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PubMed1984

Evaluation of an ion-exchange procedure for removal of hepatitis type B contamination from human gamma globulin products.

Utilizing a unique human plasma sample that contained extremely high concentrations of hepatitis B surface antigen (HBsAg), we were able in the laboratory to rapidly screen the binding efficiency for HBsAg of various ion-exchange resins that can be used to purify gamma globulin product from human plasma. 'Nonrigid' resins like QAE-Sephadex and DEAE-Sephadex showed a high affinity for HBsAg by removing greater than 10(4) of applied challenge. Much lower binding was achieved by 'rigid' resins like DEAE-Sepharose CL-6B and DEAE-Spherodex. Also examined were the usefulness of certain auxiliary procedures, special filters and resins, that offer the potential to remove HBsAg. We failed to find any significant removal of the HBsAg, and in some cases large losses of the gamma globulin product occurred. While encouraged by our findings, certain precautions are discussed concerning the conversion from large-scale production of gamma globulin by the conventional alcohol method to one utilizing chromatographic techniques.

Chromatography, Ion Exchange↗

PubMed2002

A new dipeptide, O-phosphoserylethanolamine isolated from Agkistroden blomhoffi (mamushi).

A new dipeptide was isolated from several tissues of Agkistroden blomhoffi (mamushi: a venomous snake in Japan), using ion-exchange resins and thin-layer chromatography. It was identified as O-phosphoserylethanolamine by mass spectrometry and comparison with synthetic compounds using several methods. This compound was contained in several mamushi tissues including the liver, heart, brain, bile, and muscle. The concentrations of O-phosphoserylethanolamine in the liver, brain, muscle, skin, heart, and bile were 7.17+/-3.11,16.98+/-4.25,37.37+/-7.88,37.56+/-8.97,23.93+/-6.11, and 22.21+/-5.76 micromol/g, respectively.

Agkistrodon↗

PubMedDate not supplied

[Hemoglobin J. Broussais alpha-2 90 Lys leads to Asn beta-2A (FG2) discovered in a Martinique family. Comparison of several analytical technics].

We report a new case of hemoglobin J. Broussais, present in its heterozygote form in a seven year old child from Martinque. The structural characteristics of this abnormal hemoglobin have been compared by three methods: 1. By analytical and preparative finger-printing on silica gel thin layer plates after tryptic digestion. 2. By a programmed separation of tryptic peptides on various ion exchange resins. 3. And by automatic sequencing of the peptides obtained following CNBr cleavage and separation by gel filtration chromatography. The functional behaviour of this hemoglobin was not modified by the substitution of an Asn for a Lys residue at position 90, although its presence induced slight hematological disorders in the patient. Abnormal hemoglobins which have been identified in Martinique and Guadeloupe are reviewed.

Amino Acid Sequence↗

PubMed2002

Foam fractionation of a dilute solution of bovine lactoferrin.

Lactoferrin (Lf), a protein found in human and bovine milk, tears, blood, and other secretory fluids, has been used to prevent infection from potential microbial pathogens by its ability to bind with iron (Fe3+). Currently, bovine lactoferrin can be purified from milk using ion exchange resin, which is a costly procedure making lactoferrin expensive. The purpose of this work was to investigate a low-cost foam fractionation process as the first step in separating lactoferrin from milk.

Animals↗

PubMed1983

Determination of catecholamine sulfoconjugate isomers in normal human urine by use of high-performance liquid chromatography with a photochemical detector.

A method for the determination of catecholamine sulfoconjugate isomers (CA-S) in urine was developed. The photo-induced fluorogenic reaction of CA-S with ethylenediamine reported previously was applied to the postcolumn labeling of HPLC for sensitive and selective detection. Special equipment for the reaction was made with a uv-irradiation lamp and a reaction coil of Teflon tubing inside a temperature-controlled reaction box. Lower determination limits of this system were 1 to 2 pmol. Urine samples pretreated with small ion-exchange resin columns were subjected to HPLC. Peaks corresponding to CA-S were identified quantitatively by two different separation methods. Thus, all six CA-S were first detected in the urine of normal individuals. The excretion rates of dopamine 3-sulfate, dopamine 4-sulfate, norepinephrine 3-sulfate, norepinephrine 4-sulfate, epinephrine 3-sulfate, and epinephrine 4-sulfate were 420 +/- 240, 98 +/- 55, 86 +/- 95, 15 +/- 14, 18 +/- 7, and 3 +/- 1 ng/min (+/- SD), respectively (n = 5).

Catecholamines↗

PubMed2004

Post-translocational folding of secretory proteins in Gram-positive bacteria.

