Development and validation of a new method for the simultaneous determination of PAHs and NDL-PCBs in fish by GC-SQ-MS.

A new method for the simultaneous determination of PAHs and NDL-PCBs in fish and fish products was developed and validated using a GC-SQ-MS system. The efficacy of different solvents for a quantitative extraction was studied, as was the efficacy of different sorbents for sample clean up. The developed method, which includes extraction by DCM and sample clean up by SPE using Isolute® SI cartridges, was statistically validated at two concentration levels by assessing: accuracy, precision, limit of quantification, limit of detection and matrix effect. The method was used for analysing fresh, frozen and smoked fish products from the Greek retail market. None of the samples examined showed results above the maximum limits set by EU regulations.

Development of a fast and selective method for the sensitive determination of anatoxin-a in lake waters using liquid chromatography-tandem mass spectrometry and phenylalanine-d5 as internal standard.

Anatoxin-a is a potent alkaloid neurotoxin produced by a number of cyanobacterial species and released in freshwaters during cyanobacterial blooms. Its high toxicity is responsible for several incidents of lethal intoxications of birds and mammals around the world; therefore anatoxin-a has to be regarded as a health risk and its concentration in lakes and water reservoirs should be monitored. Phenylalanine is a natural amino acid, also present in freshwaters, isobaric to anatoxin-a, with a very similar fragmentation pattern and LC retention. Since misidentification of phenylalanine as anatoxin-a has been reported in forensic investigations, special care must be taken in order to selectively determine traces of anatoxin-a in the presence of naturally occurring phenylalanine. A fast LC tandem MS method was developed by using a 1.8 microm 50 x 2.1 mm C18 column for the separation of anatoxin-a and phenylalanine, achieving a 3-min analysis time. Isotopically labelled phenylalanine-d(5) was employed as internal standard to compensate for electrospray ion suppression and sample preconcentration losses. Both compounds were preconcentrated 1,000-fold on a porous graphitic carbon solid-phase extraction (SPE) cartridge after adjustment of sample pH to 10.5. The method was validated by using lake water spiked at four different levels from 0.01 to 1 microg L(-1). Anatoxin-a recovery ranged from 73 to 97%, intra-day precision (RSD%) ranged from 4.2 to 5.9, while inter-day precision (RSD%) ranged from 4.2 to 9.1%. Limits of detection and quantification were 0.65 and 1.96 ng L(-1) respectively. The method was successfully applied for the detection of anatoxin-a in Greek lakes at concentrations ranging from less than 0.6 to 9.1 ng L(-1).

Liquid Chromatographic Determination of Biogenic Amines in Fish Based on Pyrene Sulfonyl Chloride Pre-Column Derivatization.

Monitoring of biogenic amines in food is important for quality control, in terms of freshness evaluation and even more for food safety. A novel and cost-effective method was developed and validated for the determination of the main biogenic amines: histamine, putrescine, cadaverine, spermidine and spermine in fish tissues. The method includes extraction of amines with perchloric acid, pre-column derivatization with Pyrene Sulfonyl Chloride (PSCl), extraction of derivatives with toluene, back-dissolution in ACN after evaporation and determination by reversed phase high performance liquid chromatography with UV and intramolecular excimer fluorescence detection. The structure of the pyrene-derivatives was confirmed by liquid chromatography-mass spectrometry with electrospray ionization. The standard addition technique was applied for the quantitation due to significant matrix effect, while the use of 1,7-diaminoheptane as internal standard offered an additional confirmation tool for the identification of the analytes. Method repeatability expressed as %RSD ranged between 7.4-14% for the different amines and recovery ranged from 67% for histamine up to 114% for spermine. The limits of detection ranged between 0.1-1.4 mg kg-1 and the limits of quantification between 0.3-4.2 mg kg-1. The method was applied to canned fish samples and the concentrations of the individual biogenic amines were below the detection limit up to 40.1 mg kg-1, while their sum was within the range 4.1-49.6 mg kg-1.

Determination of bisphenol A in canned food by microwave assisted extraction, molecularly imprinted polymer-solid phase extraction and liquid chromatography-mass spectrometry.

