Unambiguous detection of astaxanthin and astaxanthin fatty acid esters in krill (Euphausia superba Dana)

HPLC atmospheric pressure chemical ionization (APCI)/MS, GC MS, HPLC diode array detection (DAD), and NMR were used for the identification of astaxanthin and astaxanthin fatty acid esters in krill (Euphausia superba Dana). Matrix solid phase dispersion was applied for the extraction of the carotenoids. This gentle and expeditious extraction technique for solid and viscous samples leads to distinct higher enrichment rates than the conventional liquid-liquid extraction. The chromatographic separation was achieved employing a C30 RP column that allows the separation of shape-constrained geometrical isomers. A methanol/tert-butylmethyl ether/water gradient was applied. (all-E) Astaxanthin and the geometrical isomers were identified by HPLC APCI/MS, by coelution with isomerized authentical standard, by UV spectroscopy (DAD), and three isomers were unambiguously assigned by microcoil NMR spectroscopy. In this method, microcoils are transversally aligned to the magnetic field and have an increased sensitivity compared to the conventional double-saddle Helmholtz coils, thus enabling the measurement on small samples. The carotenol fatty acid esters were saponified enzymatically with Lipase type VII from Candida rugosa. The fatty acids were detected by GC MS after transesterification, but also without previous derivatization by HPLC APCI/MS. C14:0, C16:0, C16:1, C18:1, C20:0, C20:5, and C22:6 were found in astaxanthin monoesters and in astaxanthin diesters. (all-E) Astaxanthin was identified as the main isomer in six fatty acid ester fractions by NMR. Quantitation was carried out by the method of internal standard. (13-cis) Astaxanthin (70 microg/g), 542 microg/g (all-E) astaxanthin, 36 microg/g unidentified astaxanthin isomer, 62 microg/g (9-cis) astaxanthin, and 7842 microg/g astaxanthin fatty acid esters were found.     

High-throughput GC/MS and HPLC/MS/MS techniques for the multiclass, multiresidue determination of 653 pesticides and chemical pollutants in tea

An efficient and sensitive method has been established for simultaneous determination of 653 pesticides in teas by GC/MS and HPLC/MS/MS. The method involved extraction with acetonitrile followed by cleanup using Cleanert-TPT SPE and subsequent identification and quantitation of 490 pesticides by GC/MS and 448 pesticides by HPLC/MS/ MS. The LODs for pesticides determined by GC/MS were between 1.0 and 500 microg/kg, and those determined by HPLC/MS/MS were between 0.03 and 4820 microg/kg. At the low fortification levels of 0.01-100 microg/kg, the average recoveries of 94% of the pesticides determined by GC/MS were between 60 and 120%, 77% of which had an RSD below 20%. For 91% of pesticides determined by HPLC/MS/MS, the average recoveries were between 60 and 120%, 76% of which had an RSD below 20%. The paper also reports a novel SPE column, Cleanert TPT, which comprised graphitized carbon black (PestiCarb), polyamine silica, and amide polystyrene for purifying the tea samples. The results indicated good repeatiblity and reproducibility.     

High-performance liquid chromatography with fluorescence detection and ultra-performance liquid chromatography with electrospray tandem mass spectrometry method for the determination of indoleamine neurotransmitters and their metabolites in sea lamprey plasma

