Fast and Sensitive Screening of Oxandrolone and Its Major Metabolite 17-Epi-Oxandrolone in Human Urine by UHPLC-MS/MS with On-Line SPE Sample Pretreatment.

Oxandrolone, a synthetic testosterone analog, is used for the treatment of several diseases associated with weight loss. Unfortunately, oxandrolone is abused by many athletes and bodybuilders due to its strong anabolic effect. We have developed and validated a highly sensitive and rapid on-line SPE-UHPLC-MS/MS method for the determination of oxandrolone and simultaneous identification of its major metabolite 17-epi-oxandrolone in urine matrices. Enrichment of the analytes via an integrated solid-phase extraction was achieved using an Acquity UPLC BEH C18 Column. Subsequently, the chromatographic separation of the on-line preconcentrated sample fraction was achieved using an Acquity HSS T3 C18 Column. For the structural identification of these analytes, a high-resolution mass spectrometer Synapt-G2Si coupled to the Acquity M-class nano-LC system with ionKey source was used. A highly sensitive determination of oxandrolone was achieved using a tandem quadrupole mass spectrometer XEVO TQD. The method was successfully validated in the linear range of oxandrolone from 81.63 pg·mL-1 (limit of quantification, LOQ) to 5000 pg·mL-1 in the human urine matrix. It was applied to the analysis of real urine samples obtained from a healthy volunteer after the oral administration of one dose (10 mg) of oxandrolone. Concentration vs. time dependence was tested in the time interval of 4 h-12 days (after oral administration) to demonstrate the ability of the method to detect the renal elimination of oxandrolone from the human body. Favorable performance parameters along with successful application indicate the usefulness of the proposed method for its routine use in antidoping control labs.

A rapid method for on-line solid-phase extraction and determination of dioscin in human plasma using a homemade monolithic sorbent combined with high-performance liquid chromatography.

A phenyl-based polymer monolithic column was prepared via free radical polymerization in a stainless steel column with the size of 4.6 mm i.d. × 50 mm, using ethylene glycol phenyl ether acrylate as the monomer. The resulting monolithic column shows high porosity of 73.42% and relative uniform pore structure, as characterized by mercury porosimetry and scanning electron microscopy, respectively. The optimized polymer monolith column was used for on-line solid-phase extraction prior to the reversed phase mode HPLC-UV analysis for the determination of dioscin in human plasma, using a COSMOSIL C18 column (4.6 mm × 150 mm, 4.5 µm). Water was used to wash non-retained components from the SPE sorbent, and methanol water (80:20, V/V) was used as the mobile phase for isocratic elution of dioscin. The maximum adsorbed quantity of dioscin to the SPE column is 6.79 mg/g, which is high enough for the quantitative analysis of dioscin in plasma, due to the low content of dioscin in plasma. The method was validated by assessing the linearity, lower limit of quantification, intra- and inter-day precision, accuracy, and repeatability. The developed method was applied for the analysis of dioscin in plasma from a volunteer who had orally administered an aqueous extract of dioscorea nipponica rhizome, showing the method capable of detecting dioscin in the plasma. These results show that the developed method is a rapid method for on-line solid-phase extraction and determination of dioscin from plasma, exhibiting good selectivity with hydrogen bond interaction and hydrophobic interaction, good clean-up ability, cost-saving, and time-saving. Graphical abstract.

Molecularly imprinted vs. reversed-phase extraction for the determination of zearalenone: a method development and critical comparison of sample clean-up efficiency achieved in an on-line coupled SPE chromatography system.