Bisphenol A (BPA), a known potential endocrine disrupting compound (EDC) is expected to be present in low quantities in canned food due to its migration from the inner surface coating of cans made of epoxy resins. A selective and confirmatory analytical method, based on microwave assisted extraction (MAE), molecularly imprinted solid phase extraction (MISPE) using a polymer prepared by a non-covalent molecular imprinting technique and liquid chromatography coupled with electrospray ionization mass spectrometry (LC-ESI/MS) was developed for the determination of BPA in canned pineapple, tuna and mushrooms. First, the effect of the loading medium of hydro- organic solutions on the binding of BPA and its deuterated analogue on the MISPE sorbent was investigated. Subsequently, the effects of the experimental conditions of the microwave assisted extraction (solvent, sample mass/solvent volume, time and temperature) on the obtained recovery of BPA from canned food were assessed and the parameters were optimized to provide maximum recovery and selectivity. It was demonstrated that the combination of MAE with MISPE permits the use of a selective extraction solvent (methanol/water, 4/6, v/v), simplifying the sample preparation steps and enhancing sample clean-up of complex food matrices. The method was validated in different food matrices, using BPA-d16 as internal standard and quantitative relative recoveries were determined. The precision (RSD %) of the method ranged from 7% to 10% and the limit of detection was at low ng/g level for all food matrices. The determined concentration of BPA in commercial canned samples ranged between 7.3 and 42.3 ng/g.

Direct determination of the estrogenic compounds 8-prenylnaringenin, zearalenone, alpha- and beta-zearalenol in beer by liquid chromatography-mass spectrometry.

A novel LC-ESI-MS method for the simultaneous determination of four of the most significant estrogenic compounds naturally occurring in beer, 8-prenylnaringenin (8-PN), zearalenone (ZON), alpha-zearalenol (alpha-ZOL) and beta-zearalenol (beta-ZOL) which requires minimal sample preparation was developed using a chemometric approach. Experimental design was applied to assess the effects of the LC-ESI-MS parameters (mobile phase flow rate, drying gas flow, nebuliser pressure and capillary potential) on the obtained signal and to optimize the values in order to provide maximum sensitivity and detectability. The proposed method is simple, consisting only of degassing the beer and diluting with water (1:1, v/v) before injection. Comparison between the two internal standards used, zearalanone (ZAN) and 4,2'-dihydroxychalcone (4,2'-DHC), showed that ZAN performs better as internal standard not only for the mycotoxins but for 8-PN as well, giving lower % RSDs. Under inter-day conditions mean recoveries were 107% for ZON, 87.8% for alpha-ZOL, 72.8% for beta-ZOL, and 77.5% for 8-PN. The corresponding % RSDs ranged between 5.0 and 8.0. The method limits of detection were 1.3, 1.4, 1.0 and 0.8 ng mL(-1) for ZON, alpha-ZOL, beta-ZOL and 8-PN, respectively. The method was applied to 15 beer samples obtained from local supermarkets and the concentration of the phytoestrogen 8-PN in beer ranged between <0.8 and 38.6 ng mL(-1), while neither ZON nor its metabolites, alpha-ZOL and beta-ZOL, were detected.

Molecularly imprinted polymers for bisphenol A for HPLC and SPE from water and milk.

Molecularly imprinted polymers (MIPs) for bisphenol A (BPA) were prepared by two synthetic routes: semi-covalent and noncovalent methodology. The molecular imprinting effect was evaluated using the polymers in HPLC and SPE. Polymers prepared with noncovalent mode were proven more effective. These polymers were applied in SPE facilitating selective retention of BPA from bottled water and milk. The developed sample preparation was simple and efficient comprising only dilution of milk and MISPE prior to LC-MS analysis. Overall MISPE enhanced sample clean-up. Compared with control nonimprinted polymers and conventional C18 SPE cartridges, the MIPs exhibited selective analyte recognition. The method provided quantitative BPA recoveries, very good reproducibility (% RSDs lower than 7%), and low LOD (0.2 ng/g). MIP interacts similarly with deuterated BPA allowing its use as internal standard in LC-MS. The most critical parameters of MISPE were the organic content in loading-washing medium and the washing volume. Low flow rates in the elution step enhanced extraction recovery. Important advantages of the MIP were: the high breakthrough volumes (> 500 mL of water), high mass capacity (> 10 ng/mg of MIP sorbent), good linearity, and good stability in performance for over 35 cycles of use.

