Quantitative determination of tetracyclines in medicated feed for food-producing animals by HPLC-DAD.

Tetracyclines are a group of antibiotic substances largely administered through medicated feed to control diseases in food-producing animals. Fine dosing of antibiotics contained in medicated feed is crucial for the success of the treatment as well as minimising potential threats such as the spread of antimicrobial resistance and the transfer of antibiotic residues in food. A rapid analytical method based on HPLC with diode array detection (HPLC-DAD) was developed to quantify oxytetracycline, chlortetracycline and doxycycline in medicated feed. The reported method underwent in-house validation and was found to be suitable for the quantification of three target tetracyclines within the concentration range of 40-1000 mg kg-1 in official routine analysis. The method was applied to 103 official samples in the framework of the Italian National Plan on animal feed during the years 2021-2023 and nine non-compliant concentrations were identified in swine and fish feed samples.

A New Method for Ivermectin Detection and Quantification through HPLC in Organic Matter (Feed, Soil, and Water).

Ivermectin is a macrocyclic lactone widely used in veterinary medicine for its broad-spectrum antiparasitic properties. It has been proven to be effective and safe. The purpose of this study was to develop a high-performance liquid chromatography method with a diode array detector for ivermectin screening in feed and water for animal consumption. Furthermore, the objective was to quantify ivermectin levels that were higher than 0.5 mg/kg in solid matrixes and 0.1 mg/kg in water. Doramectin was used as process standard. Samples were extracted using solid phase extraction with silica and C-18 columns. The method involved the use of high-performance liquid chromatography (HPLC) with a diode array detector (DAD). The results were interpreted using a calibration curve built with ivermectin standards at multiple concentrations (0.5, 1, 2, 5, and 12.5 mg/kg). Statistical evaluation of data was done using ANOVA. The data analysis showed that the linear regression was highly significant (P < 0.001), the intercept values were not significantly different from zero, and the correlation coefficient values (>0.999) indicated excellent linearity. Further tests demonstrated that this method is also useful when studying soil matrixes. The soil was dried and analyzed in the same way as feed; the same recoveries were realized on the spiked samples. The method is easy, inexpensive, precise, and repeatable; it requires very small amounts of sample.

Types of pesticides involved in domestic and wild animal poisoning in Italy.

Data obtained from samples of poisoned domestic and wild animals sent for toxicological evaluation during the period between 2005 and 2014 have been analyzed. Among the 4606 tested samples, the majority of which were collected in the northern regions of Italy, 2006 (43.55%) were found positive for pesticides. Analytical detections were performed via solvent extraction followed by separation and chromatographic characterization and all the methods applied for the toxicological investigations were developed by IZSLER. Insecticides, mainly represented by acetylcholinesterase inhibitors (carbamates 17.55%, n = 352; organophosphates 15.15%, n = 304) and organochlorines (29.21%, n = 586), were found to be the first category of pesticides involved in intoxications, in both domestic and in wild animals, followed by rodenticides (anticoagulant rodenticides 21.09%, n = 423; zinc phosphide 2.59%, n = 52; chloralose 0.95%, n = 19 and thallium 0.15%, n = 3) and molluscicides (metaldehyde 6.63%, n = 133). Second and third generation anticoagulants (bromadiolone and brodifacoum) were the most represented (10.52%, n = 211) but also first generation compounds (i.e. coumatetralyl and warfarin) were still found responsible of intoxications. Even if some pesticides are frequently involved in domestic animal poisoning (i.e. metaldehyde and strychnine), they did not show the same diffusion in wild animals. In particular, unlike domestic species, cyanide and pyrethroids have not been found responsible of intoxications in wild animals. Interestingly, a great number of positive samples involved banned pesticides like α- (14.41%, n = 289) and ß- (14.16%, n = 284) endosulfan, carbofuran (5.73%, n = 115), methamidophos (9.47%, n = 190), strychnine (6.23%, n = 125) but, on the other hand, many positives were due to the exposure to commercially available products (i.e. metaldehyde and anticoagulant rodenticides). Thus, together with measures aimed to reduce illegal uses, educational campaigns and a wider range of compounds to detect would be beneficial in order to address the issue of animal poisoning, which besides has also repercussions on environmental and public health.

Pesticide incidence in poisoned baits: A 10-year report.

The incidence of pesticides in poisoned baits recovered from 2005 to 2014 in Italian northern regions of Emilia Romagna and Lombardy was analysed. A total of 956 baits tested positive for pesticides during the study period. In 9.3% (n=89) of the baits analysed more than one toxic compound was present. Insecticides (53.2%) proved to be the pesticides most commonly involved followed by rodenticides (34.3%), molluscicides (12.3%) and herbicides (0.2%). Among insecticides, the anticholinesterases (organophosphates and carbamates) were the most frequently detected (59.1%, n=412) followed by organochlorines (39.8%; n=277). Anticoagulants proved to be the rodenticides most commonly involved (78.5%; n=353) with many baits containing brodifacoum or bromadiolone, while metaldehyde was the most used molluscicide (88.2%; n=142). A high number of baits contained banned pesticides such as endosulfan, methamidophos, carbofuran and strychnine indicating that the restrictions and the bans on the use of toxic and highly toxic pesticides are not accompanied by a lower use in poisoned baits. The high incidence of banned pesticides detected in the present survey is extremely worrying and poses the question of where and how these compounds were obtained. Efforts have to be made to reduce this illegal practice and its environmental impact. Moreover, education and awareness campaigns should be instituted.

Rapid determination of the antiparasitic drugs flubendazole and febantel in feeds by HPLC with ultraviolet detection.

A rapid and very effective analytical procedure for the simultaneous HPLC determination of two anthelmintics, Flubendazole (FLUB) and Febantel (FEBA), in swine and poultry feeds was developed and tested. The ground feed samples were extracted using dimethylformamide. The extracts were directly analyzed, without any purification, on a reversed-phase ODS column (150 mm x 4.6 mm, 5 microm) with acetonitrile-phosphate buffer (pH 3; 0.017 M) as mobile phase. Ultraviolet detection of FLUB and FEBA was carried out at 300 nm. The method was validated for specificity, linearity, accuracy, repeatability, limit of detection and limit of quantification. The limits of detection (LOD) of FLUB and FEBA in feeds, based on a detector signal-to-noise ratio of 3, were 3 mg kg(-1) and the lowest levels tested in feeds by this procedure (limit of quantification, LOQ) were 10 mg kg(-1). The mean recovery of FLUB and FEBA from spiked samples, in a concentration range of 10-200 mg kg(-1), was 98% with a CV% of 4% and 99% with a CV% of 2%, respectively.