A simple and rapid ultra-high-performance liquid chromatography coupled with mass spectrometry method was developed for acyclovir and its metabolite 9-carboxymethoxymethylguanine in human serum. After precipitation of serum samples with 0.1% formic acid in acetonitrile/methanol (40:60, v/v), components were separated on a Luna Omega C18 column (1.6 μm; 2.1 × 150 mm) at 40°C. Mobile phase A (2 mmol/L ammonium acetate, 0.1% formic acid in 5% acetonitrile, v/v/v) and mobile phase B (2 mmol/L ammonium acetate, 0.1% formic acid in 95% acetonitrile, v/v/v) were used for gradient elution. A linear calibration curve was obtained over the range of 0.05-50 mg/L, and the correlation coefficients were better than 0.999. The limit of quantitation was 0.05 mg/L for both analytes. The intra- and interday accuracy and precision at three concentration levels ranged between 1.6 and 13.3%, and recoveries were achieved with a range between 92.2 and 114.2%. This method was developed and validated for the therapeutic monitoring of acyclovir in patients.
- Klíčová slova
- 9-carboxymethoxymethylguanine, acyclovir, mass spectrometry,
- MeSH
- acetonitrily chemie MeSH
- acyklovir analýza MeSH
- chemické techniky analytické normy MeSH
- dospělí MeSH
- formiáty chemie MeSH
- guanin analogy a deriváty analýza MeSH
- hmotnostní spektrometrie MeSH
- kalibrace MeSH
- lidé středního věku MeSH
- lidé MeSH
- limita detekce MeSH
- mladiství MeSH
- mladý dospělý MeSH
- reprodukovatelnost výsledků MeSH
- řízení kvality MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- 9-carboxymethoxymethylguanine MeSH Prohlížeč
- acetonitrile MeSH Prohlížeč
- acetonitrily MeSH
- acyklovir MeSH
- formiáty MeSH
- formic acid MeSH Prohlížeč
- guanin MeSH
About eight years ago, a new automation approach and flow technique called "Lab-In-Syringe" was proposed. It was derived from previous flow techniques, all based on handling reagent and sample solutions in a flow manifold. To date Lab-In-Syringe has evidently gained the interest of researchers in many countries, with new modifications, operation modes, and technical improvements still popping up. It has proven to be a versatile tool for the automation of sample preparation, particularly, liquid-phase microextraction approaches. This article aims to assist newcomers to this technique in system planning and setup by overviewing the different options for configurations, limitations, and feasible operations. This includes syringe orientation, in-syringe stirring modes, in-syringe detection, additional inlets, and addable features. The authors give also a chronological overview of technical milestones and a critical explanation on the potentials and shortcomings of this technique, calculations of characteristics, and tips and tricks on method development. Moreover, a comprehensive overview of the different operation modes of Lab-In-Syringe automated sample pretreatment is given focusing on the technical aspects and challenges of the related operations. We further deal with possibilities on how to fabricate required or useful system components, in particular by 3D printing technology, with over 20 different elements exemplarily shown. Finally, a short discussion on shortcomings and required improvements is given.
- Klíčová slova
- 3D printing of instrument elements, Lab-In-Syringe, automation of sample pretreatment, potentials and troubles, system setup and operation modes, tips and tricks in method development,
- MeSH
- chemické techniky analytické přístrojové vybavení metody normy MeSH
- injekční stříkačky * MeSH
- laboratorní automatizace * MeSH
- limita detekce MeSH
- reprodukovatelnost výsledků MeSH
- Publikační typ
- časopisecké články MeSH
Analysis of the glycosylation of proteins is a challenge that requires orthogonal methods to achieve separation of the diverse glycoforms. A combination of reversed phase chromatography with tandem mass spectrometry (RP-LC-MS/MS) is one of the most powerful tools for glycopeptide analysis. In this work, we developed and compared RP-LC and hydrophilic interaction liquid chromatography (HILIC) in nanoscale on a chip combined with MS/MS in order to separate glycoforms of two peptides obtained from the tryptic digest of hemopexin. We observed reduction of the retention time with decreasing polarity of glycans attached to the same peptide backbone in HILIC. The opposite effect was observed for RP-LC. The presence of sialic acids prolonged the retention of glycopeptides in both chromatographic modes. The nanoHILIC method provided higher selectivity based on the composition of glycan, compared to nanoRP-LC but a lower sensitivity. The nanoHILIC method was able to partially separate linkage isomers of fucose (core and outer arm) on bi-antennary glycoform of SWPAVGDCSSALR glycopeptide, which is beneficial in the elucidation of the structure of the fucosylated glycoforms.
