Determination of benzothiazole in untreated wastewater using polar-phase stir bar sorptive extraction and gas chromatography-mass spectrometry

Stir bar sorptive extraction (SBSE) was applied to extract benzothiazole (BT) from untreated wastewater using a novel polyacrylate (PA)-coated stir bar (PA Twister^®). After extraction, BT was desorbed in a thermal desorption system (TDS) and analysed by GC–MS (gas chromatography–mass spectrometry). The sample contained 30% (w/w) NaCl, the sample temperature was 30 °C and the extraction time was 240 min. Since no filtering or clean-up steps or solvents were necessary SBSE clearly performs better than all previously used extractions techniques for analysing BT in untreated wastewater in terms of easy use, sample throughput and analytical costs. In addition, matrix effects were small. The calibration curve resulting from the standard addition method was linear with a value of the stability index (R²) of 0.999 (n = 3). A good inter-day repeatability of the method was observed with a relative standard deviation (RSD) of 9.8% (n = 6). A low limit of detection (LOD) of 0.256 μg L⁻¹ was achieved using only a small sample volume of 18 mL. Small sample volumes significantly reduce sample transport costs. The concentration of BT in untreated wastewater was determined to be 1.04 μg L⁻¹.     

Non-volatile organic analysis of uranium ore concentrates

In support of nuclear safeguards and non-proliferation efforts, Oak Ridge National Laboratory is responsible for characterizing uranium ore concentrate (UOC) samples obtained from two ore mining and milling sites. A sorptive extraction method has been developed for analysis of non-volatile organic compounds that might be used to identify characteristics of the purification process by which uranium was separated from these ores. This method utilizes Gerstel Twister^® stir bars coated with polydimethylsiloxane to extract organic components from aqueous media. A slurry of UOC is extracted with the Twister^® stir bar in 20 % methanol/80 % water containing deuterated internal hydrocarbon standards. Following extraction of non-volatile organics, the Twister^® stir bar is analyzed directly in the inlet of a gas chromatograph fitted with a quadrupole mass spectrometric detector. Results have been consistent and have shown excellent recoveries of internal standards, with the average recovery being 97.5 %. Both qualitative and quantitative differences have been identified between the two sources of UOC utilizing this method. One source contained an increased concentration of amines which commonly are used in the recovery and purification of ores. Amines that were identified in this UOC source include dioctylamine, triisoctylamine, and Alamine^® 336, a common industrial complexant. Also, when comparing both sources, the same UOC source contained various decanol and C20 compounds. Based on the results from this study, non-volatile organic analysis of UOC using sorptive extraction with Twister^® stir bars and GC–MS is a tool that can be used to facilitate sourcing of unknown UOC.     

PEG modified poly(amide-b-ethylene oxide) membranes for CO2 separation

In the present work, membranes from commercially available Pebax^® MH 1657 and its blends with low molecular weight poly(ethylene glycol) PEG were prepared by using a simple binary solvent (ethanol/water). Dense film membranes show excellent compatibility with PEG system up to 50 wt.% of content. Gas transport properties have been determined for four gases (H₂, N₂, CH₄, CO₂) and the obtained permeabilities were correlated with polymer properties and morphology of the membranes. The permeability of CO₂ in Pebax^®/PEG membrane (50 wt.% of PEG) was increased two fold regarding to the pristine Pebax^®. Although CO₂/N₂ and CO₂/CH₄ selectivity remained constant, an enhancement of CO₂/H₂ selectivity (∼11) was observed. These results were attributed to the presence of EO units which increases CO₂ permeability, and to a probable increase of fractional free-volume. Furthermore, for free-volume discussion and permeability of gases, additive and Maxwell models were used.     

Optimisation of microwave-assisted extraction for the determination of nonylphenols and phthalate esters in sediment samples and comparison with pressurised solvent extraction

