Chuanlai Xu

Evaluation and interlaboratory validation of a GC-MS method for analysis of pesticide residues in teas

A method was described for simultaneous determination of nine organic heterocyclic pesticide residues by gas chromatography-mass spectrometry-selected ion monitoring. Atrazine, vinclozolin, procymidone, triflumizole, imazalil, buprofezin, propiconazole, fenarimol, and pyridaben were clearly separated from each other, extracted with acetone—hexane mixture, purified with graphitized carbon black cartridge and neutral Al2O3 cartridge, eluted with acetone—hexane mixture, simultaneously determined by GC-MS, and then quantified with an external standard method. Recoveries of pesticides ranged from 73 % to 116 % at the spiked level of 0.01–30 mg kg−1, while the relative standard deviation was between 3 % and 27 %. In addition, the limits of determination (0.01 mg kg−1 to 5.0 mg kg−1) and linearity (0.02–40 μg mL−1) revealed that simultaneous determination of multi-residues in Chinese teas (like Oolong tea, green tea, red tea, etc.) was possible. Furthermore, an interlaboratory study among 5 labs was conducted to further validate the method, and the results were satisfactory.

Synthesis of derivatives and production of antiserum for class specific detection of pyrethroids by indirect ELISA

Two series of haptens including 3-phenoxybenzoic acid (PBA) and 3-(2-chloro-3, 3,3-trifluoroprop-1-enyl)-2,2-dimethylcyclo-propanecarboxylic acid (CF3MPA) were used to prepare immunogens through attachment of 4-C or 6-C handles. Class selective antibodies were produced by immunising rabbits. Ab502 showed the highest reactivity towards tau-fluvalinate (IC50 1.3 ng mL−1), λ-cyhalothrin (IC50 2.3 ng mL−1), cyfluthrin (IC50 2.2 ng mL−1) and fenpropathrin (IC50 18.5 ng mL−1) among the antibodies in a competitive ELISA. The effects of methanol, pH and salt concentration were optimised for maximum efficiency of the ELISA (Enzyme-Linked ImmunoSorbent Assay). Ab502 (1:80000)/2-OVA-1(0.2 µg mL−1) was chosen for ELISA optimisation. Finally, 0.05 M phosphate buffered saline (PBS) at pH 6.5 containing 30% methanol (v/v) was used to dilute the standards. Target analytes in honey samples were extracted with ethyl acetate by sonication. The samples were spiked with three different concentrations of each compound (tau-fluvalinate, 0.5 ng g−1, 3 ng g−1, 12 ng g−1; λ-cyhalothrin and cyfluthrin 1 ng g−1, 5 ng g−1, 65 ng g−1). The recoveries were 36–59% at the lowest spiking concentration and 61–81% at the higher concentration. This assay might be useful to screen pyrethroid residues in honey or other matrix.

Development of colloidal gold-based immunochromatographic assay for the rapid detection of medroxyprogesterone acetate residues

Abstract One-step membrane-based competitive colloidal gold-based immunoassays in lateral-flow formats for the rapid detection of medroxyprogesterone acetate (MPA) were developed. Nitro-cellulose membrane strip was separately coated with goat anti-rabbit IgG (control line) and MPA hapten-OVA conjugate (test line). Anti-MPA polyclonal antibody labelled with colloidal gold particles was firstly incubated with MPA. A positive reaction as a result of the remaining antibody-gold conjugate combining with antigen coated on the membrane was obvious by visual detection, with detection limits for lateral flow of 5 µg/kg for detecting MPA standard solution, and the limit of detection was 10 µg/kg for detecting the MPA spiked in swine urine and liver. The assay time for test was less than 5 min, suitable for rapid testing on-site.

Effect of fructooligosaccharides and antibiotics on laying performance of chickens and cholesterol content of egg yolk

1. Nine hundred and sixty 25-week-old Lohmann Brown layers were randomly divided into 5 groups with 216 layers in each group. Layers in group one were fed a control diet, group 2 received the control diet plus 20 mg/kg zinc bacitracin and 4 mg/kg colistinsulfate, and the remaining three groups received control diet plus 2000, or 4000, or 6000 mg/kg fructooligosaccharide (FOS). 2. The results showed improvements in egg production, feed consumption and feed conversion of layers when 2000 mg/kg FOS was added to the diets. 3. The results also showed some additional improvements in the group supplemented with 2000 mg/kg FOS, including increases in egg shell thickness, yolk colour and Haugh unit, and decreases in yolk cholesterol concentration. 4. However, larger (excessive) doses of FOS did not improve the performance of layers.

A direct enzyme-linked immunosorbent assay for hexoestrol residues

Abstract A simple enzyme-linked immunosorbent assay for the detection of hexoestrol (HES) residue has been developed. The HES derivatives, hexoestrol-mono-ether-butyrate-ethyl (HES-MEBE) and hexoestrol-mono-caroxyl-propyl-ethyl (HES-MCPE) were synthesised to enable the coupling of HES and proteins together. The immunogen HES-MCPE-BSA and enzyme-linked antigen HES-MCPE-HRP were prepared by mixed anhydride methods. After the antibody was obtained by immunisation of New Zealand rabbits, a direct enzyme-linked immunosorbent assay was developed for the determination of HES residue. Several parameters were optimised. Excellent linearity (R 2=0.9905) was demonstrated from 0.01 to 8.1 ng/ml. The 50% inhibition value (IC50) and the limit of detection (LOD) were 0.23 and 0.01 ng/ml, respectively. The specificity of the immunoassay was evaluated by determining cross-reactivity of six hormones (diethylstilbestrol, dienestrol, 19-nortestosterone, medroxyprogesterone, testosterone and estrone), and all cross-reactivity rates were <0.5%. The recoveries from beef and fish ranged from 61 to 102%. The stabilisation of the coated plates was also studied. The ELISA developed was used to detect the residues in chicken muscle tissues and confirmed by high-performance liquid chromatography and tandem mass spectrometry (LC/MS/MS). This proposed method could be applied to routine residue analysis.

