F) and separated on a Pirkle‐type column, Sumichiral OA 2500(S) [(S)‐1‐naphthylglycyl‐3,5‐dinitrophenylamide silica gel] with a mobile phase of 20 mM ammonium acetate in methanol. The fluorometric detection was made at 530 nm with excitation at 470 nm. No racemization of the enantiomers occurred during the derivatization reaction. The separation factors (α) for NBD‐L‐Phe and NBD‐D‐Phe, and NBD‐L‐Leu and NBD‐D‐Leu, were 1.27 and 1.17, respectively. The detection limits were in the range of ca. 30 fmol.
Structure and ligand‐binding modes of human serum albumin studied by UV resonance raman spectroscopy
Authors: Hashimoto Shinji , Yabusaki Takahiko , Takeuchi Hideo , Harada Issei
Published: May 2005

Ultraviolet resonance Raman spectra have been measured for human serum albumin (HSA) and its complexes with three types of ligands. Tyr and Trp Raman bands of ligand‐free HSA are strongly enhanced with 240‐nm excitation compared with those of aqueous amino acids, indicating that the side chains of some Tyr residues and the unique Trp at position 214 are hydrogen bonded in hydrophobic environments. The high intensity of Tyr and Trp Raman scattering remains unchanged in a wide pH range of 3.5–9.0 and the microenvironments of the Tyr and Trp residues seem conserved in this pH range. The Tyr Raman intensity of HSA increases upon binding of palmitic acid, indicating hydrophobic interactions between the Tyr phenol ring and the aliphatic chain of the bound fatty acid. Binding of warfarin, on the other hand, does not affect the Tyr Raman intensity. Instead, an increase is observed for the Raman intensity of Trp‐214, which is located at the opening of the warfarin binding site. Concomitantly a conformation‐marker Raman band of Trp‐214 changes in frequency and the warfarin Raman intensity increases, suggesting direct hydrophobic interactions between Trp‐214 and warfarin. Complexation of ibuprofen and HSA increases the Raman intensity of Tyr but not of ibuprofen, which is ascribed to the formation of a hydrogen bond between the carboxylate of ibuprofen and the phenolic OH of Tyr‐411, the unique Tyr residue at the ibuprofen binding site. Hydrophobic interactions and hydrogen bonding of Tyr and Trp residues play a key role in ligand binding of HSA. © 1995 John Wiley & Sons, Inc.
Localized electron polarization in a substrate analog binding to the active site of enoyl‐CoA hydratase: Raman spectroscopic and conformational analyses of rotamers of hexadienoyl thiolesters
Authors: Tonge P. J. , Anderson V. E. , Fausto R. , Kim M. , Pusztai‐Carey M. , Carey P. R.
Published: May 2005

The Raman bands associated with different rotamers of hexadienoyl ethyl thiolester, CH3CHCHCHCHC(O)SCH2CH3, have been partially assigned using recent conformational and vibrational spectroscopic analyses of ethyl thiocrotonate CH3CHCHC(O)SCH2CH3.1 In particular, rotational isomers involving the CHC(O) and SCH2 axes have been characterized. The assignment of the vibrational modes of different conformers of hexadienoyl ethyl thiolester was further facilitated by variable‐temperature (+20°C to −90°C) Raman studies on the neat thiolester. High‐quality Raman spectra of hexadienoyl‐coenzyme A bound to the enzyme enoyl‐CoA hydratase were obtained using 647.1‐nm excitation and Raman difference spectroscopy. The findings provided by analysis of the ethyl thiolester model compound enable us to conclude that conformational selection occurs for the CoA analog upon binding, resulting in the presence of a single‐rotamer population in the hexadienoyl moiety on the enzyme. The hexadienoyl‐CoA was labeled with 18O in the CO group and, separately, with 13C at the C2 position. A comparison of Raman data for the free and bound ligands, isotopically labeled and unlabeled, indicates that strong π‐electron polarization occurs in only a part of the hexadienoyl chain, viz, in the CCCO fragment, upon binding. The polarization gives rise to important contributions from canonical (resonance) forms of the type C+CCO−. In contrast, the C4C5 linkage (where the carbon atom numbering is C6C5C4C3C2C1O) seems little perturbed in the bound ligand. The causes and mechanistic advantage of the observed localized polarization are discussed. © 1995 John Wiley & Sons, Inc.
Modeling the heme vibrational spectrum: Normal‐mode analysis of nickel (II) etioporphyrin‐I from resonance raman, ft‐raman, and infrared spectra of multiple isotopomers
Authors: Hu Songzhou , Mukherjee Arka , Piffat Christine , Mak Ronald S. W. , Li Xiao‐Yuan , Spiro Thomas G.
Published: May 2005