- Klíčová slova
- Glycoproteomics, Hemopexin, Hydrophilic interaction liquid chromatography, LC–MS/MS, Reversed phase chromatography,
- MeSH
- chemické techniky analytické metody normy MeSH
- chromatografie kapalinová * MeSH
- chromatografie s reverzní fází * MeSH
- glykopeptidy analýza MeSH
- hemopexin analýza MeSH
- hydrofobní a hydrofilní interakce MeSH
- polysacharidy chemie MeSH
- tandemová hmotnostní spektrometrie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- glykopeptidy MeSH
- hemopexin MeSH
- polysacharidy MeSH
Major differences in the chromatographic performance of a zwitterion ion-exchange type (ZWIX) chiral stationary phase (CSP) in supercritical fluid chromatography (SFC) and high-performance liquid chromatography (HPLC) have been observed. To explain these differences, transition from HPLC to SFC conditions has been performed. The amount of a protic organic modifier in supercritical carbon dioxide (scCO2) was stepwise increased and the effect of this change studied using acidic, basic and ampholytic analytes. At the same time, the effect of various basic additives to the mobile phase and transient acidic buffer species, formed by the reaction of scCO2 with the organic modifier and additives, was assessed. Evidence is provided that a transient acid together with the intrinsic counter-ions present in the ZWIX selector structure drive the elution of analytes even when no buffer is employed. We show that the tested analytes can be enantioseparated under both SFC and HPLC conditions; the best conditions for the resolution of ampholytes are in the so-called enhanced-fluidity mobile phase region. As a consequence, subcritical fluid and enhanced-fluidity mobile phase regions seem to be chromatographic modes with a high potential for operating ZWIX CSPs.
- Klíčová slova
- Amino acids, Enantioseparation, Enhanced-fluidity mobile phase, High performance liquid chromatography, Supercritical fluid chromatography, Transient acid,
- MeSH
- chemické techniky analytické metody normy MeSH
- iontová výměna MeSH
- kyseliny MeSH
- superkritická fluidní chromatografie * MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kyseliny MeSH
Two-dimensional liquid-phase separations have gained increasing attention for their ability to separate complex sample mixtures. Among the experimental setups used, an on-line approach is preferred to reduce the probability of sample contamination, for easier automation and high-sample throughput. The interfacing of the separation techniques in the on-line mode brings additional demands on proper optimization of the two-dimensional system. In this review, the possibilities of the on-line coupling of liquid chromatography and liquid chromatography with capillary electrophoresis in two-dimensional systems are discussed. Special attention is paid to the fraction transfer process, which includes an overview of interfaces and experimental setups applied, the compatibility issues of separation systems, and instrumental parameters. The benefits and drawbacks of using electromigration separations in combination with liquid chromatography are presented as well.
- Klíčová slova
- capillary electrophoresis, comprehensive liquid chromatography, fraction transfer, liquid chromatography, two-dimensional separations,
- MeSH
- chemické techniky analytické normy trendy MeSH
- chromatografie kapalinová * MeSH
- elektroforéza kapilární * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
Four phenyl-bonded stationary phases, differing in polar embedded group between spacer and phenyl ring, were used for the separation of flavonoids in reversed-phase conditions. In addition, the work was focused on the comparison of these stationary phases in terms of retention and nature of interactions between flavonoid solutes and both, mobile and stationary phases. The differences and similarities between the columns and between individual flavonoids were evaluated by a statistical analysis. The retention over the wider range of mobile phase composition was described using well known model suggested for partition chromatographic systems. Due to differences in polarity of flavonoids, gradient elution had to be applied to achieve appropriate conditions for the successful separation. A chromatographic optimization software was employed for establish the appropriate profiles of gradient separations using UV detection at 275 nm. The most appropriate conditions for the separation of flavonoids were apparent on the phenyl and phenoxy columns.
- Klíčová slova
- Flavonoid, Glycoside, Linear free energy relationship, Phenyl-bonded phases,
- MeSH
- chemické techniky analytické metody normy MeSH
- chromatografie kapalinová * MeSH
- flavonoidy analýza chemie MeSH
- indikátory a reagencie chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- flavonoidy MeSH
- indikátory a reagencie MeSH
A new CZE method was developed for the determination of 12 purine and pyrimidine nucleotides, two adenine coenzymes and their reduced forms, and acetyl coenzyme A in various cell extracts. As the concentration levels of these metabolites in living cells are low; CZE was combined with field-enhanced sample stacking. As a result, the separation conditions were optimised to achieve a suitable resolution at the relatively high sample volume provided by this on-line pre-concentration technique. The optimum BGE was 150 mM glycine buffer (pH 9.5). Samples were introduced hydrodynamically using a pressure of 35 mbar (3.5 kPa) for 25 s, and data were collected at a detection wavelength of 260 nm. An applied voltage of 30 kV (positive polarity) and capillary temperature of 25°C gave the best separation of these compounds. The optimised method was validated by determining the linearity, sensitivity and repeatability and it was successfully applied for the analysis of extracts from Paracoccus denitrificans bacteria and from stem cells.
