Selective adsorption of mercury ion by mercaptocarboxylic acid intercalated Mg-Al layered double hydroxide.

Intercalations of mercaptocarboxylic acid and dithiodicarboxylic acid in Mg-Al layered double hydroxide and their adsorption properties for heavy metal ions were examined. During the intercalation of mercaptocarboxylic acids, mercapto group was oxidized, and the corresponding dithiodicarboxylic acids were intercalated in the interlayer space of Mg-Al layered double hydroxide. The intercalation compounds adsorbed mercury and silver ions effectively, whereas there was no adsorption of copper ion practically.

Mouse hepatitis virus 3 binding to macrophages correlates with resistance to experimental infection.

Mouse hepatitis virus 3 (MHV3) infection of A/J and BALB/c mice has been used as a model of resistance/susceptibility. A/J mice recover from a mild disease after 4-6 days of infection and the BALB/c mice develop an acute hepatitis and die after 3-4 days of infection. In view of studying the MHV3 binding to cells or cell extracts, we performed an enzyme-linked immunosorbent assay-like virus-binding assay, preparing microplates with L929 cells, A/J or BALB/c mouse macrophages and also with proteins extracted from these cells. Higher MHV3 bindings were observed to proteins of BALB/c macrophages than to the A/J ones. The interferon-gamma (IFN-gamma) activation led to a reduction of MHV3 binding only to proteins of resistant A/J mouse macrophages. Our experiments contribute to the hypothesis that IFN-gamma activation of macrophages plays an important role against MHV3 infection by downregulating the expression of viral receptors.

Adsorption of gaseous formaldehyde and carboxylic acids by ammonium-ion-exchanged alpha-zirconium phosphate.

Ammonium-ion-exchanged alpha-Zr(HPO(4))(2)H(2)O (alpha-ZrP) was obtained as a single phase with the interlayer distance of 9.4 A by the ion-exchange of proton with ammonium ion. The ammonium ion-exchanged alpha-ZrP could adsorb ill-smelling gases, such as formaldehyde and carboxylic acids (formic acid, acetic acid, propionic acid, and butyric acid). The adsorption amounts of carboxylic acids increased in the order, butyric acid

Single strand conformation polymorphism analysis of ras oncogene by capillary electrophoresis with laser-induced fluorescence detector.

Single strand conformation polymorphism (SSCP) analysis of the N-ras oncogene was achieved by capillary electrophoresis with a laser-induced fluorescence detector (CE-LIF) using methylcellulose as a molecular sieving agent. The PCR-amplified N-ras oncogene, which is known to have a point mutation at codon 61 in the neuroblastoma, was investigated by CE-LIF combined with SSCP (SSCP-CE-LIF). A mixture of wild- and mutant-type single strand DNA fragments (103 bp) of the N-ras oncogene was separated by buffer solution containing 1.0% methylcellulose and 0.2 microM fluorescent dye (YO-PRO-1) at 25 degrees C. The SSCP-CE-LIF technique gave good resolution for wild- and mutant-type single strand DNA fragments with separation completed within 7 min. SSCP analysis using a CE system with a LIF detector was successfully applied to the detection of the one point mutation on the N-ras oncogene.

Phosphorylation of D-glucose derivatives with inorganic cyclo-triphosphate: substituent effect at the 5-CH2OH or 2-OH group.

The phosphorylation of several D-glucose derivatives has been achieved using inorganic sodium cyclo-triphosphate hexahydrate (P3m), Na3P3O9 x 6H2O, in aqueous solution. In the phosphorylation of D-glucuronic acid, 6-phosphoryl-D-glucose and D-xylose, beta-D-glucuronic acid 1-triphosphate, 6-phosphoryl-beta-D-glucose 1-triphosphate and beta-D-xylose 1-triphosphate were synthesized stereoselectively with maximum yields of 43.5, 32.8 and 41.9%, respectively. In the case of D-glucosamine and 2-deoxy-D-glucose, the main phosphorylated products were assigned to beta-D-glucosamine 1-triphosphate and 2-deoxy-beta-D-glucose 1-triphosphate by 1H-, 13C- and 31P-NMR, and the yields were 13.9 and 13.4%, respectively.

