Immunoaffinity chromatography purification and ultra-high-performance liquid chromatography-tandem mass spectrometry determination of four β-agonists in beef

A highly selective and sensitive method was developed for the simultaneous determination of four β-agonists (clenbuterol, salbutamol, ractopamine and terbutaline) in beef by immunoaffinity chromatography purification coupled to ultra-high-performance LC-MS/MS. The MS/MS conditions, ultra-high-performance LC mobile phase, injection solution, sample purification process and matrix effect were studied to optimise the operation conditions. The limits of detection (LODs) of the instrument for the studied β-agonists ranged from 0.20 to 0.25?µg?l^?1, and the LODs of the method for the studied β-agonists ranged from 0.20 to 3.00?µg?kg^?1 for beef. Calibration curves were constructed using a standard solution diluted with blank beef matrix. The linear ranges of the calibration curves ranged from 5 to 100?µg?kg^?1 and the coefficients of determination were >0.9942 (n?=?10) for all four β-agonists. Samples spiked at 5, 10 and 50?µg?kg^?1 showed recoveries >72% and RSDs <6.6%. The method is suitable for the simultaneous detection of four β-agonists at trace levels in beef.     

Rapid determination of lycopene and β-carotene in tomato by liquid chromatography/electrospray tandem mass spectrometry

BACKGROUND: The tomato fruit is a dietary source of carotenoids, bioactive antioxidant compounds that play an important role in the prevention of degenerative diseases. Several extraction and detection techniques regarding carotenoids in tomatoes can be found in the literature, mainly based on high‐performance liquid chromatography separation and ultraviolet‐visible detection. RESULTS: The best extraction conditions and tandem mass spectrometry (MS) analysis were evaluated: lycopene and β‐carotene were extracted in a cyclohexane/ethyl acetate mixture without the addition of antioxidants, next separated by liquid chromatography on a C₁₈ column and then determined through electrospray tandem MS. Ionic suppression by the matrix in negative ionisation mode did not allow the analysis of extracts, hence the positive ionisation mode was chosen. Validation parameters demonstrated the suitability for purpose of the analytical method: accuracy, precision, linearity and detection limits were adequate. The method was finally applied to different tomato samples, and differences could be easily highlighted. CONCLUSION: The method was simple, fast and appropriate for the purpose of analysing lycopene and β‐carotene in tomatoes. Copyright © 2011 Society of Chemical Industry     

Simultaneous determination of 10 mycotoxins in grain by ultra-high-performance liquid chromatography-tandem mass spectrometry using ¹³C₁₅-deoxynivalenol as internal standard

An ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for simultaneous determination of 10 mycotoxins in grain was developed. The selected mycotoxins were: deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, nivalenol, fusarenon X, moniliformin, zearalenone, zearalanone, ochratoxin A and ochratoxin B. The samples were extracted with aqueous acetonitrile (84 : 16, v/v) and purified by reliable laboratory-made mixed cartridges. The analytes were separated on an Acquity UPLC HSS T3 column (100 × 2.1 mm, 1.8 μm) and eluted with a mobile phase of water containing 0.2% aqueous ammonia and acetonitrile/methanol (90 : 10, v/v). All mycotoxins were detected with a Waters Micromass Quattro Ultima Pt tandem quadrupole mass spectrometer operating in negative electrospray ionization using multiple reaction monitoring mode. Accurate determination was achieved by employing commercial 13C??-deoxynivalenol as internal standard, which compensated for target loss and eliminated matrix effects. The established method was further validated by determining the linearity (R2 > 0.9990), average recovery (75.8-106.5%), sensitivity (limit of quantitation 0.09-8.48 μg kg?1) and precision (relative standard deviation ≤ 6.9%). It was shown to be a suitable method for simultaneous determination of 10 mycotoxins in grain. Finally, a total of 69 corn samples randomly collected from eastern and northern China were analyzed. The results showed that deoxynivalenol was the most frequently detected contaminant, whilst 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, nivalenol, zearalenone, zearalanone, fusarenon X and moniliformin also occurred frequently. Ochratoxin A and ochratoxin B were present only in trace amounts in a small number of samples.     