Nearly complete vibrational assignments have been obtained for a heme model, nickel etioporphyrin‐I (NiEPI), using variable‐wavelength resonance Raman (RR), and FT‐Raman (FT‐R), as well as infrared (IR) spectroscopy, on a series of isotopomers labeled at positions in the skeleton (15N, β‐13C, meso‐d4, 15N‐meso‐d4) and in the peripheral substituents (methyl‐d12, ethyl‐d8, and ethyl‐d12). The vibrational bands are assigned to the porphyrin skeletal and substituent modes on the basis of the mode description scheme developed for nickel octaethylporphyrin (NiOEP) with the aid of a normal‐mode analysis of NiEPI, explicitly including the peripheral substituents, i.e., the methyl and ethyl groups. The previously reported NiOEP force field was refined to account for the observed isotope shifts of NiEPI isotopomers. An important result is the requirement of relatively large, long‐range force constants for methine bridge bonds on opposite sides of the porphyrin ring. These 1–8 and 1–9 interaction force constants are required to reproduce the frequencies and isotope shifts of six Cα‐Cm stretching modes and especially to predict the relative order of the two highest‐frequency Eu modes, v(Cα‐Cm) (v38, ∼ 1570 cm−1) and v(Cβ‐Cβ) (v37, ∼ 1600 cm−1). Most of the substituent (methyl and ethyl) vibrations are located in the RR and IR spectra. Strong RR enhancement of some substituent modes can be attributed to hyperconjugative interaction of the aliphatic groups with the porphyrin a1u orbital, as well as vibrational mixing of substituent modes with the nearby skeletal modes. © 1995 John Wiley & Sons, Inc.
Fluorescence line narrowing study of the ground‐state and first‐excited‐state vibrational frequencies of Sn and Zn cytochromes c
Authors: Laberge Monique , Vanderkooi Jane M.
Published: May 2005

Fluorescence‐line‐narrowing (FLN) spectra are presented for Sn cytochrome c (Snc) obtained under Q0‐0 and Q0‐1 excitation and for Zn cytochrome c (Znc) under Q0‐1 excitation. Vibrational frequencies of the ground and first excited states are reported for Snc. Additional excited‐state Znc frequencies are also presented, thus contributing to a previous FLN Znc investigation. The spectra of Snc shows more phonon broadening than for Znc, but vibrational frequencies of the S0 and S1 states could be obtained and the distribution function of the 0, 0 transition was determined. Snc and Znc showed good agreement in their excited‐state vibrational frequencies. Some S1 vibrational frequencies of Znc are seen to downshift with respect to ground‐state vibrations, suggesting that the porphyrin S0 to S1 expansion observed in other porphyrin systems also occurs in cytochrome c derivatives. © 1995 John Wiley & Sons, Inc.
Plasma ornithine carbamyl transferase level as an indicator of ischaemic injury of rat liver
Authors: Frederiks Wilma M. , Vogels Ilse M. C. , Fronik Gerard M.
Published: May 2005

