A Low Cost Gas Sampling Valve and Gc Set-Up for Analysis of Low Concentration Gas Mixtures

Abstract

A gas sampling valve suitable for on-line injection of low pressure (< 0.5 torr) gas mixtures is described. Its advantages over the commercially available valves are enumerated. An error of < 3 per cent due to non-reproducibility in sample injection was observed. A simple arrangement of column and hot wire detector together with this valve enabled quantitative separation of CO and O₂ with a lower limit of ～ 1 × 10^?7 moles for each component.     

Targeted three-dimensional liquid chromatography: a versatile tool for quantitative trace analysis in complex matrices

Targeted multidimensional liquid chromatography (MDLC), commonly referred to as 'coupled-column' or 'heartcutting', has been used extensively since the 1970s for analysis of low concentration constituents in complex biological and environmental samples. A primary benefit of adding additional dimensions of separation to conventional HPLC separations is that the additional resolving power provided by the added dimensions can greatly simplify method development for complex samples. Despite the long history of targeted MDLC, nearly all published reports involve two-dimensional methods, and very few have explored the benefits of adding a third dimension of separation. In this work we capitalize on recent advances in reversed-phase HPLC to construct a three-dimensional HPLC system for targeted analysis built on three very different reversed-phase columns. Using statistical peak overlap theory and one of the most recent models of reversed-phase selectivity we use simulations to show the potential benefit of adding a third dimension to a MDLC system. We then demonstrate this advantage experimentally by developing targeted methods for the analysis of a variety of broadly relevant molecules in different sample matrices including urban wastewater treatment effluent, human urine, and river water. We find in each case that excellent separations of the target compounds from the sample matrix are obtained using one set of very similar separation conditions for all of the target compound/sample matrix combinations, thereby significantly reducing the normally tedious method development process. A rigorous quantitative comparison of this approach to conventional 1DLC-MS/MS also shows that targeted 3DLC with UV detection is quantitatively accurate for the target compounds studied, with method detection limits in the low parts-per-trillion range of concentrations. We believe this work represents a first step toward the development of a targeted 3D analysis system that will be more effective than previous 2D separations as a tool for the rapid development of robust methods for quantitation of low concentration constituents in complex mixtures.     

Sensitive and Selective Fluorescence Detection of Phenoxyacid Herbicides by Liquid Chromatography with On-Line Ion-Pair Extraction

Abstract

Fluorescent probes, especially a newly synthesized N-substituted 1-cyanobenz[f]-isoindole quaternary ammonium fluorophore, were used as counter ions in a reaction detector for on-line ion-pair extraction of phenoxyacid herbicides. The probe was used in an on-line post-column set-up coupled to a reversed-phase chromatographic system. After separation on an C-8-bonded silica column using an aqueous methanol (pH 2.5) mobile phase, the herbicides were on-line deprotonated by post-column addition of a 10mM sodium phosphate buffer (pH 8.0), in which the probe was dissolved. Subsequently, the ion-pairs were extracted on-line with chloroform-1-butanol (80:20, v/v) and were monitored by fluorescence detection. Using this system, at least seven herbicides could be separated. The detection limits of 2,4-dichlorophe xyacetic acid and 2,4,5-trichlorophenoxyacetic acid were 400 pg (S/N = 3). The repeatability, based on peak height measurements, for 100ng injections was about 0.5%. Calibration curves were linear over the investigated range of 1–100ng, with correlation coefficients of 0.999 for the two analytes. Application to a drinking water sample is presented.     

Simultaneous Calibration of Molecular Weight Separation and Column Dispersion of GPC by Coupling with Lallis

Abstract

It is shown theoretically and experimentally that both the calibration of the molecular weight separation and column dispersion can be evaluated simultaneously by coupling gel chromatograph with low angle light scattering photometer. The experimentally determined variation of the spreading factor with retention volume is quite similar to that obtained by Tung using reverse flow technique. A correction method is given for the lowering of inhomogeneity index printed by the data processor of the on-line GPC-LALLS.     

