Determination of flunixin in equine urine and serum by capillary electrophoresis

A capillary electrophoresis (CE) and a solid-phase extraction method was developed for the determination of flunixin in equine urine and serum. The suitable CE run conditions were described. The factors affecting flunixin recovery rates were investigated and optimum solid-phase extraction conditions for flunixin in equine urine and serum were established. Limits of detection and quantitation were 3.4 and 5.6 ng/ml for serum and 16.9 and 33.1 ng/ml for urine, respectively. The recoveries exceeded 96% for urine and 79% for serum. Urine samples from race horses and urine and serum samples from a mare administrated with flunixin were analyzed with this procedure.     

Stability measurement of oligonucleotides in serum samples using capillary electrophoresis

Five male subjects aged between 25 and 40 years were given methanol at a dose of 10 mg/kg, once orally and once intravenously, while the enzyme systems responsible for methanol oxidation were blocked by ethanol. The study assessed the duration of inhibition of methanol oxidation in relation to the blood ethanol concentration, and the elimination of methanol not influenced by ethanol. Methanol elimination was found to begin at a blood ethanol concentration of 0.04-0.13 g/kg. Elimination constants of 0.406-0.267 h-1 with corresponding half-lives of 1.71-2.60 h were established for methanol not influenced by ethanol. When data from a previous study using an identical protocol for parenteral administration were included, making the total number of subjects nine, the mean elimination constant was found to be 0.298 +/- 0.470 h-1 and the mean half-life 2.37 +/- 0.357 h, distribution being normal. No evidence of any differences in methanol elimination kinetics between alcoholics and non-alcoholics or of a significant influence of the route of administration was found. The extent of intraindividual variation in methanol elimination as indicated by the difference in each subject between the values established, expressed as a percentage of the corresponding mean values, was found to be 3-25%, which is comparable to the magnitude of intraindividual variation in the rate of ethanol elimination.     

Determination of kynurenic acid by capillary electrophoresis with laser-induced fluorescence detection

We have investigated the influence of three structurally different but functionally related compounds [1, 10 ortho-phenanthroline (phenanthroline), Rifampicin and aurin tricarboxylic acid (ATA)] on the rate and the extent of proliferation of progesterone-responsive T47D human breast cancer cells. These compounds have previously been used in this laboratory and have been shown to modulate properties of nucleic acid binding proteins. Because p53 and the progesterone receptor (PR) are both DNA binding proteins that appear to regulate proliferation of breast cells, alterations in T47D cell p53 and PR levels were examined to determine their relevance in cell proliferation. T47D cells were grown in the absence of phenol red and in the presence of 5% fetal calf serum with or without charcoal stripping in the presence of the inhibitors. The rate of proliferation of cells grown in Rifampicin containing medium exhibited nearly 70% inhibition. Phenanthroline, a known metal chelator, was an effective inhibitor of proliferation at 3 mM reducing the cell number by more than 75%. ATA (0.24-2.4 micrograms/ml) inhibited the growth of the cells by nearly 50%. Analysis of the mechanism of action of these compounds revealed that treatment with these compounds caused specific changes in the molecular composition of T47D cell PR. Whereas ATA caused increased stability of PR isoforms, Rifampicin induced a upshift in the mobility of PR in SDS gels-a phenomenon associated with hyperphosphorylation of steroid receptors (SRs). Phenanthroline treatment (> 2 mM) caused a complete down-regulation of PR and the tumor suppressor protein, p53. The downregulation of p53 paralleled the changes in the molecular composition of PR. We propose that the inhibition of T47D cell proliferation by phenanthroline, Rifampicin and ATA results from a number of cellular changes that include regulation of p53 and PR.     

