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At least 1,207 records · Page 67Linked to original sources
PubMed2000

Trace analysis of estrogenic chemicals in sewage effluent using liquid chromatography combined with tandem mass spectrometry.

A rapid, selective, sensitive, and reproducible method for the analysis of environmental estrogens, either natural or synthetic, present in samples from sewage treatment works (STW) has been developed. Isolation of these compounds from STWs was performed by applying a simple extraction procedure using an ENVI-CARB cartridge (a graphitized non porous carbon black) as the solid-phase extraction (SPE) system. Liquid chromatography coupled with atmospheric pressure chemical ionization (APCI) and tandem mass spectrometry was used for detection. For the multiple reaction monitoring (MRM) mode, the [M + H](+) ion of estrone and the [M + H-H(2)O](+) ions of 17beta-estradiol, estriol and 17alpha-ethynylestradiol were selected as the precursors for collisionally induced dissociation (CID). The average recoveries from sewage final effluent samples ranged from 84 to 93% for low levels, and from 89 to 95% for high levels. The precision of the method ranged from 11 to 8% for low level and from 9 to 7% for high level samples. The lower level of quantitation for these estrogens in STW samples was determined at 0.5 ng/L for 17beta-estradiol and 17alpha-ethynylestradiol, and 1 ng/L for estrone and estriol, based on 1-L aliquots of sewage treatment works water, using the optimum tuning parameters for each individual selected precursor ion/product ion transition. Compared to a previous gas chromatography/mass spectrometry (GC/MS) method for the analysis of ten STW samples, this method was shown to provide higher sensitivity and lower time consumption.

Calibration↗
PubMed2006

Sequential injection analysis with chemiluminescence detection for the antioxidative activity against singlet oxygen.

A sequential injection analysis (SIA) with chemiluminescence (CL) detection was developed for the measurement of antioxidative activity against singlet oxygen ((1)O2). Lactoperoxidase-hydrogen peroxide-bromide ion system was used for the generation of (1)O2. When a 100 mM sodium acetate buffer (pH 4.5) was used as a carrier solution, the SIA-CL system could be optimized with respect to the flow-rate of the carrier, concentration of reagents and their aspiration order. The antioxidative activity was expressed as an attenuation of luminol CL due to the quenching of (1)O2 by an antioxidant. The relative standard deviations of antioxidative activity (n=3) against (1)O2 for within- and between-day analyses were < or = 1.6% (20 microM Trolox). The system was successfully applied to the assay of antioxidative activities of various antioxidants including vitamin supplements at a rate of 10 samples within 15 min. The proposed SIA-CL system was rapid and reproducible with minimum consumption of the sample and of reagents, and thus was useful for the screening of compounds possessing antioxidative activity against (1)O2.

Antioxidants↗
PubMed2004

Determination of atractyloside in Callilepis laureola using solid-phase extraction and liquid chromatography-atmospheric pressure ionisation mass spectrometry.

A selective analytical method based on high-performance liquid chromatography, combined with atmospheric pressure ionisation mass spectrometry, was developed for the detection of atractyloside. The analysis was performed on an Xterra Phenyl column utilising a gradient elution profile and a mobile phase consisting of 10 mM aqueous ammonium acetate buffer-methanol-acetonitrile. The calibration curve of the method (1 ng/ml-160 microg/ml) was best described by a second order polynomial function (r2 = 0.998) but displayed good linearity in the range of 100 ng/ml-1 microg/ml (r2 = 0.999). The limit of detection for the atractyloside standard was determined and found to be 100 pg/ml and the limit of quantification of atractyloside in tuber matrix was found to be 250 pg/ml. The relative standard deviation of the method was on average below 5% (n = 8). The method was successfully applied to the analysis of Callilepis laureola tubers and unknown powdered samples for the presence of atractyloside.

Atmospheric Pressure↗
PubMed2005

Determination of cocaine and benzoylecgonine by direct injection of human urine into a column-switching liquid chromatography system with diode-array detection.