A novel photometric method for evaluation of the oxidative stability of virgin olive oils.

The Oxitester method, a novel, simple, and fast photometric method for the evaluation of the antioxidant capacity of olive oils, was validated and compared to the official oil stability index (Rancimat) method. The Oxitester method appeared to be a good alternative to the Rancimat method with adequate correlation for a wide range of virgin olive oil samples, including extrissima virgin olive oils (correlation coefficient 0.88), and extra virgin olive oils of increased acidity (free fatty acids >0.45%, correlation coefficient 0.89). Other quality factors (flavor, free fatty acids content, specific absorbance at 270 and 232 nm, peroxide value, and content of oleic, linoleic, and linolenic acids) were also measured and correlated to the antioxidant capacity values of the Oxitester and Rancimat methods. The Oxitester method, in contrast to the Rancimat method, was indicative of the flavor characteristics of the olive oils and the content of linolenic acid.

Development of a novel method based on liquid chromatography-evaporative light scattering detection for the direct determination of streptomycin and dihydrostreptomycin in raw materials, pharmaceutical formulations, culture media and plasma.

A novel method for the non-derivatization liquid chromatographic determination of streptomycin (STR) and dihydrostreptomycin (DHSTR) was developed and validated based on evaporative light scattering detection (ELSD). Utilizing a ThermoHypersil BetaBasic C18 analytical column, evaporation temperature of 50 degrees C and pressure of nebulizing gas (nitrogen) of 3.5 bar, the optimized mobile phase was 1.25 mL L(-1) TFA aqueous solution, in an isocratic mode at a rate of 1.0 mL min(-1). STR was eluted at 5.6 min and DHSTR at 7.8 min with a resolution of 4.4. Linear calibration curves were obtained from 2 to 120 microg mL(-1) (r > 0.9990) for STR and 2-75 microg mL(-1) (r > 0.9994) for DHSTR, with a LOD equal to 0.7 and 0.5 microg mL(-1), respectively. The developed method was applied for the assay of STR and DHSTR (sulfate) in pharmaceutical raw materials and formulations, while the simultaneous direct determination of sulfate was feasible (tR = 2.5 min, LOD = 1.4 microg mL(-1), double logarithmic calibration curve in the range of 4-50 microg mL(-1), r > 0.9998). Modified isocratic mobile phase (H2O-ACN, 90:10, v/v, containing 1.25 mL L(-1) TFA), was used for the determination of streptomycin B impurity in STR sulfate raw material and a gradient mobile phase (H2O-ACN containing TFA) was used for the determination of DHSTR in the presence of penicillinG procaine. The developed method was also applied for the assay of commercial formulations (STR powder and DHSTR injection solution and suspension) (%recovery 98-102, %RSD < 1.3, n = 3 x 3), for the determination of STR in bacteria culture medium (%recovery 99.6, %RSD = 0.8, n = 3 x 3), and for the determination of DHSTR in human plasma (2.0-23.0 microg mL(-1)) after solid phase extraction using carboxylate cartridges (%recovery 98.4-101.8, %RSD = 3.2, n = 3 x 3).

Determination of bisphenol A in milk by solid phase extraction and liquid chromatography-mass spectrometry.

A simple and reliable analytical method based on solid phase extraction (SPE) and liquid chromatography coupled with electrospray ionization mass spectrometry was developed for the determination of bisphenol A (BPA) in milk. The effects of the experimental parameters of the LC-ESI-MS system (mobile phase and additives, flow rate, temperature of the ionization source, cone voltage and capillary potential) on the obtained signal were assessed and the parameters were optimized to provide maximum sensitivity and detectability. In addition, the performance of three commercial SPE sorbents (C18, PS-DVB and hydroxylated PS-DVB) was evaluated using spiked water and milk, diluted with a mixture of water-methanol (8:1). By using C18 cartridges and BPA-d(16) as internal standard, the mean relative recoveries at three fortification levels ranged between 97 and 104% and the corresponding inter-day precision (RSD%) was below 6% for 50 and 500 ng/g and below 20% for 5 ng/g fortification levels. It is shown that the ion suppression during ESI, the losses from the sample preparation procedure and the inter-day instability of LC-ESI-MS were overcome by the use of the deuterated internal standard. The concentration of BPA found in commercial canned milk samples ranged from <1.7 to 15.2 ng/g.