We present a comparison of two sensitive methods, HPLC with fluorescence detector (HPLC/FLD) and UPLC with electrospray tandem mass spectrometry (UPLC/MS/MS), for the determination of indoleamine neurotransmitters (NTs) and their metabolites in sea lamprey plasma samples. Liquid–liquid extraction (LLE) and solid-phase extraction (SPE) were also tested for recovery and matrix effect. The recoveries of SPE determined by HPLC/FLD and UPLC/MS/MS ranged from 75 to 123% and 78 to 105%, respectively, while the recoveries of LLE ranged from 45 to 73% and 48 to 75%, respectively. SPE combined with HPLC/FLD and UPLC/MS/MS to determine the target analytes in plasma samples were validated of the sensitivity, reproducibility, accuracy and precision. Both methods exhibited excellent linearity in the range of 0.2–50 ng mL⁻¹ for all analytes. The limits of detection (LOD) varied from 0.04 ng mL⁻¹ to 0.13 ng mL⁻¹ for HPLC/FLD method and 0.003 ng mL⁻¹ to 0.02 ng mL⁻¹ for UPLC/MS/MS method. The inter-day accuracy ranged from 82.5 to 127.0% for HPLC/FLD and 93.0 to 113.0% for UPLC/MS/MS. The inter-day precision ranged from 9.9 to 32.3% for HPLC/FLD and 5.4 to 13.2% for UPLC/MS/MS. These results demonstrated that the values obtained by both methods were within the satisfactory range and the UPLC/MS/MS method provided more accurate and precise measurements than HPLC/FLD method. The comparison is of great importance to determine the available detectors, considering the complexity and expensiveness versus quality parameters. These two methods were applied to the analysis of four important indoleamine neurotransmitter analytes (5-hydroxytryptamine, 5-hydroxyindole-3-acetic acid, tryptamine and melatonin) in sea lamprey plasma samples.     

A new high-performance liquid chromatographic/electrospray ionization tandem mass spectrometric method for the simultaneous determination of cyclophosphamide, methotrexate and 5-fluorouracil as markers of surface contamination for occupational exposure monitoring

A new high-performance liquid chromatographic/electrospray ionization tandem mass spectrometric (HPLC/ESI-MS/MS) method was developed for the simultaneous quantification of 5-fluorouracil (5FU), methotrexate (MTX) and cyclophosphamide (CP) in environmental samples. These compounds, commonly used in the treatment of cancer, are recognized as genotoxic. In order to estimate the occupational exposure of hospital personnel handling these drugs, wipe samples were taken from the working surfaces and directly analyzed (with trophosphamide as internal standard) using a reversed-phase capillary column and MS/MS detection. This is the first HPLC/MS/MS method for the simultaneous determination of 5FU, MTX and CP. The present method offers high sensitivity, with detection limits of 1.1 microg l(-1) for MTX and CP and 33.3 microg l(-1) for 5FU, avoiding any sample preconcentration procedure. Rapidity, specificity, high accuracy (mean values between 92.4 and 99.9%) and precision (mean RSD values between 3.4 and 12.1%) make the method suitable for the routine determination of these three antineoplastic drugs.     

Determination of aliphatic and alicyclic amines in water by gas and liquid chromatography after derivatization by chloroformates

Two new methods were developed for the analysis of aliphatic (n-propylamine, pentylamine, hexylamine, heptylamine, octylamine) and alicyclic (pyrrolidine, morpholine, piperidine, piperazine) amines in water samples after derivatization and liquid-liquid-extraction. The carbamate-derivatives formed were determined by GC/MS (trichloroethyl carbamates) as well as by HPLC/fluorescence detection (9-fluorenylmethyl carbamates) in a concentration range between 0.05 and 1.0 microg/l suitable for drinking water analysis. Applications to German rivers and sewage plants show that both new methods produce corresponding results in analysing aliphatic and alicyclic amines in surface waters as well as in waste water samples.     

Determination of flavonoids and stilbenes in red wine and related biological products by HPLC and HPLC-ESI-MS-MS