A method for the determination of cocaine (COC) and benzoylecgonine (BZE) in human urine using a column-switching liquid chromatography system is reported. A homemade precolumn (20 mm x 4.6 mm i.d.) dry-packed with Alltech ODS-C18 (35-750 microm) was employed as an extraction precolumn in order to extract and concentrate the COC and BZE from the human urine sample. The analytes were continuously transferred to the analytical column (Spherisorb-C8, 250 mm x 4.6 mm i.d.; dp = 5 microm) by means of the switching arrangement in the backflush mode. Detection was carried out at 235 nm in a UV-diode array detector. The validation of the method revealed analytes quantitative recoveries (96-102%) at three concentrations in the range from 0.25 to 4.00 and from 0.5 to 12.0 microg/mL for COC and BZE, respectively. These values demonstrate the excellent extraction efficiency of the precolumn. The detection limits for COC and BZE at a signal-to-noise ratio of 3 were 0.08 and 0.15 microg/mL when a sample volume of 50 microL was injected. The overlap of sample preparation, analysis and recondition of the precolumn increases the sample throughput to four samples per hour. The proposed method has been applied to the determination of COC and BZE in human urine samples from 73 suspecting drug addicts. Urine concentrations of 1.0-118.10 microg of BZE/mL and 0.1-41.0 microg of COC/mL were found.

Chromatography, Liquid↗
PubMed1992

Direct injection analysis of carbamazepine and its active 10,11-epoxide metabolite in plasma by use of a semipermeable surface (SPS) silica column in LC.

A semipermeable surface (SPS) silica column was applied for the simultaneous determination of carbamazepine (CBZ) and its active 10,11-epoxide metabolite (EPO) in plasma following direct injection in LC. The SPS packing material consists of an ODS ligand as the hydrophobic inner phase and a polyoxyethylene network as the hydrophilic outer phase. When a 5-microliters portion of intact plasma was injected onto the column using a mobile phase of phosphate buffer (pH 7.1, ionic strength 0.1)-acetonitrile (4:1, v/v), the plasma proteins were size-excluded, whereas the drug and its metabolite were retained and separated both from each other and from other commonly co-administered drugs such as phenobarbital (PB) and phenytoin (DPH). The calibration graphs (peak area vs concentration) of CBZ, EPO and PB were linear over the therapeutic range of plasma concentration (r greater than 0.998) with good relative standard deviations (RSD less than 3.98%, n = 5). The recoveries from plasma were almost complete (greater than 96.6%). The analysis time was 17 min. The method as developed was applied in studies on the time course of plasma concentrations of unchanged CBZ and EPO after i.v. administration of CBZ to the rat.

Calibration↗
PubMed2003

Application of the 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation assay to a flow injection system for the evaluation of antioxidant activity of some pure compounds and beverages.

The 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation (ABTS(*)(+)) assay was adapted to a flow injection (FI) system to obtain a sensitive and rapid technique for the monitoring of antioxidant activity of pure compounds and complex matrixes, such as beverages and food extracts. The FI system includes a HPLC pump that flows the mobile phase (a solution of ABTS(*)(+) in ethanol) through a 20 microL loop injector, a single bead string reactor filled with acid-washed silanized beads, a delay coil and a photodiode array UV-visible detector. The technique was very sensitive, with limits of detection and of quantification of 4.14 and 9.29 micromol of Trolox/L, respectively, and demonstrated high repeatability and reproducibility. The proposed technique was then applied to the evaluation of the antioxidant activity of some pure compounds, demonstrating good agreement with published data obtained by the original spectrophotometric ABTS(*)(+) assay. Finally, the total antioxidant activity of 10 beverages was determined by both the proposed and the original method. The values ranged from 0.09 mmol L(-)(1) for cola to 49.24 mmol L(-)(1) for espresso coffee and did not result significantly different from those obtained by the original spectrophotometric ABTS(*)(+) assay (Student's paired t-test: t = 1.4074, p = 0.1929). In conclusion, the proposed FI technique seems suitable for the direct, rapid and reliable monitoring of total antioxidant activity of pure compounds and beverages and, due to the ability to operate in continuous, it allows the analysis of about 30 samples h(-)(1) making the assay particularly suitable for large screening of total antioxidant activity in food samples.