The activity of ornithine carbamyl transferase (OCT) and glutamate pyruvate transaminase (GPT) in serum has been correlated with the extent of necrosis 24 h after different periods of ischaemia in rat liver. The extent of necrosis has been quantified as the volume density of necrosis in the total ischaemic liver lobes using tetranitro BT. The GPT‐activity in serum is maximal between 1 and 5 h after different periods of ischaemia, whereas OCT reaches its maximum between 5 and 12 h after ischaemia. The total amount of leaked enzyme‐activity as well as the peak value give a linear correlation with the extent of necrosis for OCT and GPT. There is a differnece between the character of these two enzymes in that a small leakage of GPT does not indicate liver cell necrosis later on. However, the appearance of OCT in the blood, an enzyme localized in the mitochondrial matrix, has a predictive value for the extent of necrosis, likely to occur later on. GPT, an enzyme from the cytoplasm, can also occur in the blood during the reversible stage of liver cell damage.
Quantitative and qualitative changes of plasma membrane glycoproteins in the early period of liver regeneration
Authors: Enrich C. , Bachs O. , Rius E. , Serratosa J. , Domingo J.
Published: May 2005

Alterations of cell surface glycoconjugates have been observed in many developing systems and may be important in the physiological control of growth and differentiation. Liver regeneration after partial hepatectomy is a suitable model in which to study the regulatory mechanisms of cell proliferation in vivo. We have isolated the sinusoidal plasma membrane of hepatocytes at different times after partial hepatectomy. The sialic acid content and the SDS‐polyacrylamide gel electrophoresis pattern of glycoproteins were determined. A decrease of periodic acid‐Schiff‐profiles, a change in the binding capacities of 125I‐concanavalin A, a reduction of the sialic acid content and the appearance and disappearance of specific components have been observed during the pre‐replicative phase of liver regeneration. These findings during this early period are consistent with the active involvement of the plasma membrane glycoproteins in the transition of cells to the proliferative state.
Sulfur‐containing amino acids that increase renal glutathione protect the kidney against papillary necrosis induced by 2‐bromoethylamine
Authors: Thielemann L. E. , Oberhauser E. W. , Rosenblut G. , Videla L. A. , Valenzuela A.
Published: May 2005

Papillary necrosis was observed in the kidneys of rats, 72 h after receiving a single injection of bromoethylamine (BEA). This effect was associated with renal glutathione (GSH) depletion 1 h after the administration of BEA. Stimulation of renal GSH synthesis by pretreatment of the animals either with glutamine + glucine + cystine or N‐acetyl‐L‐cysteine was attempted. Low doses of these precursors administered previously to BEA, respectively, decreased or abolished the GSH depletion. Nevertheless, both pretreatments failed to modify the magnitude of renal papillary necrosis. High doses of these precursors did not modify the BEA‐induced GSH depletion, but they significantly increased GSH levels 24 h after BEA administration. At this time, although a smaller intensity of renal papillary necrosis was observed with the amino acid mixture pretreatment, N‐acetyl‐L‐cysteine pretreated rats showed no papillary necrosis. It is suggested that the observed protective effects against BEA‐induced renal papillary injury may be ascribed in some measure, to a mechanism independent of GSH.
Lack of inhibition of vasopressin‐stimulated Na+K+ATPase by atrial natriuretic factor in the rat renal medullary thick ascending limb of Henle's loop
Authors: Charlton Judith A. , Baylis Peter H.
Published: May 2005

The anti‐diuretic hormone, arginine vasopressin (AVP) stimulates the activity of Na+K+ATPase in the rat renal medullary thick ascending limb of Henle's loop (mTAL). Atrial natriuretic factor (ANF) has been suggested to exert a tubular effect on the mammalian nephron, perhaps in part, by interacting with other hormones. In the present study, we investigated the effect of rat ANF with and without AVP upon mTAL Na+K+ATPase activity using cytochemical methods. ANF alone failed to inhibit or stimulate Na+K+ATPase activity in mTAL at any of the concentrations tested (10 nmol–0·1 pmol l−1). Unlike the rat hypothalamic digitalis‐like factor, ANF (10 nmol–10 fmol l−1) did not inhibit Na+K+ATPase activity after stimulation with AVP (l fmol l−1) for either 4 or 10 min. The results suggest that ANF does not exert an effect on mTAL, either alone or in conjunction with AVP.
Studies of the regulation of renal gluconeogenesis in normal and Pi depleted proximal tubule cells
Authors: Nakagawa Takeshi , Butterworth Peter J.
Published: May 2005