Multi-residual analysis of 16 β-agonists in pig liver,kidney and muscle by ultra performance liquid chromatography tandem mass spectrometry

A delicate method was developed for simultaneous analysis of 16 illegal residual β-agonists in pork tissues including liver, kidney and meat. The samples were subjected to enzymatic hydrolysis, extracted with perchloric acid, and then dual Oasis HLB and MCX solid phase extraction (SPE) cartridges were used for cleanup. The analytes were quantified by ultra performance liquid chromatography coupled with electrospray ionisation tandem mass spectrometry (UPLC–ESI–MS/MS) operating in positive multiple-reaction mode (MRM). Matrix-fortified calibration curves were performed to compensate for the matrix effect and loss in sample preparation. CCα and CCβ of the analytes upon the method ranged from 0.02 to 0.79 μg/kg and from 0.04 to 1.62 μg/kg, respectively.     

Multiclass method for fast determination of veterinary drug residues in baby food by ultra-high-performance liquid chromatography–tandem mass spectrometry

A simple and rapid multiresidue method for the determination of different veterinary drug residues in meat-based baby food (MBF) and powdered milk-based infant formulae (PMIF) has been developed. The method involves an extraction procedure based on buffered QuEChERS (quick, easy, cheap, effective, rugged and safe) methodology, without any further clean-up step, followed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). The method has been validated in two baby food matrices (MBF and PMIF) at three different concentration levels, obtaining suitable recoveries and precision (inter and intra-day precision) values. Quantification was carried out using matrix-matched standard calibration. Furthermore, the decision limit (CCα) and the decision capability (CCβ) were evaluated, ranging from 0.5 to 16.2 μg/kg and from 1.2 to 22.4 μg/kg, respectively. Finally, the method was applied to the analysis of several kinds of baby food samples and traces of some veterinary drugs were detected.     

Determination of caramel colorants’ by-products in liquid foods by ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)

2-Methylimidazole, 4-methylimidazole (2-MI and 4-MI), 2-acetyl-4-(1,2,3,4-tetrahydroxybutyl) imidazole (THI) and 5-hydroxymethylfurfural (5-HMF) are neo-formed compounds generated during the manufacture of caramel colours and are transferred to the processed food. These contaminants are known to have a toxicological profile that may pose health risks. Hence, to characterise THI, 2- and 4-MI and 5-HMF levels in liquid foods, an ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and sample preparation was divided into two analytical strategies depending on the concentration range expected in the type of foods targeted. For the determination of the imidazole substitutes (THI, 2- and 4-MI), a sample enrichment and clean-up step by strong cation solid-phase extraction was developed. This method is capable of quantifying over a range of 5 ng ml^–1 (LOQ) to 500 ng ml^–1 with recoveries of 75.4–112.4% and RSDs of 1.5–15%. For determination of 5-HMF, a standard addition method was applied covering the linear range of 0.25–30 µg ml^–1 with RSDs from 2.8% (for intraday precision) to 9.2% (for intermediate precision). The validated analytical methods were applied to 28 liquid food samples purchased from local markets. THI was found only in the beer samples at levels up to 141.2 ng ml^–1. For 2-MI, non-quantifiable traces were observed for all samples, while 4-MI was observed in all samples with large concentration variations (from < LOQ to 563.9 ng ml^–1). 5-HMF was found at expected concentrations, except for a sherry vinegar sample (113 µg ml^–1), which required a high level of dilution before following the standard addition protocol.     

