Excretion of iodide in 24-h urine as determined by ion-pair reversed-phase liquid chromatography with electrochemical detection

A simple method is presented for the routine analysis of iodide in urine. After a one-step sample clean-up, iodide was separated by ion-pair reversed-phase liquid chromatography and detected electrochemically with a silver electrode. The coefficient of variation of a single analysis of iodide in a pooled urine sample (530 nmol/l) was 7.6%. The detection limit, derived from a signal-to-noise ratio of 3, was 3 pmol, corresponding to 0.06 mumol/l. The recovery of iodide added to urine was 96 +/- 7%. The accuracy of the method was assessed by analysing ten different samples with neutron activation analysis. The data obtained with the two methods showed a high correlation (r = 0.991) and did not differ significantly. Excretion of iodide in samples of 24-h urine from a free-living population was shown to have a log-normal distribution and to be higher in men than in women. The iodide/creatinine ratio was independent of sex and increased with age.     

Simultaneous determination of serum cortisol and cortisone by reversed-phase liquid chromatography with ultraviolet detection.

A rapid high-performance liquid chromatographic (HPLC) method for the simultaneous determination of cortisol and cortisone in a single extract of 1 ml of serum is described. The method employs meprednisone as the internal standard. The steroids were analysed isocratically by reversed-phase HPLC with an octadecylsilane-bonded (ODS) column using ultraviolet detection. The matrix effect was reduced by lowering the sample pH by adding glacial acetic acid to the sera. The samples were then filtered through regenerated cellulose membranes at 4 degrees C and extracted with diethyl ether. The dried eluates were redissolved in the mobile phase and injected into the column. The detection limit of the assay for both steroids was 500 ng/l. Cortisol was determined in twenty serum samples by both HPLC and radioimmunoassay (RIA). The results were similar. Interference by other steroids and certain steroid analogue drugs was also studied. The HPLC method yielded no cross-reactivity between the different steroids as may occur with the RIA technique. The HPLC method was technically easy to perform and it allowed us to quantify both cortisol and cortisone in a single serum extract with high specificity.     

Enantioselective determination of acylamino acid fungicides in vegetables and fruits by chiral liquid chromatography coupled with tandem mass spectrometry

An efficient and sensitive enantioselective method for simultaneous determination of three acylamino acid fungicides in vegetables and fruits was presented by high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry. The three fungicides (benalaxyl, furalaxyl, and metalaxyl) residues in samples were extracted with acetonitrile containing 1% acetic acid and an aliquot was cleaned up with Si-(CH(2) )(3) -NH-(CH(2) )(2) -NH(2) and C(18) sorbent. Complete enantioseparation of three acylamino acid fungicides enantiomers was obtained under reversed-phase conditions on a cellulose tris (4-chloro-3-methylphenylcarbamate) column at 25°C using acetonitrile-0.1% formic acid solution (45:55, v/v) as a mobile phase. The elution orders of the eluted enantiomers were determined by a circular dichroism (CD) detector. The linearity, matrix effect, recovery, and precision were evaluated. Good linearity was obtained over the concentration range of 0.5-250 μg/L for each enantiomer in the standard solution and sample matrix calibration curves. There was no significant matrix effect for three fungicides determination based on the method. The inter-day mean recoveries, intra-day repeatability, and inter-day reproducibility varied from 81.3 to 95.7%, 2.2 to 9.4%, and 2.3 to 9.6%, respectively. The method provided high selectivity and sensitivity, and limits of quantification for enantiomers of three fungicides in vegetables and fruits were both 1 μg/kg.     

Simultaneous determination of pseudouridine and creatinine in untreated urine by ion-pair liquid chromatography with diode-array ultraviolet detection

A simple procedure for the simultaneous determination of pseudouridine and creatinine in urine using ion-pair high-performance liquid chromatography with ultraviolet detection is described. It consists of simply diluting the filtered urine with mobile phase (1:20) followed by direct chromatographic injection. A single analysis takes only 10 min. This method has been applied to the analysis of urine samples from normal donors and patients with different types of cancer. The mean values, means, of the peak-area ratio of pseudouridine to creatinine were 61.79.10(-3) and 81.92.10(-3) for male and female normal donors, respectively. Out of twenty-five urine samples of patients with cancer examined, nineteen (all the forteen males included) had values higher than means + 2 sigma.     

