Long term improvement in the treatment of canine leishmaniosis using an antimony liposomal formulation.

Pharmacokinetic and clinical effectiveness of liposome-encapsulated N-methylglucamine antimoniate (LMA) was performed in dogs suffering from experimental leishmaniosis. LMA was compared with N-methylglucamine antimoniate (MGA), the same drug in its free form. Sb plasma concentrations for LMA were always higher than those for MGA. Mean residence time (MRT), half-life time (t(1/2)) and clearance (Cl) showed that Sb was eliminated slower after liposome administration. The high volume of distribution (Vd) obtained with LMA suggests that Sb could achieve therapeutic concentrations in parasite-infected tissues. Average plasma concentration at steady state (Css(ave)) shows that Sb body concentrations after LMA treatment (9.8 mg/kg Sb, each 24h) would be effective in Leishmania infantum canine infection. Comparing LMA with MGA in a 1-year follow-up we observed no relapses for LMA and total protein and gammaglobulin concentrations were within normal range, while for MGA both began to rise 3 months after treatment. Use of antimonial liposomal formulations may restore effectiveness to an existing drug and reduce toxicity.

Leishmania infantum-specific IgG, IgG1 and IgG2 antibody responses in healthy and ill dogs from endemic areas. Evolution in the course of infection and after treatment.

The expression of IgG, IgG1 and IgG2 specific antibodies for Leishmania infantum was studied in five groups of dogs in Catalonia (Spain): I, 99 asymptomatic dogs (infected and uninfected) from a highly endemic area for leishmaniosis; II, 139 untreated dogs with clinically patent leishmaniosis; III, 11 naturally infected asymptomatic dogs monitored for up to 5 years since they were found seropositive to Leishmania antigen and without treatment; IV, 25 naturally infected dogs with clinically patent leishmaniosis and treated with either meglumine antimoniate and allopurinol or allopurinol alone and V, six experimentally infected dogs, treated with meglumine antimoniate and controlled for 5 years. The levels (ELISA units) of IgG, IgG1 and IgG2 in asymptomatic dogs (group I) were very variable (24+/-33, 32+/-31 and 26+/-31, respectively), and, as expected, lower than in ill dogs (group II) (168+/-34, 84+/-71 and 172+/-31, respectively). In both groups, the correlation between IgG and IgG2 levels (r=0.95, P<0.001 in group I and r=0.63, P<0.001 in group II) was higher than between IgG and IgG1 levels (r=0.01, P>0.05 in group I and r=0.31, P<0.001 in group II). In group III, IgG and IgG2 expression increased during infection, while IgG1 expression remained the same. In dogs of group IV, IgG levels after 1 year of treatment decreased more in responsive (mean values, 163+/-42 before treatment (b.t.) and 100+/-36 after treatment (a.t.)) than in unresponsive dogs (158+/-29 b.t. and 124+/-51 a.t.), especially for IgG1 (94+/-89 b.t. and 20+/-21 a.t. in responsive dogs and 35+/-25 b.t. and 22+/-13 a.t. in unresponsive dogs) rather than for IgG2 (156+/-16 b.t. and 114+/-45 a.t. in responsive and 151+/-11 b.t. and 125+/-36 a.t. in unresponsive dogs). Similar results were observed in the evolution of experimentally infected animals after consecutive and specific treatments. Overall results show the great variation in Leishmania-specific IgG1 expression in asymptomatic and symptomatic dogs, their lack of correlation with that of IgG2 and chemotherapy is more effective in dogs with initially high expression of IgG1.

Prolongation of epidural anaesthesia in dogs with bupivacaine in a lipid emulsion.

An aqueous solution and a lipid emulsion of bupivacaine were administered epidurally in doses of 1.8 mg/kg to six beagle dogs following a randomised two-phase crossover design. The aqueous solution was absorbed rapidly and the mean (sd) peak venous plasma concentration of bupivacaine, 1.4 (0.4) microg/ml, was detected after five minutes. After administration of the lipid emulsion, the peak plasma concentration of bupivacaine, 0.6 (0.2) microg/ml, was detected after 30 minutes. The mean (sd) t1/2beta of the aqueous preparation was 149.1 (32.6) minutes, and of the lipid emulsion 119.2 (34.0) minutes. Both preparations had a similar bioavailability. The mean time to the onset of motor block after the administration of the aqueous solution, 2.3 (2.2) minutes, was significantly shorter (P=0.028) than after the administration of the lipid emulsion, 9.4 (1.9) minutes, and the duration of the motor block induced by the lipid emulsion, 217.6 (26.2) minutes, was significantly longer (P=0.043) than for the aqueous solution, 158 (48.8) minutes. During anaesthesia, the plasma concentrations of bupivacaine ranged between 1.3 and 0.2 microg/ml. Non-significant changes in systolic blood pressure and heart rate were observed which coincided with the peak plasma concentrations of bupivacaine.

