A simple and rapid in situ preconcentration method for trace ammonia nitrogen in environmental water samples using a solid-phase extraction followed by spectrophotometric determination.

A simple and rapid in situ preconcentration method for the spectrophotometric determination of trace ammonia nitrogen in environmental water samples has been developed based on solid-phase extraction using a small column packed with octadecyl group-bonded silica gel (Sep-Pak C18 cartridge). A water sample was taken into a graduated syringe for easy and simple operation and prevention of contamination immediately after sample collection. Ammonia in the sample was reacted with hypochlorite and thymol to be converted into indothymol blue; then the formed indothymol blue was collected as an ion pair between indothymol blue and tetrabutylammonium ion on a Sep-Pak C18 cartridge. The indothymol blue on the cartridge was stable for 4 days. The retained indothymol blue was easily eluted with a mixture of methanol and 0.01 mol/l sodium hydroxide solution. The color intensity due to the indothymol blue was spectrophotometrically measured at 725 nm. The proposed method was successfully applied to environmental water samples such as river water.     

A simple and rapid in situ preconcentration method for the determination of phosphate in environmental waters by use of solid-phase extraction, and its applications to brackish lake waters

A simple and rapid in situ preconcentration method for the determination of phosphate in environmental waters has been developed for field analysis. This method is based on solid-phase extraction on a zirconium-loaded Sep-Pack Accell CM cartridge (Zr-SP) and is applicable to studies in which sampling is performed by use of a graduated syringe to prevent contamination and to ensure easy operation at sampling sites. The Zr-SP cartridge was prepared by passing 0.1 mol L(-1) zirconium solution through a Sep-Pak Accell CM cartridge, packed with cation exchange sorbent based on a silica matrix. The adsorption of phosphate and its desorption depend only on the pH of the solution. A water sample containing phosphate was adjusted to pH 2 and passed through the Zr-SP cartridge to collect it. The retained phosphate was quantitatively eluted with 0.5 mol L(-1) sodium hydroxide solution. The phosphate retained in the Zr-SP cartridge was stable for at least one month. The established preconcentration method was successfully applied to brackish lake waters to investigate seasonal changes in the distribution and behavior of phosphate in a brackish lake.     

A simple and rapid in situ preconcentration method for the determination of phosphate in environmental waters by use of solid-phase extraction, and its applications to brackish lake waters

A simple and rapid in situ preconcentration method for the determination of phosphate in environmental waters has been developed for field analysis. This method is based on solid-phase extraction on a zirconium-loaded Sep-Pack Accell CM cartridge (Zr-SP) and is applicable to studies in which sampling is performed by use of a graduated syringe to prevent contamination and to ensure easy operation at sampling sites. The Zr-SP cartridge was prepared by passing 0.1 mol L^–1 zirconium solution through a Sep-Pak Accell CM cartridge, packed with cation exchange sorbent based on a silica matrix. The adsorption of phosphate and its desorption depend only on the pH of the solution. A water sample containing phosphate was adjusted to pH 2 and passed through the Zr-SP cartridge to collect it. The retained phosphate was quantitatively eluted with 0.5 mol L^–1 sodium hydroxide solution. The phosphate retained in the Zr-SP cartridge was stable for at least one month. The established preconcentration method was successfully applied to brackish lake waters to investigate seasonal changes in the distribution and behavior of phosphate in a brackish lake.     

A Comparison of Sample Preparation Techniques for the Determination of an Organo-phosphorous Pesticide from Water Using Reverse-Phase HPLC

Abstract

A methylene chloride liquid/liquid extraction and Sep-Pak C₁₈ cartridge adsorption techniques were used to quantify the pesticide, Dursban, in contaminated environmental water samples. Results showed a large disparity between Dursban levels using these two techniques, due primarily to the presence of a large adsorbed fraction of pesticide. A Sample Clarification Kit was used to isolate the particulate fraction, which can subsequently be stripped of its adsorbed pesticide compliment by means of a methanol rinse. Lastly, the filtrate from the Sample Clarification Kit may be trace enriched on a Sep-Pak C₁₈ cartridge to isolate the dissolved fraction of pesticide.     

Determination of total mercury and methylmercury in human hair by graphite-furnace atomic absorption spectrophotometry using 2,3-dimercaptopropane-1-sulfonate as a complexing agent.