Quantitative Analysis of Terbutaline (Bricanyl®) in Human Plasma with Liquid Chromatography and Electrochemical Detection Using On-Line Enrichment

Abstract

A liquid chromatographic method with electrochemical detection (LC-EC) has been developed for the quantitative analysis of terbuta-line in the range 5–50 pmole ml^?1 of human plasma. Terbutaline is isolated from 2 ml of plasma on an ion-exchange column and the eluate is concentrated on a hydrophobic precolumn on-line in the chromatographic system. The precolumn is then back-flushed for further separation onto a hydrophobic analytical column. The mobile phase is a methanol-aqueous buffer to which sodium perchlorate is added to achieve resolution from interfering peaks. A glassy carbon electrode is used for detection. Comparison has been made with gas chromatography-mass spectrometry (GC-MS) to examine the accuracy of the method.     

Transformation Studies of Some Polar Pesticides In Water By On-Line Solid-Phase Extraction and Liquid Chromatography/Particle Beam-Mass Spectrometry

ABSTRACT

An on-line automated method for phototransformation studies of alachlor, aldicarb, and methiocarb by means of liquid chromatography/mass spectrometry (LC/MS) with particle beam (PB) interface is described. Surface water samples were first spiked with 50 μg/1 of each pesticide and then exposed to the radiation of a medium-pressure mercury lamp. Next, in regular intervals of 60 min, aliquots of 50-ml sample were enriched on a solid-phase extraction (SPE) cartridge and on-line eluted by a gradient of LC eluent onto analytical column for separation, followed by MS detection. A phototransformation experiment with each pesticide was performed twice, with MS operated in electron ionisation (EI) and positive chemical ionisation (PCI) modes. Many transformation products (TPs) of parent pesticides were detected and in numerous instances, tentatively identified. Appearance of several transformation products is reported for the first time.     

Sb(III) and Sb(V) separation and analytical speciation by a continuous tandem on-line separation device in connection with inductively coupled plasma atomic emission spectrometry

A sensitive, precise and automated non-chromatographic method for Sb(III) and Sb(V) analytical speciation based on a continuous tandem on-line separation device in connection with inductively coupled plasma-atomic emission (ICP-AES) detection is proposed. Two on-line successive separation steps are included into this method: a continuous liquid-liquid extraction of Sb(III) with ammonium pyrrolidine dithiocarbamate (APDC) into methylisobuthylketone (MIBK), followed by direct stibine generation from the organic phase. Both separation steps are carried out in a continuous mode and on-line with the ICP-AES detector. Optimization of experimental conditions for the tandem separation and ICP-AES detection are investigated in detail. Detection limits for Sb(III) were 3 ng.mL^–1 and for Sb(V) 8 ng.mL^–1. Precisions observed are in the range ± 5%. The proposed methodology has been applied to Sb(III) and Sb(V) speciation in sea-water samples.     

Reversed-Phase Ion-Pairing Liquid Chromatographic Separation and Fluorimetric Detection of Polyamines

Abstract

A rapid and specific reversed-phase ion-pairing high performance liquid chromatographic procedure for putrescine, spermidine and spermine is reported. The ion-pairing reagent, heptanesulfonate, was employed and o-phthalaldehyde and 2-mercaptoethanol were used for on-line post-column derivatization and subsequent fluorescence detection. Experiments were carried out to determine the effects of several variables such as pH, concentration of the aqueous buffer, counter-ion concentration, and the percentage of organic modifier in the moving phase. The minimum detection limits for the polyamines ranged from 120 pmoles for spermine to 12 pmoles for putrescine. The method includes a gradient program which provides complete separation from amino acids and specificity for the three polyamines. The procedure was applied successfully to urine and serum samples.     