Microwave-assisted extraction (MAE) of nonylphenols (NP), nonylphenol mono- and diethoxylates (NP1EO and NP2EO, respectively) and phthalate esters was optimised using an experimental design approach. A D-optimal mixture design was used to optimise the pressure inside the extraction vessel (110-207 kPa), the extraction time (5-25 min) and the extraction solvent (methanol, acetone or n-hexane) or the solvent mixture for the microwave-assisted extraction. Percentage of microwave power (80%) and solvent volume (15 ml) were fixed in all the experiments. As a consequence, the optimum extraction of these compounds was carried out at an intermediate pressure (159 kPa) with pure methanol and during 15 min. Moreover, solid phase extraction was also optimised for the clean-up of the extracts and C-18, LiChrolut^® and Oasis^® cartridges were studied in order to obtain the best recoveries of the compounds of interest. The highest recoveries were obtained with LiChrolut^® cartridges after the elution with ethyl acetate. The cleaned extracts were analysed in a gas chromatograph with mass spectrometric detection and in a liquid chromatograph with diode array and fluorescence detection (HPLC-DAD-UV-FLD). The same sediment was also extracted twice in order to check that an exhaustive extraction of the analytes had occurred. Finally, the optimised extraction method was compared with pressurised solvent extraction (PSE), using an estuarine sediment sample.     

Extraction of europium and cerium into nitrobenzene using synergistic mixture of hydrogen dicarbollylcobaltate and polyethylene glycol PEG 600

Solvent extraction of microamounts of europium and cerium by a nitrobenzene solution of hydrogen dicarbollylcobaltate (H⁺B⁻) in the presence of polyethylene glycol PEG 600 (L) has been investigated. The equilibrium data have been explained assuming that the complexes HL⁺, H₂L²⁺ and ML³⁺ (M³⁺ = Eu³⁺, Ce³⁺; L = PEG 600) are extracted into the organic phase. The values of extraction and stability constants of the species in nitrobenzene saturated with water have been determined. It was found that the stability constants of the EuL³⁺ and CeL³⁺ cationic complex species (L = PEG 600) in water-saturated nitrobenzene are the same.     

Isolation of Pu-isotopes from environmental samples using ion chromatography for accelerator mass spectrometry and alpha spectrometry

A radiochemical method for the isolation of plutonium-isotopes from environmental samples, based on the use of specific extraction chromatography resins for actinides (TEVA^®, Eichrom Industries, Inc.), has been set up in our laboratory and optimised for their posterior determination by alpha spectrometry (AS) or accelerator mass spectrometry (AMS). The proposed radiochemical method has replaced in our lab a well-established one based on the use of a relatively un-specific anion-exchange resin (AG^® 1X8, Bio-rad Laboratories, Inc.), because it is clearly less time consuming, reduces the amounts and molarities of acid wastes produced, and reproducibly gives high radiochemical yields.In order to check the reliability of the proposed radiochemical method for the determination of plutonium-isotopes in different environmental matrixes, twin aliquots of a set of samples were prepared with TEVA^® and with AG^® 1X8 resins and measured by AS. Some samples prepared with TEVA^® resins were measured as well by AMS. As it is shown in the text, there is a comfortable agreement between AS and AMS, which adequately validates the method.     

Comparison of different solid phase extraction materials and techniques by application of multiresidue methods for the determination of pesticides in water by high-performance liquid chromatography (HPLC)

Solid phase extraction materials and techniques (C-18 EMPORE^® disks, polystyrenedivinylbenzene (SDB) EMPORE^® disks, C-18 BondElut cartridges and ENVI-Carb cartridges) are compared for the preconcentration of 33 basic/neutral and 10 acidic/phenolic pesticides and three metabolites in water. The efficiency of the different extraction procedures was investigated by application of appropriate multiresidue separation methods by reversed phase-high performance liquid chromatography with UV-diode-array detection. Calibrations were performed with multicomponent standard mixtures and recoveries, relative standard deviations and determination limits were calculated for comparing the described enrichment methods. Experiences made in practical application of the different techniques and materials were also considered for the final evaluation.     

Application of enzymatic probe sonication extraction for the determination of selected veterinary antibiotics and their main metabolites in fish and mussel samples