Antioxidants↗
PubMed2004

Ion selective electrodes for penicillin-G based on Mn(III)TPP-Cl and their application in pharmaceutical formulations control by sequential injection analysis.

The work describes the construction, evaluation and analytical application of ion selective electrodes sensitive to penicillin-G antibiotics for pharmaceutical products analysis. Different types of polymeric membranes based on PVC (poly(vinyl chloride)) and EVA (ethyl-vinyl-acetate), without internal reference solution, were prepared using 5,10,15,20-tetraphenylporphyrinate (TPP) manganese(III) (Mn(III)TPP-Cl) as electroactive material. Different additives such as tetra-n-octylammoniumbromide (cationic additive) and sodium tetraphenylborate (anionic additive) were incorporated into the membranes to evaluate their influence on electrodes performance. The comparison of the developed detectors was based on general analytical characteristics, selectivity and lifetime. To accomplish the analysis of real samples, two selective membranes composed of 33.0% (w/w) of PVC, 66.0% (w/w) of o-NPOE and 1.0% (w/w) of Mn(III)TPP-Cl (type A) and 33.0% (w/w) of PVC, 66.0% (w/w) of o-NPOE, 1.0% (w/w) of Mn(III)TPP-Cl and 10% mol (relative to the molar concentration of Mn(III)TPP-Cl) of sodium tetraphenylborate (type B) were used. Type A electrode presented a linear response between 2 x 10(-5) and 10(-1) moll(-1) for penicillin-G, a slope of about -59 mVdec(-1) and a reproducibility of about +/-0.5 mVday(-1), while type B exhibited a linear response between 5 x 10(-5) and 10(-1) moll(-1) for penicillin-G, a slope of about -61 mVdec(-1) and a reproducibility of about +/-0.3 mV day(-1). The potentiometric analysis of penicillin-G in pharmaceutical products was carried out by direct potentiometry and the results obtained were compared with those provided by the HPLC reference method. These membranes (type A and type B) were used to prepare tubular electrodes that were coupled to a sequential injection system (SIA) and presented a linear range between 2 x 10(-4) and 1 x 10(-2) moll(-1) and slopes of -59.3 +/- 0.8 and -57.3 +/- 1.2 mVdec(-1), respectively. The tubular electrode constructed using type B membrane (type TB) was used to carry out the potentiometric analysis of penicillin-G in pharmaceutical formulations. The proposed procedure enabled relative errors between 0.1% and 1.2% (n = 4) and a sampling-rate of about 25 samples per h.

Chemistry, Pharmaceutical↗
PubMed2004

On-line assay of the S-enantiomers of enalapril, ramipril and pentopril using a sequential injection analysis/amperometric biosensor system.

A sequential injection analysis/amperometric biosensor system is proposed for the enantioselective analysis of the S-enantiomer of enalapril, ramipril and pentopril. The amperometric biosensor used as detector in the sequential injection analysis was designed by immobilization of l-amino acid oxidase in carbon paste. The proposed SIA system can be utilized reliably for the enantioanalysis of the S-enantiomer from the raw materials as well as from their pharmaceutical formulations, with a rate of 75 samples per hour and R.S.D. values better than 0.1% (n = 10).

Biosensing Techniques↗
PubMed2005

Small-volume rapid-mix device for subsecond kinetic analysis in flow cytometry.

BACKGROUND: Rapid-mix flow cytometry has emerged as a powerful tool for mechanistic analysis of ligand binding, cell response, and molecular assembly. Although progress has come from improving sample delivery capabilities, little attention has been paid to the volumetric requirements associated with precious biological reagents. METHODS: By using programmable syringes, valves, and other fluidic components, we created a modular, precisely regulated rapid-mix device for the delivery of small-volume samples to the flow cytometer. The device was tested using a bead-based assay in which the binding kinetics between native biotin and fluorescein biotin-bearing beads were characterized. RESULTS: Bead suspensions and reagents paired in 35- to 45-microl aliquots were efficiently mixed by the device and delivered to the flow cytometer. Kinetic data associated with the fluorescein biotin beads were analyzed and used to calibrate the performance characteristics of the device in terms of sample delivery and mixing efficiency. CONCLUSION: The rapid-mix device is capable of detecting subsecond kinetics of biological reactions using microliter volume of samples. Dimensions of the device have been minimized, and the quantitative aspects of sample delivery and analysis have been optimized. Further, the modular design has been optimized for adaptation to a variety of experimental protocols.

