J. Vet. Med. A 50, 297–302 (2003) 2003 Blackwell Verlag, Berlin ISSN 0931–184X
CLINICAL MEDICINE
Laboratorio de Farmacologı´ a, Facultad Medicina Veterinaria, Universidad de Concepcio´n, Chilla´n, Chile
Plasma Profiles of Ivermectin in Horses following Oral or Intramuscular Administration R. Pe´rez1,5, C. Godoy1, C. Palma1, I. Cabezas1, L. Mun˜oz4, L. Rubilar1, M. Arboix2 and M. Alvinerie3 Addresses of authors: 1Facultad de Medicina Veterinaria, Universidad de Concepcio´n, Chilla´n, Chile; 2Laboratorio de Farmacologı´ a, Facultad de Veterinaria, Universidad Autonoma de Barcelona, Bellaterra, Barcelona, Espan˜a; 3Laboratoire de Pharmacologie–Toxicologie INRA, 180 Chemin de Tournefeuille, 31931 Toulouse, France; 4Regimiento de Caballeria N7 Guias, Concepcion, Chile; 5Corresponding author: E-mail:
[email protected] With 3 figures and 1 table
Received for publication: January 24, 2002
Summary A study was undertaken in order to evaluate and compare ivermectin’s (IVM) plasma disposition kinetic parameters after oral or intramuscular (IM) administration in horses. Ten clinically healthy adult horses, weighing 380–496 kg body weight (BW), were allocated to two experimental groups of five horses. Group I, was treated with an oral paste formulation of IVM at the manufacturer’s recommended dose of 0.2 mg/kg BW. Group II, was treated IM with an injectable 1% formulation of IVM at a dose of 0.2 mg/kg BW. Blood samples were collected by jugular puncture at different times between 0.5 h and 75 days post-treatment. After plasma extraction and derivatization, samples were analysed by high-performance liquid chromatography with fluorescence detection. A computerized kinetic analysis was performed, and data were compared using the Wilcoxon signed rank test. The parent molecule was detected in plasma between 30 min and either 20 (oral) or 40 (IM) days post-treatment. Significant differences were found for the time corresponding to peak plasma concentrations (tmax) and for absorption half-life. Peak plasma concentrations (Cmax) of 51.3 ± 16.1 ng/ml (mean ± SD) were obtained after oral administration and of 31.4 ± 6.0 ng/ml for the IM route. The values for area under concentration–time curve were 137.1 ± 35.9 ng day/ml for the group treated orally, and 303.2 ± 4.3 ng day/ml for the IM treated group. The mean plasma residence times were 4.2 ± 0.4 and 8.9 ± 0.7 days for oral and IM-treated groups, respectively. The results of this study show that the route of administration considerably affects the disposition of IVM. A significant difference in bioavailabilty and half-life of elimination of IVM was observed after IM administration compared with oral administration. A close relationship between pharmacokinetic profiles and the clinical efficacy of IVM was established.
Introduction Prevention of equine disease associated with the gastrointestinal parasite nematodes is largely dependent on the regular use of anthelmintics to suppress faecal egg output over the U.S. Copyright Clearance Center Code Statement:
grazing season. If correctly managed, this strategy maintains pasture infectivity within tolerable limits. The overuse of anthelmintics, however, is undesirable, as this may increase the selection pressure for resistance. Definition of the optimum interval between treatments depends on an assessment of risk factors including pasture type, grazing history, stocking density and weather. Another important consideration is the protection period afforded by the chosen anthelmintic (Jacobs et al., 1995). The availability of ivermectin (IVM) has had a major impact on the control of equine parasites. Numerous critical and controlled studies have reported its efficacy against a broad spectrum of parasites, including tissue migratory stages of Parascaris equorum, Strongylus sp., spirurid stomach worms and microfilaries of Onchocerca cervicalis (Campbell et al., 1989; Klei et al., 1993; Conder and Campbell, 1995). The initial introduction of IVM to the world market was as a sterile solution for intramuscular (IM) administration to horses (Campbell and Benz, 1984). However, because of reported septic lesions at the injection site and clostridial myositis, the parenteral formulation was withdrawn after 17 months from the US market (Pulliam and Preston, 1989). Clinical efficacy of anti-parasitic drugs is closely related to pharmacokinetic behaviour. Anthelmintic activity not only depends on interaction of the active drug ingredient to a specific receptor in the parasite target, but also on the achievement of high and sustained concentrations at the parasitic site (Lanusse and Prichand, 1993; Baggot and McKellar, 1994). Characterization of the plasma bioavailability and disposition kinetics of IVM can be useful to predict changes that could optimize its anthelmintic efficacy (Baggot and Mckellar, 1994). Factors such as animal species, dosage formulation and route of administration may substantially affect the plasma kinetics and drug concentration profile attained at the site of parasitic infection (Lanusse et al., 1997). Studies with IVM, have shown that the pharmacokinetics and hence efficacy of this compound can be substantially modified by the formulation and the route of administration (Lo et al., 1985). The
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R. Pe´rez et al.