To investigate probable health benefits of flavonoids and stilbenes in red wine a new reversed-phase (RP) high-performance liquid-chromatographic (HPLC) method with enhanced separation efficiency and improved selectivity, sensitivity, and speed has been established for determination of the flavonoids quercetin, myricetin and kaempferol and the stilbenes cis- and trans-resveratrol, in a single run . UV-absorbance, fluorescence (FLD), and mass-spectrometric (MS) detection were also evaluated. UV-absorbance detection at 320 nm for stilbenes and 377 nm for flavonoids enables their determination up to the nanogram range with a linearity of R2>0.9999 (linear range 50 ng mL(-1)-50 microg mL(-1)). Calculated values of average recoveries were between 95 and 105% for all analytes. For resveratrol, fluorescence detection was highly selective and twice as sensitive as UV detection, and linearity was satisfactory (R2>0.9996; linear range see UV detection). For the detection of the hydrophilic glycosidic compounds piceid and rutin, which are coeluted with other hydrophilic ingredients, the validated RP HPLC system was coupled to a quadrupole ion-trap mass-spectrometer (MS) via an electrospray interface (ESI) with 25% ammonia solution as sheath liquid. MS detection was, highly linear (R2>0.9878; linear range 50 ng mL(-1)-50 microg mL(-1)) for all investigated analytes and the limits of detection were in the low nanogram range. Compared with UV detection MS detection resulted in a 200% increase in signal intensity for myricetin and 400% increases for quercetin and kaempferol, but equal signal intensity for resveratrol. Calculated values of average recoveries were 102% for myricetin and 79% for piceid. Collision induced dissociation (CID) was also used to obtain characteristic fragmentation fingerprints to facilitate qualitative and quantitative analysis even in complex matrices. Finally, this hyphenated HPLC-ESI-MS method was highly suitable and an essential improvement compared with UV- and fluorescence detection.     

高效液相色谱-高锰酸钾氧化化学发光法测定水中的痕量苯二酚

基于酸性介质中甲酸对高锰酸钾-苯二酚氧化发光反应的增敏作用建立了高效液相色谱-化学发光柱后检测苯二酚的新方法。优化了高锰酸钾-苯二酚氧化发光反应及高效液相色谱分离苯二酚的条件,用甲醇-0.1 mmol/L β-环糊精水溶液(体积比为30∶70) 作为流动相可实现对水中3种苯二酚异构体的分离,且能与高锰酸钾-苯二酚氧化化学发光反应条件很好地偶合。对所测定的苯二酚异构体,方法的线性范围达两个数量级;以信噪比为3测得邻、间、对苯二酚的检出限(n=3)分别为:5.2,4.7,3.2 μg/L,对质量浓度均为0.10 mg/L的3种苯二酚混合溶液连续测定11次,邻、间、对苯二酚的相对标准偏差分别为2.8%,3.4%,6.5%。将该方法与固相萃取技术相结合,对河水中的痕量苯二酚进行了测定,加标回收率为92.1%~95.4%。     

Trace determination of alpha- and beta-endosulfan and three metabolites in human serum by gas chromatography electron capture detection and gas chromatography tandem mass spectrometry

Endosufan, alpha and beta, and three conversion products, sulphate, ether and lactone, were simultaneously determined in human serum by means of an analytical procedure which combines extraction with organic solvents, clean-up with H(2)SO(4) and by liquid column chromatography, and detection by gas chromatography (GC) using electron capture detection (ECD) and tandem mass spectrometry (MS/MS). The procedure was validated and the values of some merit figures, such as linear range, detection and quantitation limits, accuracy, precision and recovery, obtained with the GC/ECD and the GC/MS/MS methods, were compared. The lower limits of detection in GC/ECD and GC/MS/MS were 0.03 and 0.05 microg I(-1), respectively. The recovery of the pesticides at the 20 microg I(-1) concentration level was 60-65%, with the exception of endosufan alpha. Recovery studies at higher levels (100 and 200 microg I(-1)) were independent of pesticide concentration in serum samples. The application of the proposed analytical methodology to the determination of endosulfans and their metabolites in real samples was tested by analyzing serum samples from a population living in agricultural areas of Almeria (Spain). The results show the advantage of MS/MS over the ECD detector in the analysis of serum samples where matrix interferences can be confused with target pesticides.     