Rapid and Complete Separation of Inorganic Polyphosphates from Monomer to 35-Mer by Hplc

Abstract

The on-line coupling system of high-performance liquid chromatography (HPLC) and flow injection analysis (FIA) was developed for the rapid separation and sensitive detection of inorganic polyphosphates. Effects of gradient elution conditions on the chromatographic behavior were studied. The column temperature as well as a chloride concentration gradient were found to be very effective for the improvement of resolution(1). To minimize analysis time and to maximize resolution, the gradient elution conditions were optimized by use of a computer-assisted retention prediction system(2). More than 35 kinds of inorganic polyphosphates could be separated completely within 200 min under the optimum conditions as shown in Fig. 1.     

Identification of Natural and Early Synthetic Textile Dyes with HPLC and UV/Vis-Spectroscopy by Diode Array Detection

Abstract

The separation and identification of complex mixtures of natural and synthetic textile dyes was investigated using HPLC with diode array detection. Separation was carried out on a reversed phase column with acetonitrile-phosphoric acid gradient elution. The results show that the anthraquinones from madder root and the insect dye cocheneal present in ancient red dyes can easily be distinguished from azo-dyes present in later textile fibres. They further show that for an analysis of the numerous flavones and flavonoles, constituting most of the yellow natural dyestuffs, the combination of HPLC with on-line optical spectroscopy is particularly useful. Even when retention times are identical - as for instance for quercetin and luteolin - the ratio of a mixture can still be estimated by evaluating the ratio-chromatogram.     

On-line concentration by field-enhanced sample injection with reverse migrating micelles in micellar electrokinetic capillary chromatography for the analysis of triterpenoids from traditional chinese medicine

In this work, a simple, reproducible and sensitive micellar electrokinetic chromatography (MEKC) method was developed for the separation and determination of five triterpenoids, lupeol (1), 1β-hydroxy-lupeol (2), lup-3β,1α-diol (3), lup-1β,3β,11α-triol (4) and 30-norlupan-3β,11α-diol-20-one (5) in traditional Chinese medicine of Salvia roborowskii Maxim. Field-enhanced sample injection with reverse migrating micelles (FESI-RMM) was used for on-line concentration of triterpenoids. The optimum buffer contained 50 mM H₃PO₄, 160 mM SDS, 20% acetonitrile and 15% 2-propanol and pH of buffer was 2.0. The sample solution was diluted with 10 mM H₃PO₄ (pH 2.5, containing 10 mM SDS) and injected for 15 s with −8 kV after injection of 4 s water plug. The effects of concentrations of sodium dodecyl sulfate (SDS) and organic modifier, the sample matrix, the injection time of water plug, the injection voltage and injection time of sample on the separation and stacking efficiency were investigated. Under the optimum conditions, the analytes were well separated and by optimizing the stacking conditions, about 28–96-fold improvement in the detection sensitivity was obtained for triterpenoids. The contents of five triterpenoids in Salvia roborowskii Maxim were successfully determined with satisfactory repeatability and recovery.     

Anwendungsbeispiele der direkten HPLC-NMR-Kopplung mit hoher Aufl?sung

Zusammenfassung Durch HPLC-NMR on-line Kopplung ist eine gleichzeitige Trennung und Identifizierung von Komponenten eines Substanzgemisches möglich. Trennbeispiele von Aromaten und Steroiden zeigen die mögliche Anwendungsbreite der Methode durch Messung im Durchfluß-bzw. stop-flow-Verfahren. Durch Verwendung einer mathematischen Filter-Funktion kann annähernd die Qualität von herkömmlichen Routine-Spektren erzielt werden.

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Examples of application of direct HPLC-NMR-coupling with high resolution

Summary With the direct on-line coupling of HPLC-NMR, simultaneous separation and identification in the liquid phase of components in a mixture is possible. Examples carried out on some aromatic compounds as well as steroids show the wide scope of application of this method with either a continuous flow or a stop flow technique. A mathematical resolution enhancement technique is utilized to improve the on-line NMR spectra substantially.

