Determination of eleven coccidiostats in animal feed by liquid chromatography-tandem mass spectrometry at cross contamination levels.

A confirmatory multi-residue method has been developed to allow for the detection, confirmation and quantification of eleven coccidiostats in animal feed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method can be used to determine halofuginone, robenidine, nicarbazin, diclazuril, decoquinate, semduramicin, lasalocid, monensin, salinomycin, narasin, maduramicin at levels relating to unavoidable carry over as stated in Regulation 2009/8/EC. Feed samples are extracted with water and acetonitrile with the addition of anhydrous magnesium sulphate and sodium chloride. The extract then undergoes a freezing out step before being diluted and injected onto the LC-MS/MS system. The LC-MS/MS system is run in MRM mode with both positive and negative electrospray ionisation and can confirm all eleven analytes in a run time of 19 min. The sensitivity of the method allows quantification and confirmation for all coccidiostats at a 0.5% carry over level. The method was validated over three days in accordance with of European legislation; Commission Decision 2002/657/EC. Validation criteria of accuracy, precision, decision limit (CCα), and detection capability (CCß) along with measurement uncertainty are calculated for all analytes. The method was then successfully used to analyse a number of feed samples that contained various coccidiostat substances.

Development and validation of a rapid multi-class method for the confirmation of fourteen prohibited medicinal additives in pig and poultry compound feed by liquid chromatography-tandem mass spectrometry.

A confirmatory method has been developed to allow for the analysis of fourteen prohibited medicinal additives in pig and poultry compound feed. These compounds are prohibited for use as feed additives although some are still authorised for use in medicated feed. Feed samples are extracted by acetonitrile with addition of sodium sulfate. The extracts undergo a hexane wash to aid with sample purification. The extracts are then evaporated to dryness and reconstituted in initial mobile phase. The samples undergo an ultracentrifugation step prior to injection onto the LC-MS/MS system and are analysed in a run time of 26 min. The LC-MS/MS system is run in MRM mode with both positive and negative electrospray ionisation. The method was validated over three days and is capable of quantitatively analysing for metronidazole, dimetridazole, ronidazole, ipronidazole, chloramphenicol, sulfamethazine, dinitolimide, ethopabate, carbadox and clopidol. The method is also capable of qualitatively analysing for sulfadiazine, tylosin, virginiamycin and avilamycin. A level of 100 microg kg(-1) was used for validation purposes and the method is capable of analysing to this level for all the compounds. Validation criteria of trueness, precision, repeatability and reproducibility along with measurement uncertainty are calculated for all analytes.

Automated extraction of acetylgestagens from kidney fat by matrix solid phase dispersion.

A new extraction method for the acetylgestagens medroxyprogesterone acetate (MPA), chloromadinone acetate and megestrol acetate, from kidney fat, has been developed. The method is a combination of matrix solid phase dispersion and solid phase extraction and is simpler and safer than previous methods, especially as it can be automated. The recovery was estimated as 59 +/- 5% (mean +/- standard deviation) for MPA. For screening purposes detection can be achieved using a commercially available enzyme immunoassay kit giving detection limits in the range of 1.0-2.0 ng g-1.

Novel approach to the 'on-site' testing for sulfamethazine in pork carcasses.

Rapid 'on-site' methods are required by the pork industry to screen for the presence of antibiotic residues in meat and meat products. There are few rapid and easy-to-use methods suitable for application in non-analytical laboratories. This paper describes a novel approach for the screening of sulfamethazine in pork muscle using matrix solid phase dispersion, a microcolumn preconcentration step and thin-layer chromatographic detection. The characteristics of the method are reported allowing for the detection of residues at the maximum residue limit of 100 ppb. Results from the industrial evaluation of the complete method are also presented.

Application of the matrix solid phase dispersion technique for the determination of ivermectin residues in fish muscle tissue.