Analysis of isothiazolinone biocides in paper for food packaging by ultra-high-performance liquid chromatography–tandem mass spectrometry

A novel and simple method to detect isothiazolinone-type biocides (2-methyl-3-isothiazolinone (MI), 5-chloro-2-methyl-3-isothiazolinone (CMI), 1,2-benzisothiazolinone (BIT) and 2-octyl-3-isothiazolinone (OIT)) in paper used for food packaging by ultrasonic extraction coupled with UPLC-MS/MS was developed. Parameters affecting process efficiency such as extraction solvents, UPLC mobile phase, gradient elution procedure and MS/MS conditions were studied to optimise the operating conditions. Using the optimised gradient elution procedure, the retention time was less than 6?min. The limits of detection (LODs) were found to be between 0.001 and 0.010?mg?kg^?1, which was validated using actual concentrations. After diluting the standard solution with a blank matrix, the linear calibration curve ranges were 0.002–1.000?mg?kg^?1 for BIT and OIT, 0.005–1.000?mg?kg^?1 for MI, and 0.020–1.000?mg?kg^?1 for CMI, with correlation coefficients higher than 0.9985 (n?=?6). A good level of precision with a mean recovery greater than 81.3% and a relative standard deviation (RSD) less than 6.2% were also obtained. A methodology has been proposed for the analysis of isothiazolinones in paper.     

Quantification of aristolochic acids I and II in herbal dietary supplements by ultra-high-performance liquid chromatography–multistage fragmentation mass spectrometry

A rapid, selective and sensitive ultra-high-performance liquid chromatography–multistage fragmentation mass spectrometry (UHPLC-MS³) method was developed and evaluated for the determination of aristolochic acids I and II (AA I and II) in herbal dietary supplements. A hybrid triple quadrupole/linear ion-trap mass spectrometry was used to monitor MS³ ion transitions m/z 359.2 > 298.1 > 268.0 and m/z 329.2 > 268.2 > 238.0 to detect AA I and II, respectively. The extraction and clean-up of target analytes from dry powdered samples was performed using the quick, easy, cheap, effective, rugged and safe (QuEChERS) procedure. Herbal liquid extracts were analysed directly. Average recoveries ranged from 89% to 112%, with relative standard deviations (RSDs) ranging from 3% to 16%. Limits of quantification (LOQs) estimated for three selected matrices were as follows (AA I/II): 5/10 ng g^?1 (tablets); 25/50 ng g^?1 (capsules); and 2.5/5.0 ng ml^?l (liquid herbal extract). The method was applied in a limited survey of 30 herbal products marketed in the United States via the Internet. AA I and II were detected in 20% and 7%, respectively, of tested samples.     

Simultaneous determination of 10 mycotoxins in grain by ultra-high-performance liquid chromatography–tandem mass spectrometry using 13C15-deoxynivalenol as internal standard

An ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) method for simultaneous determination of 10 mycotoxins in grain was developed. The selected mycotoxins were: deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, nivalenol, fusarenon X, moniliformin, zearalenone, zearalanone, ochratoxin A and ochratoxin B. The samples were extracted with aqueous acetonitrile (84?:?16,?v/v) and purified by reliable laboratory-made mixed cartridges. The analytes were separated on an Acquity UPLC HSS T3 column (100?×?2.1?mm,?1.8?µm) and eluted with a mobile phase of water containing 0.2% aqueous ammonia and acetonitrile/methanol (90?:?10,?v/v). All mycotoxins were detected with a Waters Micromass Quattro Ultima Pt tandem quadrupole mass spectrometer operating in negative electrospray ionization using multiple reaction monitoring mode. Accurate determination was achieved by employing commercial ¹³C₁₅-deoxynivalenol as internal standard, which compensated for target loss and eliminated matrix effects. The established method was further validated by determining the linearity (R ^2?>?0.9990), average recovery (75.8–106.5%), sensitivity (limit of quantitation 0.09–8.48?µg?kg^?1) and precision (relative standard deviation?≤?6.9%). It was shown to be a suitable method for simultaneous determination of 10 mycotoxins in grain. Finally, a total of 69 corn samples randomly collected from eastern and northern China were analyzed. The results showed that deoxynivalenol was the most frequently detected contaminant, whilst 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, nivalenol, zearalenone, zearalanone, fusarenon X and moniliformin also occurred frequently. Ochratoxin A and ochratoxin B were present only in trace amounts in a small number of samples.     