Pharmacokinetics of indomethacin in poultry.

Indomethacin (INDO) is a nonsteroidal antiinflammatory drug widely used since the 1970s. The pharmacokinetic behavior of INDO (2 mg/kg) has been studied after intravenous (i.v.) and oral administration to broiler chickens. After i.v. administration, a first fast distribution phase and a later and slower elimination phase were observed. The elimination half-life and mean residence time (MRT) obtained were 1.0 hr and 0.8 hr, respectively. After oral administration, a flip-flop phenomenon was observed giving an elimination half-life and MRT approximately three times and six times higher, respectively, than the i.v. administration. The plasma concentrations after oral administration were sustained during 8-10 hr, giving an antinflammatory cover over the dose producing 50% of maximal effect during this time period.

Serological and parasitological follow-up in dogs experimentally infected with Leishmania infantum and treated with meglumine antimoniate.

Six healthy beagle dogs were infected with Leishmania infantum (MCAN/ES/92/BCN-83/MON-1) by intravenous inoculation of 5 x 10(7) promastigotes and two others were used as controls. When animals showed clinical signs of disease at 29, 37, 41 and 45 weeks post-infection (p.i.), they were treated with meglumine antimoniate (20.4 mg Sb/kg/12 h) subcutaneously for two periods of 10 days each. Sera were tested periodically for Leishmania antibodies by Dot-ELISA, ELISA and Western blot (WB). Aspirates of popliteal lymph node (PLN), peripheral blood sample (PB) and healthy skin were cultured in NNN and Schneider's medium. PLNs were positive between 8 and 20 weeks p.i. and in one animal PB was positive 6 weeks p.i. Samples of healthy skin, obtained before treatment, were also positive. Dot-ELISA and ELISA detected specific antibodies at an early stage between 4 and 12 weeks p.i and surpassed the cut-off between 16-24 weeks p.i., while the WB was positive between 10-19 weeks p.i. The pattern of bands revealed during the first stages of infection was variable and only in two cases did the positivity start with bands of low molecular weight (12-14 kD); the number of bands increased until 15-24 weeks p.i., after which sera revealed a complete pattern of bands, from 12 to 85 kD, in the antigen of Leishmania. After treatment the clinical improvement of the animals was accompanied by a decrease in antibody titers (Dot-ELISA and ELISA) although the parasites remained in the PLN. This was reflected in the WB by a decrease in the intensity of bands, especially those in the region of 12-30 kD. A new increase in the antibody levels between 3 and 5 months after terminating the therapy was detected in the WB by a restoration of the initial complete pattern of bands.

Pharmacokinetics and pharmacologic effects of the S(-) isomer of bupivacaine after intravenous and epidural administration in dogs.

OBJECTIVE: To determine pharmacokinetic variables and pharmacologic effects of the S(-) isomer of bupivacaine (S[-]-BPV) in dogs. ANIMALS: 6 adult male Beagles. PROCEDURE: Dogs received S(-)-BPV (1 mg/kg of body weight) i.v., and 15 days later, the same dogs received 1.8 mg/kg epidurally. Pharmacokinetic variables and pharmacologic effects were determined for each route of administration. RESULTS: After i.v. administration, plasma concentration versus time curves were adjusted, using biexponential equations that indicated a rapid distribution phase followed by a slower elimination phase, with a mean +/- SD half-life of 33.5 +/- 17.0 minutes. Mean plasma clearance was 21.0 +/- 10.7 ml/min/kg, and mean volume of distribution at steady state was 0.8 +/- 0.2 L/kg. After i.v. administration, mean peak plasma concentration was 2.6 +/- 0.7 micrograms/ml; after epidural administration, it was 0.9 +/- 0.5 microgram/ml at approximately 3 minutes. Half-life after epidural administration was 5 times longer than that observed after i.v. administration. Motor block began immediately after the end of epidural administration and lasted for 3 to 4 hours. Changes in systolic blood pressure and heart rate after epidural administration were slight but occurred at the same time that plasma concentration peaked. After i.v. administration, motor block or variations in physiologic variables studied were not observed. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs, the pharmacologic behavior of S(-)-BPV was similar to that of the bupivacaine racemate, but motor block attributable to S(-)-BPV lasted longer than that attributable to the racemate, with lower plasma concentrations observed at equivalent sample collection times.