高效液相色谱荧光检测法测定蔬菜中残留的甲氨基阿维菌素苯甲酸盐

建立了甘蓝和蘑菇中甲氨基阿维菌素苯甲酸盐的固相萃取-高效液相色谱荧光分析方法。蔬菜样品用乙酸乙酯提取,提取液旋转浓缩近干后用少量乙酸乙酯溶解,再经PRS固相萃取(SPE)柱净化,洗脱液经氮气吹干后用氮甲基咪唑和三氟乙酸酐衍生,衍生物用高效液相色谱分析,采用外标法定量。在添加浓度1.0～20.0 μg/kg范围内,平均添加回收率为78.6%～84.9%,日内相对标准偏差(RSD)为2.7%～6.0%,日间RSD为3.1%～8.9%,检出限为0.10 μg/kg。甲氨基阿维菌素苯甲酸盐衍生物在0.002～0.10 mg/L范围内呈良好的线性关系,相关系数为0.9999。     

A rapid, sensitive and effective quantitative method for simultaneous determination of cationic surfactant mixtures from river and municipal wastewater by direct combination of single-drop microextraction with AP-MALDI mass spectrometry

A rapid, simple, sensitive, and effective quantitative method for simultaneous determination of cationic surfactants (CS(+)) from river and municipal wastewater by direct combination of single-drop microextraction (SDME) with atmospheric pressure (AP)-MALDI mass spectrometry has been successfully demonstrated without the requirements of tedious sample pre- or post-treatment or separation by high-performance liquid chromatography (HPLC), gas chromatography (GC), and capillary electrophoresis (CE). This quantitative method can greatly enhance the signal-to-noise ratio for analysis of small molecules of CS(+) owing to the strong suppression of matrix ions by the analytes. In addition, SDME assisted in the isolation and preconcentration of CS(+) from water samples, which could effectively reduce the background interferences from the matrices present in waste and river water. The SDME conditions were optimized for achieving high extraction efficiency of CS(+) from aqueous samples, in terms of solvent selection, stirring speed, extraction time, exposure volume of acceptor phase, and salt addition. The enrichment factors for CS(+) were found to be 40-64-folds for 7 min of extraction time with no salt addition and at room temperature. This method was found to yield a linear calibration curve in the concentration range from 50 to 1500 microg/l CS(+) with a limit of detection (LOD) of 10 microg/l. The relative recoveries in river and municipal wastewater were found to be 93.8-103.6% and 91.0-98.7%, respectively. These results indicate that the combination of SDME with AP-MALDI/MS is effective for the simultaneous determination of CS(+) from river and municipal wastewater. In addition, a comparison of enrichments and LOD values for this method with hollow-fiber liquid phase microextraction (HF-LPME) was also demonstrated. The present approach is easy to operate, rapid, sensitive, and suitable for high-throughput of analysis.     

Gas chromatographic analysis of capsaicin in Gochujang

A sensitive, precise, and specific gas chromatographic (GC) method was developed for the analysis of capsaicin in Gochujang and validated by comparing with a column high-performance liquid chromatographic (HPLC) method (AOAC 995.03). The method validation parameters yielded good results, including linearity, precision, accuracy, and recovery. The GC separation was performed on a (5% phenyl)-methylpolysiloxane column [length 30 m, internal diameter (id) 250 microm, film thickness 0.25 microm] followed by flame ionization detection. The conditions of temperature programming were initially 220 degrees C for 1 min, ramp at 5 degrees C/min to 270 degrees C, and hold for 10 min. The recovery of capsaicin in Gochujang was more than 92%, and the detection limit and lower determination limit of the GC analysis were 1.0 and 5.0 microg/g, respectively. The calibration graph for capsaicin was linear from 1 to 250 microg/mL for GC and 0.5 to 50 microg/mL for HPLC. The interday and intraday precisions (relative standard deviations) were <4.02%.     

超高效液相色谱/串联质谱法分析水中的微囊藻毒素

建立了超高效液相色谱/质谱快速、准确、高灵敏度地测定水体中痕量微囊藻毒素（MCYST）的分析方法,并用于实 际样品的分析。采用固相萃取法富集净化样品。该法在5 min内即可完成4种MCYST（LR、RR、LW、LF）的分离及检测;LR 、RR、LW、LF的定量检测限、回收率分别为1.3～6.0 ng/L、91.1%～111%;工作曲线的线性相关系数大于0.99,线性范 围达3个数量级。实际样品分析表明,在所测定的水库水样中均检出了LR和RR,其质量浓度分别为0.0447～2.73 μg/L和0.0208～1.36 μg/L;而在所有的检测样品中均未检出LW和LF。     