Sample preparation prior to chromatographic separation plays an important role in the analytical process. To avoid time-consuming and manual handling sample-prep, automated on-line techniques such as on-line SPE-HPLC are therefore preferred. In this study, two different on-line extraction approaches for mycotoxin/endocrine disruptor zearalenone (ZEA) determination using either molecularly imprinted polymer (MIP) with selective cavities and binding sites for extraction or a reversed-phase sorbent C18 providing non-selective interactions have been developed, validated, and compared. The validation characteristics were compared and the two methods were evaluated as being almost equal in terms of linearity, repeatability, precision, and recovery. Recoveries were in the range of 99.0-100.1% and limits of detection were found the same for both methods (1.5 µg L-1). Method precision calculated for spiked beer samples was better for C18 sorbent (2.5 vs. 5.4% RSD). No significant differences in the selectivity of either extraction method were observed. The possible reasons and further details associated with this finding are discussed. Finally, both validated methods were applied for the determination of ZEA contamination in beer samples. Due to ZEA's native fluorescence, chromatographic separation with fluorimetric detection (λex = 270 nm and λem, = 458 nm) was selected. Graphical abstract Determination of zearalenone in beer using an on-line extraction chromatography system.

A high throughput targeted and non-targeted method for the analysis of microcystins and anatoxin-A using on-line solid phase extraction coupled to liquid chromatography-quadrupole time-of-flight high resolution mass spectrometry.

Microcystins are cyclic heptapeptide hepatotoxins produced by cyanobacteria in freshwater. Sample preparation for the analysis of these cyanotoxins in water from algal blooms can take up to several days due to the matrix complexity and the low detection limits required to comply with current legislation. Moreover, there is a large number of unknown microcystins that could potentially exist in the environment resulting from different amino acid substitutions into the microcystin skeletal structure. To tackle these problems, the present study involved the development of a high throughput method based on on-line solid phase extraction coupled to liquid chromatography that could provide quantitative results for 12 microcystin variants (LR, YR, RR, HtyR, HilR, WR, LW, LA, LF, LY, Dha7-LR, and Dha7-RR) and anatoxin-A in less than 3 h with detection limits between 0.004 and 0.01 µg L-1 and expanded uncertainty between 4 and 14%. Data-dependent acquisition was employed for the non-targeted analysis of these cyanotoxins. Filtering the data based on structure diagnostic fragments, two unknown microcystin variants not previously reported in the literature were detected. The structures Leu1-microcystin-Met(O)R and Leu1-microcystin-LY were fully characterized by accurate mass measurement, collision-induced dissociation, and fragmentation prediction software.

Targeted lipidomics profiling of acute arsenic exposure in mice serum by on-line solid-phase extraction stable-isotope dilution liquid chromatography-tandem mass spectrometry.

Lipidomics is the area of study dedicated to the comprehensive analysis and characterization of the functions and metabolism of lipids in biological samples. One of the most comprehensively studied classes of lipids is polyunsaturated fatty acids (PUFAs). Eicosanoids are a series of bioactive lipid mediators that are metabolized from PUFAs and generated majorly from the precursor arachidonic acid. This study identified the profiles of target eicosanoids after acute exposure to arsenic. The principle objective was to determine and validate 10 eicosanoids in mouse serum using on-line solid-phase extraction integrated with liquid chromatography-electrospray tandem mass spectrometry. Intra-day and inter-day repeatability was 82.4-119.2 and 86.7-124.4%, respectively. The limit of detection and limit of quantification were from 0.003 to 0.288 ng/mL and from 0.009 to 0.962 ng/mL, respectively. The levels of 7 of the 10 eicosanoids-namely 8-isoPGF2α, PGF2α, PGE2, 13(s)-HODE, 15(s)-HETE, 12(s)-HETE, and 5(s)-HETE-in mouse serum significantly and dose-dependently increased after arsenic exposure compared with the levels in the vehicle control group. To our knowledge, this is the first study to quantify eicosanoids in mouse serum. This approach provides simple sample preparation, small sample volumes, and a precise and accurate method for determining changes in the profile of these eicosanoids in mouse serum after acute exposure to arsenic.

Developments in coupled solid-phase extraction-capillary electrophoresis 2013-2015.