Pi depletion of proximal tubule cells isolated from mouse kidney results in a decrease in the cell content of fructose‐2,6‐bisphosphate and an increase in the rate of gluconeogenesis from pyruvate, malate and succinate. Gluconeogenesis from glycerol is unaffected by Pi depletion. Introduction of fructose‐2,6‐bisphosphate into the cytosol of ATP‐permeabilized cells is accompanied by a fall in gluconeogenesis.

The presence of external Ca2+ stimulates gluconeogenesis. When cytosolic Ca2+ is raised to 1·8 μM by permeabilization, the resealed cells still require 2·5 mM Ca2+ in the bathing medium in order to perform gluconeogenesis at the maximum rate. Cells permeabilized in the presence of cAMP show a decreased rate of glucose production. Phorbol ester stimulates gluconeogenesis provided that the phorbol treatment is performed in the absence of Ca2+ ions.

It is suggested that Pi depletion may stimulate pyruvate carboxylase activity and facilitate the entry of certain gluconeogenic substrates into mitochondria. It is also proposed that important aspects of the control of renal gluconeogenesis by parathyroid hormone are mediated by protein kinase C.
Evidence of the receptor nature of the binding sites induced in tetrahymena by insulin treatment. A quantitative cytofluorimetric technique for the study of binding kinetics
Authors: Kovács P. , Csaba G.
Published: May 2005

Tetrahymena pyriformis GL cells pretreated (imprinted) and not pretreated with insulin showed dissimilar quantitative relations of FITC‐insulin binding. Displacement of FITC‐insulin by unlabelled insulin was considerably less in the control than in the imprinted series. The curve for saturation of the binding sites with FITC‐insulin resembled a true saturation curve. The imprinted cells bound considerably more hormone in a shorter time than the control cells at identical levels of exposure. The dissociation of bound hormone from the imprinted cells increased over the control at 23°C, and to a still greater degree at 4°C. The effect of the pH of the medium on the dissociation of bound FITC‐insulin also differed between the imprinted and not imprinted cells. Thus the proposed cytofluorimetric assay of binding kinetics demonstrated the actual conditions of receptor activity, and indicated that the induced insulin binding sites of Tetrahymena behaved similarly to ‘classical’ receptors.
Temporal adaptation of human neutrophil metabolic responsiveness to the peptide formylmethionyl‐leucyl phenylalanine: A comparison between human neutrophils and granule‐depleted neutrophil cytoplasts
Authors: Dahlgren Claes
Published: May 2005

When polymorphonuclear leukocytes (neutrophils) and soluble or particulate matter interact, the cells produce superoxide anions (O2−) and hydrogen peroxide (H2O2). The chemotactic peptide formylmethionyl‐leucylphenylalanine (FMLP) induced a very weak response in normal neutrophils. The cellular response was changed, however, as a result of in vitro aging of the cells, i.e. the magnitude of the response was increased following storage of the cells at 22°C for up to 120 min, in the absence of any stimulus, and before the addition of the peptide. When phorbol myristate acetate was used as a stimulus, there was a pronounced production of O2− and H2O2, but no change in magnitude as a result of in vitro aging. When neutrophil cytoplasts (granule‐free vesicles of cytoplasm enclosed by plasmalemma) were exposed to the peptide FMLP of PMA, the vesicles produced both O2− and H2O2. There was, however, no increase in oxidative metabolite production in cytoplasts as a result of in vitro aging when either FMLP or PMA was used as a stimulus. The results thus indidate that mere incubation at room temperature primed the cells to increase their production of oxidative metabolites as a result of spontaneous exposure of hidden receptors. The fact that no such effects were observed with cytoplasts indicates that spontaneous receptor recruitment is a granule‐dependent process.
The inhibitory effect of neonatal rat smooth muscle cells and elastic extracellular matrix on development of the newly seeded cell population in culture
Authors: Hausmann J. , Horáčková J. , Krausová J. , Verbitzki V.
Published: May 2005