Determination of type A trichothecenes by high-performance liquid chromatography with coumarin-3-carbonyl chloride derivatisation and fluorescence detection

A method for the analysis of type A trichothecenes T-2 toxin, HT-2 toxin, neosolaniol and diacetoxyscirpenol by high-performance liquid chromatography with fluorescence detection using coumarin-3-carbonyl chloride has been developed. Different parameters concerning the analytical procedure such as stability of both the reagent and derivatised analytes, time and temperature of the derivatisation reaction, were studied and optimised. Three different clean-up procedures (solid-phase extraction with silica gel or C-18 cartridges, and liquid–liquid partition between toluene and dihydrogen phosphate buffer) were tested in order to remove the excess reagent peaks. The last procedure gave the best results when the buffer pH was 3–5.5, and is therefore recommended. Separations were performed on a stainless steel LiChrospher 100 C-18 reversed-phase column with pre-column of the same phase. The mobile phase was acetonitrile/water (65:35, v/v) containing 0.75% acetic acid at a flow-rate of 1.0 ml/min. The proposed method provides good separation between the four trichothecenes and good reproducibility (RSD of calibration standards <5%). The limits of detection of the studied trichothecenes at a signal-to-noise ratio of 3:1, with an injection volume of 20 μl were 10 ng/g sample for T-2 toxin and about 15 ng/g sample for the remaining mycotoxins. The calibration curve was linear between 10 and 2000 ng for the four trichothecenes assayed. The method was applied to the analysis of these mycotoxins in fungal cultures (corn and rice) of Fusarium sporotrichioides, and is also perfectly suitable for the quantification of type A trichothecenes in contaminated cereals.     

Determination of Diethylstilbestrol in Aquatic Products by High Performance Liquid Chromatography with Fluorescence Detection

A sensitive method for the determination of diethylstilbestrol (DES) in aquatic products by high performance liquid chromatography (HPLC) using a fluorescence detector (FLD) was developed. By means of sulfonation in concentrated sulfuric acid and hydrolysis, DES was converted into a reaction product that can emit fluorescence and determined by HPLC. For the first time, HPLC with FLD was applied to the determination of DES and sensitivity comparable to that of LC‐MS gained. The limit of detection was 0.1 µg·kg^?1, and the recoveries were above 86% with relative standard deviations less than 6.2%. The proposed method was successfully applied to the determination of DES in aquatic products.     

Capillary electrophoresis for the analysis of paralytic shellfish poisoning toxins in shellfish: Comparison of detection methods

Paralytic shellfish toxins (PSTs) are produced by marine and freshwater microalgae and accumulate in shellfish including mussels, oysters, and scallops, causing possible fatalities when inadvertently consumed. Monitoring of PST content of shellfish is therefore important for food safety, with currently approved methods based on HPLC, using pre‐ or postcolumn oxidation for fluorescence detection (HPLC‐FLD). CE is an attractive alternative for screening and detection of PSTs as it is compatible with miniaturization and could be implemented in portable instrumentation for on‐site monitoring. In this study, CE methods were developed for C⁴D, FLD, UV absorption detection, and MS—making this first report of C⁴D and FLD for PSTs detection. Because most oxidized toxins are neutral, MEKC was used in combination with FLD. The developed CZE‐UV and CZE‐C⁴D methods provide better resolution, selectivity, and separation efficiency compared to CZE‐MS and MEKC‐FLD. The sensitivity of the CZE‐C⁴D and MEKC‐FLD methods was superior to UV and MS, with LOD values ranging from 140 to 715 ng/mL for CZE‐C⁴D and 60.9 to 104 ng/mL for MEKC‐FLD. With the regulatory limit for shellfish samples of 800 ng/mL, the CZE‐C⁴D and MEKC‐FLD methods were evaluated for the screening and detection of PSTs in shellfish samples. While the CZE‐C⁴D method suffered from significant interferences from the shellfish matrix, MEKC‐FLD was successfully used for PST screening of a periodate‐oxidized mussel sample, with results confirmed by HPLC‐FLD. This confirms the potential of MEKC‐FLD for screening of PSTs in shellfish samples.     

