SCX Cartridge Research Articles

Determination of Flubendazole and Metabolite in Livestock Products Using LC-MS/MS

This study proposes a method to determine flubendazole and metabolite R35475 in livestock products using tandem mass spectrometry coupled with positive ion electrospray ionization. Acetone is used to extract flubendazole and metabolite R35475 from the livestock samples. These extracts were purified using an SCX cartridge column (500 mg). Furthermore, high-performance liquid chromatography was performed on an Inertsil ODS-4 column with a gradient formed using methanol and water, both of which contain 5 mmol/L of ammonium acetate. The recovery tests using bovine muscle, fat, liver, milk, and egg fortified at the maximum residue limits of analytes or 0.005 mg/kg revealed that the trueness (n=5) of flubendazole and metabolite R35475 ranged from 89.4 to 106.4% with a repeatability rate of 1.7-7.8%.

Fluoride derivatization-enabled sensitive and simultaneous detection of biomarkers for nitrogen mustard in human plasma and urine via gas chromatography tandem mass spectrometry.

Methyl-diethanolamine (CAS: 105-59-9), ethyl-diethanolamine (CAS: 139-87-7), and triethanolamine (CAS: 102-71-6) were identified as the degradation products and bio-markers of nitrogen mustard exposure. Sensitive and convenient detection methods for amino alcohol are of great importance to identify nitrogen mustard exposure in forensic analysis. Herein, analytical methods including gas chromatography-tandem mass spectrometry combined with heptafluorobutyryl derivatization and solid phase extraction were established for retrospective detection of the biomarkers in human plasma and urine samples. The efficiency of the method was improved by optimizing the conditions for sample preparation and the GC-MS/MS method. The optimization included the derivatization temperature, reaction time, reagent dosage and solid phase extraction cartridges, eluent and pH of the loading sample. The results indicated that the SCX cartridge resulted in better enrichment and purification effects, and the best recovery could be obtained with pH = 3-4 for the loading samples and an eluent of 2 mL 10% NH4OH/MeOH. The GC-MS/MS parameters were also optimized for better specificity and sensitivity. The established method was fully validated for each analyte both in plasma and urine matrixes. The linear range of analytes in plasma was 1.0-1000 ng mL-1 with a correlation parameter (R2) of ≥0.994, intra-day/inter-day accuracy of 93.7-117%, and relative standard deviation (RSD) of ≤6.5%. Meanwhile the results in urine were 1.0-1000 ng mL-1 with R2 of ≥0.996, intra-day/inter-day accuracy of 94.3-122%, and RSD of ≤6.6%. The detection limit of the analytes was 1.0 ng mL-1. The method was applied for the detection and identification of trace amino alcohols present in urine samples dispatched by the Organization for the Prohibition of Chemical Weapons (OPCW) and the results were confirmed to be correct.

Simultaneous quantifications of four purine derivatives biomarkers in cow milk by SPE HPLC-DAD

In this study, simultaneous quantification of allantoin, uric acid, xanthine, and hypoxanthine in cow milk by solid phase extraction (SPE) and high performance liquid chromatography-diode array detection (HPLC-DAD) method was perform. Five different SPE cartridges were tested in order to evaluate the isolation of purine derivatives (PD) from cow milk. Chromatography was carried out on ODS-2 Hypersil column and 0.05 M (NH<sub>4</sub>)<sub>2</sub>HPO<sub>4</sub> buffer solution (pH = 7.76) as mobile phase. The HPLC-DAD validated method showed a linearity with regression coefficients higher than 0.999 and the limits of detection and quantification with values in the range 0.09–0.74 µg mL<sup>–1</sup> and 0.27–2.24 µg mL<sup>–1</sup>, respectively. The method showed good precision with a relative standard deviation (RSD) below 4.48%, while the accuracy ranged from 95.34 to 104.47% for all analytes. The best recovery degree of PD by SPE were obtained on Strata SCX cartridge for xanthine (87.79%) and hypoxanthine (89.02%); on Strata NH<sub>2</sub> for allantoin (35.09%) and on Strata C8 for uric acid (101.08%). Finally, the HPLC-DAD method with SPE on SCX cartridges was applied to quantify the PD in a batch of thirty cow milk samples.