Determination of hyaluronidase activity in venoms using capillary electrophoresis

The technique used in this study was based on the addition of a known quantity of hyaluronic acid (HA) to an aliquot of crude venom sample, followed by obtaining capillary electrophoresis profiles both immediately after the mixing and after a known period of incubation. The presence of hyaluronidase (HYASE) and the degree of its activity were determined from the change in the abundance (peak height) of intact HA at its retention time. Quantitative and/or comparative enzyme activity could also be obtained from determining the incubation time needed either to achieve a selected percentage decrease in the size of the intact HA peak or to complete the digestion process as determined by the attainment of a constant profile of the oligosaccharide end products. The detection limit was 3 x 10(-6) U/microliter HYASE, defined as the decrease of the peak height of the added standard quantity of HA (0.008 mg/ml) from the initial signal-to-noise ratio of 6 down to 2; with respect to sample size, 1.5 x 10(-8) U of HYASE could be detected in 5 nl of incubated sample. The utility of the technique was illustrated by the rapid detection of HYASE activity in HYASE standards, crude bee venom and several snake venoms, the HYASE content of which has not yet been reported, and in collected high-performance liquid chromatography-separated venom fractions.     

Determination of peptidoglycan-associated protein in Escherichia coli NCIB 8545 by capillary zone electrophoresis

An efficient reliable and sensitive capillary zone electrophoresis assay for the six major bacterial peptidoglycan-associated proteins of Escherichia coli NCIB 8545 is described. The method provides the facility to determine quantitatively the effect of antibacterials on bacterial peptidoglycan-associated protein synthesis and thus to further elucidate the mechanism of antibacterial action of such drugs as the antifolates which recently have been shown to adversely affect peptidoglycan synthesis.     

DNA sequencing by capillary electrophoresis

Capillary electrophoresis has been under development for DNA sequencing since 1990. This development has traveled down two parallel tracks. The first track studied the details of DNA separation by gel electrophoresis. Early work stressed rapid separations at high electric fields, which reached the extreme of a 3.5 min sequencing run at 1200 V/cm. While fast separations are useful in clinical resequencing applications for mutation detection, long read-length is important in genomic sequencing. Unfortunately, sequence read-length degrades as electric field and sequencing speed increases; this tradeoff between read-length and sequencing speed appears to be a fundamental result of the physics of DNA separations in a polymer. The longest sequence sequencing read-lengths have been obtained at modest electric fields, high temperature, and with low concentration noncrosslinked polymers. In parallel with our understanding of DNA separations, the second track of DNA sequencing development considered the design of large-scale capillary instruments, wherein hundreds of DNA samples can be sequenced in parallel. Real-world application of these very high throughput capillary electrophoresis systems will require significant investment in sample preparation technology.     

Determination of bupivacaine and three of its metabolites in rat urine by capillary electrophoresis

A capillary electrophoretic (CE) method for the analysis of urinary extracts of the local anesthetic, bupivacaine, and its three main metabolites, desbutylbupivacaine, 3'-hydroxybupivacaine, and 4'-hydroxybupivacaine, in rat urine has been developed. The limits of detection were 0.22 microM for desbutylbupivacaine and bupivacaine, 0.15 microM for 3'-hydroxybupivacaine, and 0.16 microM for 4'-hydroxybupivacaine. The linear range was from 0.7 microM to 16.8 microM for all four compounds. Migration time and peak height reproducibilities, and extraction efficiencies were determined for all four compounds. Peak height reproducibilities (n = 5) for the overall method were improved through the use of prilocaine as an internal standard. Peak height reproducibilities were 5.6% RSD for desbutylbupivacaine and bupivacaine, and 9.9% RSD for 3'-hydroxybupivacaine and 4'-hydroxybupivacaine. Migration time reproducibilities (n = 5) were 2.4% for all compounds. Urine samples were collected from rats administered therapeutic doses of bupivacaine and extracted using a solid-phase extraction method (SPE). Separation of bupivacaine and its metabolites was achieved in 15 min.     

Determination of association constants of dansyl-amino acids and beta-cyclodextrin in N-methylformamide by capillary electrophoresis

The use of nonaqueous background electrolytes in capillary electrophoresis (CE) is a promising new trend which should widen the scope of this technique. We demonstrate the chiral separation of dansyl-amino acids (Dns-AAs) in N-methylformamide (NMF) using beta-cyclodextrin (beta-CD) as chiral selector. The solubility of beta-CD is much better in NMF than in water, allowing high concentration of the chiral selector and successful enantioseparation despite the weak host-guest interaction between the Dns-AAs and beta-CD. The association constants for the complexation between Dns-AAs and beta-CD could be calculated from the electrophoretic mobilities, with attention paid to the change in viscosity of the electrolyte upon addition of beta-CD. The association constants ranged between 2 and 13 M(-1).     