Biotin↗
PubMed2001

Enzyme microgels in packed-bed bioreactors with downstream amperometric detection using microfabricated interdigitated microsensor electrode arrays.

In this article, we describe the use of pH- responsive hydrogels as matrices for the immobilization of two enzymes, glucose oxidase (GOx) and glutamate oxidase (GlutOx). Spherical hydrogel beads were prepared by inverse suspension polymerization and the enzymes were immobilized by either physical entrapment or covalent immobilization within or on the hydrogel surface. Packed-bed bioreactors were prepared containing the bioactive hydrogels and these incorporated into flow injection (FI) systems for the quantitation of glucose and monosodium glutamate (MSG) respectively. The FI amperometric detector comprised a microfabricated interdigitated array within a thin-layer flow cell. For the FI manifold incorporating immobilized GOx, glucose response curves were found to be linear over the concentration range 1.8-280 mg dL(-1) (0.1-15.5 mM) with a detection limit of 1.4 mg dL(-1) (0.08 mM). Up to 20 samples can be manually analyzed per hour, with the hydrogel-GOx bioreactor exhibiting good within-day (0.19%) precision. The optimized FI manifold for MSG quantitation yielded a linear response range of up to 135 mg dL(-1) (8 mM) with a detection limit of 3.38 mg dL(-1) (0.2 mM) and a throughput of 30 samples h(-1). Analysis of commercially produced soup samples gave a within-day precision of 3.6%. Bioreactors containing these two physically entrapped enzymes retained > 60% of their initial activities after a storage period of up to 1 year.

Aspergillus niger↗
PubMed1993

Direct injection analysis of atenolol enantiomers in plasma using an achiral/chiral coupled column HPLC system.

A novel HPLC system was developed with an achiral/chiral coupled column for the direct injection analysis of atenolol (AT) enantiomers in plasma. The system consists of a size-exclusion column, an ODS silica column and a newly developed beta-cyclodextrin perphenylcarbamate(ph-beta-CD)-bonded silica column connected in a series via two switching valves. The neat plasma sample was directly injected onto the size-exclusion column, and the deproteinized fraction of AT enantiomers was concentrated on the ODS silica column. The enantiomers were then transferred and separated mutually on the ph-beta-CD silica column. The calibration line for each of the AT enantiomers was linear in the range of plasma concentration of 10-200 ng/ml (r > 0.9997) with good reproducibility (CV < 9.7%, n = 20). The recoveries from plasma were almost complete (> 97.4%) for both enantiomers. One analysis finished within 30 min. The developed system was applied to the enantioselective determination of plasma concentration-time curve of AT after the oral administration of racemic AT to a healthy volunteer.

Atenolol↗
PubMed2003

Determination, by dynamic surface-tension analysis, of the molar mass of proteins denatured in guanidine thiocyanate.

A drop-based dynamic surface-tension detector (DSTD) has been used to study the dynamic surface tension behavior of proteins denatured in guanidine thiocyanate (GndSCN). The dynamic surface tension at the air-liquid interface is obtained by measuring the internal pressure of drops that grow and detach at a specified rate. In the method the sample of interest is injected and subsequently flows to the DSTD-sensing capillary tip. For this work, a novel DSTD calibration procedure utilizing two distinct mobile phases is applied. Here, the mobile phases are aqueous with different constituents, for example GndSCN and phosphate buffer, either added or omitted. The dual-mobile phase calibration procedure gives the analyst the capability of making protein measurements in a GndSCN-phosphate buffer mobile phase, while measuring a calibration standard in another mobile phase, such as water, in which the surface tension of the calibration standard is readily available. Results are presented with drop volumes of either 2 microL (i.e. 2-s drops) or 7 microL (i.e. 7-s drops) for proteins varying in molar mass from 12,000 to 330,000 g mol(-1). We demonstrate that the DSTD can be used to determine the molar mass of proteins denatured in GndSCN. The method applies a regime where the denatured protein is detected by surface-active properties, and selectivity with regard to molar mass is contained in the dynamic component of the DSTD signal. The dynamic surface pressure signals of the denatured proteins suggest that diffusion plays a large role in the kinetics of the surface activity. The limit of detection for the denatured proteins studied ranged from 3 mg L(-1) to 14 mg L(-1). The DSTD, coupled with the novel dual-mobile phase calibration procedure, can be used to investigate the fundamental properties of proteins. Insight into the behavior at the air-liquid interface for native and denatured proteins is achieved; this is a novel tool for studying protein denaturation, complementary to other common approaches such as spectroscopy and calorimetry. Furthermore, the reported method could be widely applied to the study of effects on the interfacial properties of proteins after a variety of chemical and physical modifications that are possible with the dual-mobile phase calibration procedure.