Development and Optimization of an Indirect Enzyme‐Linked Immunosorbent Assay for Thiamphenicol

Abstract The specificity of an enzyme‐linked immunosorbent assay (ELISA) was modified by changing the chemical structure of the haptenated coating antigen. A competitive indirect immunoassay was developed, evaluated, and validated to measure thiamphenicol (TAP) in eels. The working range of the assay was placed between 0.45 and 8.35 µg/L. Moreover, the influence of several physicochemical parameters, such as incubation time, ionic strength, detergent concentration, and pH, were studied. The specificity of this immunoassay was evaluated using 23 structurally related compounds. Finally, a good correlation was observed between the chromatographic and immunoassay techniques while analyzing five certified eel samples.

Comparative Analysis of Medroxyprogesterone Acetate Residue in Animal Tissues by ELISA and GC‐MS

Abstract Medroxyprogesterone acetate is a xenobiotic growth promoter of meat‐producing animals. In this study, the liver, kidney, and muscle of rabbits were obtained after the medroxyprogesterone acetate was administrated successively for 5 days at 0.8 mg/kg and withdrawn for 7 days before slaughter. The medroxyprogesterone acetate residues were determined by ELISA method. The average residue in muscle, liver, and kidney, as 7, 6, and 23 µg/kg, respectively. Residues were confirmed by GC/MS by monitoring four ions, 373, 283, 265, and 145 m/z. A good correlation was observed between the GC/MS and ELISA methods.

Separation and identification of synthetic antigens of hexoestrol residue in animal derived food by HPLC-MS

Abstract Hexoestrol and its derivatives, hexoestrol-mono-ether-butyrate-ethyl (HES-MEBE), hexoestrol-di-ether-butyrate-ethyl (HES-DEBE) were separated and identified using liquid chromatography and spectrometry (ESI). The conditions were as follows: Methyl alcohol and water as mobile phase, 0.3 ml/min of flow rate, elution with linear gradient concentrations from 60–100% of methanol for 25 min on a Lichrospher C-18 column. Synthetic antigen was prepared by coupling hexoestrol-mono-caroxyl-propyl-ethyl (HES-MCPE) with carrier protein by the mixed anhydride method, and the conjugation rate, analysed by UV scanning, is 36.

Simultaneous Determination 9 Anabolic Steroids Residues in Animal Muscle Tissues by Gas Chromatography‐Mass Spectrometry

Abstract A method was developed for determining 9 anabolic steroids (ASs) residues in animal muscle tissues by gas chromatography‐mass spectrometry (GC/MS). After undergoing enzymolysis, being homogenized, and then being picked‐up by ultrasound the sample was extracted with tert‐Butyl methyl ether, cleaned up through solid‐phase extraction (SPE) based on the reverse phase (RP) principle, and after derivatization of the analyst of interest, analyzed by GC/MS under selective ion monitoring (SIM). The limits of the detection (LOD) of the GC/MS method used for testing epitestosterone (ETS), nandrolon (17β‐NT), 17α‐methyl‐testosterone (MTS), testosterone 17‐propionate (PTS), 17β‐estradiol 3‐benzoate (BES), estrone (ESN), 17β‐estradiol (17β‐ES), 17α‐ethynylestradiol (EES), and estriol (EST) in animal muscle ranged from 0.10 to 0.33 µg/kg and the limits of quantification (LOQ) were from 0.24 to 0.82 µg/kg. The experiments using spiked samples, such as pork, beef, chicken, and fish, made it clear that at addition level of 2.0 µg/kg, the average recovery of the ASs ranged from 62.5% to 80.5%, and the coefficient of variation ranged from 12.5% to 26.8%, while at an addition level of 5.0 µg/kg, the average recovery was from 70.8% to 86.4%, and the coefficient of variation was from 8.8% to 18.4%.

Determination of 19-nortestosterone residues in aquaculture tissues by enzyme-linked immunosorbent assay and comparison with liquid chromatography and tandem mass spectrometry

19-Nortestosterone (17β-NT) was oximated by carboxymethoxylamine and then coupled with bovine serum albumin (BSA) in a mixed-anhydride reaction in order to produce an antibody. The conjugate rate of 17β-NT and BSA was estimated to be 24 by ultraviolet spectrophotometry. Polyclonal antibody of 17β-NT was acquired from the animal immunized with the conjugate. Through an indirect enzyme-linked immunosorbent assay (ELISA), which demonstrated that the synthesis of immunogen was successful, the titre of antiserum was found to be 6.4 × 105. Based on the purified antibody, a competitive indirect ELISA was developed. ELISA revealed that the limit of detection (LOD) was 0.07 ng g−1, the recovery (in edible tissues) was 71–89%, and the working range was 0.05–31.25 ng g−1. The preliminary evaluation of assay performance through specificity, sensitivity, precision, and accuracy revealed that this ELISA method could be used in the practical detection of 17β-NT in tissue samples. Moreover, this method was compared with high-performance liquid chromatography tandem mass spectrometry, for which the transition for quantification of 17β-NT was 275.4/109.1.