Neonatal smooth muscle cells were seeded in standard plastic Falcon flasks, on top of another 2‐month‐old culture of the same cell population or on top of an acellular matrix prepared by removal of these cells. The effect of both complete and acellular layers on the production of elastin, collagen and total extracellular matrix (EM) proteins as well as on cell division was measured. Compared with the standard population grown on plastic, the complete cell layer almost completely prevented the newly seeded cells from dividing. The acellular matrix did not affect cell doubling but caused a distinct decrease in the production of EM components.
Activation of phospholipase A2 by 1,25 (OH)2 vitamin D3 and cell growth in monocytic U937 and mono mac 6 cells
Authors: Aepfelbacher Franz C. , Weber Peter C. , Aepfelbacher Martin
Published: May 2005

Soluble phospholipase A2 activity was characterized in two human monocytic cell lines, U937 and Mono Mac 6. The enzyme showed an absolute requirement for Ca++, an alkaline pH optimum and Michaelis‐Menten kinetics in both cell lines. Differentiation of U937 and Mono Mac 6 cells with 1,25 (OH)2 vitamin D3 (10 nM, 72 h) enhanced PLA2 activity by 82 per cent and 56 per cent, respectively. Furthermore, kinetic experiments revealed that enzyme activity increased within 3 h when cells were brought from the nonproliferative phase of growth to the start of a new cycle of cell proliferation. This initial activation of PLA2 could be inhibited by cycloheximide and actinomycin D, indicating the requirement of gene transcription. Taken together, these results suggest a role of cytosolic, Ca++‐dependent PLA2 in differentiation and growth of monocytic cells.
Guinea pig peritoneal macrophages. Differential effects of lectins on interaction with IgG immunoglobulins
Authors: Sokal Izabela , Kułacz Agata , Gorczyca Wojciech , Janusz Maria , Lisowski Jozef
Published: May 2005

Guinea pig peritoneal macrophages have on their surface two receptors, one (Fcγ1/γ2R) binding both guinea pig IgG1 and IgG2 and the second (Fcγ2R) binding only IgG2 immunoglobulins. We have previously shown that treatment of macrophages with neuraminidase or glycosylation inhibitors affects, in a different way, the binding of guinea pig IgG1, IgG2, and rabbit IgG. In the present study we have shown that pretreatment of guinea pig macrophages with lectins (Con A, WGA, and PNA) also has a different effect on the interaction of the cells with IgG. The lectins increased the binding of guinea pig IgG1, whereas rabbit IgG and guinea pig IgG2 were bound with a lower efficiency than in the case of control cells. Since sialic acid residues seem to modulate the activity of receptors and WGA interacts with sialylated oligosaccharides, we determined the IgG‐binding characteristics for WGA‐pretreated macrophages. We found that the increase in IgG1‐binding ability was caused by an increase in the value of Kapp, but the number of IgG‐binding sites was lower than in the control cells. In the case of rabbit IgG and guinea pig IgG2 we observed a decrease of both the value of Kapp and the number of IgG‐binding sites. WGA did not interact directly with the Fcγ receptor. The results of our former papers and the different effects of lectins of various specificities described in this paper suggest different positions of Fcγ1/γ2 and Fcγ2R in the plane of the plane of the macrophage membrane in respect to various membrane glycoconjugates. Interaction of IgG with macrophage Fcγ receptors depends in a different way on glycoconjugates on the surface of the macrophage. Our results suggest that changes in glycosylation of macrophage surface glycoconjugates may be used by the cell for regulating the binding activities of the macrophage Fcγ receptors.
Platelet glycohydrolase activities: Characterization and release
Authors: Emiliani Carla , Martino Sabata , Orlacchio Antonio , Vezza Roberta , Nenci Giuseppe G. , Gresele Paolo
Published: May 2005

Granules containing acid hydrolases have been detected in human platelets but have not been thoroughly characterized.