298 study reported here was designed to compare the pharmacokinetics of IVM in horses following either oral or IM administration and to establish the relative bioequivalence for the two routes of administration. The complex connections between route of administration, formulation, physicochemical properties of the drug and the resultant pharmacokinetic behaviour of IVM need to be fully understood to optimize parasite control and minimize selection for drug resistance (Alvinerie et al., 2000).
Materials and Methods
Parasitological procedures Quantitative pre- and post-treatment FEC were performed using a modified McMaster technique (Zajac, 1994) on days )1, 0, 1, 3, 4, 10, 20, 30, 40, 50, 60, 75, 90, 105 and 120. All fecal samples were obtained per rectum or from freshly voided feces. The egg reappearance period (ERP) was measured in both groups of horses treated with IVM. For the purpose of this study, the ERP was defined in agreement with Jacobs et al. (1995), as Ôthe interval between pre-treatment and the time when more than half of the horses in the group had egg counts ‡200 eggs per gram (EPG)Õ. For comparative purposes FECs were transformed to ln (x + 1).
Animals Ten crossbred adult saddle horses, clinically healthy, weighing 380–490 kg body weight (BW) were selected for this study. The trial was conducted from May to September of 2000, on horses belonging to a cavalry regiment of the Chilean army stationed in Concepcion (Chile). During the experimental period, the horses were kept stabled in individual stalls during the night and subjected to exercise during the day. They were fed daily with a mix of ryegrass and clover hay and 3 kg of oats. Feed was supplied three times per day at 8:00, 12:00 AM and 18:00 PM. The day before treatment the ration supply corresponding to 18:00 PM, was withdrawn and the horses were fasted for a period of at least 14 h before drug administration. After treatment the animals were fed normally according to their established feeding time. All horses were weighed before the treatments by means of a digital scale (Ruddweig Australasia Pty Ltd, Guyra New South Wales). This study was approved by the Animal Care and Use Committee of the Faculty of Veterinary Medicine, University of Concepcio´n, Chile. Treatments Faecal exams were performed on all horses to determine parasite faecal egg counts (FEC). Following these determinations horses were allocated to groups of five horses to provide a random distribution with respect to sex, BW and FEC. Group I, was treated with an oral paste formulation of IVM (IVM 1.87% oral paste, Eqvalan; Merck, Sharp & Dome Agvet, Rahway, NJ, USA) at the recommended therapeutic dose of 0.2 mg/kg BW. Group II, was treated with an injectable formulation of IVM (IVM 1.0% injectable, Ivomec; Merck, Sharp & Dome B. V., Haarlem, the Netherlands), administered in the musculature of the neck, at the dose level of 0.2 mg/kg BW (Di Pietro et al., 1982). The animals were dosed early in the morning before they received their daily ration of feed. Sampling Heparinized blood samples were collected from the jugular vein prior to the treatments and at 0.5, 1.0, 2.0, 4.0, 8.0, 12.0 and 24.0 h and 1.5, 2.5, 3.0, 4.0, 6.0, 8.0, 12.0, 15.0, 20.0, 25.0, 30.0, 40.0, 50.0, 60.0 and 75.0 days post-treatment. Plasma was separated by centrifugation and stored at )18C until analysis. After the treatments, the animals were observed continuously for a 4-h period and at least twice daily within 2 days of treatment for any sign of adverse reaction.
Analytical procedures Ivermectin was assayed by high-performance liquid chromatography (HPLC) with fluorescence detection after solid-phase extraction, according to procedures previously described by De Montigny et al. (1990). Drug extraction and derivatization Drug-free plasma samples (1 ml) were fortified with IVM to reach the following final concentrations of 0.1, 0.5, 1.0, 5.0, 10.0, 25.0 and 50.0 ng/ml. Fortified and experimental plasma samples were homogenized and solid-phase extraction was performed after 15 min of incubation at room temperature. Briefly, 1 ml of acetonitrile and 0.25 ml of water were added to 1 ml of plasma. After mixing for 20 min, the samples were centrifuged at 2000 g for 5 min, and the supernatant transferred to a Supelco C18 cartridge (Supelco Inc., Bellefonte, PA, USA). After washing with water, IVM was eluted with 1.0 ml MeOH. The eluate was evaporated to dryness under a gentle stream of nitrogen, and the residue dissolved in 100 ll of N-methylimidazole solution in acetonitrile (1 : 2 v/v). To initiate the derivatization, 150 ll of trifluoroacetic anhydride solution in acetonitrile (1 : 2 v/v) was added. After completion of the reaction (