Tandem Cation/Anion Exchange SPE Cartridge Method for Sample Desalting for HPLC Analysis of Soluble Dietary Fiber: Development and Inter-laboratory Validation.

In HPLC analyses of soluble dietary fiber, desalting processes using open, mixed-bed ion-exchange columns are time-consuming and labor-intensive. We developed and validated a simple desalting method using tandem cation/anion exchange SPE cartridges. We found that combining Bond Elut Jr SCX (upstream) and Bond Elut PSA (downstream) cartridges provided adequate desalting of test solutions. The developed method was then validated in an inter-laboratory study. Five test samples were prepared by mixing food matrixes with purified soluble dietary fiber and treated to generate solutions to test the desalting process. These solutions were then analyzed by eight different laboratories. The results demonstrated that the developed method is simple and reliable for desalting samples containing 140 to 945 mg/100 mL of soluble dietary fiber in preparation for HPLC analysis of soluble dietary fiber.

Determination of Zilpaterol in Livestock Products by LC-MS/MS

An analytical method for the determination of zilpaterol in livestock products was developed. The sample was stirred with n-hexane and n-hexane saturated acetonitrile, and zilpaterol in the sample was extracted with acetonitrile. The extract was cleaned up on a ODS cartridge column (1 g) and SCX cartridge column (500 mg). The LC separation was carried out using an Inertsil ODS-4 column and linear gradient elution with 0.1%formic acid and acetonitrile containing 0.1% formic acid as mobile phase. Detection of MS was carried out positive ion electrospray ionization mode. Average recoveries (n=5) of zilpaterol from 6 kinds of livestock products fortified at the MRLs (0.01 mg/kg) were 87.0-99.4%, and the relative standard deviations were 2.4-6.3%. The limits of quantitation were 0.01 mg/kg.

Quantitative analysis of pyrrolizidine alkaloids in Gynura procumbens by liquid chromatography–tandem quadrupole mass spectrometry after enrichment by PCX solid-phase extraction

ABSTRACTGynura procumbens is commonly consumed as a vegetable and has been approved as an ingredient for food and dietary supplements in China. However, Gynura species are known to contain toxic pyrrolizidine alkaloids (PAs) and the PA profiles in G. procumbens are not known. This study was to extract and enrich PA from G. procumbens or health care products using PCX solid-phase extraction (SPE) cartridges identify the main PAs in the herb and to develop an liquid chromatography–mass spectrometry (LC-MS/MS) method for assaying the PA contents in the plant and its derived products. Upon using multiple reaction monitoring (MRM) acquisition together with comparison to the characteristics of mass fragmentations and retention times of reference standards, 11 PAs were identified as the main PAs in the plant. After clean-up and enrichment with PCX solid-phase extraction (SPE) cartridges, which resulted in better PA recoveries than either C18 or SCX cartridges, the LC-MS/MS method was subjected to validation in terms of linearity, repeatability, limits of quantification (LOQ) and recovery. The validated method was applied to quantify PAs in 12 plant samples and 7 commercial finished products. The total amounts of targeted PAs were found to vary from 15.6 to 848 μg/kg in the herbs and from 9.9 μg/kg to 33.9 mg/kg in the commercial products. The present work was the first to demonstrate that G. procumbens contained PAs in the herb and its derived products and the PA contents might exceed the daily dose limits in food and herbal medicinal products proposed by the European Medicines Agency (i.e. 0.35 μg PA per day for 50 kg adult).

