Evaluation of a chromogenic assay to measure the factor Xa inhibitory activity of unfractionated heparin in canine plasma

BACKGROUND: Unfractionated heparin (UFH) has a complex pharmacologic profile that necessitates patient monitoring to prevent inadequate anticoagulation or overdosage and hemorrhage. Factor Xa inhibitory assays (to measure anti-Xa activity) are used to adjust UFH dosage and define safe and effective regimens for specific thrombotic disorders in humans. OBJECTIVE: In this study, the accuracy, linearity, and clinical utility of a chromogenic assay were assessed for monitoring UFH anti-Xa activity in canine plasma samples. METHODS: A commercial assay (Rotachrom Heparin, Diagnostica Stago, Parsippany, NJ, USA) was used to measure anti-Xa activity in canine plasma samples spiked with different concentrations of UFH. Background absorbance and assay linearity were compared for canine and human plasmas. Percentage recovery of UFH anti-Xa activity and intra- and interassay imprecisions were investigated by multiple measurements of canine plasma to which known amounts of UFH were added. The spiked plasma samples also were used to determine the heparin sensitivity of an activated partial thromboplastin time (aPTT) test. RESULTS: Canine plasma samples were assayed at a higher dilution than were human plasma samples (3:8 versus 4:8) to eliminate higher background anti-Xa activity in canine plasma. Using this modification, the recovery of anti-Xa activity in canine plasma was linear (R2 > .9) at concentrations of 0 - 0.75 U/mL UFH. Intra- and interassay imprecisions for plasma samples containing 0.5 U/mL UFH were <10%, whereas samples containing 0.25 U/mL UFH had imprecisions of 13% and 24%, respectively. The anti-Xa activity range of 0.5 - 0.75 U/mL caused prolongation of aPTTs to 1.5 - 2.5 times the assay mean. CONCLUSION: Plasma anti-Xa activity of dogs treated with UFH can be accurately monitored using this commercially available chromogenic assay.     

Pharmacokinetics of single‐dose sildenafil administered orally in clinically healthy dogs: Effect of feeding and dose proportionality

Basic information related to the pharmacokinetics of sildenafil in dogs is scarce. This study aimed to describe the pharmacokinetic properties of oral sildenafil and determine the effect of feeding and dose proportionality. The effect of feeding on pharmacokinetics of sildenafil (1 mg/kg) was investigated using a crossover study with six dogs. In addition, the dose proportionality of sildenafil ranging 1–4 mg/kg was evaluated using five dogs in the fasted states. The plasma concentrations of sildenafil were determined using high‐performance liquid chromatography, and pharmacokinetic parameters were calculated using a noncompartmental analysis. Sildenafil administrations were well tolerated in all studies. Feeding reduced the area under the curve extrapolated to infinity (AUC_inf) and the maximum plasma concentration (C_max) significantly. The elimination half‐life (T_1/2) did not differ between the fasted and the fed states. For dose proportionality, nonproportional increases in AUC_inf and C_max at 1–4 mg/kg doses were detected by a power model analysis.     

Pharmacokinetics and pharmacodynamics of propofol with or without 2% benzyl alcohol following a single induction dose administered intravenously in cats

ObjectiveTo compare the pharmacokinetics and pharmacodynamics of propofol with or without 2% benzyl alcohol administered intravenously (IV) as a single induction dose in cats.Study designProspective experimental study.AnimalsSix healthy adult cats, three female intact, three male castrated, weighing 4.8 ± 1.8 kg.MethodsCats received 8 mg kg⁻¹ IV of propofol (P) or propofol with 2% benzyl alcohol (P28) using a randomized crossover design. Venous blood samples were collected at predetermined time points to 24 hours after drug administration to determine drug plasma concentrations. Physiologic and behavioral variables were also recorded. Propofol and benzyl alcohol concentrations were determined using high pressure liquid chromatography with fluorescence detection. Pharmacokinetic parameters were described using a 2-compartment model. Pharmacokinetic and pharmacodynamic parameters were analyzed using repeated measures anova (p < 0.05).ResultsPlasma concentrations of benzyl alcohol were below the lower limits of quantification (LLOQ) at all time points for two of the six cats (33%), and by 30 minutes for the remaining four cats. Propofol pharmacokinetics, with or without 2% benzyl alcohol, were characterized by rapid distribution, a long elimination phase, and a large volume of distribution. No differences were noted between treatments with the exception of clearance from the second compartment (CLD2), which was 23.6 and 38.8 mL kg⁻¹ minute⁻¹ in the P and P28 treatments, respectively. Physiologic and behavioral variables were not different between treatments with the exception of heart rate at 4 hours post administration.Conclusions and clinical relevanceThe addition of 2% benzyl alcohol as a preservative minimally altered the pharmacokinetics and pharmacodynamics of propofol 1% emulsion when administered as a single IV bolus in this group of cats. These data support the cautious use of propofol with 2% benzyl alcohol for induction of anesthesia in healthy cats.     