Determination of glycyrrhizin and glycyrrhetinic acid in traditional Chinese medicinal preparations by capillary electrophoresis

A simple, rapid, accurate and reproducible capillary electrophoretic method was developed for the assay of glycyrrhizin and glycyrrhetinic acid in traditional Chinese medicinal preparations. The buffer solution used in this method was acetonitrile and 0.02 M sodium dihydrogen-phosphate solution adjusted to pH 7.5 with 0.05 M sodium hydroxide. The linear calibration range was 0.04-2.00 mg/ml (r = 0.9988) for glycyrrhizin and 0.007-0.35 mg/ml (r = 0.9985) for glycyrrhetinic acid and recoveries were 98.1-101.3% for glycyrrhizin and 98.5-101.4% for glycyrrhetinic acid. The relative standard deviations were 1.02% (n = 6) for glycyrrhizin and 0.91% (n = 6) for glycyrrhetinic acid. The content of these two acids in Glycyrrhizae Radix and Glycyrrhizae Radix-containing Chinese medicinal preparations was successfully determined within 10 min.     

Determination of polyamines in serum by high-performance capillary zone electrophoresis with indirect ultraviolet detection

A method for determining polyamines in serum by capillary zone electrophoresis (CZE) with indirect ultraviolet detection was established. The concentrations of polyamines in the sera of six healthy adults were determined and the results were in accordance with those obtained previously by high-performance liquid chromatography (HPLC). However, the CZE method is superior to HPLC in that it has high sensitivity, small sample consumption and easy sample pretreatment.     

Analysis of antibiotics by capillary electrophoresis

The broad category of antibiotics encompasses some of the most widely prescribed pharmaceuticals in the world. As is the case with any pharmaceutical, an antibiotic must be characterized in terms of its potency or activity, and the presence and quantity of impurities. Additionally, any residue or metabolite that may be present as a result of its use must be monitored. Many capillary electrophoretic techniques have been utilized in the analysis of antibiotics, addressing the various aspects of their quantitation, profiling, and monitoring. Some of the more recent applications are summarized in this review article.     

Analysis of nucleotides by capillary electrophoresis

Capillary electrophoresis is a useful tool for the analysis of nucleotides. Methods have been optimized for both CZE and MECC modes. A variety of CZE buffers, such as borate, carbonate and phosphate were used successfully. The pH of the buffer changes the charge on the nucleotides. Therefore, the selectivity of the analytes can be controlled by the acidity of the buffer solution. CE separations of nucleotides have been performed at all pH levels, in both CZE and MECC modes. SDS was the most commonly used modifier in MECC separations, but other additives have been added to optimize selectivity. In addition, nucleotides have been quantified in different matrices, including tissue and cell extracts and several DNA and RNA sources. This paper summarizes the methods used for the optimization of nucleotides by CE and includes the most recent techniques to improve selectivity, reproducibility and sensitivity. A summary of CE methods is used in analyses of nucleotides in biological matrices is included.     

Determination of aminopeptidase X activity in tissues of normo- and hypertensive rats by capillary electrophoresis

We investigated the immunohistochemical localization of the macrophage migration inhibitory factor (MIF) in the rat brain. In addition to epithelial ependymal cells lining the ventricular wall, tanycytes in the basomedial hypothalamus were heavily immunostained. The immunoreactive processes of tanycytes made contacts to sinusoidal capillaries and reached the pial surface forming an immuno-positive structure at the floor of the hypothalamus. Other immunoreactive cells contained the subcommissural organ in the roof of the third ventricle and the epithelial lamina of the choroid plexus. The localization of MIF in cells which have contact with cerebrospinal fluid and blood vessels suggests that MIF might play a role as a humoral factor in the brain.     