Determination of paraquat in rat brain using ion-pair solid-phase extraction and reversed-phase high-performance liquid chromatography with ultraviolet detection

A simple and sensitive procedure for the measurement of N-methylisoquinolinium ion (NMIQ+), a putative neurotoxin, was devised using high-performance liquid chromatography (HPLC) with fluorescence detection. Separation of NMIQ+ was carried out by gel filtration and reversed-phase HPLC on a column of hydrophilic polymer gels (Asahipak GS-302H). The method was sensitive enough to measure 50 fmol of NMIQ+. Uptake of NMIQ+ into rat striatal slices was confirmed by this method.     

Determination of benzimidazolic fungicides in fruits and vegetables by supramolecular solvent-based microextraction/liquid chromatography/fluorescence detection

A supramolecular solvent consisting of vesicles, made up of equimolecular amounts of decanoic acid (DeA) and tetrabutylammonium decanoate (Bu₄NDe), dispersed in a continuous aqueous phase, is proposed for the extraction of benzimidazolic fungicides (BFs) from fruits and vegetables. Carbendazim (CB), thiabendazole (TB) and fuberidazole (FB) were extracted in a single step and no clean-up or concentration of extracts was needed. The high extraction efficiency obtained for BFs was a result of the different types of interactions provided by the supramolecular solvent (e.g. hydrophobic and hydrogen bonds) and the high number of solubilisation sites it contains. Besides simple and efficient, the proposed extraction approach was rapid, low-cost, environment friendly and it was implemented using conventional lab equipments. The target analytes were determined in the supramolecular extract by LC/fluorescence detection. They were separated in a Kromasil C₁₈ (5 μm, 150 mm × 4.6 mm) column using isocratic elution [mobile phase: 60:40 (v/v) 50 mM phosphate buffer (pH 4)/methanol] and quantified at 286/320 nm (CB) and 300/350 nm (TB and FB) excitation/emission wavelengths, respectively. Quantitation limits provided by the supramolecular solvent-based microextraction (SUSME)/LC/fluorescence detection proposed method for the determination of CB, TB and FB in fruits and vegetables were 14.0, 1.3 and 0.03 μg kg⁻¹, respectively, values far below the current maximum residue levels (MRLs) established by the European Union, i.e. 100-2000 μg kg⁻¹ for CB, 50-5000 μg kg⁻¹ for TB and 50 μg kg⁻¹ for FB. The precision of the method, expressed as relative standard deviation, for inter-day measurements (n = 13) was 3.3% for CB (50 μg kg⁻¹), 3.5% for TB (10 μg kg⁻¹) and 2.8% for FB (0.5 μg kg⁻¹) and recoveries for fruits (oranges, tangerines, lemons, limes, grapefruits, apples, pears and bananas) and vegetables (potatoes and lettuces) fortified at the μg kg⁻¹ level were in the interval 93-102%.     

Simultaneous determination of four 5-hydroxy polymethoxyflavones by reversed-phase high performance liquid chromatography with electrochemical detection

Accumulating evidence has suggested the potential health-promoting effects of 5-hydroxy polymethoxyflavones (5-OH-PMFs) naturally existing in citrus genus. However, research efforts are hampered by the lack of reliable and sensitive methods for their determination in plant materials and biological samples. Using reversed-phase high performance liquid chromatography (HPLC) equipped with electrochemical (EC) detection, we have developed a fast and highly sensitive method for quantification of four 5-OH-PMFs, namely 5-hydroxy-6,7,8,3′,4′-pentamethoxyflavone, 5-hydroxy-3,6,7,8,3′,4′-hexamethoxyflavone, 5-hydroxy-6,7,4′-trimethoxyflavone, and 5-hydroxy-6,7,8,4′-tetramethoxyflavone. The method was fully validated in terms of linearity, accuracy and precision. The limit of detection (LOD) was determined as being between 0.65 and 1.8 ng/mL (ppb), demonstrating an over 160 times higher sensitivity in comparison with the previously reported method using UV detection. The recovery rate of the method was between 96.17% and 110.82%, and the precision for the retention times and peak areas was all below 13%. The method was successfully used to quantify 5-OH-PMFs with a wide range of abundance in the citrus products and preparations, such as orange juice, citrus peel, and dried tangerine peel. The quantification method for 5-OH-PMFs developed herein could be useful for the nutritional and pharmacological studies of these compounds in future.     