Stimulating effect of both 4’‐O‐methylnorbelladine feeding and temporary immersion conditions on galanthamine and lycorine production by Leucojum aestivum L. bulblets

Bulb cultures of Leucojum aestivum and L. aestivum ‘Gravety Giant’ were subcultured in medium containing the precursor 4’‐O‐methylnorbelladine (MN) at various concentrations [0 (control), 0.15 and 0.3 g/L]. The cultures were conducted in bioreactor RITA® and lasted for 15, 30, 40 and 50 days. The growth rate and the alkaloid accumulation in bulblets were studied. For this latter purpose, a purification method was developed. It comprised a highly selective solid phase extraction using on the one hand, UPTI‐CLEAN SI and SCX cartridges for plant extracts and on the other hand, 2H cartridges for culture media. Pure alkaloidal fractions were, thus, analyzed by LC‐ESI‐MS allowing the quantitative evaluation of galanthamine and lycorine from culture extracts. Precursor feeding along with temporary immersion conditions was found to significantly improve the accumulation of both galanthamine and lycorine. The maximal concentrations of galanthamine (0.81 mg/g DW) and lycorine (0.54 mg/g DW) in L. aestivum bulblets were reached, respectively, after 40 days of culture with 0.15 g/L of precursor and after 30 days of culture with 0.3 g/L of precursor. In L. aestivum ‘Gravety Giant’ bulb cultures, 0.3 g/L of precursor was the best condition for both galanthamine (0.6 mg/g DW after 50 days) and lycorine (1.13 mg/g DW after 30 days).

Fully automated synthesis of 11C-acetate as tumor PET tracer by simple modified solid-phase extraction purification

IntroductionAutomated synthesis of 11C-acetate (11C-AC) as the most commonly used radioactive fatty acid tracer is performed by a simple, rapid, and modified solid-phase extraction (SPE) purification. MethodsAutomated synthesis of 11C-AC was implemented by carboxylation reaction of MeMgBr on a polyethylene Teflon loop ring with 11C-CO2, followed by acidic hydrolysis with acid and SCX cartridge, and purification on SCX, AG11A8 and C18 SPE cartridges using a commercially available 11C-tracer synthesizer. Quality control test and animals positron emission tomography (PET) imaging were also carried out. ResultsA high and reproducible decay-uncorrected radiochemical yield of (41.0±4.6)% (n=10) was obtained from 11C-CO2 within the whole synthesis time about 8min. The radiochemical purity of 11C-AC was over 95% by high-performance liquid chromatography (HPLC) analysis. Quality control test and PET imaging showed that 11C-AC injection produced by the simple SPE procedure was safe and efficient, and was in agreement with the current Chinese radiopharmaceutical quality control guidelines. ConclusionThe novel, simple, and rapid method is readily adapted to the fully automated synthesis of 11C-AC on several existing commercial synthesis module. The method can be used routinely to produce 11C-AC for preclinical and clinical studies with PET imaging.

Simultaneous determination of serine enantiomers in plasma using Mosher's reagent and stable isotope dilution gas chromatography‐mass spectrometry

D-Serine is a co-agonist of the N-methyl-D-aspartate receptor in glutamate neurotransmission and has been proposed as a potential therapeutic agent for schizophrenia. However, D-serine also acts as a nephrotoxic substance in rats at high doses. To investigate the pharmacokinetics and toxicokinetics of D-serine, a method for the stereoselective determination of serine enantiomers in rat plasma was developed using GC-MS with selected ion monitoring (GC-MS-SIM). DL-[(2)H(3)]Serine was used as an internal standard to account for losses associated with the extraction, derivatization and chromatography. Serine enantiomers were purified by cation-exchange chromatography using BondElut SCX cartridge and derivatized with HCl in methanol to form methyl ester followed by subsequent N,O-diacylation with optically active (+)-α-methoxy-α-trifluoromethylphenylacetyl chloride to form epimeric amide. Quantitation was performed by SIM of the molecular-related ions of the epimers in the chemical ionization mode. The intra- and inter-day reproducibility of the assay was less than 5% for D-serine and 3% for L-serine. The method was successively applied to study the pharmacokinetics of D-serine in rats.

