Yijuan Su

Novel cyromazine imprinted polymer applied to the solid-phase extraction of melamine from feed and milk samples

A water compatible molecularly imprinted polymer (MIP) using cyromazine as a mimic template, methacrylic acid as the functional polymer and ethylene glycol dimethacrylate as the cross-linker was synthesized and used to extract melamine from feed and milk samples via a molecularly imprinted solid-phase extraction (MISPE) protocol. Optimum retention of melamine on the MISPE cartridge was achieved using methanol, and the interferences in the samples were effectively washed out. The binding capacity of the polymer toward melamine was found to be about 500 microg of melamine/g of polymer. The recoveries of 2 microg and 20 microg melamine standard spiked into water extract of blank feeds and milk samples were between 83.4% and 103%, with relative standard deviation <5.6%. The cyromazine-MIP demonstrated high cross-reactivity for melamine and low affinity to cyanuric acid. The ionic bond interaction was regarded as the main factors that dominated the retention of the melamine on the MISPE cartridge.

Solid-phase extraction of melamine from aqueous samples using water-compatible molecularly imprinted polymers.

A method for molecularly imprinted SPE of the melamine from environmental water samples was investigated. Cyromazine-imprinted polymers were synthesized in water-methanol systems for the selective extraction of melamine from aqueous samples, followed by HPLC analysis. Molecular recognition properties and binding capability to melamine were evaluated by adsorption test and Scatchard analysis, which showed the dissociation constant (KD) and the maximum binding quantity (Qmax) were 0.032 micromol/L and 4.77 micromol/g for high affinity binding site, and 0.26 micromol/L and 19.10 micromol/g for lower affinity binding site, respectively. Under the optimum extraction protocol, the method can be successfully applied to selectively extract and enrich melamine in environmental water. The linearity was ranged from 0.500 to 100.0 ng/mL (r > 0.999) in tap water, lake water, and seawater analysis. When 50 mL of the water samples loaded, the LODs of the method were ca. 0.1 ng/mL, and the LOQs were ca. 0.5 ng/mL. The mean recoveries of melamine from blank water samples spiked at 0.5, 5.0, and 50 ng/mL were more than 86.3%, with the RSD less than 8.8%.

Matrix Effects in Analysis of β-Agonists with LC-MS/MS: Influence of Analyte Concentration, Sample Source, and SPE Type

The synergistic influences of analyte concentration, sample source, and solid-phase extraction (SPE) type on matrix effects in the multiresidue analyses of eight β-agonists with LC-ESI-MS/MS were evaluated. Porcine muscle and liver extracts and urine from diverse sources were purified by strong or mixed-mode cation exchange and molecularly imprinted polymer SPE cartridges, respectively. Three spiked concentrations (2, 10, and 20 ng/mL) of eight β-agonists in the purified matrices and the different sample sources were analyzed. The results show that for most β-agonists there are significant differences in matrix effects between analyte concentrations or sample sources (P < 0.05), whereas there is no significant difference in matrix effects between different SPE cartridges (P > 0.05). Results from main effects testing indicated that analyte concentration was the main effector.

High-throughput screening and confirmation of 22 banned veterinary drugs in feedstuffs using LC-MS/MS and high-resolution Orbitrap mass spectrometry.

A new analytical strategy based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) combined with accurate mass high-resolution Orbitrap mass spectrometry (HR-Orbitrap MS) was performed for high-throughput screening, confirmation, and quantification of 22 banned or unauthorized veterinary drugs in feedstuffs according to Bulletin 235 of the Ministry of Agriculture, China. Feed samples were extracted with acidified acetonitrile, followed by cleanup using solid-phase extraction cartridge. The extracts were first screened by LC-MS/MS in a single selected reaction monitoring mode. The suspected positive samples were subjected to a specific pretreatment for confirmation and quantification of analyte of interest with LC-MS/MS and HR-Orbitrap MS. Mean recoveries for all target analytes (except for carbofuran and chlordimeform, which were about 35 and 45%, respectively) ranged from 52.2 to 90.4%, and the relative standard deviations were <15% except for 20% for carbofuran. The decision limits (CCαs) for target analytes in formulated feed were between 2.6 and 23 μg/kg, and the detection capabilities (CCβs) were between 4.2 and 34 μg/kg. The method was successfully applied to screening of real samples obtained from local feed markets and confirmation of the suspected target analytes. It provides a high-throughput, sensitive, and reliable screening, identification, and quantification of banned veterinary drugs in routine monitoring programs of feedstuffs.

