Quantitative chromatographic determination of several benzimidazole anthelmintic molecules in parasite material.

A specific, precise and accurate high-performance liquid chromatographic (HPLC) analytical method has been developed for the quantitative determination of different benzimidazole (BZD) anthelmintics in parasite material (Moniezia benedeni). Mebendazole (MBZ), oxibendazole (OBZ), flubendazole (FLBZ), albendazole (ABZ) ricobendazole (RBZ), albendazole sulphone (ABZSO(2)), fenbendazole (FBZ), oxfendazole (OFZ) and fenbendazole sulphone (FBZSO(2)) were measured simultaneously in M. benedeni, a sheep and cattle cestode parasite used as a model of the biological matrix. The recovery, linearity, precision, accuracy, limits of detection and quantification of the method were determined. Drug extraction from the parasite's tissue homogenate was performed using methanol (liquid phase extraction), and after solvent evaporation, the residual material was cleaned up by solid phase extraction prior to analysis by reversed-phase HPLC. The resolution of all the BZD molecules assayed was obtained on a C(18) reversed-phase (5 microm, 250 mm x 4.6 mm) column using acetonitrile and ammonium acetate as the mobile phase and ultraviolet (UV) detection at 292 nm. Regression analyses for all the BZD compounds assayed were linear at concentrations ranging from 1.61 to 64.21 nmol/100mg protein (triplicate determinations) showing correlation coefficients greater than 0.9922. The developed method is efficient for the simultaneous determination of several benzimidazole anthelmintic molecules in parasite material and useful for the ex vivo and in vivo characterisation of the kinetics of drug uptake/diffusion in target parasites, which seems to be relevant to optimise parasite control both in human and veterinary medicine.

Disposition of doramectin milk residues in lactating dairy sheep.

Doramectin (DRM) is a broad spectrum macrocyclic lactone antiparasitic drug not approved for use in dairy animals. However, DRM and other endectocide compounds are widely used extra-label to control endo- and ectoparasites in dairy sheep. The plasma disposition kinetics and the pattern of DRM excretion in milk were characterized following its subcutaneous administration to lactating dairy sheep. DRM concentration profiles were measured in plasma and milk samples after validation of a specific HPLC-based methodology. DRM was detected between 1 h and 30 days post-treatment. DRM concentrations of 0.48 ng.mL(-1) (plasma) and 1.03 ng.mL(-1) (milk) were measured at 30 days post-treatment. DRM was extensively distributed from the bloodstream to the mammary gland, and large concentrations were excreted in milk. The peak concentrations and total amount of DRM recovered in milk (expressed as area under the concentration versus time curve) were 3-fold higher than those measured in plasma; 2.44% of the total DRM dose was excreted in milk. The long persistence of DRM milk residues should be seriously considered before its extra-label use in dairy animals is recommended.

Measurement of triclabendazole and its metabolites in liver flukes: method development and full validation.

The trematode parasite Fasciola hepatica is still the cause of considerable loss in livestock production all over the world. The knowledge of the pharmacological properties of the available flukicidal drugs is critical to control this parasite. Triclabendazole is an halogenated benzimidazole anthelmintic with potent flukicidal activity. A simple reversed-phase high-performance liquid chromatographic analytical method has been developed, validated and applied for the quantitative determination of the flukicidal drug triclabendazole and its sulpho- and hydroxy-metabolites in the liver fluke, F. hepatica. Triclabendazole, triclabendazole sulphoxide, triclabendazole sulphone, hydroxy-triclabendazole, hydroxy-triclabendazole sulphoxide and hydroxy-triclabendazole sulphone were measured simultaneously in this trematode parasite. Linearity, resolution, precision, accuracy, recovery, limits of detection and quantification of the method were determined. Drug extraction from the parasite's tissue homogenate was effectively performed using liquid extraction (acetonitrile), avoiding clean up by solid phase extraction, prior to analysis by reversed-phase high-performance liquid chromatography. The resolution of all the halogenated benzimidazole thiol molecules assayed was obtained on a C(18) reversed-phase (5 microm, 250 mm x 4.6 mm) column using acetonitrile and ammonium acetate as the mobile phase and ultraviolet detection at 300 nm. Regression analyses were linear over the concentration range examined (from 0.272 to 16.331 nmol/100 mg trematode protein) and the correlation coefficients of the calibration curves ranged between 0.996 and 1.000. The calculated limits of detection of the proposed method for the parent drug and its metabolites ranged between 0.007 and 0.079 nmol/100 mg trematode protein. The extraction efficiency for the different analytes from the parasite material was greater than 71%. The results obtained indicated that the developed chromatographic method was selective, accurate and easy to reproduce. The developed procedure was successfully applied to quantify triclabendazole/metabolites in F. hepatica incubated under ex vivo conditions, demonstrating to be efficient for the determination of the most extensively used flukicidal drug available for veterinary medicine, and its metabolites. The analytical method described here is an useful tool for the measurement of this flukicidal compound in different studies addresses to evaluate drug influx/eflux and metabolism in its main target parasite.

Vector-borne diseases--constant challenge for practicing veterinarians: recommendations from the CVBD World Forum.

The human-animal bond has been a fundamental feature of mankind's history for millennia. The first, and strongest of these, man's relationship with the dog, is believed to pre-date even agriculture, going back as far as 30,000 years. It remains at least as powerful today. Fed by the changing nature of the interactions between people and their dogs worldwide and the increasing tendency towards close domesticity, the health of dogs has never played a more important role in family life. Thanks to developments in scientific understanding and diagnostic techniques, as well as changing priorities of pet owners, veterinarians are now able, and indeed expected, to play a fundamental role in the prevention and treatment of canine disease, including canine vector-borne diseases (CVBDs).The CVBDs represent a varied and complex group of diseases, including anaplasmosis, babesiosis, bartonellosis, borreliosis, dirofilariosis, ehrlichiosis, leishmaniosis, rickettsiosis and thelaziosis, with new syndromes being uncovered every year. Many of these diseases can cause serious, even life-threatening clinical conditions in dogs, with a number having zoonotic potential, affecting the human population.Today, CVBDs pose a growing global threat as they continue their spread far from their traditional geographical and temporal restraints as a result of changes in both climatic conditions and pet dog travel patterns, exposing new populations to previously unknown infectious agents and posing unprecedented challenges to veterinarians.In response to this growing threat, the CVBD World Forum, a multidisciplinary group of experts in CVBDs from around the world which meets on an annual basis, gathered in Nice (France) in 2011 to share the latest research on CVBDs and discuss the best approaches to managing these diseases around the world.As a result of these discussions, we, the members of the CVBD Forum have developed the following recommendations to veterinarians for the management of CVBDs.