Simultaneous determination of selenomethionine enantiomers in biological fluids by stable isotope dilution gas chromatography–mass spectrometry

A method for the stereoselective determination of d- and l-enantiomers of selenomethionine in mouse plasma was developed using gas chromatography–mass spectrometry with selected-ion monitoring (GC–MS-SIM). dl-[ 2H 3, 82Se]selenomethionine was used as analytical internal standard to account for losses associated with the extraction, derivatization and chromatography. Selenomethionine enantiomers in mouse plasma were purified by cation-exchange chromatography using BondElut SCX cartridge and derivatized with HCl in methanol to form methyl ester followed by subsequent N-acylation with optically active (+)-α-methoxy-α-trifluoromethylphenylacetyl chloride to form diastereomeric amide. Quantification was performed by SIM of the molecular-related ions of the diastereomers on the chemical ionization mode. The intra- and inter-day precision for d- and l-selenomethionine spiked to mouse plasma gave good reproducibility with relative standard deviation of 3% and 3% for d-selenomethionine and 6% and 3% for l-selenomethionine, respectively. The estimated amounts were in good agreement with the actual amounts spiked, the intra- and inter-day relative error being 5% and 2% for d-selenomethionine and 2% and 1% for l-selenomethionine, respectively. The present method is sensitive enough to determine pharmacokinetics of selenomethionine enantiomers.

Synthesis of O-(2-[18F]fluoroethyl)-l-tyrosine based on a cartridge purification method

O-(2-[(18)F]fluoroethyl)-L-tyrosine (FET) is widely used as a positron emission tomography tracer for brain tumors. Usually, a high-performance liquid chromatography (HPLC) purification at the end of the two-step synthesis is applied. In this work, we report an automatic radiosynthesis of FET with a purification procedure based on standard cartridges. O-(2-[(18)F]fluoroethyl)-L-tyrosine was prepared by [(18)F]fluoroethylation of L-tyrosine by a two-step synthesis using a modified [(11)C]methionine module (Nuclear Interface). In the first reaction step, we synthesized [(18)F]fluoroethyltosylate starting from [(18)F]fluoride. After a purification step, L-tyrosine was [(18)F]fluoroethylated with [(18)F]fluoroethyltosylate. The final reaction mixture was purified by means of solid phase extraction. The FET was trapped on an SCX cartridge, eluted with saline solution and trapped again on an HRX cartridge. For a second purification step, the FET was eluted from the HRX cartridge with ammonium acetate buffer and collected on two SCX cartridges followed by a washing step with water. The final product was eluted with saline solution and neutralised with 450 μl NaHCO(3) solution (8.4%). The synthesis was finished after 50 min and delivered the FET in a range of 3-16 GBq. The synthesis typically yielded 41% (21 experiments) of FET (d.c.) without an HPLC purification step. The radiochemical purity ranged between 97% and 100%. We present a radiosynthesis of FET where the usually used HPLC purification procedure has been substituted by a purification step based on standard cartridges. This method is useful for automatic modules without an expensive HPLC purification unit and for the routine production of FET.

Determination of antihypertensive and anti-ulcer agents from surface water with solid-phase extraction–liquid chromatography–electrospray ionization tandem mass spectrometry

Pharmaceuticals are emerging contaminants in surface water and they must be measured to follow their effects on the aquatic environment. We developed a solid-phase extraction and liquid chromatography–electrospray ionization tandem mass spectrometry (SPE-LC–ESI-MS/MS) method for the determination of twenty-six pharmaceutical compounds – which belong to antihypertensive and anti-ulcer agents – from surface water samples. The selection of pharmaceuticals was based on usage frequency in Hungary. During method development Oasis HLB, SampliQ Polymer SCX and Si-SCX SPE cartridges were tested. As LC eluent ammonium formate, ammonium acetate buffers at pH 3 and 5 were investigated and for quantitation both matrix-matched and internal standard calibration was used. For matrix effect assessment post-extraction spike method was applied which can separate the extraction efficiency from ion suppression for better determination of recovery. Method detection limits (MDLs) varied between 0.2 and 10ng/L. Precision of the method, calculated as relative standard deviation (RSD), ranged from 0.2 to 14.6% and from 1.2 to 22.4% for intra- and inter-day analysis, respectively. The method was applied to analyze Danube water samples. Measured average concentrations varied between 2 and 39ng/L for eleven compounds and another one could be detected under LOQ.