An overview of the design and application of coupled solid-phase extraction-capillary electrophoresis (SPE-CE) systems reported in the literature between July 2013 and June 2015 is provided in this paper. The present article is a continuation of our previous review papers on this topic which covered the time period 2000-2013 (Electrophoresis 2008, 29, 108-128; Electrophoresis 2010, 31, 44-54; Electrophoresis 2012, 33, 243-250; Electrophoresis 2014, 35, 128-137). The use of in-line and on-line SPE-CE approaches is treated and outlined in this review. Recent advancements, such as, for example, the use of aptamers as affinity material for in-line SPE-CE, the use of a bead string design for in-line fritless SPE-CE, and new interfacing techniques for the on-line coupling of SPE to CE, are outlined. Selected examples demonstrate the applicability of the coupled SPE-CE systems for biomedical, pharmaceutical, environmental, and food studies. A complete overview of the recent SPE-CE studies is given in table format, providing information on sample type, SPE sorbent, coupling mode, detection mode, and LOD. Finally, some general conclusions and perspectives are provided.

Ultra-trace levels analysis of microcystins and nodularin in surface water by on-line solid-phase extraction with high-performance liquid chromatography tandem mass spectrometry.

An on-line solid phase extraction coupled with high-performance liquid chromatography in tandem with mass spectrometry (on-line SPE/HPLC/MS-MS) method for the determination of five microcystins and nodularin in surface waters at submicrogram per liter concentrations has been optimized. Maximum recoveries were achieved by carefully optimizing the extraction sample volume, loading solvent, wash solvent, and pH of the sample. The developed method was also validated according to both UNI EN ISO IEC 17025 and UNICHIM guidelines. Specifically, ten analytical runs were performed at three different concentration levels using a reference mix solution containing the six analytes. The method was applied for monitoring the concentrations of microcystins and nodularin in real surface water during a sampling campaign of 9 months in which the ELISA method was used as standard official method. The results of the two methods were compared showing good agreement when the highest concentration values of MCs were found. Graphical abstract An on-line SPE/HPLC/MS-MS method for the determination of five microcystins and nodularin in surface waters at sub µg L(-1) was optimized and compared with ELISA assay method for real samples.

Simultaneous, rapid, and sensitive quantification of 8-hydroxy-2'-deoxyguanosine and cotinine in human urine by on-line solid-phase extraction LC-MS/MS: correlation with tobacco exposure biomarkers NNAL.

Cigarette smoke can increase oxidative DNA damage. The main component in cigarette smoke is nicotine. Nicotine is metabolized to cotinine, which can be regarded as a biomarker for measuring exposure to tobacco smoke. A sensitive, simple, and robust method based on on-line solid-phase extraction liquid chromatography with tandem mass spectrometry (on-line SPE LC-MS/MS) has been developed and validated for the simultaneous determination of 8-OHdG and cotinine. The matrix effects of 8-OHdG and cotinine were measured at 97.1 and 91.7 %, with values for CV at 4.4 and 4.2 %, respectively. The limits of detection of 8-OHdG and cotinine were 10.0 and 5.5 pg mL(-1), and the limits of quantification were 40.0 and 20.0 pg mL(-1), respectively. The total run time was 12 min. We quantified 8-OHdG and cotinine in the urine of 80 male subjects. The results showed the levels of 8-OHdG and cotinine in smokers were significantly higher than that in non-smokers. Furthermore, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and its glucuronide conjugate (defined as total NNAL) are the nitrosation metabolites of nicotine. In this study, urinary levels of 8-OHdG and cotinine were well correlated with urinary levels of total NNAL. This is also the first study to focus on the future risk of oxidative stress from exposure to cigarette smoke based on the relationship between 8-OHdG levels, cotinine levels, and total NNAL concentrations in the urine of humans. Graphical Abstract On-line SPE LC-MS/MS for the simultaneous determination of 8-OHdG and cotinine in human urine.