A new method based on enzymatic probe sonication extraction prior to high-performance liquid chromatography (HPLC) has been developed for the determination of 11 antibiotics (drugs) and the main metabolites of five of them in fish tissue and mussel samples. The analytes belong to four different classes of antibiotics (sulfonamides, tetracyclines, penicillins and amphenicols). The analysed compounds were sulfadiazine (SDI) and N⁴-acetylsulfadiazine (NDI) metabolite, sulfamethazine (SMZ) and N⁴-acetylsulfamethazine (NMZ), sulfamerazine (SMR) and N⁴-acetylsulfamerazine (NMR), sulfamethoxazole (SMX), trimetroprim (TMP), amoxicillin (AMX) and its main metabolite amoxicilloic acid (AMA), ampicillin (AMP) and its main metabolite ampicilloic acid (APA), chloramphenicol (CLF), thiamphenicol (TIF), oxytetracycline (OXT) and chlortetracycline (CLT).The main factors affecting the extraction efficiency (type of enzyme, type and volume of extractant, ultrasounds power and extraction time) were optimised in tissue of hake (Merluccius merluccius), anchovy (Engraulis encrasicolus), mussel (Mytilus sp.) and wedge sole (Solea solea). The extraction was carried out using an extraction time of 5 min with 5 mL of water and subsequent clean-up with dichloromethane.High-performance liquid chromatography (HPLC) with diode array (DAD) and fluorescence (FLD) detectors was used for the determination of the antibiotics. The separation of the analysed compounds was conducted by means of a Phenomenex^® Gemini C₁₈ (150 mm × 4.6 mm I.D., particle size 5 μm) analytical column with LiChroCART^® LiChrospher^® C₁₈ (4 mm × 4 mm, particle size 5 μm) guard-column. Analysed drugs were determined using formic acid 0.1% (v/v) in water and acetonitrile in gradient elution mode as mobile phase. The proposed method was also evaluated by a laboratory assay consisting of the determination of the targeted analytes in samples of Cyprinus carpio which had previously administered the antibiotics.     

Determination of inorganic selenium species in water and garlic samples with on-line ionic liquid dispersive microextraction and electrothermal atomic absorption spectrometry

A non-chromatographic separation and preconcentration method for Se species determination based on the use of an on-line ionic liquid (IL) dispersive microextraction system coupled to electrothermal atomic absorption spectrometry (ETAAS) is proposed. Retention and separation of the IL phase was achieved with a Florisil^®-packed microcolumn after dispersive liquid-liquid microextraction (DLLME) with tetradecyl(trihexyl)phosphonium chloride IL (CYPHOS^® IL 101). Selenite [Se(IV)] species was selectively separated by forming Se-ammonium pyrrolidine dithiocarbamate (Se-APDC) complex followed by extraction with CYPHOS^® IL 101. The methodology was highly selective towards Se(IV), while selenate [Se(VI)] was reduced and then indirectly determined. Several factors influencing the efficiency of the preconcentration technique, such as APDC concentration, sample volume, extractant phase volume, type of eluent, elution flow rate, etc., have been investigated in detail. The limit of detection (LOD) was 15 ng L⁻¹ and the relative standard deviation (RSD) for 10 replicates at 0.5 μg L⁻¹ Se concentration was 5.1%, calculated with peak heights. The calibration graph was linear and a correlation coefficient of 0.9993 was achieved. The method was successfully employed for Se speciation studies in garlic extracts and water samples.     

Solvent extraction of lead using a nitrobenzene solution of strontium dicarbollylcobaltate in the presence of polyethylene glycol PEG 400

Summary From extraction experiments and γ-activity measurements, the extraction constant corresponding to the equilibrium Pb²⁺(aq)+SrL²⁺ (nb)↔PbL²⁺ (nb)+Sr²⁺ (aq) taking place in the two-phase water-nitrobenzene system (L=PEG 400; aq = aqueous phase, nb = nitrobenzene phase) was evaluated as logK_ex(Pb²⁺, SrL²⁺)=2.0±0.1. Further, the stability constant of the PEG 400 - lead complex in nitrobenzene saturated with water was calculated for a temperature of 25 °C: log β_nb(PbL²⁺)=12.9±0.1.     

Poly(ethylene glycol)-supported α,α,α-trifluoroacetophenone in dioxirane mediated alkene epoxidation reactions

Poly(ethylene glycol) (PEG) was used for the immobilization of α,α,α-trifluoroacetophenone and the utility of this supported ketone has been examined in dioxirane mediated epoxidation of alkenes. The PEG-ketone reagent was found to be an effective homogeneous catalyst for the epoxidation of a variety of alkenes in the presence of Oxone^® and was readily recovered from the reaction mixtures and reused.     

An assay for identification and determination of toxic rodenticide valone in serum by ion chromatography-electrospray ionization tandem mass spectrometry with ion trap detector

Valone has a chronic and toxic anticoagulant rodenticide that has widely used in China and has resulted in some accidental and intentional intoxication in recent years. The literature reported so far lacks sensitive and selective method for the confirmation of valone. The purpose of this study was to establish a novel assay for the identification and quantification of valone in serum by ion chromatography-electrospray ionization tandem mass spectrometry (IC-MS/MS). After serum sample was extracted with methanol/acetonitrile (10:90, v/v) and cleaned by Oasis^® HLB solid-phase extraction cartridge, chromatographic separation was performed on an Ionpac^® AS11 column with an eluent of methanol/30 mmol/L KOH (10:90, v/v). The overall extraction efficiency was >81.0%, and the limit of quantification was 0.5 ng/mL for valone. Regression analysis of the calibration data revealed good correlation (r² > 0.99) for valone. Intra- and inter-day precisions for quality-control samples were less than 8.0 and 13.7%, respectively. The proposed method enables the identification and quantification of valone in both clinical and forensic specimens.     