Separation of tetracyclines by high-performance capillary electrophoresis and capillary electrochromatography

Separations of various tetracycline mixtures by high-performance capillary electrophoresis (HPCE) and a new form of electrochromatography (CEC) are compared. The new CEC method involves etching the inner wall of the capillary surface with an appropriate reagent (ammonium dihydrogen fluoride) in order to produce a significant increase in surface area. The etched surface is then modified by a silation/hydrosilation reaction sequence to first produce a hydride intermediate which is then further reacted to attach a C18 moiety. The bare and hydride capillaries are tested under HPCE conditions while the C18 capillary functions in the CEC mode. The effects of pH and the presence of an organic modifier (methanol) are also studied. Detection limits ( < 10 micrograms/ml) are comparable to previous HPLC and HPCE results. Resolutions for mixtures which simulate real analytical problems are equal to or better than those reported for separations on polymeric and diol columns by HPLC and in earlier studies by HPCE and MECC.     

Urine protein analysis by capillary electrophoresis

Increased concentration of proteins in urine as well as abnormal patterns are seen in many disorders such as various renal disorders and light chain disease. At the wavelength of 214 nm used for detection of the peptide bond, numerous compounds interfere in the analysis of urinary proteins. We show that either adsorptive filtration with a wash step or cold ethanol precipitation are two methods which can eliminate many of the interferences. The wash step is rapid, greatly reduces the interfering substances, and decreases the effect of sample matrix. Both of these methods yield results comparable to the traditional agarose method. Capillary electrophoresis (CE) is faster and more cost-effective than agarose electrophoresis.     

Detection of 2'-deoxyoxanosine by capillary electrophoresis

We have investigated capillary electrophoresis separation of oxanine (Oxa), a base moiety of 2'-deoxyoxanosine (Figure 1), from various bases. Oxa was not well separated from the others by micelle electrokinetic chromatography which is a general method for separating of the bases. On the other hand, capillary zone electrophoresis showed much improved separation when used pH 12 where the six-membered ring of Oxa is opened.     

Advances in capillary electrophoresis

This review summarizes the advancement in operational modes and selected applications of the title technique over the past five years. Regarding operational modes particular emphasis is put upon increasing selectivity and resolution, hyphenation of capillary electrophoresis with techniques based on other than electromigration principles, the so-called chip technology and new ways of detection. In applications selected examples of chiral separation and separation of biopolymers (proteins, nucleic acids) are emphasized. It is demonstrated that capillary electrophoresis represents a complementary technique to high-performance column chromatography and in a number of cases it offers better separations than standard chromatographic procedures.     

Characterization of DNA standards by capillary electrophoresis

We have used capillary electrophoresis to evaluate commercial DNA size standards and have found that it can provide an efficient assessment of size. However, the accuracy of the determination is adversely affected by anomalous migration times due to specific interactions of the DNA with the gel matrix as well as conformational differences in the DNA due to sequence heterogeneity. These anomalous migration times are strongly dependent on the choice of gel matrix. For example, the anomalous migration times that are observed in a 1 kilobase standard DNA ladder can be minimized using nongel hydroxyethylcellulose. In addition, the peak resolution can be increased and the anomalous migration can be reduced by the addition of the intercalating dye, ethidium bromide. However, in the case of the D1S80 allelic ladder, some of the DNA fragments possess nucleotide sequences which do not interact equivalently with the dye and produce irregular migration times. These measurements yield preliminary information useful in evaluating DNA size standards which may be used for a wide range of DNA diagnostic applications.     

差示分光光度法测定铁粉置换铜溶液中铁离子含量

以磺基水杨酸为显色剂,Fe3+标准溶液作为参比溶液,通过差示分光光度法在419 nm处测定 铁粉置换铜溶液中铁离子总量;研究不同波长、显色剂用量、氨水用量、显色时间对溶液吸光度的影 响.在1 h内最佳测量条件为:吸收波长为419 nm,20 %磺基水杨酸用量为15 mL,(1+1)氨水用 量为15 mL.得到线性回归方程为:C=0.009 4Abs+0.015 95,相关性系数R2=0.999 4.加标回收率 为99.38 %~99.61 %,平均回收率达到99.50 %.      

Separation of chiral compounds by capillary electrophoresis

This review presents the different chiral selectors used in capillary electrophoresis (CE) for the separation of enantiomers. The use of charged cyclodextrins, crown ethers, polysaccharides, proteins, natural and synthetic micelles, macrocyclic antibiotics and ergot alkaloids is discussed in detail. Neutral native and derivatized cyclodextrins are not treated because several review articles have already been published on this topic. Recent developments like the application of two chiral selectors in the same background electrolyte are highlighted.