For the determination of total mercury in hair, an amount (25.0 mg) of hair sample was digested with conc. HNO3 (400 microl) at 90 degrees C for 10 min in a 7-ml teflon microreaction vessel. After digestion, the pH of the acidic hair mixture was adjusted to 5.0-6.0 by NaOH and was then passed through a clean-up Sep-Pak C18 cartridge. To the eluate, 2,3-dimercaptopropane-1-sulfonate (DMPS) and sodium acetate buffer (pH = 6.0) were added to form a mercury-DMPS complex. This complex was preconcentrated on two Sep-Pak C18 cartridges in series, and each cartridge was eluted with methanol and adjusted to 2.00 ml. A portion (50 microl) was introduced into a graphite cuvette and then atomized according to a temperature program. The method detection limit (MDL, 3sigma) was 0.064 (microg g(-1)); the calibration graph was linear up to 7.52 microg g(-1). Good accuracies were obtained when testing two human hair certified reference materials (GBW 09101 and BCR-397). Six real samples were analyzed, and the recoveries were 95.8 - 98.2% with a relative standard deviation (RSD, n = 3) < 2.1%. For the determination of methylmercury (CH3Hg+), 25.0 mg of hair sample was extracted with 2.0 mol dm(-3) HCl (1.0 ml) by ultrasonicating for 1 h. The supernatant solution was used for CH3Hg+ analysis and the hair residue was used for the analysis of inorganic mercury (Hg2+). The MDL of CH3Hg+ was 0.068 microg g(-1); the calibration graph was linear up to 6.00 microg g(-1). Six real samples were analyzed, and the recoveries were 96.0-99.2% with RSD (n = 3) < 2.3%. The sum of the concentrations of CH3Hg+ and Hg2+ was very close to that of the total mercury measured with a relative error within 3.6%. The proposed method can be accurately applied to the measurement of CH3Hg+, Hg2+, and total mercury in hair samples.     

Selective determination of urinary free cortisol by liquid chromatography after solid-state extraction

We have developed a selective and precise high-performance liquid chromatographic method for urinary free cortisol with an improved and efficient sample clean-up using C18 Sep-Pak cartridges. The urine sample (2 ml), with 11-deoxycortisol as internal standard, is applied to the Sep-Pak, which is then sequentially washed with acetone-water (1:4, v/v), water and hexane. Cortisol is eluted with diethyl ether, evaporated to dryness and redissolved in 2 ml of water. The wash cycle is repeated once using the same Sep-Pak cartridge. This double extraction greatly improves sample clean-up and allows modification of the mobile phase (tetrahydrofuran-methanol-water) so that cortisol is rapidly eluted as a single well resolved peak at 13 min. Chromatography is performed isocratically on a reversed-phase column with detection at 254 nm. Detection limits for urinary free cortisol by this procedure were two or three times lower than those obtained with two commercial radioimmunoassay kits. The chromatographic method was used successfully in the diagnosis of patients with hypercortisolism and Cushing's syndrome.     

Determination of Organophosphorus Pesticides in Water Using C18 or Florisil Sep-Pak Cartridge and Gas Chromatography with Flame-Photometric Detection

Simple methods for the concentration and clean-up of fifteen organophosphorus pesticides in water using a C₁₈ Sep-Pak cartridge or a Florisil Sep-Pak cartridge and subsequently determining the pesticides by gas chromatography with flame photometric detection have been developed. The conditions for stepwise or simultaneous desorption of these pesticides from the Sep-Pak cartridges are given.     

Fast solid phase extraction of polychlorobiphenyls and chlorinated pesticide residues from mussels using sep-pak cartridges