The transport of proteins from their site of synthesis in the cytoplasm to their functional location is an essential characteristic of all living cells. In Gram-positive bacteria the majority of proteins that are translocated across the cytoplasmic membrane are delivered to the membrane-cell wall interface in an essentially unfolded form. They must then be folded into their native configuration in an environment that is dominated by a high density of immobilised negative charge-in essence an ion exchange resin. It is essential to the viability of the cell that these proteins do not block the translocation machinery in the membrane, form illegitimate interactions with the cell wall or, through intermolecular interactions, form insoluble aggregates. Native Gram-positive proteins therefore have intrinsic folding characteristics that facilitate their rapid folding, and this is assisted by a variety of folding factors, including enzymes, peptides and metal ions. Despite these intrinsic and extrinsic factors, secretory proteins do misfold, particularly if the cell is subjected to certain types of stress. Consequently, Gram-positive bacteria such as Bacillus subtilis encode membrane- and cell wall-associated proteases that act as a quality control machine, clearing misfolded or otherwise aberrant proteins from the translocase and the cell wall.

Bacterial Proteins↗

PubMed1981

Purification of a human progesterone receptor.

The cytoplasmic progesterone receptor form human uterus has been purified to apparent homogeneity by a combination of ammonium sulfate fractionation and affinity chromatography. Affinity resins prepared by conventional means were compared to those prepared by a modified method. The latter give more reproducible results. A consistent finding was that low capacity resins gave the highest fold purification of the receptor. The pure receptor sedimented at 3.6 S on sucrose density gradient centrifugation, was eluted as a single band by 0.2 M KCl from DEAE-cellulose, and migrated as a single band of molecular weight 42 000 on NaDodSO4-polyacrylamide gel electrophoresis. Molecular weight determinations, obtained from Strokes' radii and sucrose gradient centrifugation, the receptors' behavior on ion exchange resins, and hormone binding specificity were all similar to those of the receptor found in crude cytosol. When the crude cytosol receptor was photoaffinity labeled by using 3H-labeled 17,21-dimethyl-19-norpregna-4,9-diene-3,20-dione followed by NaDodSO4-polyacrylamide gel electrophoresis, only protein of Mr 42 000 was labeled. This is consistent with our previous findings that alkylation of the pure receptor using 11-deoxycorticosterone bromo[3H]acetate showed labeling of a single protein of Mr 42 000. These properties confirm that the identity and integrity of the receptor have been maintained throughout its purification.

Cytosol↗

PubMed1995

[Cyclic AMP-specific nucleotide phosphodiesterase from the insoluble fraction of the human brain].

The cAMP-specific cyclic nucleotide phosphodiesterase (PDE) from pellet fraction of human brain has been identified. Two alternative methods for isolation and purification of the enzyme are presented. The first method consists of three chromatographic steps (separation on the DEAE-Toyopearl column followed by gel filtrations on AcA-34 and G3000 SW columns) and results in purification of several forms of PDE. One of those is weakly inhibited by cGMP and is insensitive to Ca(2+)-calmodulin or cyclic nucleotides. The second procedure consisting of the chromatographic separation on blue agarose and DEAE-TSK-5PW ion-exchange resin leads to an isolation of several peaks of cAMP-specific PDE. Both methods produce different cAMP-specific forms of PDE with molecular masses ranging from 36 to 47 kDa. These forms differ in kinetic properties (Km = 2.0-7.3 microM, Ki = 1480-5300 microM for cGMP). However, the KicGMP/KmcAMP ratio remains constant for all the peaks of the cAMP-specific enzyme.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1991

Analytical studies on the chiral separation and simultaneous determination of pantothenic acid and hopantenic acid enantiomers in rat plasma by gas chromatography-mass fragmentography.

The chiral separation and simultaneous determination of D- and L-pantothenic acids and D- and L-hopantenic acids in rat plasma using gas chromatography-mass fragmentography are described. The method is based on deproteinization by ion-exchange resin, extraction with ethyl acetate under acidic conditions, and derivatization to form several interesting compounds. After methyl esterification for carboxylic acid of D- and L-pantothenic acids, D- and L-hopantenic acids could be derivatized to trifluoroacetate, cyclic sulphite, and cyclic n-butylboronate for hydroxy groups. D- and L-forms of these derivatives were completely separated by mass fragmentographic technique with quasi-molecular ions. Calcium salt of D-5-[(2,4-dihydroxy-3,3-dimethyl-1-oxobutyl)amino]pentanoic acid was used as an internal standard for the determination of DL-pantothenic acids and DL-hopantenic acids. The detection limits of DL-pantothenic acids and DL-hopantenic acids in this method were 5 and 12 ng, respectively. This method could be applied to the study of plasma levels of D-, L-pantothenic acids and D-, L-hopantenic acids in rat.