Determination of sulbactam in raw bovine milk by isotope dilution–ultra-high-performance liquid chromatography–tandem mass spectrometry

We developed a sensitive and selective isotope dilution ultra-high-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) method for the determination of sulbactam residue in raw bovine milk. Sulbactam and internal standard, sulbactam-d₅, were extracted from raw bovine milk via liquid-liquid extraction and enriched with strong anion exchange solid-phase extraction cartridges and finally analyzed by using UPLC-MS/MS with multiple reaction monitoring mode. The method was validated according to European regulations. The calibration curve showed good linearity, with a correlation coefficient of 0.9998. Decision limit and detection capability of sulbactam were determined by matrix calibration curve and were 0.0445 and 0.0517 μg/L, respectively. The recoveries of sulbactam in fortified raw bovine milk ranged from 72.1 to 91.5%, with the intra- and interday relative standard deviations ranging from 3.0 to 18.9%. Furthermore, the developed method was applied to analyzing real raw bovine milk samples collected from dairy farms in Beijing, China. Sulbactam was not determined in all samples. The proposed method could ultimately serve as a methodological foundation for the determination of sulbactam in different types of raw milk and dairy products.     

Microwave pretreatment and gas chromatography–mass spectrometry determination of herbicide residues in onion

A rapid multi-residue method was developed for the determination of 16 herbicides in onion. The analytical procedure was based on preventing formation of sulfur-containing compounds in onion by microwave inactivation of the enzyme alliinase. The onion samples which had been pretreated were extracted with acetonitrile and cleaned by solid-phase extraction. The herbicide residues in onion were detected by gas chromatography/mass spectrometry with selected ion monitoring. The recoveries of 16 herbicides ranged from 69.2% to 105.0% with the relative standard deviations (RSD) below 10.7%. The limit of quantitation (LOQ) ranged from 0.003 to 0.015 mg kg⁻¹. The method was applied to the analysis of herbicide residues in onion samples.     

Microwave-assisted extraction and determination of cyanuric acid residue in infant formula samples by liquid chromatography–tandem mass spectrometry

In the present work, microwave-assisted extraction method in combining with liquid chromatography–tandem mass spectrometry (LC–MS/MS) was proposed for the determination of cyanuric acid (CYA) in infant formula samples. The separation was performed on a MERCK ZIC HILIC column (150 × 2.1 mm i.d., 5 μm) with gradient elution of 20 mM ammonium acetate solution – acetonitrile. The method could respond linearly with cyanuric acid at concentrations from 1.0 to 50 ng mL⁻¹ with a quantification limit of 0.25 mg kg⁻¹. The intra- and inter-day precision was less than 4% and the recovery of the assay was in the range of 86.7–93.1%. In the analysis of practical spiked infant formula samples, the new method yielded satisfactory results. Due to its simplicity and accuracy the straightforward method is particularly suitable for routine cyanuric acid detection.     

Analysis of heterocyclic amines and β-carbolines by liquid chromatography–mass spectrometry in cooked meats commonly consumed in Korea

Heterocyclic amines (HAs), which form in meats during heating and cooking, are recognized as mutagenic and carcinogenic compounds. In this study, 13 HAs and 2 β-carbolines (BCs) were analyzed in cooked Korean meat products, including griddled bacon, griddled pork loin, boiled pork loin, boiled chicken meat, chicken meat stock, chicken breast for salad and chicken patty. The samples were either cooked in the laboratory or purchased from local fast-food restaurants. The HAs and BCs in the samples were separated using solid-phase extraction and were analyzed by high performance liquid chromatography–mass spectrometry (HPLC–MS). The most frequently detected HAs and BCs in the cooked meats were harman (1-methyl-9H pyrido[4,3-b]indole; 990.9 ng g^?1), norharman (9H-pyrido[4,3-b]indole; 412.7 ng g^?1) and PhIP (2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; 258.2 ng g^?1). The griddled pork loin and bacon contained higher levels of norharman, harman and PhIP than the other cooked meats. PhIP, which is classified as a Group 2B carcinogen by the International Agency for Research on Cancer, had levels of 258.2 and 168.2 ng g^?1 in the griddled pork loin and griddled bacon, respectively. The griddled bacon was the only sample containing TriMeIQx (2-amino-3,4,7,8-tetramethylimidazo[4,5-f]quinoxaline; 79.9 ng g^?1). IQ (2-amino-3-methyl imidazo[4,5-f]quinoline), 7,8-DiMeIQx (2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline), 4,8-DiMeIQx (2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline) and AαC (2-amino-9H-pyrido[2,3-b]indole) were detected at trace levels in all samples.     