NOR and nitroxide-based HAS in accelerated photooxidation of carbon-chain polymers; Comparison with secondary HAS: An ESRI and ATR FTIR study

Understanding processes resulting in heterogeneous degradation in polymers is of extreme importance for improving their stabilization and minimizing negative impact of photooxidation on the material properties. We adopted modern physical techniques for studies of spatial distribution of intermediates and products of photodegradation during accelerated ageing of four commodity polymers, polypropylene (PP), polyethylene (PE), polystyrene (PS) and poly(ethylene-co-norbornene) (Topas^®, TP) stabilized with hindered amine stabilizer (HAS). Concentration profiles of nitroxides inside polymer plaques along the direction perpendicular to their surface were determined by electron spin resonance imaging (ESRI) as a function of the duration of the accelerated photooxidation. We present data characterizing stabilization activity of three alkoxyamine derivatives of HAS (Tinuvin^® NOR 123, Tinuvin^® NOR 371, Flamestab^® NOR 116), Chimasorb^® 119 structurally similar to Flamestab^® NOR 116, and nitroxide-based HAS Dastib^® 1045 and compare them with the data characterizing stabilization activity of the secondary HAS (>NH) Tinuvin^® 770. ESRI data are complemented by ATR FTIR spectroscopic detection of oxidation products on the surface and inside the plaques and by data characterizing diffusive optical transmittance of the polymer plaques in the spectral region 280-400 nm (terrestrial range of the solar UV radiation).     

Aspects of the thermal and photostabilisation of high styrene-butadiene copolymer (SBC)

The thermal and photo-oxidative stabilisation of high styrene-butadiene copolymer (SBC) with high styrene content (K-Resin) has been studied using a variety of analytical and spectroscopic methods including yellowness, luminescence and FT-IR spectroscopy coupled with hydroperoxide analysis in order to understand the nature and effectiveness of the processes involved. The next stage of the program was to evaluate the effects of various chemical/solvent treatments on the role of metal ions/residual catalysts and hydroperoxides in the thermal and photostabilisation of SBS as well as combinations of phenolic antioxidants and phosphites/phosphonites. Other additives, such as HALS and a metal deactivator, were also added to the combinations of phenolic and phosphite antioxidants in order to study their behaviour and efficiency. The chemical treatments appeared to stabilise SBS against thermal oxidation to a greater or lesser extent. Phosphoric acid treatment via reflux and zinc dithiocarbamate treatments showed better performances than the rest of the treatments, the latter was particularly effective at inhibiting the discolouration. During photo-oxidation, on the other hand, chemical treatments involving phosphoric acid and pre-thermal effects showed the importance of catalyst effects. The addition of phenolic antioxidants, phosphites/phosphonites, metal deactivator and HALS was found to stabilise the SBS against thermal and photo-oxidation. In thermal oxidation, the combination of Irganox^® 1010/Irgafos^® 168 was found to effectively stabilise the polymer when the finalisation of the polymerisation was with adipic acid. When the same antioxidants were used, but with polymer finalised with BHT, strong yellowing was observed and a higher amount of hydroperoxides and oxidation products. Increasing the amount of antioxidants did not increase the stabilisation efficiency. The stabilisation efficiency of Irganox^® 1010 combined with Alkanox^® P-24 was found to be more effective than when it was combined with Irgafos^® 168. The formulations containing Irgafos^® 168/Irganox^® 1010 and Irgafos^® 168/Irganos^® 1330 were more effective in colour protection and retarding the formation of oxidation products than the combinations of Irgafos^® 168/Irganox^® 3114 and Irgafos^® 168/Lowinox^® 1790. The effect of the addition of HALS, such as Tinuvin^® 770, Tinuvin^® 622 and Chimassorb^® 944, and a metal deactivator, such as Irganox^® MD 1024, to the combination of Irgafos^® 168/Irganox^® 1010 was found to be antagonistic. In photo-oxidation, a combination of Irganox^® 1010/Irgafos^® 168 protected the polymer efficiently, when the polymerisation of the polymer was finalised with adipic acid. When the polymerisation was finalised with BHT, a higher amount of hydroperoxides and oxidation products was found. An increase in the amount of antioxidants did not enhance the stability of the polymer. The addition of Alkanox^® P-24 exhibited an opposite effect to that seen in thermal oxidation, as the stabilisation efficiency was less effective than with Irgafos^® 168. The formulation containing Irgafos^® 168/Irganox^® 1010 was found to be the most efficient compared with the other phenolic antioxidants. The addition of Tinuvin^® 770 to the formulation Irgafos^® 168/Irganox^® 1010 was found to have a synergistic effect. The addition of polymeric HALS or Irganox^® MD 1024, a metal deactivator, had an antagonistic effect on the stabilisation of the polymer. Disruption of the excimer sites in the styrenic phase also correlated with stabilisation effects.     