Animals↗

PubMed2006

Overexpression, purification and crystallization of a choline-binding protein CbpI from Streptococcus pneumoniae.

The choline-binding protein CbpI from Streptococcus pneumoniae is a 23.4 kDa protein with no known function. The protein has been successfully purified initially using Ni-NTA chromatography and to homogeneity using Q-Sepharose ion-exchange resin as an affinity column. CbpI was crystallized using PEG 3350 as a precipitant and X-ray crystallographic analysis showed that the crystals belonged to the tetragonal space group P4, with unit-cell parameters a = b = 83.31, c = 80.29 angstroms, alpha = beta = gamma = 90 degrees. The crystal contains two molecules in the asymmetric unit with a solvent content of 55.7% (V(M) = 2.77 angstroms3 Da(-1)) and shows a diffraction limit of 3.5 angstroms.

Bacterial Proteins↗

PubMedDate not supplied

Sequential isotopic determination of plutonium, thorium, americium, strontium and uranium in environmental and bioassay samples.

A procedure has been developed to provide sequential analysis of 238Pu, 230Th, 241Am, 238U, and 90Sr in environmental and bioassay samples. Tracers and/or carriers (242Pu, 243Am, 232U, and stable strontium) are added into the sample as chemical yield monitors, and then, plutonium, thorium, strontium, americium, and uranium are sequentially separated and purified by Dowex ion-exchange resin, EiChroM Sr-resin, EiChroM TRU-resin, and Chelate-100 resin, respectively. The radioactivities of 90Sr and the actinides are measured using the liquid scintillation counter and alpha-particle spectrometer, respectively. Acidified water, glass-fiber air filter, soil, synthetic urine and synthetic feces samples of US National Institute of Standard and Technology Radiochemical Intercomparison Program(NRIP) were analyzed to verify this method. All the analytical results of 238Pu, 230Th, 90Sr, 241Am and 238U meet the traceability limit per ANSI N42.22, and when appropriate, evaluation of radiobioassay measurement bias and precision per ANSI N13.30.

Chromatography, Ion Exchange↗

PubMed2003

Continuous clavulanic acid adsorption process.

Adsorption kinetics and equilibrium data of clavulanic acid, a beta-lactam antibiotic, on ion-exchange resin Amberlite IRA 400 were utilized to carry out the modeling and simulation of a continuous adsorption process. These simulations allowed the estimation of yield, concentration, and purification factors of the process utilizing the product final concentration. Experimental runs of this process were carried out using the conditions pointed out by simulation studies. Comparison of the experimental results and those calculated by the proposed model showed that the model could describe very well the main features of the continuous process.

Adsorption↗

PubMed2004

Development of new toxicity identification evaluation by assessing toxicity reduction on adsorption and ion-exchange treatments.

A new toxicity identification evaluation method for water exhibiting toxicity has been suggested by evaluating toxicity reduction resulting from adsorption and ion-exchange treatments. Adsorption using various adsorbents such as activated carbon, chitosan and zeolite, and ion-exchange using cationic and anionic ion-exchange resins were performed. In addition, toxicity was evaluated by a cell survival test using human liver cancer origin cells (HepG2), and the dose response data was applied to treatment characteristics. The amount of toxicity adsorbed by the various adsorbents was defined based on adsorption isotherm. Parameters of the toxicity adsorption isotherm provided information that allowed us to identify the toxicity-controlling chemicals in environmental water containing various chemicals. The method is promising for water quality management because it can be used to identify toxicity-controlling chemicals among various environmental pollutants.

Adsorption↗

PubMed2004

The monitoring of tap waters in Milano: planning, methods and results.

The radiometric monitoring of tap waters has been carried out, in Northern Italy, since late the 1980s as part of the activities headed by the National Surveillance Network on environmental radioactivity. Measurements were accomplished by gamma spectrometry over large samples of water that were drained in a single place and concentrated by ion exchange resin treatment. More recently a regular, periodical monitoring was started using liquid scintillation measurements. In this manner, gross alpha and beta activities, uranium, (226)Ra and (222)Rn concentrations are determined at present. Nevertheless, a single-point sampling may be not fully representative of the whole Milano aqueduct, significantly extended and fed by a number of wells. As a control, an extended monitoring of the aqueduct was planned and variations of water radioactivity in different Milano areas have been assessed.

Chromatography, Ion Exchange↗

PubMed1999

Aldehyde dehydrogenase from rat intestinal mucosa: purification and characterization of an isozyme with high affinity for gamma-aminobutyraldehyde.