A one-step phosphorylation of D-aldohexoses and D-aldopentoses with inorganic cyclo-triphosphate.

The phosphorylation reaction by inorganic cyclo-triphosphate (P3m) having a six-membered ring was examined for D-aldohexoses and D-aldopentoses in aqueous solution. Similar to the process for D-glucose, D-galactose, D-xylose or D-allose were phosphorylated with P3m to give stereoselectively beta-D-galactopyranosyl 1-triphosphate, beta-D-xylopyranosyl 1-triphosphate or beta-D-allopyranosyl 1-triphosphate with maximum yields of 31.3, 32.5 or 32.1%, respectively. On the other hand, in the reaction of D-ribose, D-lyxose, D-mannose or D-arabinose with P3m, the yields of beta-D-ribopyranosyl 1-triphosphate, alpha-D-lyxopyranosyl 1-triphosphate, alpha-D-mannopyranosyl 1-triphosphate or alpha-D-arabinopyranosyl 1-triphosphate were 8.0, 16.5, 9.6 or 14.1%, respectively. The phosphorylation mechanism of D-aldopyranoses with P3m was also discussed.

Simultaneous analysis of genes by capillary electrophoresis with a laser-induced fluorescence detector using a stepwise field strength gradient.

A mixture of polymerase chain reaction (PCR) products, 100, 105, 300, 310, 485, and 500 base pair (bp) DNA fragments, was analyzed by capillary electrophoresis equipped with a laser-induced fluorescence detector (CE-LIF) using a stepwise gradient of electric field strength. The optimum condition for the analysis of PCR products was 0.5% methylcellulose and 160 V/cm from 0 to 10 min and 270 V/cm from 10 to 17 min. The length (bp) of DNA could be estimated from the relationship between the relative migration time and bp length. The relative standard deviation (R.S.D.) of DNA size (bp) was less than 3.5% and the difference from the true value was only 2.4 bp.

Analysis of variable number of tandem repeats of human genome D1S80 locus using capillary electrophoresis with laser-induced fluorescence detector.

The conventional method of identifying individuals by DNA in the field of forensic medicine is slab gel electrophoresis, which is time-consuming, labor-intensive, and nonquantitative. Accordingly, the use of capillary electrophoresis with a laser-induced fluorescence detector (CE-LIF), human genome D1S80 locus, a DNA marker which has a variable number of tandem repeats (VNTR) on chromosome 1, was examined to improve DNA analysis for identification. Using an internal standard, fragment size of VNTR was accurately and rapidly determined.

Relationship between spontaneous aminoglycoside resistance in Escherichia coli and a decrease in oligopeptide binding protein.

Changes in the amount of oligopeptide binding protein (OppA) in spontaneous kanamycin-resistant mutants of Escherichia coli were investigated. Among 20 colonies obtained from 10(8) cells cultured in the presence of 20 microgram of kanamycin/ml, 1 colony had no detectable OppA and 7 colonies were mutants with reduced amounts of OppA. Sensitivity of wild-type cells to kanamycin increased slightly by transformation of the oppA gene, but the sensitivity of the mutants increased greatly by the transformation. A mutant with no OppA was found to be a nonsense mutant of the oppA gene at amino acid position 166. In a mutant having a reduced level of OppA, the reduction was due to the decrease in OppA synthesis at the translational level. These mutants were also resistant to other aminoglycoside antibiotics, including streptomycin, neomycin, and isepamicin. Isepamicin uptake activities decreased greatly in these two kinds of mutants. The results support the proposition that aminoglycoside antibiotics are transported into cells by the oligopeptide transport system, and that transport is an important factor for spontaneous resistance to aminoglycoside antibiotics.

Enhanced separation of DNA sequencing products by capillary electrophoresis using a stepwise gradient of electric field strength.