Direct determination of sinigrin in mustard seed without desulfatation by reversed-phase ion-pair liquid chromatography

Reversed-phase ion-pair liquid chromatography has been investigated for directly analyzing sinigrin in mustard seed without desulfatation. After extraction by phosphate buffer (pH 7.0) from the grind-pastes of inactivated-myrosinase mustard seeds, sinigrin was first isolated through deproteinization and centrifugation, followed by filtration and injection into the chromatographic system. A reversed-phase C18 column was used to separate the sinigrin with an eluent of acetonitrile (ACN)-water (20:80) containing 0.02 M tetrabutylammonium (TBA) as the counter ion at pH 7.0. Detection was carried out with an UV detector operated at 227 nm. Factors affecting the chromatographic separation and quantitative determination, such as concentrations of TBA and ACN, and pH, were studied. The linear dynamic range is larger than three orders of magnitude and the detection limit is 0.045 mg/L. The RSD is around 3% and the recovery is 85% (3% RSD, n = 3).     

Improved determination of sulpiride in plasma by ion-pair liquid chromatography with fluorescence detection

A specific and sensitive high-performance liquid chromatographic method for the measurement of sulpiride in plasma is described. The internal standard used was veralipride, a structurally related substituted benzamide. A fluorescence detector with maximum excitation at 300 nm and maximum emission at 365 nm was used for quantitation. After an alkaline extraction procedure, the benzamides were separated on a 5-micron ODS column using a large organic counter ion in the mobile phase. The detector response was linear from 10 to 1000 ng/ml and the detection limit was 10 ng/ml, which is sensitive enough for pharmacokinetic studies. The suitability of the method for the analysis of biological samples was tested by studying the variation with time of plasma concentrations of sulpiride in normal human volunteers after a single therapeutic 200-mg oral dose of three different formulations of sulpiride.     

Selective clean-up applicable to aqueous acetone extracts for the determination of carbendazim and thiabendazole in fruits and vegetables by high-performance liquid chromatography with UV detection

Fungicide residues in vegetables (benomyl, carbendazim, thiabendazole) are analyzed through a clean-up procedure that uses a portion of the aqueous acetone extract prepared for multiresidue methodology. A portion of the aqueous acetone extract (equivalent to 5 g of vegetables) is loaded onto an Extrelut-20 cartridge (the cartridge is filled with a coarse, large-pore diatomaceous material). Then, acetone is partially removed by an upward stream of nitrogen at 2l/min for 30 min. Benzimidazolic fungicides are recovered by percolating the cartridge with 100 ml of 0.1 M phosphoric acid solution, which also serves to convert benomyl to carbendazim. The percolating acid solution is drained on-line through a strong cation-exchange (SCX) solid-phase extraction cartridge with the aid of a slight vacuum. Benzimidazolic fungicides are retained on the SCX cartridge. The phosphoric acid solution is discarded together with the washings of the SCX cartridge, i.e., water followed by methanol-water (75:25), that remove unwanted coextractives. Finally, benzimidazolic fungicides are recovered by eluting the SCX cartridge with methanol-ammonium formate buffer (75:25). The final extract is then analyzed by reversed-phase HPLC with UV detection. Recoveries from crops such as apples, lettuce, strawberries and citrus fruits are generally greater than 80% and no interferences were observed. The clean-up is simple and straightforward, requires only disposable items, water solutions and a few milliliters of solvent and a minimum number of manipulations, and does not require concentration steps or electrical equipment.     