A fully automated and fast method using direct sample injection combined with fused-core column on-line SPE-HPLC for determination of ochratoxin A and citrinin in lager beers.

A new fast and sensitive method based on on-line solid-phase extraction on a fused-core precolumn coupled to liquid chromatography with fluorescence detection has been developed for ochratoxin A (OTA) and citrinin (CIT) determination in lager beer samples. Direct injection of 100 µL filtered beer samples into an on-line SPE-HPLC system enabled fast and effective sample extraction including separation in less than 6 min. Preconcentration of OTA and CIT from beer samples was performed on an Ascentis Express RP C18 guard column (5 × 4.6 mm), particle size 2.7 µm, with a mobile phase of methanol/0.5% aqueous acetic acid pH 2.8 (30:70, v/v) at a flow rate of 2.0 mL min(-1). The flow switch from extraction column to analytical column in back-flush mode was set at 2.0 min and the separation was performed on the fused-core column Ascentis Express Phenyl-Hexyl (100 × 4.6 mm), particle size 2.7 µm, with a mobile phase acetonitrile/0.5% aqueous acetic acid pH 2.8 in a gradient elution at a flow rate of 1.0 mL min(-1) and temperature of 50 °C. Fluorescence excitation/emission detection wavelengths were set at 335/497 nm. The accuracy of the method, defined as the mean recoveries of OTA and CIT from light and dark beer samples, was in the range 98.3-102.1%. The method showed high sensitivity owing to on-line preconcentration; LOQ values were found to be 10 and 20 ng L(-1) for OTA and CIT, respectively. The found values of OTA and CIT in all tested light, dark and wheat beer samples were significantly below the maximum tolerable limits (3.0 µg kg(-1) for OTA and 2000 µg kg(-1) for CIT) set by the European Union.

Study of spatial and temporal distribution of antimicrobial in water and sediments from caging fish farms by on-line SPE-LC-MS/MS.

An on-line solid phase extraction liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) method for the determination of 12 antimicrobials in sediment and surface water was developed and validated. Furthermore, the spatial and temporal antimicrobials distributions in the sediment and in the water of four fish farms located in the hydroelectric dam of Ilha Solteira Reservoir in Brazil were investigated over four seasons in three sampling sites: at the fish cages, 100 and 1,000 m downstream far from the cages. The method was performed using an Agilent Zorbax 80 SB-C8 column (9.4 × 15 mm, 5 µm) as the loading column, and the Agilent Zorbax Eclipse Plus C18 column (3.0 × 100 mm, 3.5 µm) as a separation column within a run time of 13 min. The limits of quantification were less than 9 ng·L(-1) for the antibiotics in water and 16 µg·kg(-1) in sediment; the recovery ranged from 80 to 119%, with a variation coefficient less than 11%, and the repeatability was lower than 15%. Oxytetracycline was found in the water in all sample seasons. However, florfenicol was identified in April and October 2013 and January 2014, and tetracycline was present in July 2013. Regarding the sediment, oxytetracycline and tetracycline were found in all sampling periods, but chlortetracycline was only identified in January 2014. The spatial distribution of antimicrobials showed that the main pollution source came from the fish farms. This study demonstrated that the proposed method is reliable for the monitoring of antimicrobials in water and sediments and it showed contamination in both matrices from Ilha Solteira Reservoir.

Developments in coupled solid-phase extraction-capillary electrophoresis 2011-2013.

This article presents an overview of the design and application of coupled SPE-CE systems that have been reported in the literature between January 2011 and June 2013. The present paper is an update of three previous review papers covering the years 2000-2011 (Electrophoresis 2008, 29, 108-128; Electrophoresis 2010, 31, 44-54; Electrophoresis 2012, 33, 243-250). The use of in-line and on-line SPE-CE approaches is described in this review. Emerging technological developments, such as the use of in-line frit-free SPE and chip-based SPE for extraction of sample components prior to CE analysis, are outlined. Selected examples illustrate the applicability of SPE-CE in biomedical, pharmaceutical, and environmental analysis. A complete overview of recent SPE-CE studies is given in table format, providing information on sample type, SPE sorbent, coupling mode, detection mode, and LOD. Finally, some general conclusions and future perspectives are provided.