液相色谱-二极管阵列检测器/离子阱质谱法检测水中的5种微囊藻毒素

建立了一种液相色谱-二极管阵列检测器(LC-DAD)/离子阱质谱(IT MS)对水中5种微囊藻毒素(microcystins)的分析方法。水中的微囊藻毒素经固相萃取富集和净化,经LC分离后,采用DAD和IT MS定性分析,DAD定量分析。在优化的条件下,水中5种微囊藻毒素的检出限为0.1 μg/L; 3个质量浓度加标水平(0.2、0.8和5 μg/L)的平均回收率为52.2%～115.2%,相对标准偏差为1.2%～10.0%。该方法从紫外吸收光谱和质谱角度同时进行定性定量分析,可用于地表水和饮用水中多种微囊藻毒素的检测。     

Determination of butylated hydroxytoluene in food samples by high-performance liquid chromatography with ultraviolet detection and gas chromatography/mass spectrometry

A reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed and compared with a gas chromatography/mass spectrometry (GC/MS) method for determining butylated hydroxytoluene (BHT) in foodstuffs as a result of migration from plastic packaging. Similar extraction procedures were used in both methods. BHT was quantitated using an external standard in the HPLC method and an internal standard in the GC/MS method. Both methods presented good linearity (r(2) > or = 0.9917) and low detection limits. Recoveries obtained with the HPLC method (chicken meat, 95.8%, and Gouda cheese, 83.9%) were better than with the GC/MS method (chicken meat, 85.6%, and Gouda cheese, 71.3%).     

Determination of polycyclic aromatic hydrocarbons in water by solid-phase microextraction and liquid chromatography.

This study describes the determination of polycyclic aromatic hydrocarbons (PAHs) in water using high-performance liquid chromatography (HPLC) coupled with fluorescence detection (FLD). Because individual PAHs are generally present in water only at trace levels, a sensitive and accurate determination technique is essential. The separation and detection of five PAHs were run completely within 25 min by the HPLC/FLD system with an analytical C18 column, a fluorescence detection, and acetonitrile-water gradient elution. Calibration graphs were linear with very good correlation coefficients (r > 0.9998), and the detection limits were in the range of 2-6 ng/l for five PAHs. Solid phase microextraction (SPME) was performed for sample pretreatment prior to HPLC-FLD determination, and the governing parameters were investigated. Compared to conventional methods, SPME has high recovery, saves considerable time, and reduces solvents waste. The extraction efficiencies of five PAHs were above 88% and the extraction times were 35 min in one pretreatment procedure. One particular discovery is that 1.5 M sodium monochloroactate (ClCH2COONa) can improve the extraction yield of PAH compounds more than other inorganic salts. The SPME-HPLC-FLD technique provides a relatively simple, convenient, practical procedure, which was here successfully applied to determine five PAHs in water from authentic water samples.     

Determination of melamine in pet food by enzyme immunoassay, high-performance liquid chromatography with diode array detection, and ultra-performance liquid chromatography with tandem mass spectrometry

Melamine in pet food (fortified or originally contaminated) was determined by enzyme immunoassay (EIA), high-performance liquid chromatography with diode array detection (HPLC-DAD), and ultra-performance liquid chromatography with tandem mass spectrometry (UPLC-MS/MS). The limits of detection (LOD) for EIA and HPLC-DAD were 0.02 and 0.1 microg/mL, respectively. The linear ranges of the calibration curves for EIA and HPLC-DAD were 0.02-0.5 and 0.1-500 microg/mL, respectively. The coefficient of determinations (r2) of the standard curves for EIA and HPLC were 0.9991 and 0.9999, respectively. Coefficient of variations from both inter- and intra-assay were <9.31%, and recovery range for all concentrations was between 71 and 105%. The r2 values between the EIA and HPLC-DAD methods for melamine analysis of the fortified and originally contaminated samples were 0.9973 and 0.9885. The r2 values for UPLC-MS/MS with HPLC-DAD and with EIA were 0.9566 and 0.9489, respectively.