Sex differences in the disposition of albendazole metabolites in sheep.

Sex differences in the disposition of albendazole metabolites in sheep after oral administration of 20 mg/kg of netobimin have been studied. Some kinetic parameters of both metabolites show statistical differences between sexes; the sulphoxide and sulphone t1/2beta and MRT were lower in male animals than in females. Peak concentrations and AUC of sulphone metabolites were higher in males suggesting a greater oxidation rate compared with females. Urine excretion of albendazole metabolites, sulphoxide, sulphone, and amino sulphone appeared to be greater in female sheep than in males, mainly the sulphoxide metabolite. These differences between sexes can be caused by male sexual hormones, because testosterone and progesterone can induce or inhibit the microsomal Cytochrome P450 metabolism. Plasma protein-binding of albendazole sulphoxide and albendazole sulphone has been studied between male and female sheep, also their binding to sheep albumin and globulins. Both albendazole metabolites readily bind to sheep albumin and globulins. Male animals show a significantly lower binding of albendazole metabolites than females. These differences could be responsible for the non-esterified fatty acids (NEFA) present in the plasma. Males have significantly higher plasma levels of NEFA than females and which may compete with albumin for binding to albendazole metabolites.

Study of haemostatic disorders in experimentally induced leishmaniasis in Beagle dogs.

Haemostatic alterations in dogs experimentally infected with Leishmania infantum were studied before and after therapy with meglumine antimonate. Haemostatic function tests including platelet count, collagen-induced platelet aggregation, prothrombin time, activated partial thromboplastin time, thrombin time, plasma fibrinogen determination, and serum fibrinogen/fibrin degradation products concentration were performed. In the course of infection and before treatment, moderate thrombocytopenia (P<0.00001), decreased collagen induced platelet aggregation (P=0.0003), prolonged thrombin time (P=0.0117) and increased fibrinogen/fibrin degradation products were observed. Statistically significant differences of plasma fibrinogen concentration, prothrombin time, and activated partial thromboplastin time were not encountered. Haemostatic parameters returned to normal values after therapy. The results indicate that Leishmania infection may impair haemostasis suggesting induction of disseminated intravascular coagulation (DIC), and that treating dogs in an early stage of infection may potentially avoid the possibility of developing an uncompensated DIC.

Determination of indomethacin residues in poultry by high-performance liquid chromatography.

A HPLC method using a C18 column and UV detection (254 nm) is described for the determination of indomethacin residues in chicken tissues (liver, muscle and fat). Drug extraction from tissue homogenate in phosphate buffer (pH 3.5) was performed with dichloromethane. Mobile phase was acetonitrile-acetic acid (0.5% in water) (50:50). Indomethacin detection limit was 20 ng/g for the studied tissues. After administration of an oral dose of indomethacin (2 mg/kg), only three of the eight poultry studied showed drug tissue levels, in those cases the levels were below 50 ng/g.

Pharmacokinetics of meglumine antimoniate after administration of a multiple dose in dogs experimentally infected with Leishmania infantum.

Pharmacokinetics of meglumine antimoniate in dogs with experimentally induced leishmaniosis has been investigated. After infection, dogs received a dose of 75 mg kg-1 of meglumine antimoniate twice daily by subcutaneous injection for 10 days. Blood samples were collected throughout the treatment. No statistical differences were found in the kinetic behaviour of the drug administered as a single dose to healthy dogs and that administered as a multiple dose to infected animals. However, peak plasma concentrations (Cmax) of 30.8 +/- 12.8 micrograms ml-1 found after this dosage regimen were higher than those observed after the single dose administration of 100 mg kg-1 24 h-1. Furthermore, sustained antimony concentrations of 1.14 +/- 0.52 micrograms Sb ml-1 were detected throughout the treatment. No signs of toxicity were found in the animals treated indicating that this regimen would be very appropriate to treat canine leishmaniosis.

Pharmacokinetics of liposome-encapsulated meglumine antimonate after intramuscular and subcutaneous administration in dogs.