Development and validation of a novel LC non-derivatization method for the determination of amikacin in pharmaceuticals based on evaporative light scattering detection.

A novel method for the direct determination of the aminoglycoside antibiotic amikacin and its precursor component kanamycin was developed and validated, based on reversed phase LC with evaporative light scattering detector (ELSD). ELSD response to amikacin was found to be enhanced by: (a) use of ion-pairing acidic reagents of increased molecular mass, (b) increase of mobile phase volatility and (c) decrease of peak width and asymmetry (obtained by controlling the mobile phase acidity and/or ratio of organic solvent to water). Utilizing a Thermo Hypersil BetaBasic C(18) column, the selected optimized mobile phase was water-methanol (60:40, v/v), containing 3.0 mll(-1) nonafluoropentanoic acid (18.2mM) (isocratic elution with flow rate of 1.0 mlmin(-1)). ELSD experimental parameters were: nitrogen pressure 3.5 bar, evaporation temperature 50 degrees C, and gain 11. Amikacin was eluted at 8.6 min and kanamycin at 10.4 min with a resolution of 1.5. Logarithmic calibration curves were obtained from 7 to 77 microgml(-1) (r>0.9995) for amikacin and 8 to 105 microgml(-1) (r>0.998) for kanamycin, with a LOD equal to 2.2 and 2.5 microgml(-1), respectively. In amikacin sulfate pharmaceutical raw materials, the simultaneous determination of sulfate (t(R)=2.3 min, LOD=1.8 microgml(-1), range 5-40 microgml(-1), %R.S.D.=1.1, r>0.9997), kanamycin and amikacin was feasible. No significant difference was found between the results of the developed LC-ELSD method and those of reference methods, while the mean recovery of kanamycin from spiked samples (0.5%, w/w) was 97.3% (%R.S.D.

Determination of L-carnitine in food supplement formulations using ion-pair chromatography with indirect conductimetric detection.

A novel method for the determination of L-carnitine in food supplement formulations was developed and validated, using ion-pair chromatography with indirect conductimetric detection. The chromatographic method was based on a non-polar (C18) column and an aqueous octanesulfonate (0.64 mM) eluent, acidified with trifluoroacetic acid (5.2 mM). The retention time was 5.4 min and the asymmetry factor 0.65. A linear calibration curve from 10 to 1000 microg/ml (r= 0.99998), with a detection limit of 2.7 microg/ml (25 microl injection volume), a repeatability %RSD of 0.8 (40 microg/ml, n = 5) and reproducibility %RSD of 2.6 were achieved. The proposed method was applied for the determination of carnitine in oral solutions and capsules. No interference from excipients was found and the only pretreatment step required was the appropriate dilution with the mobile phase. Recovery from spiked samples was ranged from 97.7 to 99.7% with a precision (%RSD, n = 3) of 0.01-2.1%.

Enhancement of evaporative light scattering detection in high-performance liquid chromatographic determination of neomycin based on highly volatile mobile phase, high-molecular-mass ion-pairing reagents and controlled peak shape.

In the frame of the development of a novel HPLC-ELSD (evaporative light scattering detection) method for the determination of the aminoglycoside antibiotic neomycin sulfate, the influence of mobile phase composition and peak broadening on ELSD response was evaluated. ELSD response was enhanced by: (a) increase of mobile phase volatility (solvents examined: water, acetonitrile, methanol and acetone), (b) increase of molecular mass of ion-pairing species [acidic reagents tested: formic, acetic, trifluoroacetic, trichloroacetic and heptafluorobutyric acid (HFBA)], and (c) decrease of peak width and asymmetry obtained by controlling the concentration of the ion-pairing acidic reagent (HFBA). Utilizing a Waters ODS-2 C18 Spherisorb column, evaporation temperature of 45 degrees C and nitrogen pressure of 3.5 bar, the optimized mobile phase was water-acetone (50:50), containing 11.6 mM HFBA, in an isocratic mode at a rate of 1.0 ml/min. Neomycin was eluted at 4.9 min, with asymmetry factor 1.3. Logarithmic calibration curve was obtained from 2 to 50 microg/ml (r > 0.9997). Limit of detection (LOD) was 0.6 microg/ml and R.S.D. = 1.7% (n = 3, 3.3 microg/ml). In raw materials, the simultaneous determination of sulfate (LOD = 3 microg/ml, R.S.D. = 1.7%, r> 0.9998) and of minor impurities was feasible. The developed method was also applied for the determination of neomycin in pharmaceutical formulations (powder, aerosol and cream) without any interference from excipients (recovery from spiked samples ranged from 99 to 102%) and a %R.S.D. of <2.1 (n = 3). The HPLC-ELSD method was also found applicable in the determination of neomycin in animal feeds (LOQ=0.2%) without any interference from the feed matrices.