Summary A rapid method for the determination of chlorinated pesticides and polychlorinated biphenyls in mussels (Mytilus sp.) is reported. The mussel sample is homogenized and extracted with acetonitrile. The organic solution is concentrated and successively diluted with distilled water solution (12 g L⁻¹ NaCl). The organic compounds from water solution are adsorbed onto a NH₂ Sep-Pak cartridge. The clean-up step, in which the polychlorobiphenyls and chiorinated pesticides are separated in different eluates, is achieved by passing 25 mL of a 40% methanol aqueous solution through the NH₂ Sep-Pak and the C₁₈ Sep-Pak cartridges connected in series. The polychloroblphenyls are desorbed from the NH₂ Sep-Pak cartridge whilst the chlorinated peslicides are recovered from the C₁₈ Sep-Pak cartridge. In the separation of polychlorobiphenyls from the chlorinated pesticides tested in this work, only aldrin, hepatachlor and 4,4′-DDD are partially adsorbed with the polychlorobiphenyls onto the NH₂ Sep-Pak cartridge. The average recovery is ≥95.0% with a relative standard deviation ≤5.0%. The limits of detection for different pesticides and polychlorobiphenyl congeners are 0.01 and 0.008 μg Kg⁻¹. The final determination is carried out by capillary gas chromatography with ECD.     

Use of solid-phase extraction in various of its modalities for sample preparation in the determination of estrogens and progestogens in sediment and water.

The environmental analysis of estrogens and progestogens at physiologically active concentrations (low ng/l range) requires the use of very sensitive and selective methods, which, in most cases, make necessary an extraction/purification step. In this study, various procedures for the determination of several estrogens (estriol, estradiol, ethynyl estradiol, estrone, and diethylstilbestrol) and progestogens (progesterone, norethindrone, and levonorgestrel) in environmental matrices, including water and river sediment, are described. In all procedures, final analysis of the target compounds is performed by reversed-phase liquid chromatography-diode array detection-mass spectrometry, whereas sample preparation always includes a solid-phase extraction (SPE) step. For this SPE step. various types of sorbents, protocols, and devices have been used, and their respective advantages and disadvantages are discussed. For the off-line SPE of estrogens and progestogens from water samples, a syringe type cartridge LiChrolut RP-18 (500 mg) was selected out of two other sorbents--LiChrolut EN (200 mg) and Isolut ENV (500 mg)--for use with the automated sample preparation instrument ASPEC XL. For the on-line SPE and analysis of water samples the 10 mm x 2 mm I.D. HySphere-Resin-GP cartridge, was preferred to the C18 Baker, the PLRP-S, and the Oasis HLB. for use with the Prospekt system. A completely manual protocol based on the use of Sep-Pak C18 Plus cartridges was developed for purification of sediment extracts. All procedures were shown to be linear over a wide range of concentration, exhibited satisfactory repeatability and accuracy, and reached limits of detection usually in the low ng/l and ng/g range. Comparatively, the on-line method was shown to be advantageous in terms of automation and general method performance.     

Direct determination of p-hydroxymethamphetamine glucuronide in human urine by high-performance liquid chromatography

High-performance liquid chromatography (HPLC) with UV detection for the simultaneous determination of the free form of p-hydroxymethamphetamine (p-OHMA) and its metabolite, glucuronide (p-OHMAG) was accomplished for the first time. We achieved this by employing 1) an ion pair reagent for retention of sample to a solid-phase extraction (SPE) cartridge, Sep-Pak Light C18 and 2) a simple two-step stepwise elution technique for subsequent ion pair RP-HPLC. The proposed method was optimized for resolution of p-OHMAG, p-OHMA and MA. The method was successfully applied to urine samples collected from MA abusers.     

Separation and characterization of the metabolic products of lappaconitine in rat urine by high-performance liquid chromatography

The separation and characterization of the metabolic products of lappaconitine in rat urine by high-performance liquid chromatography with electrochemical and ultraviolet detection are described. Urine samples from rats intravenously administered lappaconitine hydrobromide were extracted with chloroform and then purified on a Sep-Pak C18 cartridge. The subsequent resolution into individual compounds was achieved by high-performance liquid chromatography. Identification of these compounds was based on comparisons of the chromatographic behaviour and the detector response with those of authentic samples. Changes in the ratio of lappaconitine to its metabolites in rat urine with time after dosing led to a proposal for one of the probable metabolic pathways of lappaconitine in the rat.     

Rapid screening method for methamphetamine in urine by colour reaction in a Sep-Pak C18 cartridge

A simple screening method for methamphetamine in urine by colour reaction was developed. Methamphetamine, which is quantitatively retained in a Sep-Pak C18 cartridge, is (after a clean-up procedure) coloured by Simon's reagent (consisting of sodium nitroprusside solution, sodium carbonate solution and acetaldehyde gas). The detection limit was 0.5 microgram/ml using 5 ml of urine sample. The results of the screening method agreed with those of thin-layer chromatography and gas chromatography-mass spectrometry.     