Multi-residue method for the determination of organochlorine pesticides in fish feed based on a cleanup approach followed by gas chromatography–triple quadrupole tandem mass spectrometry

A multi-residue method for the determination of organochlorine pesticides in fish feed samples was developed and optimized. The method is based on a cleanup step of the extracted fat, carried out by liquid–liquid extraction on diatomaceous earth cartridge with n-hexane/acetonitrile (80/20, v/v) followed by solid phase extraction (SPE) with silica gel–SCX cartridge, before the identification and quantification of the residues by gas chromatography–triple quadrupole tandem spectrometry (GC–MS/MS). Performance characteristics, such as accuracy, precision, linear range, limits of detection (LOD) and quantification (LOQ), for each pesticide were determined. Instrumental LODs ranged from 0.01 to 0.11 μg L −1, LOQs were in the range of 0.02–0.35 μg L −1, and calibration curves were linear ( r 2 > 0.999) in the whole range of explored concentrations (5–100 μg L −1). Repeatability values were in the range of 3–15%, evaluated from the relative standard deviation of six samples spiked at 100 μg kg −1 of fat, and in compliance with that derived by the Horwitz's equation. No matrix effects or interfering substances were observed in fish feed analyses. The proposed method allowed high recoveries (92–116%) of spiked extracted fat samples at 100 μg kg −1, and very low LODs (between 0.02 and 0.63 μg kg −1) and LOQs (between 0.05 and 2.09 μg kg −1) determined in fish feed samples.

Highly sensitive spectrophotometric determination of cationic surfactants in ground waters as their Cu(II)–TPPS aggregates preceded by solid-phase fractionation

A highly sensitive spectrophotometric determination of cationic surfactants in ground waters was established by forming their Cu(II)–TPPS aggregates, preceded by solid-phase extraction with an SCX cartridge. Cationic surfactants (CSs) were quantitatively trapped and isolated by the SCX solid phase. The use of Cu(II)–TPPS anionic chromophore could reduce the interference by unintentional metal ions coexisting in surrounding waters. The method was very sensitive in the determination of CSs less than 10 −5 M levels with acceptable recovery and calibration data. The colorimetric sensitivity was very dependent on the alkyl-chain length of the surfactants, and a CS having 23 carbon atoms gave the highest sensitivity. Overall recoveries were 95–97% with R.S.Ds. less than 3% in the cases over 10 −6 M levels. In the cases in 10 −7 M levels, however, a portion of the analyte would be adsorbed by reservoir walls, which could seriously affect the trace determination. The preliminary addition of 4,4′-bipyridyl into the sample solution was effective in decreasing such unintentional analyte losses, leading up to 73% recovery. The developed method was applied to the analysis of river water at ppb levels of CSs with a fractional concentration through a SCX solid phase subjected by 500-mL aliquots of sample.

Determination of ractopamine and clenbuterol in feeds by gas chromatography–mass spectrometry

A simple, sensitive and reproducible gas chromatographic–mass spectrometric method was developed for monitoring ractopamine (RAC) and clenbuterol (CLB) in feeds. Feed samples were extracted with 0.1 M perchloric acid, centrifuged, neutralized, followed by liquid–liquid extraction with ethyl acetate-isopropanol (9:1, v/v). The concentrated extracts were dissolved in 0.02 M NH 4Ac (pH 5.2), and applied to a solid phase extraction SCX cartridge for cleanup. The drugs were eluted with 3% (v/v) ammonia hydroxide in methanol, and the eluate was evaporated to dryness. The residue was derivatized with N, O- bis (trimethylsilyl) trifluoroacetamide at 80 °C for 1 h, and cooled, then analyzed by gas chromatography–mass spectrometry. The selected ions monitoring mode was performed at m/z 179, 250, 267 and 502 for RAC, and m/z 86, 243, 262 and 277 for CLB. Recoveries of RAC and CLB from concentrated feeds and premix fortified at 10, 100 and 5000 μg/kg were between 64.6 and 84.2%, with relative standard deviations of less than 15%. The limits of detection were about 4 μg/kg for RAC and 2 μg/kg for CLB.