Determination of trichothecenes in cereals and cereal-based products by liquid chromatography–tandem mass spectrometry

A sensitive, accurate and precise method for the simultaneous determination of nivalenol (NIV), deoxynivalenol (DON), T-2 toxin (T-2) and HT-2 toxin (HT-2) in different food matrices, including wheat, maize, barley, cereal-based infant foods, snacks, biscuits and wafers, has been developed. The method, using liquid chromatography coupled with atmospheric pressure chemical ionization triple quadrupole mass spectrometry (LC–APCI–MS/MS), allowed unambiguous identification of the selected trichothecenes at low µg per kg levels in such complex food matrices. A clean-up procedure, based on reversed phase SPE Oasis® HLB columns, was used, allowing good recoveries for all studied trichothecenes. In particular, NIV recoveries significantly improved compared to those obtained by using Mycosep® #227 columns for clean-up of the extracts. Limits of detection in the various investigated matrices ranged 2.5–4.0 µg kg^?1 for NIV, 2.8–5.3 µg kg^?1 for DON, 0.4–1.7 µg kg^?1 for HT-2 and 0.4–1.0 µg kg^?1 for T-2. Mean recovery values, obtained from cereals and cereal products spiked with NIV, DON, HT-2 and T-2 toxins at levels from 10 to 1000 µg kg^?1, ranged from 72 to 110% with mean relative standard deviation lower than 10%. A systematic investigation of matrix effects in different cereals and cereal products was also carried out by statistically comparing the slopes of standard calibration curve with matrix-matched calibration curve for each of the four toxins and the eight matrices tested. For seven of the eight matrices tested, statistically significant matrix effects were observed, indicating that, for accurate quantitative analysis, matrix-matched calibration was necessary. The method was applied to the analysis of 57 samples of ground wheat originated from South Italy and nine cereal food samples collected from retail markets.     

Application of dispersive liquid–liquid microextraction for the determination of selected organochlorine pesticides in honey by gas chromatography–mass spectrometry

Dispersive liquid–liquid microextraction (DLLME) is a rapid and easy technique that consumes minute amounts of organic solvents. In this work, we present chemometric study on optimization of DLLME parameters for the extraction of aldrin, endrin, lindane, α-endosulfan, 4,4′-DDT and its metabolites from honey matrix. Method quantification limits (MQLs) vary between 0.3 ng/g for 2,4′-DDE and 4,4′-DDE to 13.2 ng/g for α-endosulfan and enable determination at levels below EU-established Maximum Residue Limits. The developed method is linear (R ² > 0.994) in the investigated range (MQL—100 ng/g), with preconcentration factors of 13.2–30.5 and good repeatability (CV ≤ 17%). A comparison with other available methods reported in the last decade is provided. The method has been applied to 19 real samples from Poland, and the results show that organochlorine pesticides (OCPs) are present in analysed honeys at levels not posing threat to human health (below 14 ng/g for sum of 4,4′-DDT and metabolites and below 5 ng/g for aldrin, endrin and lindane). To the best of our knowledge, this is the first reported application of DLLME for the determination of OCPs in honey.     

Confirmatory analysis of steroids in muscle using liquid chromatography–tandem mass spectrometry

A method is described for screening and confirmation of synthetic and endogenous steroids in muscle tissue. The method is sensitive, selective, and rapid and the consumption of organic solvents is low, compared to previously published methods. The procedure involves hydrolysis, defattening with heptane and final clean up with SPE using C₁₈ cartridge. After filtration, the analytes are analysed by LC/MS/MS and quantification is performed using deuterated internal standards. Decision limits (CCα) varied from 0.02 to 0.33?µg?kg^?1 and the detection capabilities (CCβ) were <0.50?µg?kg^?1. The mean within-laboratory reproducibility ranged 5–22% (%RSD_IR). Endogenous steroids (e.g. testosterone, epitestosterone and androstenedione) have been included in the method, to provide an insight into their levels, as the presence of these steroids was detected several times during analysis of imported meat.     

