Xiangmei Li

Development of an immunochromatographic strip test for rapid detection of melamine in raw milk, milk products and animal feed.

A simple, rapid and sensitive immunogold chromatographic strip test based on a monoclonal antibody was developed for the detection of melamine (MEL) residues in raw milk, milk products and animal feed. The limit of detection was estimated to be 0.05 μg/mL in raw milk, since the detection test line on the strip test completely disappeared at this concentration. The limit of detection was 2 μg/mL (or 2 μg/g) for milk drinks, yogurt, condensed milk, cheese, and animal feed and 1 μg/g for milk powder. Sample pretreatment was simple and rapid, and the results can be obtained within 3-10 min. A parallel analysis of MEL in 52 blind raw milk samples conducted by gas chromatography-mass spectrometry showed comparable results to those obtained from the strip test. The results demonstrate that the developed method is suitable for the onsite determination of MEL residues in a large number of samples.

Residue depletion of gentamicin in swine tissues after intramuscular administration.

A sensitive and robust high-performance liquid chromatographic method with fluorescence detection (HPLC-FLD) was developed for the determination of gentamicin (GEN) residues in swine tissues. The limit of quantification (LOQ) of the method was 50 ng/g for muscle and 100 ng/g for liver and kidney. Mean recoveries at all fortification levels ranged from 82.34 to 93.20% with coefficient of variation (CV) below 5.39%. Residue depletion study of GEN in swine was performed after intramuscular injections twice daily at a dose of 4 mg/kg of bw with 12 h intervals for 5 consecutive days. The concentrations of GEN were determined in injection site, muscle, liver, and kidney by the HPLC-FLD method. The highest GEN concentration was measured in kidney, indicating that kidney was the primary target tissue for GEN residue. GEN concentrations in all examined tissues were below the maximum residue limit (MRL) recommended by the European Union (EU) at 50 days post-treatment.

Development of a Monoclonal Antibody-Based ELISA for the Detection of Sulfaquinoxaline in Chicken Tissues and Serum

Abstract A monoclonal antibody (Mab) was produced by using sulfaquinoxaline-human serum albumin (SQX-HSA) conjugate as immunogen. The anti-SQX Mab exhibited negligible cross reactivity with other commonly used sulfonamides. Using this Mab, a competitive indirect enzyme-linked immunosorbent assay (ciELISA) was developed to detect SQX in chicken tissues and serum. The ciELISA showed a 50% inhibition (IC50) value of 2.60 ng/mL. The recoveries of SQX from spiked chicken muscle, liver, and serum at levels of 5–50 µg/kg were 82.6–96.5%, 75.3–94.5%, and 69.7–89.3%, respectively. The coefficient variations (CVs) were 6.22–7.17%, 4.9–8.9%, and 1.20–10.15%, respectively. Detection limits were 1.29 µg/kg in muscle, 1.32 µg/kg in liver, and 2.44 µg/kg in serum.

Pharmacokinetics of a novel amoxicillin/colistin suspension after intramuscular administration in pigs

An amoxicillin (AMO) or colistin (COS) oil suspension was developed and corresponding pharmacokinetics studies were conducted in pigs after i.m. injection. The combination product is a white- to cream-colored oil suspension which is easy to be re-dispersed. Settling volume ratio, syringeability and flowability of the product is well consistent with the technical standards set by the Ministry of Agriculture of Peoples Republic of China. Two studies were conducted to investigate the pharmacokinetics of the combination product in swine. First, the pharmacokinetics of the combination product was compared with those of the same products merely removing either AMO or COS. No significant change in the major pharmacokinetic parameters (C(max) , T(max) , MRT, t(1/2λ) , AUC and AUMC) was observed when either component was removed from the combination product, indicating that AMO and COS do not interfere with each other in their absorption and distribution in the tissue when used as a combination. Second, the pharmacokinetics of the combination product was compared with that of their respective single products. It was found that the apparent elimination half-lives (t(1/2λ) ) of AMO and COS in combination product were 6.38 and 8.09 h, which were 2.40 and 2.38 times longer than the single products, respectively. Thus, the novel AMO/COS suspension extended significantly the half-life of both drugs to maintain a longer drug residence time in pigs when compared to their single products.

Pharmacokinetics of doramectin in rabbits after subcutaneous administration.