Influence of magnetic field on aqueous two-phase extraction of horse ferritin in the polyethylene glycol/hydroxyethyl starch system

The presented experiments show the model of expectation of equine spleen ferritin extraction in a new aqueous two-phase system which was formed by mixing polyethylene glycol (PEG) and hydroxyethyl starch (HES). The tendency of the protein to migrate in the analyzed systems was dependent on the concentrations of HES and PEG as well as PEG molecular weight. The highest concentration of ferritin in the top phase (rich in PEG) was recorded in the system composed of 6% PEG 3000 and 3% HES. The obtained concentration was 0.88 mg mL⁻¹. The lowest concentration was 0.42 mg mL⁻¹ in the system composed of 5% PEG 6000 and 1% HES.     

Extraction of radiostrontium from the mixture of radionuclides in milk using dicarbolide of cobalt

The possibilities of radiostrontium extraction from milk after the isolation of radiocesium were investigated. The polyhedral complex of the type H⁺[(π-/3/-1,2-C₂B₉H₁₁)₂Co⁻], further referred to as dicarbolide-H⁺, in nitrobenzene was used for the extraction. The increase of values and larger differentiation of distribution ratios in the extraction of ion associates of radionuclides were achieved by hydrophobizing with polyethyleneglycol (PEG). The changes of the distribution ratios of⁹⁰Sr,⁸⁹Sr and⁹⁰Y with PEG concentration were studied, and ranges of the highest synergic effect, changing with different extraction agent concentrations in extraction from water solutions and a fresh milk were determined. The influence of milk dilution and nitric acid concentration upon the value of distribution ratio in radiostrontium extraction using dicarbolide-H⁺ and the optimum PEG concentration were investigated. Under suitably selected conditions of radiostrontium isolation, the distribution ratios of some potential contaminants were determined. The influence of nonisotopic carrier Ca²⁺ upon radiostrontium extraction was examined. The possibilities of purification of isolated radiostrontium using reextraction were investigated. A selective and quick analytical procedure was suggested for radiostrontium isolation after the separation of radiocesium from a mixture of fission and activated radionuclides in milk using extraction by dicarbolide-H⁺. The chemical yield is about 98% and⁹⁰Sr is determined by liquid scintillation technique.     

Polyethylene glycol-based ultrasound-assisted extraction of magnolol and honokiol from Cortex Magnoliae Officinalis

In this study, a kind of green solvent named polyethylene glycol (PEG) was developed for the ultrasound-assisted extraction (UAE) of magnolol and honokiol from Cortex Magnoliae Officinalis. The effects of PEG molecular weight, PEG concentration, sample size, pH, ultrasonic power and extraction time on the extraction of magnolol and honokiol were investigated to optimise the extraction conditions. Under the optimal extraction conditions, the PEG-based UAE supplied higher extraction efficiencies of magnolol and honokiol than the ethanol-based UAE and traditional ethanol-reflux extraction. Furthermore, the correlation coefficient (R²), repeatability (relative standard deviation, n = 6) and recovery confirmed the validation of the proposed extraction method, which were 0.9993–0.9996, 3.1–4.6% and 92.3–106.8%, respectively.     