Calibration↗
PubMed2000

On the kinetics of distamycin binding to its target sites on duplex DNA.

Distamycin A is a well known polyamide antibiotic that can bind in the minor groove of duplex DNA primarily at AT-rich sequences both as a monomer or as a side-by-side antiparallel dimer. The association phase of the distamycin binding reaction has not been studied in either of its binding modes, because of the lack of an adequate UV or CD signal at the low concentrations needed to monitor the fast bimolecular reaction. We report a significant increase in fluorescence amplitude, accompanied by a small red shift, on binding distamycin to its specific target sites. This signal can be used to monitor drug binding in steady-state and time-resolved processes. Distamycin shows extremely fast association with the 1:1 binding site, with a bimolecular rate of 7 x 10(7) M(-1) small middle dots(-1) and also fairly rapid dissociation ( approximately 3 s(-1)). When DNA is in excess, there is a slow component in the association reaction whose rate decreases strongly with increasing DNA concentration. Binding of the drug to the 2:1 site occurs in two distinct steps: fast, sequential binding of each drug molecule to the DNA with a bimolecular rate comparable to that at the 1:1 site, followed by a slow ( approximately 4 s(-1)) equilibration to the final population. Dissociation from the 2:1 site is approximately 40-fold slower than from the 1:1 site. This study provides the groundwork for analysis of the binding kinetics of longer polyamides and covalently linked polyamides that have recently been shown to inhibit transcription in vivo.

Anti-Bacterial Agents↗
PubMed1999

Sequential injection technique for automated titration: spectrophotometric assay of vitamin C in pharmaceutical products using cerium(IV) in sulfuric acid.

For the first time sequential injection analysis (SIA) technique has been employed for titrimetry. A new SI titrimetric spectrophotometric method for the assay of vitamin C in drug formulations was explored. The method is based on the oxidation reaction of vitamin C with cerium(IV) in sulfuric acid media using a spectrophotometer as a detector with the wavelength monitored at 410 nm. A 2(3) factorial design chemometric approach was employed to study the interaction effect of the chemical and system variables, mainly cerium(IV), sulfuric acid concentrations and the flow rate. The results of the chemometric optimization revealed that the optimum operating conditions for the SI titrimetric analysis of vitamin C were 7.0 x 10(-3) M cerium(IV), 0.455 M sulfuric acid and 28.9 microL s-1 flow rate. A linear calibration plot for the determination of vitamin C was obtained in the concentration range between 30 to 200 ppm. The method was applied to the determination of vitamin C in pharmaceutical preparations and no excipient was found to pose any interference, thus rendering the method suitable for the determination of the drug in pharmaceutical preparations. The SIA method is found to be accurate when the results were statistically compared with the results obtained by the BP standard method. The SIA method is superior when compared to the conventional titration method, the BP standard method and previous methods with respect to precision and automation in solution handling.

Ascorbic Acid↗
PubMed2002

Determination of yeast assimilable nitrogen content in wine fermentations by sequential injection analysis with spectrophotometric detetection.