In vitro heparinization of canine whole blood with low molecular weight heparin (dalteparin) significantly and dose‐dependently prolongs heparinase‐modified tissue factor‐activated thromboelastography parameters and prothrombinase‐induced clotting time

Background: Low‐molecular‐weight heparin (LMWH) is being used increasingly in veterinary medicine for both treatment and prophylaxis of thromboembolic disease, but no predictable patient‐side method exists to monitor its effect. Objectives: The aim of this study was to evaluate thromboelastography (TEG) and prothombinase‐induced clotting time (PiCT) assays for detecting hemostatic alterations following in vitro heparinization of canine whole blood with dalteparin (Fragmin). Methods: Citrated whole‐blood samples were collected from 7 clinically healthy dogs. Dalteparin was added at concentrations of 0, 0.156, 0.625, 1.25, and 2.5 U/mL of whole blood. TEG was performed using heparinase cups with tissue factor (TF, 1:50,000) and kaolin as activators. Reaction time (R), clotting time (K), angle (α), and maximum amplitude (MA) were recorded. PiCT and anti‐FXa activity were measured in plasma. Results: With TF, increasing concentrations of dalteparin significantly prolonged R and K and significantly decreased α and MA. K, α, and MA ratios were significantly different from baseline at all dalteparin concentrations and R was significantly different from baseline at concentrations of 0.625, 1.25, and 2.5 U/mL. With kaolin, only R was significantly different from baseline at dalteparin concentrations of 0.625 and 2.5 U/mL. PiCT detected dalteparin concentrations ≤ 0.625 U/mL, with a good linear correlation (r²=.96, P<.0001). Conclusion: These results suggest that TF‐activated TEG and PiCT assays should be further evaluated as promising new methods for evaluating the effect of LMWH, using doses in the recommended clinical range and prospective clinical studies.     

Pharmacokinetics and pharmacodynamics of alfaxalone after a single intramuscular or intravascular injection in mallard ducks (Anas platyrhynchos)

Pharmacokinetics and pharmacodynamics of alfaxalone was performed in mallard ducks (Anas platyrhynchos) after single bolus injections of 10 mg/kg administered intramuscularly (IM; n = 10) or intravenously (IV; n = 10), in a randomized cross‐over design with a washout period between doses. Mean (±SD) C_max following IM injection was 1.6 (±0.8) µg/ml with T_max at 15.0 (±10.5) min. Area under the curve (AUC) was 84.66 and 104.58 min*mg/ml following IV and IM administration, respectively. Volume of distribution (V_D) after IV dose was 3.0 L/kg. The mean plasma clearance after 10 mg/kg IV was 139.5 (±67.9) ml min^?1 kg^?1. Elimination half‐lives (mean [±SD]) were 15.0 and 16.1 (±3.0) min following IV and IM administration, respectively. Mean bioavailability at 10 mg/kg IM was 108.6%. None of the ducks achieved a sufficient anesthetic depth for invasive procedures, such as surgery, to be performed. Heart and respiratory rates measured after administration remained stable, but many ducks were hyperexcitable during recovery. Based on sedation levels and duration, alfaxalone administered at dosages of 10 mg/kg IV or IM in mallard ducks does not induce clinically acceptable anesthesia.     

Pharmacokinetics of marbofloxacin after a single oral dose to loggerhead sea turtles (Caretta caretta)

The single-dose disposition kinetics of marbofloxacin (MBX) were determined in clinically healthy loggerhead sea turtles (n = 5) after oral (PO) administration of 2 mg kg⁻¹ bodyweight. Marbofloxacin plasma concentrations were determined by DAD–HPLC (LOD/LOQ 0.015/0.05 μg ml⁻¹). Data were subjected to non-compartmental analysis. Following PO administration, marbofloxacin achieved maximum plasma concentrations of 11.66 ± 2.53 mg L⁻¹ at 15.00 ± 3.00 h. The absence of general adverse reactions in the turtles of the study, and the favourable pharmacokinetic properties (long half-life and high maximum plasma concentration) of MBX administered PO at the single-dose of 2 mg kg⁻¹ suggest the possibility of its safe and effective clinical use in loggerhead sea turtles.     

Cardiovascular effects of a proprietary l-methadone/fenpipramide combination (Polamivet) alone and in addition to acepromazine in healthy Beagle dogs