The effect of the electric field strength gradient on the separation of DNA sequencing fragments was investigated. We demonstrate that the stepwise gradient of electric field improves the separation of DNA sequencing fragments more than 500 bases in size and diminishes the analysis time for DNA sequencing of lager DNA fragments. The use of the electric field strength gradient induces an increase in the theoretical plate number as predicted by the theoretical formulation discussed in this paper.

Analysis of a variable number of tandem repeats in a heart disease gene by capillary electrophoresis with laser-induced fluorescence detector.

Capillary electrophoresis with laser-induced fluorescence detector (CE-LIF) was successfully applied to the analysis of the variable number of tandem repeats (VNTR) in a human apolipoprotein B gene (APOB). Apolipoprotein B VNTR alleles containing more than 35 repeat units are a significant risk factor for heart disease. Thus, we developed a method for accurately determining the number of repeat units (16 bp) in the VNTR using capillary electrophoresis. The CE-LIF technique gave excellent resolution of APOB alleles differing by 2 or 4 repeat units over the range 600 to 1000 bp. The recommended conditions for the analysis of APOB VNTR loci by CE-LIF are as follows: effective length of capillary (100 microns i.d., 360 microns o.d.), 50 cm; running buffer, 50 mM Tris-borate 0.5% methylcellulose and 0.1 microM fluorescent dye YO-PRO-1; electric field, 150 V/cm.

Rapid typing of variable number of tandem repeat locus in the human apolipoprotein B gene for DNA diagnosis of heart disease by polymerase chain reaction and capillary electrophoresis.

The apolipoprotein B (apoB) variable number of tandem repeat (VNTR) alleles containing larger repeat units is a risk factor for coronary heart disease. Capillary electrophoresis (CE) in entangled polymer solution was applied to the analysis of polymerase chain reaction (PCR) amplified apoB VNTR locus for DNA diagnosis of heart disease. The CE separation gives an excellent resolution of two alleles differing by one or two 16 bp repeat units in the DNA size range up to 600 bp with high speed. The apoB alleles differing in length by 2 or 4 repeat units are readily distinguishable by CE in the DNA size range from 600 to 1000 bp. The plate number achieved was 1 million plates per meter. CE combining with PCR provides an excellent technique for accurate determination of the number of repeat units of apoB VNTR alleles and differentiation of heterozygous from homozygous individuals. Using the CE technique, the apoB VNTR loci from some individuals in genotyping were examined towards precise DNA diagnosis for coronary heart disease.

High-resolution separation of PCR product and gene diagnosis by capillary gel electrophoresis.

High-resolution separation of a PCR product from a mixture of DNA restriction fragments was achieved using capillary gel electrophoresis. The capillary gel electrophoretic separation gives an excellent resolution of two fragments of the 500-bp PCR product and the 506-bp DNA fragment, which differs by only 6 bp, and the complete separation of a broader chain length range of DNA afragments up to 12 kbp within 20 min. The plate number of gel-filled capillary was achieved to be 2 million plates per meter. Capillary gel electrophoresis is applied to the gene diagnosis for heart disease through apolipoprotein E genotyping. The advantages and limitations of capillary gel electrophoresis in the application to PCR analysis and DNA diagnosis are discussed.

High-resolution separation of DNA restriction fragments by capillary electrophoresis in cellulose derivative solutions.

The performance and the efficiency of several cellulose derivatives as a molecular sieving agent for the capillary electrophoretic separation of DNA restriction fragments were investigated. All fragments up to 12,000 base pairs (bp) in the 1-kbp DNA ladder were resolved using linear polyacrylamide-coated capillaries filled with a buffer solution containing 0.5% cellulose derivative and the separation was completed within 17 min. High-concentration (0.7%) cellulose derivative solutions are effective for the complete separation of small fragments (50-1000 bp) of a HincII and a HaeIII digest of phi X174 DNA. A plate number of 0.5-1 x 10(6) plates per metre was achieved. The migration time and the resolution of DNA fragments were manipulated by varying several parameters, such as the size (viscosity) and the concentration of cellulose derivatives and the applied field strength. Some guidelines are presented for choosing these parameters, depending on the size of the DNA fragments being separated.

