Pharmacokinetics of hemoglobin-based oxygen carrier hemoglobin-glutamer-200 bovine (oxyglobin) in the horse

Oxyglobin (OXY) is a hemoglobin‐based oxygen carrier (HBOC) made of glutaraldehyde‐polymerized bovine hemoglobin (bHb). Products similar to OXY are under development for use as temporary blood substitutes in trauma, shock and anemia. Since they all may increase blood O₂‐carrying capacity and thus, possibly tissue oxygenation, they may also be used to enhance performance of both equine and human athletes. That is why HBOCs are banned from use in athletic competition. Our goal was to determine the pharmacokinetics of OXY after intravenous (IV) infusion to horses. Blood and urine samples were collected from adult horses that received an IV dose of 32.5 g of OXY. Concentrations of OXY in plasma and urine were quantified using a newly developed LC/Q‐TOF‐MS/MS detection technique. Level of quantification (LOQ) was 50 μg mL^–1. The decline of the plasma concentration‐time curve of the HBOC was described by a 2‐compartment model (C1 and C2). The median distribution alpha (t1/2k1,0) and elimination beta (t1/2k2,0) half‐lives were 1.3 and 12.0 hours, respectively. The bHb molecules in OXY are not of uniform size and vary substantially in molecular weight (MW). Of the OXY molecules 53% were eliminated in C1, which represented the smaller MW molecules and 47% in C2, which represented the larger MW bHb. The maximal 0‐time plasma concentration was 662.0 μg/mL and declined to 97.1 μg mL^–1 at 24 h. The area below the plasma concentration‐time curve was 5143 μg h^–1 mL^–1. The volumes of C₁ and C₂ were 86.9 and 63.9 mL kg^–1, respectively. Oxyglobin was not detected in urine. This study shows the detection and quantification in equine plasma of a HBOC following IV infusion and demonstrates the short half‐life of about 50% of infused bHb molecules.     

SCINTIGRAPHIC EVALUATION OF THE DISTAL TARSAL REGION IN HORSES WITH DISTAL TARSAL PAIN

Distal tarsal pain is a common reason for hind limb lameness, but diagnosis cannot always be made on radiographic examination. Scintigraphy may allow detection of subtle changes undetected by other diagnostic methods. We hypothesized that (1) distal tarsal pain would be associated with a loss of the expected pattern of radiopharmaceutical uptake (RU) detected in normal horses, (2) distal tarsal RU would be greater in limbs with tarsal pain than without pain, (3) RU in painful tarsi with radiographic evidence of osteoarthritis (OA) would be greater than in distal tarsal pain with no radiographic evidence of OA. The study aimed to describe radiopharmaceutical distribution in the distal tarsal region of horses with distal tarsal pain, and to compare this with the contralateral limb and results from horses without tarsal pain. Retrospective evaluation of scintigraphic images of the distal tarsal region was performed for 52 horses with distal tarsal pain: 15 with no radiographic evidence of OA (Group 1) and 37 with radiographic evidence (Group 2). The images were assessed using vertical and horizontal profile analysis across the distal tarsal region and regions of interest comparisons between the distal tarsal region and tibia within each horse (RU ratio). Painful limbs in unilaterally lame horses from Groups 1 and 2 had a significantly greater RU ratio than the respective contralateral limbs, and were significantly greater than the RU ratio in normal horses. On plantar images, mean region of interest counts were greater in the lame than the contralateral limb in Group 2 but not in Group 1. Although there was a positive correlation between lame and contralateral limb RU ratio in group 1, this was lost in group 2 horses. In lame limbs, the normal vertical activity profile was lost in 85% of group 1 and all of group 2, and the normal horizontal activity profile was lost in all of group 1 and 96% of group 2. There was a significant effect of lameness, but not of group on sites of peak activity on all profiles. The results of this study indicate that distal tarsal pain is associated with loss of the expected pattern of RU detected in normal horses. The findings also suggest that distal tarsal RU in lame limbs is greater than in limbs without pain, and that painful limbs with radiographic evidence of OA have a greater RU than painful limbs without radiographic evidence of OA.     