Molecularly imprinted solid-phase extraction for the selective HPLC determination of ractopamine in pig urine.

A method was developed for the determination of ractopamine in pig urine using molecularly imprinted solid-phase extraction (MISPE) as the sample clean-up technique combined with high-performance liquid chromatography. The molecularly imprinted polymer (MIP) was synthesized in acetonitrile-triethylamine system using ractopamine (RAC) as the template and acrylamide as the monomer. The binding capacity of the polymer toward RAC was found to be about 2.57 mg of ractopamine/g of polymer. The optimal procedures for MISPE consisted of conditioning with 3 mL methanol, equilibrating with 3 mL of water, loading volume of <10 mL of aqueous sample (pH 7), washing with 3 mL water and 3 mL methanol, and eluting with 5 mL of 5% ammonia in methanol. In the four spiked samples with the levels of 0.01, 0.1, 1.0 and 5.0 μg/mL, the mean recoveries of analyte on the MIP were higher than 90% with relative standard deviation <10%, and significant differences between imprinted and non-imprinted materials were observed. The MIP selectivity was evaluated by checking 11 drugs with similar and different molecular structures to that of RAC. The characteristics of three-dimensional cavities and hydrogen bond interaction were regarded as the main factors that dominated the retention of RAC on the MISPE cartridge.

Development of a modified QUick, Easy, CHeap, Effective, Rugged and Safe method for the determination of multi-class antimicrobials in vegetables by liquid chromatography tandem mass spectrometry.

A modified quick, easy, cheap, efficient, rugged and safe (QuEChERS) method coupled with liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed for rapid determination of 26 veterinary antimicrobials in vegetables. Samples were extracted by single-phase extraction with acetonitrile-methanol (85:15, v/v) and citric buffer solution, followed by liquid-liquid partitioning with the addition of anhydrous magnesium sulfate and sodium chloride. A dispersive solid-phase extraction with primary secondary amine was applied for cleanup. Concentration and solvent exchange was performed prior to LC-MS/MS analysis. All matrix-matched calibration curves were linear with correlation coefficients (r) over 0.99. Recoveries for all the analytes spiked at 0.5 (1 or 1.5), 5 and 50 ng/mL were in the range of 60.0-98.0%, except for sulfaquinoxaline, sulfaclozine and doxycycline, with relative standard deviations below 25% for the low concentration level, 20% for the medium and 15% for the high. The decision limits and the detection capabilities of the analytes ranged from 0.005 to 0.5 μg/kg and from 0.02 to 1.5 μg/kg, respectively. The method was developed and validated in accordance with romaine lettuce matrix, and higher recovery rates were obtained from the other five kinds of vegetables including white radish, Chinese cabbage, cucumber, string bean and green pepper. Matrix effects of different vegetables were evaluated and signal suppression effect was observed for the majority of 26 analytes. Finally, the method was applied to the analysis of real samples collected from the agricultural areas in the vicinity of local pig farms, and the phenomenon of vegetables contaminated by antimicrobials residues is provoking.

Simultaneous determination of quinoxaline-1,4-dioxides in feeds using molecularly imprinted solid-phase extraction coupled with HPLC

A simple, selective, and reproducible molecularly imprinted SPE coupled with HPLC method was developed for monitoring quinoxaline-1,4-dioxides in feeds. Molecularly imprinted polymers were synthesized in methanol using mequindox (MEQ) as template molecule and acrylamide as functional monomer by bulk polymerization. Under the optimum SPE conditions, the novel polymer sorbents can selectively extract and enrich carbadox, MEQ, quinocetone, and cyadox from a variety of feeds. The molecularly imprinted SPE cartridge was better than nonimprinted, C(18) , and HLB cartridges in terms of both recovery and precision. Mean recoveries of four quinoxaline-1,4-dioxides from six kinds of feeds spiked at 1.0, 10, and 100 mg/kg ranged between 75.2 and 94.7% with RSDs of less than 10%. The decision limits (CCαs) and the detection capabilities (CCβs) of four analytes were 0.15-0.20 mg/kg and 0.44-0.56 mg/kg, respectively. The class selectivity of the polymers was evaluated by checking three drugs with different molecular structures to that of MEQ.