Fast analysis of short-chain and ultra-short-chain fluorinated organics in water by on-line extraction coupled to HPLC-HRMS.

A rapid on-line solid-phase extraction liquid chromatography high-resolution mass spectrometry (on-line SPE-LC-HRMS) method was developed to analyze 11 ultra-short and short-chain PFAS in surface water. Analytical optimization involved screening 7 chromatographic columns and 5 on-line SPE columns, as well as evaluating SPE loading conditions, filters, sample acidification, chromatographic mobile phases, and SPE loading mobile phases. The optimized method was then applied to 44 river water samples collected in Eastern Canada, including sites near airports with fire-training areas. Among the 11 targeted PFAS, the most frequently detected were trifluoroacetic acid (TFA, 4.6-220 ng/L), perfluorobutanoic acid (PFBA, 0.85-33 ng/L), perfluoropentanoic acid (PFPeA, 1.2-2100 ng/L), trifluoromethane sulfonic acid (TMS, 0.01-4.3 ng/L), and perfluorobutane sulfonic acid (PFBS, 0.07-450 ng/L). Levels of C3-C5 perfluoroalkyl carboxylic acids (PFCAs), C2-C4 perfluoroalkyl sulfonates (PFSAs) and n:3 polyfluoroalkyl acids (n = 2,3; n:3 acids) were significantly higher in water bodies near fire-training area sites compared with rivers in urban areas. In contrast, TFA, TMS, and 1:3 acid were not significantly elevated, likely reflecting atmospheric deposition or other diffuse sources for these compounds. Nontarget and suspect screening analysis revealed an abundance of other ultra-short and short-chain PFAS in AFFF-impacted water bodies. Perfluoroalkyl sulfonamides (FASA, C2, C3, and C5), perfluoroalkyl sulfonamide propanoic acids (FASA-PrA, C1-C2) and n:3 acids (n = 1, 4, and 5) were detected for the first time in environmental surface waters.

Development and validation of a bioanalytical method for the quantification of methotrexate from serum and capillary blood using volumetric absorptive microsampling (VAMS) and on-line solid phase extraction (SPE) LC-MS.

High-dose methotrexate is part of the polychemotherapy protocols for the treatment of Acute lymphoblastic leukaemia (ALL) with therapeutic drug monitoring (TDM) to adjust leucovorin rescue. An immunoassay is commonly used to analyse serum samples collected via venous blood sampling. However, immunoassays cannot distinguish between the parent drug and its metabolites. Besides, the blood volume required by venous blood sampling is high. Therefore, the aim of this project was to develop a fast, simple, reliable and cost-efficient micro sampling bioanalytical method using capillary blood to minimize the harm of children and to analyse both methotrexate and its metabolites. To achieve this aim, a LC-MS method with on-line solid phase extraction (SPE) for the simultaneous detection of methotrexate and its metabolites from capillary blood using volumetric-absorptive-microsampling (VAMS) technology was developed and fully validated. Besides, the method was also validated and modified for serum samples to compare the results with the immunoassay. A single-quadrupole MS detector was used for detection. Through the use of on-line SPE technology, a lower limit of quantitation of 0.03 µM for MTX and 7-OH-MTX and of 0.05 µM for DAMPA from a 10 µL capillary blood sample was achieved. The accuracy is between 90.0 and 104% and the precision between 4.7 and 12% for methotrexate and its metabolites, respectively. Because of the cross reactivity of the immunoassay a cross-validation was not successful. Besides, a correlation factor of 0.46 for MTX between plasma and whole-blood was found. A fast, simple, reliable and cost-efficient extraction and analysis LC-MS method could be developed and validated, which is applicable in ambulatory and clinical care.