Ion-chromatographic determination of carbocisteine in pharmaceuticals based on non-suppressed conductimetric detection.

A novel method for the determination of carbocisteine (S-CMC), a mucolytic and expectorant drug with an acidic amino acid structure, was developed and validated, using non-suppressed ion-chromatographic system with conductimetric detection, and anion or cation exchange columns. Among the various combinations of column type and eluent composition tested, a cation exchange column with a 0.25 mM tri-fluoroacetic acid (TFA) as eluent in isocratic mode at 1.2 ml/min gave the best results. S-CMC was very well separated from all common amino acids (resolution > 2.6). The retention time was 3.5 min and the asymmetry factor 1.1. A linear calibration curve from 17 to 400 microg/ml (r = 0.99994), with a detection limit of 0.14 microg (5.6 microg/ml-25 microl injection volume) and a precision of 1.5% R.S.D. (100 microg/ml, n = 3) was achieved. The proposed method was applied for the determination of S-CMC content in intensely colored commercial formulations (syrups). No interference from excipients was found and the only pretreatment step was the appropriate dilution with the mobile phase. Recovery from standard additions was ranged from 96.0 to 104.9% and precision (R.S.D., n = 3) 1.8-3.6%.

Comparison of performance of C18 monolithic rod columns and conventional C18 particle-packed columns in liquid chromatographic determination of Estrogel and Ketoprofen gel.

The performance of monolithic HPLC columns Chromolith (made by Merck, Germany) and conventional C18 columns Discovery (Supelco, Sigma-Aldrich, Prague, Czech Republic) was tested and the comparison for two topical preparations Ketoprofen gel and Estrogel gel was made. The composition of mobile phases - for Ketoprofen analysis a mixture of acetonitrile, water and phosphate buffer adjusted to pH 3.5 (40:58:2) and for Estrogel analysis a mixture of acetonitrile, methanol, water (23:24:53) - was usually not optimal for analyses at all types of columns. Thus an adjustment of components ratio was necessary for sufficient resolution of the compounds analysed. Various flow rates (1.0-5.0 ml/min) and mobile phases (usually increasing ratio of water content) were applied. Determination of active substances, preservatives and impurities and comparison of retention times and system suitability test parameters was accomplished. For Estrogel gel, following chromatographic conditions were found: using Chromolith Flash RP-18e monolith column, mobile phase was acetonitrile, methanol, water (13:24:63, v/v/v) and flow-rate 3.0 ml/min. Using monolith column ChromolithSpeedROD RP-18e, the mobile phase was acetonitrile, methanol, water (18:24:58, v/v/v) and flow-rate 4.0 ml/min. For the monolith column Chromolith Performance RP-18e, the mobile phase was acetonitrile, methanol, water (23:24:53, v/v/v), flow-rate 3.0ml/min. Analysis of Ketoprofen gel gave the best results using following analytical conditions: for monolith column Chromolith Flash RP-18e, mobile phase as a mixture of acetonitrile, water, phosphate buffer pH 3.5 (30:68:2, v/v/v) was used, at flow-rate 2.0 ml/min. For ChromolithSpeedROD RP-18e monolith column, acetonitrile, water, phosphate buffer pH 3.5 (35:63:2, v/v/v) was used as a mobile phase at flow-rate 3.0 ml/min. Chromolith Performance RP-18e gave the best results using mobile phase acetonitrile, water, phosphate buffer pH 3.5 (30:68:2, v/v/v) at the flow-rate 5.0 ml/min. It was proved that monolith columns, due to their porosity and low back-pressure, can save analysis time by about a factor of three with sufficient separation efficiency. Thus, for example 11 min long analysis can be performed in 4 min with comparable results.