Simultaneous determination of cyadox and its metabolites in plasma by high-performance liquid chromatography tandem mass spectrometry

Cyadox is a novel antimicrobial growth-promoter of the quinoxalines. For food safety and pharmacokinetic studies, a convenient, sensitive and reproducible LC-ESI-MS/MS method was developed for the simultaneous determination of cyadox and its major metabolites, quinoxaline-2-carboxylic acid, 1,4-bisdesoxycyadox, cyadox-1-monoxide and cyadox-4-monoxide in chicken plasma. Plasma sample was subjected to a simple deproteinisation with acetonitrile. Analysis was performed on a C18 column by detection with mass spectrometry in multiple reaction monitoring mode. A gradient elution program with 0.2% formic acid, methanol and acetonitrile was performed at a flow rate of 0.2 mL/min. The decision limits (CCαs) of five analytes in plasma ranged from 1.0 to 4.0 μg/L, and the detection capabilities (CCβs) were <10 μg/L. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The extraction recoveries of five analytes were between 87.4 and 93.9% in plasma at the spiked levels of 5 (10)-200 μg/L with the relative standard deviations <10% for each analyte. The developed method demonstrated a satisfactory applicability in real plasma samples.

Matrix effects caused by phospholipids in multi-residue analysis for beta-agonists with liquid chromatography-electrospray tandem mass spectrometry

It was first confirmed that 5 glycerophospholipids (GPs, (15.6 mg L−1)), including lysophosphatidylcholine (LPC C18:0), lysophosphatidylcholine, LPC (C16:0), phatidylcholine, PC (C16:0/C18:2), phatidylcholine, PC (C16:0/C18:1) and phatidylethanolamine, PE (C18:0/C20:4), could significantly suppress the ionization of nine beta-agonists (2 μg L−1) using post-column infusion during liquid chromatography-electrospray tandem mass spectrometry analyses. Under optimal experimental conditions, most beta-agonists such as fenoterol, clorprenaline, tulobuterol, clenbuterol and especially penbuterol were co-eluted with GPs, and a positive linear correlation (R2 > 0.85) between the amount of GPs and the extent of ionization suppression was observed for the five analytes. In addition, the linear correlation was statistically significant (p < 0.001). GPs including LPC C18:0 (m/z 496.2), PC C16:0/C18:2 (m/z 758.5), PC C16:0/C18:1 (m/z 760.5), PE C18:0/C20:4 (m/z 768.6) and plasmalogen phosphatidylcholine, PLPC C18 (Plasm)/C18:1 (m/z 772.6) were detected in the final extracts of porcine liver sample. These endogenous GPs may be the main effectors that cause matrix effects in beta-agonist residue detection in porcine liver.

Quick Multi-Class Determination of Residues of Antimicrobial Veterinary Drugs in Animal Muscle by LC-MS/MS

On the basis of the highly sensitive and selective liquid chromatography-tandem mass spectrometry technique, a generic extraction solvent and a sample dilution method was developed for the residue analysis of different polar veterinary drugs known as fluoroquinolones, sulfonamides, macrolides, and tiamulin in chicken muscle. The results showed that the matrix-matched calibration curves of all 10 compounds were in an effective linear relationship (r2 ≥ 0.997) in the range of 0.2–100 μg L−1. At three spiking levels of 2 (5), 50, and 100 μg kg−1, average recoveries of analytes were between 67.1% and 96.6% with relative standard deviations of intra-day and inter-day below 20%. The limits of detection and limits of quantification of the method were in the range of 0.3–2.0 μg kg−1 and 2.0–5.0 μg kg−1, respectively, which were significantly lower than their maximum residue limits. In addition, the intensity of the target analytes and its corresponding matrix effects were obviously related to the sample dilution times (matrix concentration). There were no significant differences (p > 0.05) in the average content of almost any of the analytes in medicated chickens between this method and the method in the literature for determining analytes. Lastly, the proposed method was successfully applied for the simultaneous analysis of 10 common veterinary drugs in food animal muscle tissues.