Simultaneous determination of water-soluble vitamins excreted in human urine after eating an overdose of vitamin pills by a HPLC method coupled with a solid phase extraction

We have applied a quick and convenient method for determining water-soluble vitamins excreted in human urine. We found that the Sep-Pak C(18) cartridge was useful for preconcentration and recovery of water-soluble vitamins in urine with minimized loss of vitamins. The recovery of vitamins was well over 90%. The separation was carried out by gradient elution with 90/10 (v/v%) methanol/water with 0.1% trifluoroacetic acid (TFA) and water with 0.1% TFA on a muBondapak C(18) column. The separation was completed within 15 min. We measured concentrations of water-soluble vitamins excreted in urine after swallowing an overdose of vitamin pills on purpose, and found that the concentration of each vitamin increased rapidly to the maximum in 2-3 h and decreased swiftly.     

Determination of tetracyclines in bovine and porcine muscle by high-performance liquid chromatography using solid-phase extraction.

A method is presented for the determination of the three tetracyclines oxytetracycline, tetracycline and chlortetracycline in muscle, spiked at 100 ng/g, using high-performance liquid chromatography (HPLC). The concentration and extraction steps are carried out using Waters Environmental Sep-Pak cartridges. The principal steps involve homogenizing the sample in EDTA-McIlvaine buffer followed by centrifugation and precipitation of the supernatant using trichloroacetic acid. After further filtration and concentration on a Sep-Pak cartridge, the sample is eluted and analysed by HPLC with UV detection and confirmation by diode-array. The column used is a Nova-Pak C18 (4 microns) cartridge (10 cm x 8 mm I.D.). A phosphate-citrate-acetonitrile buffer, utilizing ion suppression, is the mobile phase. The analytes are detectable at levels down to 10 ng/g. The analyte identity can be confirmed at 20 ng/g by the use of diode-array detection and spectral library comparison.     

Specific mass fragmentographic assay for 25,26-dihydroxyvitamin D in human plasma using a deuterated internal standard

A specific mass fragmentographic assay for the measurement of 25,26-dihydroxyvitamin D3 [25,26(OH)2D3] in human plasma, using a stable isotope labelled internal standard ([26,27-2H5]25,26(OH)2D3), is described. Plasma samples (5 ml) were extracted with acetonitrile and applied to a C18 Sep-Pak cartridge, from which the vitamin D metabolites were eluted with methanol. The metabolites were then applied to a Sep-Pak SIL cartridge and three fractions were collected. The most polar fraction, containing the polyhydroxylated metabolites, was further purified by high-performance liquid chromatography on Zorbax SIL. The eluent containing 25,26(OH)2D3 was collected, and the 25,26-n-butylboronate cyclic ester 3-trimethylsilyl ether derivative was formed. Gas chromatography-mass spectrometry was carried out, monitoring the intensities of the ions at m/z 449 and m/z 454 (for the internal standard). These ions represent the loss of a methyl group and the 3-silanol group, (M-90-15)+. The minimum limit of detection of the assay was estimated to be approximately 0.05 microgram/l. Inter-assay (3.7%) and intra-assay (8.0%) precision was acceptable and added 25,26(OH)2D3, over the concentration range 0.5-1.5 microgram/l, was recovered quantitatively. The plasma 25,26(OH)2D3 level was estimated in 26 healthy volunteers and ranged from 0.05 to 1.30 microgram/l, with a mean value of 0.54 microgram/l.     

Liquid chromatographic determination of imazosulfuron in drinking water and in soil using ultraviolet detection.

Reversed-phase liquid chromatography (LC) is used to determine a relatively new sulfonylureic herbicide, imazosulfuron, 1-(2-chloroimidazo-[1,2-a] pyridin-3-ylsulfonyl)-3-(4,6-dimethoxy-2-pyrimidinyl)-urea (TH-913), in drinking water and in soil. TH-913 is extracted from water using solid-phase extraction on C18 bonded silica. Soil samples (20 g) are extracted with 300 ml of methanol-water (50:50) and the acidified extracts are transferred onto Sep-Pak C18 and processed as described for water samples. Off-line desorption is done with 20 ml of methanol-water (50:50). The eluate is evaporated to dryness, the residue dissolved in acetonitrile and analysed by LC with UV detection at 238 nm. The recoveries of TH-913 from water were over 95% (at 0.05 microg/l level) and from soil over 90% (at 0.005 mg/kg level).     