Doramectin (DRM), 25-cyclohexyl-5-O-demethyl-25-de (1-methylpropyl) avermectin B1, is an exceptionally potent anthelminthin with a broad spectrum of activity against nematode and arthropod parasites acting at very low dosage (Goudie et al., 1993). Initial in vivo evaluation of DRM is carried out against the sheep parasite, Trichostrongyle colubriformis, and the rabbit ear mite, Psoroptes cuniculi. It is commercially available since 1993 (Goudie et al., 1993) and injectable oil-based formulations of DRM are particularly optimized for their pharmacodynamic efficacy (Wicks et al., 1993). DRM is licensed for use in cattle, swine, and sheep in USA and many European countries as well as in China (Flajs et al., 2005). The pharmacokinetic behavior of DRM after various routes of administration have been studied in domestic animals and livestock (Lanusse et al., 1997; Toutain et al., 1997; Escudero et al., 1999; Gayrard et al., 1999; Atta & Abo-Shihada, 2000; Pérez et al., 2002; Barber et al., 2003; Flajs et al., 2005; Zhang et al., 2005, 2007; Gokbulut et al., 2006). However, its off-label use for the treatment of parasite in rabbits has been increasing in recent years. Up to now, studies on pharmacokinetics of DRM in rabbits have not been reported. The aim of this study was to describe the plasma kinetics profile of DRM in rabbits over a 35-day period following subcutaneous administration at a dose of 400 lg ⁄ kg b.w. Six 5-month-old German white rabbits weighting 2.8 to 2.9 kg (2.89 ± 0.04 kg) were selected for this study. Animals were clinically healthy and parasite-free and had not been treated with doramectin for at least 2 months before the beginning of the experiment. During a 2-week acclimation period and subsequent experimental period, the rabbits were fed commercial food three times per day at 8:00 h, 12:00 h and 18:00 h. Water was available ad libitum. Rabbits were subcutaneously administrated 1% DRM oil injection (Zhejiang Haizheng Co., Zhejiang Provence, PRC) at a dosage of 400 lg per kg body weight in the jugular area. Heparinized blood samples (2 mL) were collected from the marginal ear vein immediately before and at 0.5, 1, 2, 4, 8, 12 and 24 h and at 2, 3, 4, 5, 6, 7, 9, 11, 14, 17, 20, 24 and 35 days after administration. Samples were centrifuged at 2000 g for 20 min and the recovered plasma was transferred to vacutainer tubes and frozen at )20 C until analysis. The sample extraction and cleanup procedures were performed following a little modification of the methods described by Zhang et al. (2007) and Hou et al. (2007), respectively. The extract was mixed with 0.5 mL de-ionized water before passing though the C18 cartridge and the ratio of acetonitrile-water which was prepared to wash the cartridge was 60:40 (v ⁄ v). The derivertization procedure was carried out following a little modification of a previous method described by Hou et al. (2007). The derivertization sample was diluted to 500 lL with methanol and filtered through a 0.2 lm filter. The sample was allowed to stablilize for 1 h before being analyzed by HPLC. The HPLC system includes a Waters 2695 separations module and a Waters 2475 fluorescence detector with an autosampler (Waters Co., Milford, MA, USA). The mobile phase was acetonitrile-water (92:8, v ⁄ v), and the flow rate was 1 mL ⁄ min. The chromatographic column was a reverse phase C18 column (Symmetry Shield RP C18 column, 4.6 mm i.d. · 250 mm, 5 lm, Waters Co., Ireland). The column temperature was set at 35 C. The fluorescence detector settings were an excitation wavelength of 365 nm and an emission wavelength of 475 nm. The injection volume was 50 lL. The calibration curve in the range of 0.15–50 ng ⁄ mL was linear with the correlation coefficient (r) of 0.9999. For all fortified samples at four concentration levels (0.3, 5, 20, 50 ng ⁄ mL), mean recoveries of the analyte were in the range of 91.5–97.7%, J. vet. Pharmacol. Therap. 32, 397–399, doi: 10.1111/j.1365-2885.2008.01044.x. SHORT COMMUNICATION

Determination of Tilmicosin by Fluorescence-Based Immunochromatography

ABSTRACT Tilmicosin has been widely used in the dairy industry, but this compound is hazardous to human health. Therefore, it is urgent to develop a rapid, convenient, sensitive, cost-effective, and user-friendly method to determine tilmicosin. A fluorescent microsphere-based immunochromatographic assay was developed for this analyte. A 200-nm red fluorescent microspheres were selected to be the label, and the mass of the labeled antibody was 4.48 µg/mL. 1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide was used to catalyze carboxyl groups on the surface of the microspheres. The optimum reaction time between the antibodies and fluorescent microspheres was 30 min, and under these conditions, high coupling ratios and stable conjugations were obtained. The linear dynamic range of the developed protocol was from 0.02 to 2.10 µg/L. The half maximal inhibitory concentration was 0.19 µg/L. The recoveries were from 87 to 98% with a coefficient of variation less than 7.14%. In comparison with a colloidal gold immunochromatographic assay, the developed method offered a higher coupling rate, lower amounts of antibodies, and higher sensitivity. This approach is simple, sensitive, and suitable for on-site applications on farms for rapid screening.