Comparing adsorption performance of microfibers and nanofibers with commercial molecularly imprinted polymers and restricted access media for extraction of bisphenols from milk coupled with liquid chromatography.

Advanced solid phase extraction (SPE) fibrous sorbents including polyethylene, polypropylene poly (hydroxybutyrate), and polyamide 6 nanofibers, polycaprolactone microfibers/nanofibers, polycaprolactone microfibers/polyvinylidene difluoride nanofibers, and poly (hydroxybutyrate) microfibers/polypropylene microfibers composites, as well as commercial molecularly imprinted polymers and restricted access media sorbent were compared in terms of bisphenols extraction from milk and their clean-up efficiency. Three on-line SPE-HPLC methods were completely validated for the extraction and detection of bisphenols A, AF, C, A diglycidyl ether, and F diglycidyl ether in bovine milk. Polycaprolactone composite nanofibers compared favorably to restricted access media, enabled excellent clean-up of bisphenols from the proteinaceous matrix, and yielded recoveries 98.0-124.5% and 93.0-115.0%, respectively, with RSD less than 10%. Total analysis time including on-line SPE step lasted only 12 min, which represents a significant reduction in time compared with previously reported as well as official European Union and AOAC methods defined for the determination of bisphenols in various matrices.

Detection, occurrence, influencing factors and environmental risks of paralytic shellfish toxins in seawater in a typical mariculture bay.

Paralytic shellfish toxins (PSTs) producing algae are widely distributed in the global coastal aquatic environment, posing a threat to coastal ecosystem health and mariculture safety. However, the levels and potential environmental risks of PSTs frequently detected in shellfish remain largely unexplored in seawater of mariculture zones. In this study, a new method for trace detection of 13 common PSTs (<1.0 ng/L) in seawater was established based on off-line solid phase extraction (SPE) and on-line SPE-liquid chromatography-tandem mass spectrometry (on-line SPE-LC-MS/MS), and a systematic investigation of PSTs in seawater of the Laizhou Bay, a typical aquaculture bay in China, was conducted to understand their pollution status, environmental impact factors and ecological risks for the first time. Eleven PSTs were detected in the seawater of Laizhou Bay with total concentrations ranging from 0.75 to 349.47 ng/L (mean, 176.27 ng/L), which indicates the rich diversity of PSTs in the mariculture bay and demonstrates the reliability of the proposed analytical method. C1, C2, GTX2, GTX3, dcGTX2, and dcGTX3 were found to be the predominant PSTs, which refreshed the knowledge of PST contamination in the coastal aquatic environment. PST levels in seawater exhibited the highest levels in the southeastern mouth of Laizhou Bay and decreased toward the inner bay. Correlation analyses showed that climatic factors, nutrient status and hydrological conditions had significant effects on the distribution of PST in mariculture bay. Preliminary environmental risk assessments revealed that aquatic organisms throughout the waters of Laizhou Bay are at risk of chronic PST toxicity. These findings imply that the risk of PST in seawater of mariculture bay has previously been grossly underestimated, and that the coastal aquatic environment in North China and even the world may be at more serious risk of PST pollution, which should be taken seriously.

Mass Spectrometry Approaches for Detection and Determination of Prostaglandins from Biological Samples.

Accurate determination of prostaglandins (PGs) from biological samples is critical for understanding their biological functions and interactions during physiological and pathological processes. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a highly sensitive, accurate, and high-throughput approach for simultaneous detection of ultra-trace PGs from a single biological sample. Here we describe LC-MS/MS techniques and related sample pretreatment methods including both off-line and on-line SPE for the determination of PGs in biological samples.

[Application progress of on-line sample preparation techniques coupled with liquid chromatography-mass spectrometry system in the detection of food hazards].