ObjectiveTo determine the cardiovascular effects of a proprietary l-methadone/fenpipramide combination (Polamivet) alone and in addition to acepromazine in dogs.Study designProspective, randomized, experimental crossover study.AnimalsFive adult healthy Beagle dogs (one male and four females, weighing 12.8–16.4 kg).MethodsDogs were instrumented for haemodynamic measurements whilst anaesthetized with isoflurane. Three hours after recovery dogs received 0.025 mg kg^?1 acepromazine (AP) or saline (SP) IM followed by 0.5 mg kg^?1L-methadone/ 0.025 mg kg^?1 fenpipramide IV after 30 minutes. Cardiac output using thermodilution, heart rate, mean arterial pressure (MAP), central venous pressure (CVP), mean pulmonary artery pressure (MPAP), pulmonary artery occlusion pressure (PAOP), haemoglobin concentration, arterial and mixed-venous blood gas analysis were measured and sedation evaluated at baseline (BL), 30 minutes after acepromazine or saline IM (A/S), 5 minutes after L-methadone/fenpipramide IV application (35), every 15 minutes for 1 hour (50, 65, 80, 95 minutes) and every hour until baseline cardiac output was regained. Standard cardiovascular parameters were calculated. Data were analyzed by repeated measures anova and paired t-tests with p < 0.05 considered significant.ResultsBaseline measurements did not differ. Cardiac index decreased after acepromazine administration in treatment AP (p = 0.027), but was not significantly influenced after l-methadone/fenpipramide injection in either treatment. In both treatments heart rate did not change significantly over time. Stroke volume index increased after A/S in both treatments (p = 0.049). Systemic vascular resistance index, MAP, CVP, MPAP, and pulmonary vascular resistance index did not change significantly after either treatment and did not differ between treatments. Dogs were deeply sedated in both treatments with a longer duration in treatment AP.Conclusions and clinical relevanceIn healthy dogs the dose of l-methadone/fenpipramide used in this study alone and in combination with acepromazine induced deep sedation without significant cardiovascular changes.     

Effects of 6% hetastarch (600/0.75) or lactated Ringer's solution on hemostatic variables and clinical bleeding in healthy dogs anesthetized for orthopedic surgery

ObjectiveTo evaluate and compare hemostatic variables and clinical bleeding following the administration of 6% hetastarch (600/0.75) or lactated Ringer’s solution (LRS) to dogs anesthetized for orthopedic surgery.Study designRandomized blinded prospective study.AnimalsFourteen, healthy adult mixed-breed hound dogs of either sex, aged 11–13 months, and weighing 20.8 ± 1.2 kg.MethodsThe dogs were randomly assigned to receive a 10 mL kg^?1 intravenous (IV) bolus of either 6% hetastarch (600/0.75) or LRS over 20 minutes followed by a maintenance infusion of LRS (10 mL kg^?1hour^?1) during anesthesia. Before (Baseline) and at 1 and 24 hours after bolus administration, packed cell volume (PCV), total protein concentration (TP), prothrombin time (PT), activated partial thromboplastin time (APTT), von Willebrand’s factor antigen concentration (vWF:Ag), factor VIII coagulant activity (F VIII:C), platelet count, platelet aggregation, colloid osmotic pressure (COP) and buccal mucosal bleeding time (BMBT) were measured. In addition a surgeon who was blinded to the treatments assessed bleeding from the incision site during the procedure and at 1 and 24 hours after the bolus administration.ResultsFollowing hetastarch or LRS administration, the PCV and TP decreased significantly 1-hour post-infusion. APTT did not change significantly compared to baseline in either treatment group, but the PT was significantly longer at 1-hour post-infusion than at 24 hours in both groups. No significant change was detected for vWF:Ag, FVIII:C, platelet aggregation or clinical bleeding in either group. The BMBT increased while platelet count decreased significantly at 1-hour post-infusion in both groups. The COP decreased significantly in both treatment groups 1-hour post-infusion but was significantly higher 1-hour post-infusion in the hetastarch group compared to the LRS group.Conclusions and clinical relevanceAt the doses administered, both hetastarch and LRS can alter hemostatic variables in healthy dogs. However, in these dogs undergoing orthopedic surgery, neither fluid was associated with increased clinical bleeding.     

A phase II study to evaluate the toxicity and efficacy of alternating CCNU and high‐dose vinblastine and prednisone (CVP) for treatment of dogs with high‐grade,metastatic or nonresectable mast cell tumours

Safety and efficacy of a protocol of alternating 1‐(2‐chloroethyl)‐3‐cyclohexyl‐1‐nitrosourea (CCNU; 70 mg m^?2) and vinblastine (3.5 mg m^?2), and prednisone (1–2 mg kg^?1; CVP) in dogs with mast cell tumours (MCT) were evaluated. A total of 17 dogs had nonresectable MCTs and 35 received CVP as adjunctive treatment to locoregional control of metastatic MCTs or grade III MCTs. Neutropenia with fever occurred in 8% of dogs after treatment with vinblastine and in 2% after treatment with CCNU. Persistent elevation of serum alanine transaminase, suggestive of hepatotoxicity, occurred in 9% of the dogs. Response rate in dogs with nonresectable MCTs was 65%; five achieved a complete response (median, 141 days) and six achieved a partial response (median, 66 days). Overall median progression‐free survival (PFS) time in dogs treated in the adjuvant setting was 489 days. Dogs with grade III MCTs had shorter PFS compared with dogs with metastatic grade II MCTs (190 days versus 954 days; P < 0.001). Phase III studies are needed to provide reliable information about the comparative efficacy of this protocol.