A modified high-performance liquid chromatographic method has been developed for the determination of ivermectin (IVM) residues in fish muscle tissue. The extraction and clean-up procedure is based on the matrix solid phase dispersion technique. Control and IVM-fortified fish muscle samples (0.5 g) are blended with octadecyl (C18 end-capped) packing material. A column made from the C18-fish tissue blend is washed with hexane (8 ml) and the IVM is eluted with 8 ml of dichloromethane-ethyl acetate (3 + 1). Ivermectin is derivatized and analyzed by high-performance liquid chromatography (HPLC) with fluorescence detection. The recovery from fortified samples was greater than 80% in the concentration range 20-100 ng g-1 of tissue. The limit of determination was 10 ng g-1 of tissue. This method and another, using solvent extraction and clean-up of the extract on a solid phase extraction cartridge, gave comparable results for an incurred sample containing IVM residue. This method incorporates a rapid sample pre-treatment step which makes it an attractive and useful method for routine analysis of IVM in fish muscle tissue.

Comparison of matrix solid phase dispersion (MSPD) with a standard solvent extraction method for sulphamethazine in pork muscle using high performance liquid and thin layer chromatography.

A rapid and simple extraction/clean-up procedure (matrix solid phase dispersion, MSPD) for the determination of sulphamethazine (SMZ) in pork muscle tissue is compared with a solvent extraction method. Extracts of samples fortified with SMZ or of incurred samples were found to be free from interfering compounds when chromatographed using HPLC or TLC separation systems. Recovery of SMZ from fortified samples is greater than 80% and residue levels of incurred samples found using the MSPD procedure compare favourably with results obtained using the solvent extraction method. Use of aqueous back extraction of SMZ from dichloromethane is also reported as an alternative step to solvent evaporation for ease of use in both the laboratory and in industry (i.e. at slaughter plants).

Reversed-phase high-performance liquid chromatographic determination of dexamethasone in bovine tissues.

A sensitive and selective high-performance liquid chromatographic procedure is described for the determination of the synthetic corticosteroid dexamethasone (DXM), in bovine muscle, kidney, liver and fat tissues, using methylprednisolone as the internal standard. Following extraction with ethyl acetate (muscle, kidney and liver) or diethyl ether (fat) and clean-up of the tissue extract, the drug residue was isolated using a C18 solid-phase extraction column. Separation of DXM was achieved by reversed-phase high-performance liquid chromatography with ultraviolet detection at 254 nm. By using this procedure, DXM levels as low as 0.01 mg kg-1 can be detected in muscle, kidney, liver and fat.

High performance liquid chromatographic separation of cisplatin and its hydrolysis products on alumina and application to studies of their interaction with cysteine.

An alumina stationary phase has been assessed in the study of the retention behaviour of the anticancer drug cisplatin and its major hydrolysis products. Parameters such as buffer concentration in the mobile phase, pH, organic modifier and competing ion have been investigated in order to optimize chromatographic separation with ultraviolet detection. The separation scheme developed has been used to monitor the hydrolysis of cisplatin in aqueous and saline media, and to monitor the interaction of hydrolysed solutions of cisplatin with the amino acid cysteine. A new peak was observed in the chromatograms of such mixtures when they had been allowed to stand for periods of greater than 16 h and, from analysis of the data obtained, it was concluded that this new peak was due to a complex formed between the mono-aquo hydrolysis product of cisplatin and the amino acid.

Adsorptive voltammetric investigation of the interaction of cisplatin with cystine and human serum albumin.

The interaction of the anti-cancer drug cisplatin with human serum albumin and cystine has been investigated using differential pulse adsorptive voltammetry. Based on an understanding of the voltammetric behaviour of these biological molecules, which rely on the presence of the disulphide groups within their molecular structure for their electroactivity, it has been postulated that binding of cisplatin to these molecules occurs at the disulphide bond. A fractional coefficient for the binding of cisplatin to human serum albumin at pH 7.4 was calculated to be 0.32. The reactivity of hydrolysis products of cisplatin was shown to be greater than that of the parent drug.