Determination of d- and l-enantiomers of methionine and [ 2H 3]methionine in plasma by gas chromatography–mass spectrometry

A method for the stereoselective determination of d- and l-enantiomers of both methionine and [ 2H 3]methionine in rat plasma was developed using gas chromatography–mass spectrometry with selected-ion monitoring (GC–MS-SIM). dl-[ 2H 7]Methionine was used as analytical internal standard to account for losses associated with the extraction, derivatization and chromatography. The amino acids were purified by cation-exchange chromatography using BondElut SCX cartridge and derivatized with HCl in methanol to form methyl ester followed by subsequent N-acylation with optically active (+)-α-methoxy-α-trifluoromethylphenylacetyl chloride to form diastereomeric amide. Quantification was performed by SIM of the molecular-related ions of the diastereomers on the chemical ionization mode. Endogenous l-methionine concentrations in 50 μl of rat plasma were measured with relative intra- and inter-day precision of 4.0 and 6.3%, respectively. The intra- and inter-day reproducibility in the amounts of d- and l-[ 2H 3]methionine determined were in good agreement with actual amount added.

Analysis of N7‐(benzo[a]pyrene‐6‐yl)guanine in urine using two‐step solid‐phase extraction and isotope dilution with liquid chromatography/tandem mass spectrometry

Analysis of urinary N7-(benzo[a]pyren-6-yl)guanine (BP-6-N7Gua), a DNA adduct induced by benzo[a]pyrene, may serve as a risk-associated biomarker for exposure to polyaromatic hydrocarbons (PAHs). In this study a highly sensitive and specific analytical method, incorporating on-line sample preparation coupled with isotope-dilution liquid chromatography and tandem mass spectrometry (LC/MS/MS), was developed to quantitate this adduct in human urine. In order to achieve accurate quantitation, 15N5-labeled BP-6-N7Gua was synthesized to serve as the internal standard, and a two-step solid-phase extraction (SPE) procedure using C8 and SCX cartridges was used for sample cleanup. BP-6-7-N7Gua was analyzed using positive ion LC/MS/MS operated in multiple reaction monitoring (MRM) mode. The [M+H]+ ions at m/z 402 and 407 and the common fragment ion of [M+H]+ at m/z 252 were monitored for quantification. The recovery of this analyte after two-step SPE was 90%, and the limit of detection was 2.5 fmol/mL in 10 mL of urine. This highly specific and sensitive method for BP-6-N7Gua in urine may be applied to assess exposure to PAHs in coke-oven workers for future molecular epidemiology studies on health effects of PAHs.

Spectrophotometric Determination of Polyoxyethylene Nonionic Surfactants in Ground Waters Using Triple-Stage Solid-Phase Extraction Followed by an Improved Ferric Thiocyanate Complexation Method

A practical spectrophotometric determination method for polyoxyethylene nonionic surfactants in ground waters was established, which is based on a ferric thiocyanate complexation colorimetric method preceded by a triple-stage solid-phase extraction technique using SCX, SAX, and C18 cartridges interconnected. Cationic and anionic surfactants interfere with the determination and were therefore effectively trapped and isolated by the SCX and SAX solid phases, respectively. Nonionic surfactants (NSs) were finally introduced and concentrated in the C18 cartridge. The analyte was quantitatively eluted from the C18 sorbent, and the residue was subjected to the colorimetric determination. The calibration line was linear (r2 = 0.9997) up to 200 µg L−1 of heptaoxyethylenedodecylether when analyzing sample sizes of 100 mL. Overall recoveries were 95–97% with an RSD of less than 3%. The method was applied to the analysis of river water, and 6.4 µg L−1 of NSs as heptaoxyethylenedodecylether was found by means of the standard addition method. The proposed method is very practical and features minimum consumption of chemicals.