Controlling canine leishmaniasis may reduce the incidence of human leishmaniasis, which affect immunocompromised persons, especially those with human immunodeficiency virus infection. Thus, the pharmacokinetics of liposome-encapsulated meglumine antimonate (LMA) in dogs was studied after intramuscular (I.M.) and subcutaneous (S.C.) administration. Serum concentration-time data for both forms of administration were best described by a triexponential open model. The absorption phase showed statistically significant differences between I.M. and S.C. administrations (K01(I.M.) = 0.046/min, K01(S.C.) = 0.025/min). The first phase of decrease of plasma concentrations showed a longer half-life for S.C. than for I.M. administration, with the delay being caused by the slow absorption process after S.C. injection. Mean terminal phase half-lives after administration of I.M. and S.C. were 904.1 min and 637.4 min, respectively. Peak plasma concentrations after administration of I.M. (Cmax = 43.8 microg/ml) and S.C. (Cmax = 24.9 microg/ml) were detected at 42.8 min and 79.8 min, respectively. Urinary excretion of antimony for both routes surpassed 80% during the first 6 hr, with the rest of the drug being excreted slowly over the following 18 hr. The results obtained with this formulation suggest that for treating canine leishmaniasis, it would be more advisable to inject LMA intramuscularly if we assume that the significantly higher Cmax observed after I.M. administration is more relevant to dog's clinical outcome than is maintenance of concentrations over longer periods.

Disposition of netobimin, albendazole, and its metabolites in the pregnant rat: developmental toxicity.

Netobimin (NTB), a benzimidazole prodrug with a good anthelmintic spectrum, was administered orally to female rats at a dose of 59.5 mg NTB/kg, to study its pharmacokinetic behavior and the disposition of its most important metabolites, albendazole (ABZ), albendazole sulfoxide (ABZSO), and albendazole sulfone (ABZSO2). ABZ was found in plasma after 6 hr. Peak plasma concentrations (Cmax) and areas under curves (AUC) of ABZSO were eight- and fourfold higher, respectively, than those of ABZSO2. To study NTB disposition in pregnant rats, three different drug doses (50, 59.5, and 70.7 mg/kg) were given. No significant differences were found between plasma concentrations for each metabolite at the three doses studied. Only ABZ concentrations rose slightly as dose increased. ABZ, ABZSO, and ABZSO2 were found in amniotic sacs and embryos at concentrations that were higher than plasma at the same times. The fetuses obtained after administration of each of the doses of NTB were studied to detect developmental toxicity. A significant correlation was found between rate of developmental toxicity and metabolite concentration. ABZSO embryo concentrations could be the main factor accounting for toxicity.

Thrombocytopathia and light-chain proteinuria in a dog naturally infected with Ehrlichia canis.

A 6-year-old dog was presented for evaluation of recurrent epistaxis. Platelet counts, biochemical tests, and coagulation tests were within the normal range, but a mucosal bleeding time was prolonged; there was hyperproteinemia and a monoclonal gammopathy. Heterogeneity of light chains appeared in urine, however, thus suggesting that the gammopathy was polyclonal. Platelet aggregation tests showed decreased responsiveness to collagen. An Ehrlichia canis indirect fluorescent-antibody titer was positive (1:40). Treatment with tetracycline, melphalan, and prednisone resulted in a rapid clinical improvement that persisted for at least 3 years.

Disposition of antimony after the administration of N-methylglucamine antimoniate to dogs.

A study was carried out in dogs to define the pharmacokinetic profile of antimony and to define a better therapeutic protocol for the treatment of canine leishmaniasis. Six healthy beagle dogs received 100 mg/kg of N-methylglucamine antimoniate containing 27.2 per cent of antimony intravenously, intramuscularly and subcutaneously. After intravenous administration the plasma concentration of antimony decreased rapidly and after 240 minutes it was lower than the ED50 values suggested for Leishmania donovani. The pharmacokinetic parameters and bioavailability of antimony were calculated after each route of administration in each dog. The curves of plasma concentrations vs time were best described by a triexponential open model with a mean (sd) half life t1/2 alpha of 9.4 (4.4) min, a t1/2 beta of 45.3 (4.5) min and a t1/2 gamma of 618.0 (93.5) min. The mean volume of distribution at steady state was 0.25 (0.03) litres/kg and the total body clearance was 0.25 (0.04) litres/h/kg. The peak plasma concentration (Cmax) after intramuscular administration was 27.2 (3.1) micrograms/ml, and after subcutaneous administration it was 25.5 (4.5) micrograms/ml; they were reached after 73.6 (11.9) min and 85.6 (11.3) min, respectively. The bioavailabilities after intramuscular and subcutaneous administration were 91.7 (7.1) and 92.2 (7.1) per cent, respectively. More than 80 per cent of the antimony was excreted in the urine in the first nine hours.