     

Comparison of alpha1-adrenergic receptor cell-membrane stationary phases prepared from expressed cell line and from rabbit hepatocytes

A G-protein-coupled receptor-cell-membrane stationary phase (CMSP) has been prepared by immobilizing cell membranes on the surface of silica, as carrier. The resulting HEK293 α _1A adrenoceptor cell-membrane stationary phase can be used for rapid on-line chromatographic determination of potential subtype-selective α ₁ -adrenoceptor ligand-binding affinities for α ₁ -adrenoceptor subtypes. The objective of the research was to study whether cell lines stably overexpressing subtype receptors could improve the sensitivity and specificity of cell-membrane chromatography (CMC) compared with use of homogenized tissue and cells in primary culture. Effects of mobile-phase ionic strength, sample concentration, and the presence of competitive agents on ligand-receptor interaction in CMSP were also evaluated. We found that cell lines stably overexpressing subtype receptors led to improved sensitivity and specificity in CMC. The technique leads to useful procedures-cell-membrane stationary phases may, for example, facilitate exploration of ligand-receptor interaction and determination of ligand-receptor binding affinity in initial screening and separation of lead compounds or active components in Chinese traditional natural medicine and herbs. This might eventually be an important contribution and an addition to our collection of techniques.     

Tandem on-line continuous separation and determination of arsenic by inductively coupled plasma atomic emission spectrometry

The principle of tandem on-line continuous separation techniques as an alternative means of introducing samples into plasmas was applied to the development of a sensitive, selective and convenient method for the determination of arsenic by inductively coupled plasma atomic emission spectrometry (ICP-AES). Arsenic is continuously extracted as AsI₃ into xylene from the sample dissolved in 0.1 M potassium iodide solution in 7.2 M hydrochloric acid. The xylene phase (containing the analyte) is continuously mixed on-line with NaBH₄ in dimethylformamide and acetic acid solutions. Arsine is thus continuously generated directly from the organic phase and is separated in a gas—liquid separation device which prevents most of the xylene phase vapour from reaching the ICP. The system was optimized for the continuous extraction of AsI₃, the direct generation of arsine from xylene and the final ICP determination of arsenic. Finally, the tandem on-line continuous separation ICP detection system was applied to the determination of arsenic in real samples (white metal, cast iron, cupro-nickel and orchard leaves standard materials). Very good agreement between the experimental results and the certified values was obtained.     

The Application of UV-Radioactivity High Performance Liquid Chromatography to the Study of Hypoxanthine Transport in Human Erythrocytes

Abstract

A procedure is presented for the simultaneous measurement of concentrations of labeled and non labeled hypoxanthine by HPLC in order to study hypoxanthine transport in erythrocytes. A radioactivity detector connected on-line to the high performance liquid chromatograph in series with a UV detector provides on-line quantitative monitoring of hypoxanthine in erythrocytes or incubation medium. The procedure provides a rapid, sensitive and convenient means for the study of hypoxanthine transport.     

Characterization and application of SuperLig<Superscript>®</Superscript> 620 solid phase extraction resin for automated process monitoring of <Superscript>90</Superscript>Sr

Characterization of SuperLig^® 620 solid phase extraction resin was performed in order to develop an automated on-line process monitor for ⁹⁰Sr. The main focus was on strontium separation from barium, with the goal of developing an automated separation process for ⁹⁰Sr in high-level wastes. High-level waste contains significant ¹³⁷Cs activity, of which ^137mBa is of great concern as an interference to the quantification of strontium. In addition barium, yttrium and plutonium were studied as potential interferences to strontium uptake and detection. A number of complexants were studied in a series of batch K_d experiments, as SuperLig^® 620 was not previously known to elute strontium in typical mineral acids. The optimal separation was found using a 2 M nitric acid load solution with a strontium elution step of ~0.49 M ammonium citrate and a barium elution step of ~1.8 M ammonium citrate. ⁹⁰Sr quantification of Hanford high-level tank waste was performed on a sequential injection analysis microfluidics system coupled to a flow-cell detector. The results of the on-line procedure are compared to standard radiochemical techniques in this paper.     