Simultaneous determination of sulfadiazine and trimethoprim in animal feed by liquid chromatography with UV and tandem mass spectrometric detection

A method for the simultaneous determination of sulfadiazine and trimethoprim in animal feed was developed and validated. The chromatography was performed both on a liquid chromatography–ultraviolet detection (LC–UV) and LC–tandem mass spectrometric detection (MS/MS) instrument, intended to be applied for two separate purposes. The LC–UV method was used to examine the quality requirements for medicated premixes incorporated into animal feedingstuffs. This comprised e.g. the homogeneity testing in medicated feed at the mg kg −1 level. The LC–MS/MS method was used to quantify residues of sulfadiazine and trimethoprim in unmedicated feed at the μg kg −1 level. Sulfadiazine and trimethoprim were extracted from animal feed with 50% methanol in water and with ethylacetate. Clean-up was performed on a benzenesulfonic acid SCX cartridge. The LC–UV analysis of the extracts was done on a Hypersil ODS column using a gradient of 0.5% acetic acid, 0.25% triethylamine and 5% methanol in water (A) and 5% methanol in acetonitrile (B). Detection was at 240 nm for trimethoprim and 270 nm for sulfadiazine. The same extracts were also injected for quantitation onto an ion-trap LC–MS n instrument, operating in the atmospheric pressure chemical ionization tandem MS mode (APCI–MS/MS). The elution was performed isocratically with 0.1% acetic acid in water and methanol. Both the LC–UV and LC–MS/MS methods were validated for their linearity, trueness, precision, LOD (limit of detection), LOQ (limit of quantification) and extraction recovery. All results fell within the ranges specified. Both methods were useful, but the LODs of the LC–MS/MS method were about 200-times lower than with UV detection, i.e. 0.052 mg kg −1 sulfadiazine and 0.022 mg kg −1 trimethoprim. Therefore, the LC–MS/MS method was applied for monitoring residues as a possible result from carry-over during feed production.

Simultaneous determination of N-oxides and free bases of pyrrolizidine alkaloids by cation-exchange solid-phase extraction and ion-pair high-performance liquid chromatography

Cation-exchange solid-phase extraction using LiChrolut SCX (Merck, Darmstadt) cartridges filled with polymeric strong cation-exchanger enabled efficient isolation of both N-oxides and free bases of pyrrolizidine alkaloids (PAs). The recoveries were about 80% for retrorsine- N-oxide, 90% for retrorsine and 100% for senkirkine and were assessed both by TLC–densitometry and ion-pair high-performance liquid chromatography (HPIPC) on Hypersil BDS C 8 stationary phase and hexane-1-sulfonic acid as ion-pairing agent. The applied HPIPC gradient procedure was suitable for separation of PAs with various types of structures ( N-oxides, free bases, otonecine-PAs). The method limits of detection and quantitation, respectively, ranged from 0.06 ng/μl (senecionine) and 0.2 ng/μl (senkirkine) to 0.1 and 0.35 ng/μl for retrorsine- N-oxide. For each component calibrated by linear regression method, correlation coefficients were higher than 0.9995 (six-point calibration from 4 to 100 μg/ml). The elaborated procedure was used in searching for PAs in plant derived samples from Symphytum sp. (comfrey), Petasites hybridus and Petasites albus (butterbur), Tussilago farfara (coltsfoot), Emilia coccinea (tassel flower) and Doronicum columnae (leopard’s bane). For the last three samples macrocyclic PAs (senecionine, senecionine- N-oxide, senkirkine) have been detected for the first time. Details of precision of the analyses are also included.