An automated method for measuring the primary amino acid concentration in wine fermentations by sequential injection analysis with spectrophotometric detection was developed. Isoindole-derivatives from the primary amino acid were formed by reaction with o-phthaldialdehyde and N-acetyl- L-cysteine and measured at 334 nm with respect to a baseline point at 700 nm to compensate the observed Schlieren effect. As the reaction kinetic was strongly matrix dependent the analytical readout at the final reaction equilibrium has been evaluated. Therefore four parallel reaction coils were included in the flow system to be capable of processing four samples simultaneously. Using isoleucine as the representative primary amino acid in wine fermentations a linear calibration curve from 2 to 10 mM isoleucine, corresponding to 28 to 140 mg nitrogen/L (N/L) was obtained. The coefficient of variation of the method was 1.5% at a throughput of 12 samples per hour. The developed method was successfully used to monitor two wine fermentations during alcoholic fermentation. The results were in agreement with an external reference method based on high performance liquid chromatography. A mean-t-test showed no significant differences between the two methods at a confidence level of 95%.

Amino Acids↗
PubMed2001

Reagentless chemiluminescence flow sensor for the determination of riboflavin in pharmaceutical preparations and human urine.

A novel continuous-flow sensor based on chemiluminescence (CL) detection was developed for the determination of riboflavin at pg ml(-1) levels by the immobilization of the reagents. It was found that the CL intensity from the oxidation between luminol and periodate could be enhanced in the presence of riboflavin. The increase of CL emission was correlated with the riboflavin concentration in the range from 0.04 to 200 ng ml(-1), and the detection limit was 0.02 ng ml(-1) (3s). Considering the effective reaction ions, luminol and IO4- was immobilized on anion-exchange resin. The system could produce an evident CL signal by water as eluant and it was also shown that the flow sensor could greatly improve the selectivity and sensitivity for determination of riboflavin with a high signal-to-noise ratio. A complete analysis, including sampling and washing, could be performed in 0.5 min with a relative standard deviation of less than 3.0%. The flow sensor was applied successfully to the determination of riboflavin in pharmaceutical preparations and human urine samples.

Flow Injection Analysis↗
PubMed2006

Fast BIA-amperometric determination of isoniazid in tablets.

This paper proposes a new, fast and precise method to analyze isoniazid based on the electrochemical oxidation of the analyte at a glassy carbon electrode in 0.1M NaOH. The quantification was performed utilizing amperometry associated with batch injection analysis (BIA) technique. Fast sequential analysis (60 determinations h(-1)) in an unusually wide linear dynamic range (from 2.5 x 10(-8) to 1.0 x 10(-3)M), with high sensitivity and low limits of detection (4.1 x 10(-9)M) and quantification (1.4 x 10(-8)M), was achieved. Such characteristics allied to a good repeatability of the current responses (relative standard deviation of 0.79% for 30 measurements), were explored for the specific determination of isoniazid in isoniazid-rifampin tablet.

Electrochemistry↗
PubMed2006

Sequential injection analysis system for the sandwich hybridization-based detection of nucleic acids.

A sequential injection analysis lab-on-valve (SIA-LOV) system was developed for the specific detection of single-stranded nucleic acid sequences via sandwich hybridization of specific DNA probes to the target sequence. One DNA probe was tagged with fluorescein; the other was biotinylated and immobilized to streptavidin-coated porous beads. The system was optimized with respect to buffer composition, length of hybridization and wash steps, and volumes and concentrations of components used. On-bead oligonucleotide hybridization was studied using UV detection at 260 nm, while a final dose response curve was quantified using fluorescence detection. A dynamic range of 1-1000 pmol was obtained for a synthetic DNA sequence that was homologous to a segment in the B. anthracis atxA mRNA. A within-day variation of 7.2% and a day-to-day variation of 9.9% was observed. Each analysis was completed within 20 min. Subsequently, the system was applied to the detection of atxA mRNA expressed in a surrogate organism and amplified using NASBA. The SIA-LOV will find its application in routine laboratory-based analysis of specific single-stranded DNA/RNA sequences. Future improvements will include the integration of dye-encapsulating liposomes for signal enhancement used in lieu of the single fluorophore-labeled probe in order to lower the limit of detection.

Biosensing Techniques↗