Extraction of actinides and fission products from salt solutions using polyethylene glycols (PEGs)

The extraction behavior of several metal ions viz., Am³⁺, Eu³⁺, UO₂ ²⁺, Th⁴⁺, Sr²⁺ and Cs⁺ was investigated from sulphate medium employing phosphotungstic acid (PTA) and polyethylene glycol (PEG). The influence of various parameters such as pH, PTA concentration, PEG concentration and salt concentration was studied. The order of extraction followed the trend: Am³⁺>Eu³⁺>>Th⁴⁺>UO₂ ²⁺>Sr²⁺>Cs⁺ which deviate significantly from the reported order with conventional solvents. The relatively poor extraction of UO₂ ²⁺, Sr²⁺ and Cs⁺ was ascribed to their lack of interaction with the phosphotungstate anion. The separation behaviour of Am³⁺ vis-a-vis Eu³⁺ was also investigated under different experimental conditions.     

Generation and characterization of polyclonal antibodies against microcystins-Application to immunoassays and immunoaffinity sample preparation prior to analysis by liquid chromatography and UV detection

Polyclonal antibodies against microcystin-LR (MC-LR), a cyclic heptapeptide toxin, were generated in rabbits using MC-LR-BSA. An enzyme-linked immunosorbent assay (ELISA) was developed for the characterization of the antibodies and their potential use for analytical purposes. The concentration of MC-LR that inhibits 50% of antibody-antigen binding (IC₅₀) was 0.5 μg L⁻¹ for the indirect ELISA format and 0.9 μg L⁻¹ for the direct ELISA, using MC-LR-horseradish peroxidase conjugate. The limit of detection corresponding to IC₈₀ was found to be 0.06 μg L⁻¹, well below the Word Health Organization level for drinking water of 1 μg L⁻¹. The direct competitive ELISA was applied to water samples and was shown useful for screening purposes. The developed anti-microcystin antibodies were immobilized on solid supports for use in selective solid phase extraction (SPE) systems, prior to liquid chromatography (LC) quantification. An immunoaffinity cartridge (IAC), a Sepharose^®-based cartridge incorporating 2 mg of antibodies allowed the selective and quantitative recovery of a mixture of 0.2 μg of MCs showing potential use in sample preparation of real matrices. When applied to water and green algae samples, average recoveries from Sepharose^®-based cartridges were in the range of 86-113% for water samples and 85-92% for blue-green algae samples. Selectivity of the IAC clean-up was proven by comparison with non-specific solid phase extraction using octadecylsilica (ODS) sorbent. Results obtained using LC/UV after IAC clean-up agreed well with results obtained using liquid chromatography and mass spectrometry detection (LC/MS and LC/MS/MS) after SPE-C18 clean-up, allowing therefore to validate the resulting technique.     

Simultaneous determination of 11 antibiotics and their main metabolites from four different groups by reversed-phase high-performance liquid chromatography-diode array-fluorescence (HPLC-DAD-FLD) in human urine samples

A new, accurate and sensitive reversed-phase high-performance liquid chromatography (RP-HPLC) as analytical method for the quantitative determination of 11 antibiotics (drugs) and the main metabolites of five of them present in human urine has been worked out, optimized and validated. The analytes belong to four different groups of antibiotics (sulfonamides, tetracyclines, penicillins and anphenicols). The analyzed compounds were sulfadiazine (SDI) and its N⁴-acetylsulfadiazine (NDI) metabolite, sulfamethazine (SMZ) and its N⁴-acetylsulfamethazine (NMZ), sulfamerazine (SMR) and its N⁴-acetylsulfamerazine (NMR), sulfamethoxazole (SMX), trimetroprim (TMP), amoxicillin (AMX) and its main metabolite amoxicilloic acid (AMA), ampicillin (AMP) and its main metabolite ampicilloic acid (APA), chloramphenicol (CLF), thiamphenicol (TIF), oxytetracycline (OXT) and chlortetracycline (CLT). For HPLC analysis, diode array (DAD) and fluorescence (FLD) detectors were used. The separation of the analyzed compounds was conducted by means of a Phenomenex^® Gemini C₁₈ (150 mm × 4.6 mm I.D., particle size 5 μm) analytical column with LiChroCART^® LiChrospher^® C₁₈ (4 mm × 4 mm, particle size 5 μm) guard column. Analyzed drugs were determined within 34 min using formic acid 0.1% in water and acetonitrile in gradient elution mode as mobile phase. A linear response was observed for all compounds in the range of concentration studied. Two procedures were optimized for sample preparation: a direct treatment with methanol and acetonitrile and a solid phase extraction procedure using Bond Elut^® Plexa™ columns. The method was applied to the determination of the analytes in human urine from volunteers under treatment with different pharmaceutical formulations. This method can be successfully applied to routine determination of all these drugs in human urine samples.