Direct determination of trace aluminum with quercetin by reversed-phase high performance liquid chromatography

The determination of trace levels of aluminum by high performance liquid chromatography (HPLC) with spectrophotometric detection using quercetin, a bioactive substance as a pre-column reagent, is developed in this paper. The Al-quercetin chelate was separated on a reversed-phase ODS column with a mobile phase consisting of 70% water (pH 1.0 with perchloric acid) and 30% methanol, and detected at its maximum of 415 nm. The response was linear over the 1.0×10⁻⁷ to 8.0×10⁻⁵ M concentration range with a detection limit of 5.0×10⁻⁸ M and a relative standard deviation of 1.0% at the 5×10⁻⁶ M level. The analysis was free from common ions except iron, which could be successfully screened by 1,10-phenanthroline. This method has been employed to the determination of Al in environmental and biological samples. Moreover, direct speciation of labile monomeric Al, the toxic form of Al, in natural water by the present technique was explored. The coordination ratio of Al complex with quercetin was also elucidated by HPLC combined with molar ratio method. Only a 1:1 complex was formed. Because quercetin exists in body, a preliminary thinking of in vivo determination of Al is provided in this study.     

Simple and rapid method for the determination of ethylenebisdithiocarbamate fungicides in fruits and vegetables using liquid chromatography with tandem mass spectrometry

A simple and rapid method for determining ethylenebisdithiocarbamates (EBDCs; mancozeb, maneb, and zineb) in fruits and vegetables is described. EBDCs are transformed into dimethylethylenebisdithiocarbamate (EBDC-dimethyl) by methylation after their decomposition with ethylenediaminetetraacetic acid (EDTA). These processes were performed simultaneously in this method. Dimethyl sulfate was used as the methylation reagent, and acetonitrile extracts obtained from partitioning with anhydrous magnesium sulfate and sodium chloride were subjected to dispersive solid-phase extraction with the primary secondary amine sorbent. Liquid chromatography with tandem mass spectrometry in the positive heated-electrospray ionization mode was used for the determination of EBDC-dimethyl produced from EBDCs. The method was validated at levels of 10, 50, and 100 ng g⁻¹ maneb as a representative EBDC. The recoveries of the present method were between 71 and 101%. The limits of detection and quantification were 0.24 and 0.8 ng g⁻¹ maneb, respectively.     

Gunshot residue determination by high-performance liquid chromatography with electrochemical detection

A procedure for the detection of gunshot residue via the organic constituent diphenylamine is described. The method incorporates high-performance liquid chromatography with electrochemical detection.     

Multiresidue determination of 19 fungicides in processed fruits and vegetables by capillary gas chromatography after gel permeation chromatography.

A gas chromatographic (GC) method was developed for simultaneous determination of 19 fungicides (chlorothalonil, vinclozolin, dichlofuanid, triadimefon, penconazole, chlozolinate, captan, procymidone, triadimenol, folpet, hexaconazole, myclobutanil, cyproconazole, propiconazole, nuarimol, captafol, iprodione, fenarimol, and bitertanol) and the acaricide tetradifon in tomato puree, peach nectar, orange juice, and canned peas. Samples were extracted with acetone, partitioned with ethyl acetate-cyclohexane (50 + 50, v/v), and cleaned using gel permeation chromatography with ethyl acetate-cyclohexane (50 + 50, v/v) as eluant. The final extract was analyzed by GC with ion trap mass spectrometry (ITMS) using a DB5 capillary column. Recoveries from fortified samples ranged from 74.6 to 99.3%, except for triadimenol and bitertanol. Quantitation limits for most analytes were between 0.005 and 0.050 mg/kg. The purified extracts were analyzed further by GC with electron capture and nitrogen phosphorus detection, and the results were compared with those obtained by ITMS.     

Identification of cosmetic dyes by ion-pair reversed-phase high-performance liquid chromatography

A method based on ion-pair reversed-phase high-performance liquid chromatography with detection at four wavelengths between 400 and 600 nm is reported for the separation and identification of the most common synthetic colour additives in cosmetic products. All the dyes generally employed in the U.S.A. and almost all those in current use in cosmetics in the European Community have been taken into account. The chromatography was performed on a C8 bonded silica packed column, with a 60-min gradient changing from 10 to 95% acetonitrile in water containing 10(-2) M sodium perchlorate (pH 3.0) as mobile phase (flow-rate 2.5 ml/min). Detection limits are in the range 20-100 ng for all dyes investigated. The method has been applied to the analysis of commercial lipsticks.