Development and validation of a novel LC/ELSD method for the quantitation of gentamicin sulfate components in pharmaceuticals.

The equivalent response of evaporative light scattering detector (ELSD) for compounds of similar structure is exploited to develop an LC/ELSD method for the simultaneous quantitation of the four main components of gentamicin sulfate, using as external standard the one main component kanamycin. A C18 column was used along with a mobile phase consisting of H2O (containing 35.4 microg/ml of trichloroacetic acid and 0.89 microl/ml of trifluoroacetic acid)-methanol-acetonitrile (990:5:5, v/v/v), in an isocratic mode at 1.1 ml/min. Parameters of ELSD were 50 degrees C for evaporation temperature and 3.0 bar for pressure of carrier gas (N2). A logarithmic calibration curve was obtained for sulfate (tR = 1.9 min) from 4.2 to 150 microg/ml (r > 0.994) with a precision of 0.18%R.S.D. Kanamycin and the four gentamicin components (C(1a), C2, C(2a) and C1) were eluted at 3.2, 4.6, 5.9, 7.1 and 8.7 min, respectively, with good resolution (Rs > 1.5). Logarithmic calibration curve was obtained for each component (r > 0.99) with statistically equal slopes varying from 2.457 to 2.558. The mass range of total gentamycin was 35-240 microg/ml. The proposed method was applied for the determination of gentamicin components and sulfate in raw materials and pharmaceutical formulations (injection, drops and cream) without any pretreatment except cream, for which liquid-liquid extraction was required. Recovery from standard addition experiments in commercial formulations was 99-100% regarding total gentamicin and 89-108% regarding individual components, with a precision (%RSD, n = 4) 0.7-5.8%.

A novel derivatization method for the determination of Fosfomycin in human plasma by liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometric detection via phase transfer catalyzed derivatization.

An analytical method employing novel sample preparation and liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometric detection (LC-APCI/MS) was developed for the determination of fosfomycin in human plasma. Sample preparation involves derivatization through phase transfer catalysis (PTC) which offers multiple advantages due to the simultaneous extraction, preconcentration and derivatization of the analyte. Using a PT catalyst, fosfomycin was extracted from plasma in an organic phase and, then converted to a pentafluorobenzyl ester with the use of pentafluorobenzyl bromide (PFBBr) derivatization reagent. The method was fully optimized by taking into account both PTC and derivatization parameters. Several catalysts, in a wide range of concentrations, with different counter ions and polarities were tested along with different extraction solvents and pH values. Thereafter, the derivatization procedure was optimized by altering the amount of the derivatization reagent, the temperature of the reaction and finally, the derivatization duration. As internal standard (I.S.) ethylphosphonic acid was chosen and underwent the same pretreatment. The derivatives were separated on a pentafluorophenyl (PFP)-C18 analytical column, which provides unique selectivity, using an isocratic elution with acetonitrile-water (70-30, v/v). The method was validated according to US Food and Drug Administration (FDA) guidelines and can be used for a bioequivalence study of fosfomycin in human plasma. The correlation coefficient (r(2)) of the calibration curve of spiked plasma solutions in the range of 50-12000 ng/mL was found greater than 0.999 with a limit of quantitation (LOQ) equal to 50 ng/ml (for 500 µL plasma sample).

A generic screening methodology for horse doping control by LC-TOF-MS, GC-HRMS and GC-MS.

In the present study a general screening protocol was developed to detect prohibited substances and metabolites for doping control purposes in equine sports. It was based on the establishment of a unified sample preparation and on the combined implementation of liquid and gas chromatographic MS analysis. The sample pretreatment began with two parallel procedures: enzymatic hydrolysis of sulfate and glucuronide conjugates, and methanolysis of the 17ß-sulfate steroid conjugates. The extracts were treated for LC-TOF-MS, GC-HRMS and GC-MS assays. The majority of the prohibited substances were identified through a high mass accuracy technique, such as LC-TOF-MS, without prior derivatization. The sample preparation procedure included the formation of methylated and trimethylsilylated derivatives common in toxicological GC-MS libraries. The screening method was enhanced by post-run library searching using automated mass spectral deconvolution and identification system (AMDIS) combined with deconvolution reporting software (DRS). The current methodology is able to detect the presence of more than 350 target analytes in horse urine and may easily incorporate a lot of new substances without changes in chromatography. The full scan acquisition allows retrospective identification of prohibited substances in stored urine samples after reprocessing of the acquired data. Validation was performed for sixty representative compounds and included limit of detection, matrix interference - specificity, extraction recovery, precision, mass accuracy, matrix effect and carry over contamination. The suitability of the method was demonstrated with previously declared positive horse urine samples.