Food safety has received increased attention, and food detection is of great significance. The food matrix is complex, and diverse food hazards have been identified. Thus, the detection methods and sample preparation techniques for food matrices must be continuously optimized and updated. Several steps are usually required when a chromatographic system is used to determine food hazards: sample preparation, that is, the separation of targets from different substrates using a suitable preprocessing method and target-substance separation and purification, which is usually achieved using chromatographic separation. The selection of an appropriate detector for qualitative and quantitative analyses is usually based on the properties of the target compound. The sample preparation procedure is considered the most time-consuming aspect of the entire food-analysis process. It is also prone to analytical errors. Therefore, optimization of the sample preparation process is a key issue in the field of chemical analysis. Researchers have developed a series of new, efficient, and accurate sample preprocessing methods, and an on-line sample-preparation system has been found to be a feasible approach. On-line sample preparation coupled with liquid chromatography-mass spectrometry (LC-MS) presents many advantages. First, manual operation could reduce analytical errors to ensure good accuracy and repeatability. It could also reduce the consumption of chemical reagents and avoid cross-contamination between samples. Furthermore, an on-line sample-preparation system could shorten the sample-preparation time and improve the detection efficiency. On-line sample preparation coupled with LC-MS has been widely applied in the fields of environment, biology, and food. On-line sample preparation systems coupled with LC-MS are divided into two modules: the first modules involves sample preparation and the second module involves the LC system. The first module remove impurities and isolates the target compounds in preparation for their qualitative and quantitative detection. The coupling of these two modules depends mainly on valve switching. In this paper, we introduce the most frequently used on-line sample-preparation techniques, including on-line solid phase extraction (on-line SPE), in-tube solid phase microextraction (in-tube SPME), and turbulent chromatography (TFC). We then describe the basic principles and coupling equipment of these three on-line analytical technologies in detail. The coupling equipment establishes a physical connection between the two modules. Next, we discuss the properties of different purification fillers in an on-line sample-preparation column. The applications and research progress of on-line systems for pesticide residues, veterinary drug residues, and biotoxins are also discussed. Compared with offline sample preparation, on-line analytical systems present several advantages. On-line analytical systems can not only greatly reduce the analysis time and solvent consumption but also improve the detection sensitivity and accuracy. Such systems can be used to determine food hazards to ensure food safety. Finally, the existing problems and development trends of on-line analytical systems are discussed and prospected. To promote the applications of on-line analytical technology in food-safety detection, we suggest that the following three aspects be considered. First, more on-line purification columns with novel fillers, in addition to C18 or polymer fillers, should be developed. Second, compared with ordinary detectors, high-resolution MS detectors have better precision and accuracy. Coupling on-line analytical technologies with a high-resolution mass spectrometer may be beneficial for the further development of on-line analyses. Third, different food matrices should be compared and evaluated to continuously optimize the detection process and improve the efficiency of on-line analytical systems. As concerns regarding food safety issues have increased, the applications of on-line analytical technologies for food detection can be expected to become increasingly important.

Fast screening of saxitoxin, neosaxitoxin, and decarbamoyl analogues in fresh and brackish surface waters by on-line enrichment coupled to HILIC-HRMS.

The proliferation of harmful cyanobacterial algal blooms is of concern due to the associated release of toxins affecting ecosystems and human health. The paralytic shellfish poison saxitoxin (STX) is a small polar alkaloid that can occur in inland and marine aquatic environments. Here, we optimized a fast and sensitive analytical method for the determination of STX, neosaxitoxin (NeoSTX), and their decarbamoyl analogues in surface waters. The method involves a simple filtration, addition of isotope-labelled internal standard (ILIS), and analysis by on-line solid-phase extraction coupled to hydrophilic interaction liquid chromatography high-resolution mass spectrometry (on-line SPE-HILIC-HRMS). Except glass fiber filters, other tested materials (e.g., nylon, nitrocellulose) provided suitable filtration performance. Time-dependent adsorptive losses occurred during the LC-MS batch sequence if glass autosampler vials were used, while no such effect was observed for polypropylene autosampler vials. Matrix effects were evaluated for 4 different quantification scenarios, including external vs. internal curves and neat reagent water vs. matrix-matched curves. Matrix-matched calibration with ILIS correction (NeoSTX-15N7) provided the best performance overall. The analytical method was validated in freshwater lake water and estuarine brackish water (30‰ salinity), with suitable determination coefficients (R2 > 0.9975), matrix spike accuracy (90-107%), and intraday/interday precision (RSD of 0.61-16%). Method limits of detection (LOD in lake water: 0.72-3.9 ng/L) are also improved over most of the recent literature. The method was applied to a set of 302 surface water samples collected in Canada, France, and the United Kingdom, and positive detections were reported for STX (max: 98 ng/L), decarbamoyl-STX (max: 15 ng/L), and NeoSTX (max: 87 ng/L).