In rat adrenal gland and gastric mucosa putrescine is efficiently oxidized to GABA via gamma-aminobutyraldehyde (ABAL) by action of diamine oxidase and aldehyde dehydrogenase. Having turned our attention on the rat intestinal mucosa, where putrescine uptake and diamine oxidase are active, we have purified and characterized an aldehyde dehydrogenase optimally active on gamma-aminobutyraldehyde. A dimer with a subunit molecular weight of 52,000, the native enzyme binds ABAL and NAD+ with high affinity: at pH 7.4, Km values are equal to 18 and 14 microM, respectively. Affinity for betaine aldehyde is much lower (Km = 285 microM), but the efficiency is equally good, thanks to a high value of V. Unaffected by disulfiram and Mg2+, the enzyme is activated by high NAD+ concentrations (Vnn = 1.6 x Vn) and is competitively inhibited by NADH. According to the best fitting model, the dimeric enzyme only binds one NADH and the mixed complex enzyme-NAD(+)-NADH is inactive. The increase of activity promoted by NAD+ can therefore be ascribed to an allosteric effect, rather than to the activation of a second reaction center. Highly stable at pH 6.8 in the presence of dithiothreitol and high phosphate concentrations, ABALDH is inactivated by ion-exchange resins and by cationic buffers. Our results show that the enzyme can be effectively involved in the metabolism of biogenic amines and, with a K(m) for ABAL lower than 20 microM, in the synthesis of GABA.

Aldehyde Dehydrogenase↗

PubMed1991

[Purification of sodium channel from squid mantle and reincorporation in planar lipid bilayers].

The purification of voltage-dependent sodium channels from mantle muscles of the squid is monitored through increase in the specific binding of tritiated saxitoxin (3H-STX) of membrane extracts solubilized with Lubrol-PX, chromatographed over an ion-exchange resin then through an affinity column coupling lectins (WGA) and Sepharose. The achieved purification factor is 300 and the electrophoretic pattern of purified fractions shows a single band at 260 kD. Reincorporation of these fractions into planar lipid bilayers yields two conductance levels (in 0.5 M NaCl). The most frequently observed level, at 15 pS, is in agreement with in situ recordings. The other level, at 150 pS, equally blocked by tetrodotoxin, could be explained by some flexibility of the open state or alternatively by cooperative interactions between channels.

Animals↗

PubMed1996

Regulation of rat proximal tubule Na/H exchange by protein kinase C.

The effect of protein kinase C (PKC) activation on fluid and bicarbonate transport in renal tubules has been discussed controversially. Stimulation and inhibition have been shown to depend on factors such as experimental model and exposure time to the mediator of enzyme activation. We studied the role of PKC activation by phorbol-12-myristate-13-acetate (PMA) and by 1,2-dioctanoyl glycerol (DOG) in proximal bicarbonate reabsorption (JHCO3-) by 'in vivo' stationary microperfusion and ion-exchange resin microelectrode determination of luminal pH. Both PMA (10(-8) mol/l) and DOG (10(-3) mol/l) added to lumen or to peritubular capillaries reduced the net JHCO3- significantly. When added to lumen, the inhibition was 44 and 32%, respectively. This reduction did not involve changes in lumen stationary pH, but was mediated by a marked increase in the halftime of luminal bicarbonate disappearance; from 4.22 +/- 0.23 to 6.27 +/- 0.51 s with PMA and from 3.90 +/- 0.25 to 6.33 +/- 0.48 s with DOG. This effect was intensified by 10(-6) mol/l okadaic acid, a phosphatase inhibitor (inhibition of JHCO3- increased to 61%), and reduced by 30% by 10(-6) mol/l H7, an inhibitor of PKC. H89, a protein kinase A inhibitor, did not affect the inhibitory action of PMA. Our data suggest that PKC activation reduces the rate of H ion secretion (bicarbonate reabsorption) in convoluted segments of rat renal proximal tubules and that phosphorylation of the Na+/H+ exchanger by this kinase is the cause of the reduction in net secretion of H ions.

Acids↗

PubMed1961

Citric acid fermentation of sugars purified with chelating resin.

Noguchi, Yuichi (University of Wisconsin, Madison), and Marvin J. Johnson. Citric acid fermentation of sugars purified with chelating resin. J. Bacteriol. 82:538-541. 1961.-A new, independently isolated strain of Aspergillus niger capable of giving high yields of citric acid in submerged culture was found to show the same behavior toward iron, zinc, and manganese as a previously studied strain. Citric acid accumulation did not occur in the presence of manganese. Best citric acid production was obtained in the presence of limited amounts of iron and zinc. Use of a chelating ion exchange resin was found to be an excellent method of removing polyvalent metals from sugars, either for analytical purposes or for fermentation. Commercial glucose, after resin treatment, gave citric acid yields of more than 80% when supplemented with iron and zinc. Unpurified glucose was converted to citric acid in good yields by mycelium grown on resin-treated glucose.

Aspergillus↗