Measurement of ascorbic acid and dehydroascorbic acid in gastric juice by HPLC

New methods are presented for measuring total vitamin C and the ascorbic acid/dehydroascorbic acid ratio in gastric juice. Extracts are prepared from a gastric juice which are suitable for direct injection onto a Waters Nova-pak C18 Radial-pak cartridge for high performance liquid chromatography (HPLC) using ultraviolet absorbance at 270 nm for detection. Both enable removal of interfering mucus and mucopolysaccharide breakdown products in a novel way. The first uses mini-columns of Sephadex G-50, run in acidic conditions to remove large molecular weight material while maintaining the ascorbic acid/dehydroascorbic acid ratio as it was in the fresh sample. Addition of dithiothreitol converts the dehydroascorbic acid quantitatively to ascorbic acid, thus enabling measurement of both components. The second method converts all the dehydroascorbic acid to ascorbic acid at the outset. A perchloric acid extract is neutralized and passed through a Sep-Pak C18. A new internal standard, reductic acid, is introduced for ascorbic acid analysis which behaves identically on Sep-Pak C18. Samples are analysed by ion-pair chromatography using 0.02 M NH4H2PO4 buffer (pH 7.1): methanol (80:20 v/v) containing 0.62 g/L tetrapentylammonium bromide. The detection limit was 1 ng ascorbic acid, and chromatography was completed in 5 min. The values obtained by the two independent HPLC methods were in good agreement with each other and with those obtained by the 2,4-dinitrophenylhydrazine colorimetric method.     

Determination of organochlorine pesticide residues in honey, applying solid phase extraction with RP-C18 material

In this study, a new clean up method was developed for the routine multiresidue determination of organochlorine pesticide residues in honey. The analytical procedure requires sample extraction with methanol, followed by a clean up step through a C18 Sep-Pak cartridge. Finally, pesticides are eluted with hexane. The determination of organochlorine pesticide residues was performed by capillary gas chromatography with electron capture detection. The mean recoveries of 18 organochlorine pesticides were estimated at various concentrations and found very efficient in most cases. The detection limits were found to be between 0.05 and 0.20 microgram kg-1.     

Determination of ethylenediaminetetraacetic acid in sea water by solid-phase extraction and high-performance liquid chromatography

The chelating agent EDTA is widely used, and as a result is showing up widely in the aquatic environment. Here we describe a preconcentration procedure for measuring EDTA concentration in sea water samples by HPLC. The procedure consists of forming an Fe(III) complex followed by solid-phase extraction using an activated carbon cartridge. After the preconcentration, EDTA was quantified by HPLC with ultraviolet detection (260 nm). The enrichment permitted the determination of EDTA at concentrations as low as 1 nM. Good recoveries were obtained for both brackish and full-strength sea water with high repeatability (RSD < 6%). The method was applied to sea water samples taken from near the mouth of the Oyabe River in Japan.     

Reversed-Phase High Performance Liquid Chromatography (HPLC) of Nucleosides in Cell Extracts with Special Reference to Deoxythymidine

Abstract

A reversed-phase high performance liquid chromatography (HPLC) system was designed to assay the nucleoside concentrations, especially deoxythymidine, of tissue cultured cells. Concentration and purification of the acid extracted cell samples were achieved by utilizing a Sep-Pak C₁₈ cartridge. After adsorption of the sample, the cartridge was washed with 0.5 ml of water followed by 2 × 0.5 ml of 2.5% methanol. The compounds of interest were subsequently eluted in 2 × 0.5 ml of methanol. The cartridge procedure was found to be fast, inexpensive and showed good recoveries for most tested nucleosides. The nucleosides uridine, deoxythymidine and adenosine could be detected in green monkey kidney cells. Ribonucleotides and deoxyribonucleotides could be separated from each other with the HPLC system used.