- Klíčová slova
- Metabolomics, Nucleotides, Paracoccus denitrificans, Stem cells, System biology,
- MeSH
- acetylkoenzym A analýza MeSH
- adenosintrifosfát analýza MeSH
- chemické techniky analytické metody normy MeSH
- cytidintrifosfát analýza MeSH
- embryonální kmenové buňky chemie MeSH
- guanosintrifosfát analýza MeSH
- lidé MeSH
- limita detekce MeSH
- Paracoccus denitrificans chemie MeSH
- reprodukovatelnost výsledků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- acetylkoenzym A MeSH
- adenosintrifosfát MeSH
- cytidintrifosfát MeSH
- guanosintrifosfát MeSH
A reference material for the biological monitoring of occupational exposure to toluene, benzene and phenol was prepared. O-cresol and hippuric acid (metabolites of toluene) are used for the biological monitoring of occupational exposure to toluene. Phenol, a metabolite of benzene, is used for the biological monitoring of exposure to benzene, but phenol can of course also be used as an indicator of exposure to phenol as well. The reference material (RM) used for the determination of these metabolites was prepared by freeze-drying pooled urine samples obtained from healthy persons occupationally exposed to toluene and those taking part in an inhalation experiment. Tests for homogeneity and stability were performed by determining urine concentrations of o-cresol, hippuric acid, creatinine and phenol. To investigate the stability of the RM, the urinary concentrations of o-cresol and phenol were monitored for eighteen months using GC and HPLC, while those of hippuric acid and creatinine were followed for five and six years, respectively, using HPLC. Analysis of variance showed that the concentrations did not change. The certified concentration values (and their uncertainties) of the substances in this reference material (phenol concentration c=6.46+/-0.58 mg l(-1); o-cresol concentration c=1.17+/-0.15 mg l(-1); hippuric acid concentration c=1328+/-30 mg l(-1); creatinine concentration c=0.82+/-0.10 g l(-1)) were evaluated via the interactive statistical programme IPECA.
- MeSH
- analýza moči metody normy MeSH
- benzen analýza MeSH
- chemické techniky analytické metody normy MeSH
- fenol analýza MeSH
- hippuráty analýza MeSH
- kalibrace MeSH
- kreatin moč MeSH
- kresoly analýza MeSH
- lidé MeSH
- moč chemie MeSH
- reprodukovatelnost výsledků MeSH
- toluen moč MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- zmrazování MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- multicentrická studie MeSH
- práce podpořená grantem MeSH
- Názvy látek
- 2-cresol MeSH Prohlížeč
- benzen MeSH
- fenol MeSH
- hippuráty MeSH
- hippuric acid MeSH Prohlížeč
- kreatin MeSH
- kresoly MeSH
- toluen MeSH
Several sets of reference air filters were prepared as part of an IAEA evaluation of the performance of laboratories involved in air-pollution studies. Each set comprised three polycarbonate membrane filters, two of which were loaded with urban air particulate matter (APM) obtained in Vienna or Prague, and one unloaded filter. The filters were loaded by filtration of a suspension of the APM materials in water. The homogeneity both of bulk APM materials and of the loaded filters was evaluated and found suitable by determining several elements by instrumental neutron-activation analysis (INAA), proton-induced X-ray emission (PIXE), and micro-X-ray energy-dispersive fluorescence analysis (micro-EDXRF). After evaluation of the homogeneity, INAA, PIXE, EDXRF, atomic absorption spectrometry (AAS), inductively coupled plasma optical emission spectrometry (ICP-OES), and ICP mass spectrometry (ICP-MS) were used to characterize the filter materials and establish "target values" and their associated standard deviations for 15 elements. Problems encountered during the preparation of these unique, simulated air filters and the criteria for setting both the target values and standard deviations are presented.
- MeSH
- chemické techniky analytické přístrojové vybavení metody normy MeSH
- filtrace přístrojové vybavení MeSH
- kovy analýza MeSH
- laboratoře normy MeSH
- látky znečišťující vzduch analýza normy MeSH
- monitorování životního prostředí přístrojové vybavení metody normy MeSH
- referenční standardy * MeSH
- řízení kvality MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kovy MeSH
- látky znečišťující vzduch MeSH
Lyophilized calf uterine cytosol standards were prepared for quality control of estrogen receptor (ER) determination, and lyophilized cytosols and tissue powders were used for quality control of progesterone receptor (PR) analysis. Two series of four samples were analyzed either for ER or PR contents, twice within one month, by 7 laboratories in 5 countries. Coefficient of variation (CV) of the between-laboratory averages assayed in a single run of ER-positive (ER+) and PR-positive (PR+) standards varied from 29.6 to 61.8% and from 32.4 to 76.2%, respectively. All laboratories, with the exception of a single value, could recognize samples of low, medium, an high ER level, as well as a negative sample. Most laboratories evaluated properly also the level of PR samples. The average between-laboratory CV values of protein determination in the relevant standards were 23%.
- MeSH
- chemické techniky analytické normy MeSH
- cytosol chemie MeSH
- dextrany MeSH
- dřevěné a živočišné uhlí MeSH
- imunohistochemie MeSH
- mezinárodní spolupráce MeSH
- receptory pro estrogeny analýza MeSH
- receptory progesteronu analýza MeSH
- reprodukovatelnost výsledků MeSH
- řízení kvality MeSH
- skot MeSH
- steroidní receptory analýza MeSH
- uterus ultrastruktura MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
- Názvy látek
- dextrany MeSH
- dřevěné a živočišné uhlí MeSH
- receptory pro estrogeny MeSH
- receptory progesteronu MeSH
- steroidní receptory MeSH