We have studied the activity and characteristics of glycohydrolases present in normal human platelets, evaluated their release upon stimulation with thrombin, and assessed the contribution of platelet ‐ released lysosomal contents to the glycohydrolase activity present in normal serum. Platelets contained a remarkable glycohydrolase activity with a prevalence of β ‐ N‐acetylhexosaminidase. All glycohydrolases were released to some extent upon stimulation with thrombin and contributed to the glycohydrolase activity found in human serum. α‐Mannosidase and α‐galactosidase were partially inactivated after release by a mechanism as yet undefined. In addition, thrombin stimulation affects the intraplatelet isoenzyme pattern of β‐N‐acetylhexosaminidase by producing the appearance of a new form.
Experimental design in chemometrics
Authors: Deming Stanley N. , Palasota John A. , Palasota Josephine M.
Published: May 2005

Chemometrics is defined as the application of mathematical and statistical methods to chemical systems. Systems theory is seen to be useful for organizing and categorizing the inputs to and outputs from chemical systems. Advances in measurement science in the 1950s and 1960s, particularly in analytical chemistry, created a need for a multivariate approach to data analysis. Early chemometrics emphasized the use of structure‐finding methods for existing data sets. In many instances, data sets can be obtained from designed experiments. Such data sets are more likely to contain the desired information and the data can usually be acquired at less cost. Renewed interest in statistical process control will provide many new, more robust data sets in the future.
The quality of mean and variance estimates for normal and lognormal data when the underlying distribution is misspecified
Authors: Blackwood L. G.
Published: May 2005

Theoretical and simulation results are employed to evaluate mean and variance estimates for normal data when a lognormal distribution is assumed and for lognormal data when a normal distribution is assumed. Misspecifying the distribution leads to the use of suboptimal estimation methods. However, the results show that the suboptimal methods still produce estimators of good quality (low bias and variance) relative to the minimum variance unbiased estimators for each distribution, at least when practical efficiency is considered.
Comparison of two screening methods for the detection of volatile organic compounds in ground water
Authors: Rajagopal R. , Li Ping‐Chi
Published: May 2005

It is shown that the presence of 31–35 commonly measured volatile organic compounds (VOCs) in ground water can be detected with small error rates by using screening methods which analyze for a subset of such VOCs. A study of selected data sets indicates that analytical determinations of only from two to eight VOCs will suffice to detect 95% of all VOC hits. It is also shown that a serially optimal algorithm for selecting the VOCs for screening is very nearly as accurate as a globally optimal algorithm and much easier to implement. These conclusions are supported by empirical evidence from two drinking‐water data sets and one hazardous waste site data set. Additional research areas are also outlined.
Theory of medium‐rank second‐order calibration with restricted‐Tucker models
Authors: Smilde Age K. , Wang Yongdong , Kowalski Bruce R.
Published: May 2005

If an analytical instrument or instrumental method gives a response matrix when analyzing a pure analyte, the instrument or instrumental method is called a second‐order method. Second‐order methods that generate a response matrix for a pure analyte of rank one are called rank‐one second‐order methods. If the response matrix of a pure analyte is not rank one, essentially two cases exist: medium rank (between two and five) and high rank (greater than five). Subsequently, medium‐ and high‐rank second‐order calibration tries to use medium‐ and high‐rank second‐order methods to analyze for analytes of interest in a mixture. A particular advantage of second‐order methods is the ability to analyze for analytes of interest in a mixture which contains unknown interferences. Keeping this advantage is the challenge on moving away from rank‐one second‐order calibration methods. In this paper a medium‐rank second‐order calibration method is proposed based on least‐squares restricted Tucker models. With this method the second‐order advantage is retained.
A graphical technique for assessing differences among a set of rankings
Authors: Hirst David , Naes Tormod
Published: May 2005

A graphical method of assessing differences between sets of rankings based on cumulative ranks is developed. The method can be used to identify rankings that differ over all or just part of the range of objects ranked. The method is applied to an example of sensory evaluation of green peas in which ten assessors scored six attributes on each of 60 samples.