Sample preparation and antibiotic quantification in vinasse generated from sugarcane ethanol fuel production.

Vinasse - liquid organic residue derived from the production of sugarcane ethanol fuel, has been applied as soil amendment via fertigation for decades in Brazil. This organic residue is an important source of nutrients and water for sugarcane crops. Through fertigation, approximately 400 billion liters of vinasse are recycled annually. Despite the economic and agronomic importance of this practice, vinasse-based fertigation can be a source of antibiotic contamination in the environment. The present work reports the application of solid phase extraction (SPE), salting-out liquid-liquid extraction (SALLE), and on-line solid phase extraction (on-line SPE) as sample preparation techniques for the analysis of the following antibiotics (contaminants) in vinasse sample: monensin, penicillin G, virginiamycin M1, virginiamycin S1, tetracycline and erythromycin. The study also employed a totally automated quantitative method based on on-line SPE and liquid chromatography coupled to quadrupole linear ion trap tandem mass spectrometry (LC-QqLIT-MS/MS) for the analysis of these contaminants in vinasse. The application of the aforementioned sample preparation techniques led to the successful extraction of the analytes, and a comparative analysis of the techniques showed that the on-line SPE technique was the most advantageous among the techniques investigated. The quantitative analytical method applied yielded well-defined chromatographic peaks, working range of 1.0-370.0 ng·mL-1, apparent recovery ranging from 80 to 110% for most compounds, repeatability between 3 and 16%, and limits of detection ranging from1.0 to 10 ng·mL-1. The analysis of six vinasse samples from different ethanol producing plants led to the detection of monensin at the concentration of 14.3 ng·mL-1 in their compositions. The results obtained show that fertigation with vinasse is a source of antibiotic contamination in the environment.

Trace determination of multiple hydrophilic cyanotoxins in freshwater by off- and on-line solid phase extraction coupled to liquid chromatography-tandem mass spectrometry.

Hydrophilic cyanotoxins (HCTs), such as paralytic shellfish toxins (PSTs), anatoxin-a (ATX-a), and cylindrospermopsin (CYN) are highly toxic and toxin-producing algae are widely distributed worldwide. However, HCTs, especially PSTs, are rarely reported in freshwater due to analytical limitations. This may result in an underestimation of the ecological risks and health risks. This study developed a new method to detect ATX-a, CYN, and thirteen common PSTs in freshwater simultaneously by using off-line solid phase extraction (SPE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The limits of detection (LODs) of all targets were lower than 0.05 µg/L, which could meet the regulatory requirements for monitoring of HCTs in drinking water in different countries and regions. To improve the detection sensitivities for trace PSTs, a method based on off-line SPE and on-line SPE-LC-MS/MS was established with LOD around 0.001 µg/L. GTX1&4, GTX2&3, and GTX5 were detected in freshwater in China for the first time, highlighting that overall communities are facing potential risks of exposure to various PSTs in China. High concentrations of ATX-a and CYN were also detected in freshwater from Northern China. The proposed method helps to understand the pollution status of HCT in water bodies, especially during the non-algal bloom period.