Improved method for the determination of 12 non-nutritive sweeteners and monitoring in various foods using liquid chromatography–tandem mass spectrometry

An improved and highly sensitive method was developed and validated for the determination of 12 (7 permitted and 5 non-permitted in Korea) non-nutritive sweeteners in various foods using liquid chromatography-electrospray ionisation-tandem mass spectrometry. The chromatographic separation was performed on an Xbridge BEH C18 column (3 mm × 100 mm, 2.5 μm) with gradient elution using 10 mM ammonium acetate in water and 10 mM ammonium acetate in methanol. Sample preparation consisted of simple dilution, homogenisation, centrifugation and purification with a C18 cartridge prior to analysis. The relative matrix effect (%ME) was within ±20% for all sweeteners. The method also showed good linearity (R² > 0.99). The limit of detection and limit of quantification values in sample were in the range of 0.02–2.66 and 0.06–8.05 mg kg^?1, respectively. The recoveries at three concentration levels ranged between 80% and 119%, with relative standard deviation values below 10%. In addition, the expanded uncertainties determined for 12 sweeteners in 5 different food matrices were confirmed to be <14%. Finally, the method was successfully applied to the analysis of sweeteners in 681 food samples purchased in Korea, Australia and Turkey. These results demonstrate that the method is suitable for the simultaneous determination of multiple-sweeteners in a variety of foods.     

Simultaneous determination of kasugamycin and validamycin-A residues in cereals by consecutive solid-phase extraction combined with liquid chromatography–tandem mass spectrometry

Two polar aminoglycosides, kasugamycin and validamycin-A, were determined in cereals (brown rice, wheat and corn) by high-performance liquid chromatography–tandem mass spectrometry. The analytes were extracted from samples using methanol and water (70:30, v/v) at pH 5.5, purified using both a hydrophilic–hydrophobic-balanced cartridge and a strong cation-exchange cartridge, and then analysed using multiple reaction monitoring in positive electrospray ionisation mode with a special ReproSil 100 C₁₈ high-performance liquid chromatography column. This newly proposed method yielded good sensitivity and excellent chromatographic performance. The limits of quantification for kasugamycin and validamycin-A were 4.1 µg/kg and 1.0 µg/kg, respectively. The recoveries for both compounds at three fortification levels (4, 100 and 500 µg/kg for kasugamycin; 1, 10 and 100 µg/kg for validamycin-A) ranged from 75% to 110%, and the relative standard deviations were below 15%.     

Simultaneous determination of bisphenol A,octylphenol, and nonylphenol by pressurised liquid extraction and liquid chromatography–tandem mass spectrometry in powdered milk and infant formulas

A new analytical method, using pressurised liquid extraction (PLE) and liquid chromatography–tandem mass spectrometry (LC–MS/MS), was developed for the simultaneous determination of bisphenol A (BPA), octylphenol (OP) and nonylphenol (NP) in powdered infant formulas (IF) and powdered skimmed milk (PM). The analytes were extracted by PLE, using this optimised conditions: ethyl acetate as solvent, 70 °C of temperature, reversed-phase silica C₁₈ as dispersing agent and three cycles of extraction. The extracts were then injected in LC–MS/MS using a Gemini C₁₈ column and a mixture of 5% water and 95% methanol/acetonitrile, both with 0.1% ammonia, as a mobile phase. Recoveries at different fortification levels (0.5 and 0.05 mg kg⁻¹), were between 89% and 92% for BPA, 84 and 98% for OP, and 93% and 101% for NP. The method was applied to the analysis of samples of PM and IF, bought in Italian and Spanish markets. In positive samples, phenols concentration ranged from 0.07 to 1.29 mg kg⁻¹ for BPA, from 0.028 to 1.55 mg kg⁻¹ for OP and from 0.026 to 1.47 mg kg⁻¹ for NP.