Recent trends in biomonitoring of bisphenol A, 4-t-octylphenol, and 4-nonylphenol.

Bisphenol A (BPA), 4-t-octylphenol (4-t-OP), and 4-nonylphenol (4-NP) are man-made alkylphenolic environmental contaminants possessing controversial endocrine disruption properties. Nowadays, an increased interest is raised for their accurate determination in biological media in order to estimate the exposure to these compounds and the associated health risk. The aim of this review is to present the available analytical methodologies for biomonitoring these three EDCs in human population. In non-occupational human exposure, they are detected in human matrices in trace level concentrations, commonly lower than 1ng/mL. The use of mass spectrometry based methods is particularly emphasized due to their well known superiority over sensitivity, selectivity and precision, even in difficult matrices, such as blood plasma and serum. Recent and most applicable sample preparation techniques are thoroughly presented. The benefits of solid phase extraction (SPE) and expected developments are demonstrated. Recent results from exposure assessment and epidemiologic studies for BPA, 4-t-OP and 4-NP are summarized and future trends are discussed.

Optimization and comparison of ESI and APCI LC-MS/MS methods: a case study of Irgarol 1051, Diuron, and their degradation products in environmental samples.

A systematic and detailed optimization strategy for the development of atmospheric pressure ionization (API) LC-MS/MS methods for the determination of Irgarol 1051, Diuron, and their degradation products (M1, DCPMU, DCPU, and DCA) in water, sediment, and mussel is described. Experimental design was applied for the optimization of the ion sources parameters. Comparison of ESI and APCI was performed in positive- and negative-ion mode, and the effect of the mobile phase on ionization was studied for both techniques. Special attention was drawn to the ionization of DCA, which presents particular difficulty in API techniques. Satisfactory ionization of this small molecule is achieved only with ESI positive-ion mode using acetonitrile in the mobile phase; the instrumental detection limit is 0.11 ng/mL. Signal suppression was qualitatively estimated by using purified and non-purified samples. The sample preparation for sediments and mussels is direct and simple, comprising only solvent extraction. Mean recoveries ranged from 71% to 110%, and the corresponding (%) RSDs ranged between 4.1 and 14%. The method limits of detection ranged between 0.6 and 3.5 ng/g for sediment and mussel and from 1.3 to 1.8 ng/L for sea water. The method was applied to sea water, marine sediment, and mussels, which were obtained from marinas in Attiki, Greece. Ion ratio confirmation was used for the identification of the compounds.

Hydrophilic interaction vs ion pair liquid chromatography for the determination of streptomycin and dihydrostreptomycin residues in milk based on mass spectrometric detection.

Streptomycin (STR) and dihydrostreptomycin (DHSTR) are two of the most common aminoglycoside antibiotics used in veterinary medicine. The physicochemical properties of both substances, make their determination challenging. In the present study the development of methods based on ion-pair chromatography (IPC) and on hydrophilic interaction chromatography (HILIC), for the determination of the above mentioned aminoglycosides in the range of 100-1000 µg L(-1) is described. The two methods were validated according to EU requirements for residues in food. The recoveries for the IPC method were 69.3% and 56.5% of STR and DHSTR, respectively, and for HILIC method 85.5% and 72.3%, respectively. The intra- and inter-day precision, studied at 100, 200 and 300 µg kg⁻¹ levels in milk samples, gave %RSD ≤ 13 for both methods. LOQs for the HILIC method were 14 µg kg⁻¹ for both analytes and for the IPC method were 109 and 31 µg kg⁻¹, for STR and DHSTR, respectively. The sensitivity of the HILIC method is 80 and 210 times greater than that of the ICP method, for STR and DHSTR, respectively.