Effect of urea concentration on the base-specific separation of oligodeoxynucleotides in capillary affinity gel electrophoresis.

Base-specific separations of oligodeoxynucleotides were achieved with high resolution by electrophoresis, using a urea-gel capillary, in which poly(9-vinyladenine) (PVAd) was utilized as an affinity ligand. The migration behaviour and the plate number of oligodeoxynucleotides were investigated as a function of urea concentration between 2 and 10 M in capillary gel electrophoresis (CGE) as well as capillary affinity gel electrophoresis (CAGE). The migration time in CGE separation increases as urea concentration increases. An increase in the viscosity of the gel buffer medium as well as a change in the conformation of oligodeoxynucleotides is found to be a predominant factor for an increase in the migration time. The migration time and the plate number of oligothymidylic acids, which interact with PVAd in CAGE, is manipulated by differing urea concentration, which leads to the change in the dissociation process of a specific hydrogen bonding between oligothymidylic acids and PVAd. The migration time of oligodeoxyadenylic acids, which do not interact with PVAd in CAGE, increases with an increase in urea concentration as in CGE separation. The plate number of oligodeoxyadenylic acid was not affected by the urea concentration.

Specific base recognition of oligodeoxynucleotides by capillary affinity gel electrophoresis using polyacrylamide-poly(9-vinyladenine) conjugated gel.

Poly(9-vinyladenine) was synthesized and utilized as an affinity macroligand entrapped within the gel matrix. Base-specific separation of oligodeoxynucleotides was achieved with high resolution and high speed by electrophoresis, using capillaries filled with conjugated polyacrylamide-poly(9-vinyladenine) gel. Oligothymidylic acids were selectively separated from the mixture of oligothymidylic and oligodeoxyadenylic acids by utilizing a specific hydrogen bonding between poly(9-vinyladenine) and oligothymidylic acids. Migration time and resolution of oligodeoxynucleotides were influenced by several parameters, such as the size of poly(9-vinyladenine), capillary temperature, and concentrations of poly(9-vinyladenine) and urea. Some guidelines are presented, based on the theoretical formulation of the effect of these parameters, in order to find optimum electrophoretic conditions. Analytical capillary affinity gel electrophoresis was developed for the selective and sensitive base recognition of oligodeoxynucleotides with efficiencies as high as several 10(6) plates/m by using a urea-gel capillary with poly(9-vinyladenine) and temperature-programming.

Recognition of (d(TTTATT) and d(TTATTT) by capillary affinity gel electrophoresis (CAGE) using poly(9-vinyladenine)-polyacrylamide conjugated gel.

Sequence-specific recognition of oligodeoxynucleotide isomers (TTTATT and TTATTT) was achieved by using polyacrylamide-poly(9-vinyladenine) conjugated gel filled capillary affinity gel electrophoresis. It was found that the interaction between poly(9-vinyladenine) and the isomers was dependent on the sequential thymidylic acids of them.

High-performance separation of polynucleotides by capillary gel electrophoresis.

Capillary gel electrophoresis was applied to the high speed separation of DNA and RNA. Factors affecting resolution and speed were optimized for the single base resolution of polynucleotides. Polynucleotides up to 350 bases were completely resolved within 38 min under optimum conditions.

Comparative studies of nifurtimox uptake and metabolism by drug-resistant and susceptible strains of Trypanosoma cruzi.

1. Nifurtimox uptake and metabolism by epimastigote forms of three strains of Trypanosoma cruzi (Basileu, Y, YuYu) with different drug responsiveness in mice experimental infections were compared. 2. Statistical analysis of the results demonstrated no correlation between the ability of the strains to catalyze nifurtimox redox-cycling (Basileu = Y = YuYu) nor nifurtimox multiple electron reduction (Basileu = Y greater than Y) and drug susceptibility (Basileu greater than Y greater than YuYu). 3. A partial correlation however, was observed between drug responsiveness and nifurtimox uptake (Basileu greater than Y = YuYu). 4. The results suggest that drug uptake may be more important than drug metabolism in modulating resistance to nifurtimox in T. cruzi strains.