Automation of radiochemical analysis by flow injection techniques: Am-Pu separation using TRU-resin™ sorbent extraction column

A rapid automated flow injection analysis (FIA) procedure was developed for efficient separation of Am and Pu from each other and from interfering matrix and radionuclide components using a TRU-resin^™column. Selective Pu elution is enabled via on-column reduction. The separation was developed using on-line radioactivity detection. After the separation had been developed, fraction collection was used to obtain the separated fractions. In this manner, a FIA instrument functions as an automated separation workstation capable of unattended operation.     

Comprehensive two-dimensional liquid chromatography coupled to the ABTS radical scavenging assay: a powerful method for the analysis of phenolic antioxidants

The on-line combination of comprehensive two-dimensional liquid chromatography (LC?×?LC) with the 2,2′-azino-bis(3-ethylbenzothiazoline)-6 sulphonic acid (ABTS) radical scavenging assay was investigated as a powerful method to determine the free radical scavenging activities of individual phenolics in natural products. The combination of hydrophilic interaction chromatography (HILIC) separation according to polarity and reversed-phase liquid chromatography (RP-LC) separation according to hydrophobicity is shown to provide much higher resolving power than one-dimensional separations, which, combined with on-line ABTS detection, allows the detailed characterisation of antioxidants in complex samples. Careful optimisation of the ABTS reaction conditions was required to maintain the chromatographic separation in the antioxidant detection process. Both on-line and off-line HILIC?×?RP-LC–ABTS methods were developed, with the former offering higher throughput and the latter higher resolution. Even for the fast analyses used in the second dimension of on-line HILIC?×?RP-LC, good performance for the ABTS assay was obtained. The combination of LC?×?LC separation with an on-line radical scavenging assay increases the likelihood of identifying individual radical scavenging species compared to conventional LC–ABTS assays. The applicability of the approach was demonstrated for cocoa, red grape seed and green tea phenolics.

Flow Injection On-Line Two-Stage Solvent Extraction Preconcentration Coupled with Et-Aas for Determination of Bismuth in Biological and Environmental Samples

ABSTRACT

A robust flow injection/sequential injection(FIA/SIA) on-line two-stage solvent extraction separation/preconcentration procedure coupled to electrothermal atomic absorption spectrometry (ETAAS) is described and demonstrated for the determination of trace-levels of bismuth. In 1-4% (v/v) nitric acid medium, diethyldithiophosphate (DDPA) chelates bismuth and the chelating complex formed is extracted into isobutyl methyl ketone (IBMK), which is separated from the aqueous phase by means of cavity gravitational phase separation. Both single and two-stage separations are used. In each case, 40 μl of the concentrate is entrapped and metered in a loop, and subsequently delivered via air-segmentation into a pyrolytically coated graphite tube. The ETAAS determination is performed in parallel with the preconcentration process of the ensuing sample. Enrichment factors of 17.8 and 31.5, detection limits of 12 ng 1^?1 and 10.5 ng 1^?1 for single and two-stage extractions, respectively, along with a sampling frequency of 13 h^?1 were obtained with 100 s of sample loading time at a sample flow rate of 5.4 ml min^?1. The relative standard deviations were 2.8% and 1.9% for single and two-stage extractions, respectively. The procedure was validated by determining the bismuth content in a certified reference material and i8543220 human urine samples.     

On-line screening and identification of polyphenolic antioxidant compounds of Convolvulus trabutianus

Abstract

Convolvulus trabutianus Schweinf. & Muschl. is an endemic plant from northern Sahara used in folk medicine. Herein we report, the isolation, characterization and evaluation of the radical scavenging properties of twenty three compounds from different extracts of this species by on-line HPLC-ABTS^?+ screening. These compounds include nine phenolic acids: 2, 6, 10–16, two phytosterols: 3–4, four coumarins: 5, 7–9, two quinic acids: 21 and 22 and six flavonoids: 1, 17–20 and 23 among which the most active were: 10, 16, 21 and 22. All the extracts showed a significant antioxidant activity on-line. These results were validated off-line by ORAC and TEAC assays. Four compounds: 1, 5, 18 and 19 were described for the first time from the Convolvulaceae family, whereas compounds 2, 6, 8, 10, 13 and 21 were new for the genus Convolvulus.