Effects of dietary fat and sire breed on puberty, weight, and reproductive traits of F1 beef heifers

Prepubertal F1 heifers (n = 246; from crossbred dams bred to either Hereford [H], Limousin [L], or Piedmontese [P] sires) were fed 1.9% (LF) or 4.4% (HF) dietary fat from 254+/-4 d of age until they reached puberty or the breeding season started. Safflower seeds (37% oil with 79% linoleic acid) were the added fat source. Blood samples and backfat thickness measurements were obtained from 60 randomly selected heifers representing the sire breeds and diets studied. In addition, five H-sired heifers from both diets were serially bled at 28-d intervals. Total gain, ADG, body condition score, and backfat thickness were affected by sire breed (P < 0.001) but not diet. Backfat thickness was affected (P < 0.01) by the diet x time on feed interaction. Diet did not affect pubertal age (P > 0.10) but tended (P = 0.08) to affect the percentage of heifers pubertal by the beginning of breeding (June 4). Sire breed effects on puberty age at beginning of breeding, percentage pubertal at the beginning of breeding, and puberty age during the entire study were all highly significant. The effect of the diet x sire breed interaction on percentage of heifers pubertal at beginning of breeding (P < 0.05) was 74.4 vs 76.3% in H-sired, 69.8 vs 60.5% in L-sired, and 76.2 vs 97.6% in P-sired heifers (LF vs HF, respectively). Number of AI services per pregnancy and final pregnancy percentage were not affected by diet or the diet x sire breed interaction. Diet affected progesterone (P < 0.05) and cholesterol (P < 0.001) concentrations, and sire breed tended to affect (P = 0.06) cholesterol concentrations. The effect of the diet x time on feed interaction on cholesterol concentrations was highly significant. There were no effects of diet or sample period on insulin or growth hormone concentrations in serially collected blood samples. We conclude that effects of supplemental dietary fat may be breed-dependent and hypothesize that a feeding period of approximately 60 d duration may be more appropriate than the 162 d used in this study.     

Methodology for the analysis of benzimidazole anthelmintics as drug residues in animal tissues

Methodology for the analysis of 8 benzimidazoles as residues in bovine liver is reported. Spiked tissues were extracted by homogenization in saline and ammonium hydroxide and blending with diatomaceous earth. This matrix was packed into a column, and the benzimidazoles were eluted with ethyl acetate. After the sample was further purified, benzimidazoles were separated and quantitated by liquid chromatography with ultraviolet detection (290 nm). Liver tissue samples obtained from cattle which had undergone a drug depletion study of fenbendazole administered per os were analyzed using these methods. The results of these analyses and the application of this approach to multiresidue analysis of drugs in animal tissues are discussed.     

Cimetidine inhibits nitric oxide associated nitrate production in a soft-tissue inflammation model in the horse

Cimetidine (CIM) is an H2-receptor antagonist that has been used in racehorses in an attempt to reduce the occurrence of stress-related gastric ulceration. It has also been shown to produce several useful effects other than its gastric acid suppression properties. Further, it is a well documented antagonist of cytochrome P-450 (CYP) mediated oxygenation reactions. Nitric oxide (NO), a recently discovered mediator or modifier of numerous physiological functions, is generated by several forms of nitric oxide synthase (NOS), one of which is inducible (iNOS). Inducible NOS, expressed in neutrophils and macrophages as part of the inflammatory response to noxious stimuli, contains both a CYP and a CYP reductase domain. Because of the similarity of structure of iNOS and CYP, it was decided to determine whether CIM could reduce NO production, using a carrageenan inflammation model in the horse. Two experiments were conducted. In Trial 1, six female Thoroughbred horses each had three tissue chambers inserted subcutaneously on the sides of the neck. The study was divided into three treatments: 0.9% NaCl (NaCI), CIM (3 mg/kg), and aminoguanidine (AG; 25 mg/kg), an inhibitor of iNOS. Each mare received three i.v. injections 12 h apart prior to instillation of 1 mL of carrageenan into the test chamber. Blood and tissue chamber fluid (TCF) were collected serially. Concentrations of NO3- (the major metabolite of NO), albumin, total protein, CIM and AG were measured and complete cell counts and differentials were conducted. Trial 2 also used six female Thoroughbred horses implanted with at least two tissue chambers inserted subcutaneously on the sides of the neck. The study was divided into two treatments: NaCl (0.9%) and CIM (6 mg/kg). Each mare received seven i.v. injections of either NaCl or CIM 8 h apart prior to instillation of 1 mL of carrageenan into the test chamber. Blood and TCF were collected serially as before, and analysed for NO3- and CIM content. Areas under the curve (AUC) of the different parameters were calculated for the periods of -1-1, -1-3 and -1-7 days (Trial 1) and -2-1 for Trial 2. Absolute values were also compared at 4, 8 and 12 h postcarrageenan. Saline treatment did not reduce the elevated concentrations of NO3- in either plasma or TCF. Plasma, test chamber and control chamber NO3-concentrations rose from 0 to 12 h, and were very similar in all three sampled fluids. Cimetidine significantly (P< or =0.05) decreased NO3- production in plasma over the periods of -1-1, -1-3, and -1-7 days post inflammation when compared to NaCl treatment in Trial 1. Aminoguanidine and CIM decreased NO3-production in TCF for the periods -1-1, 1-3, and -1-7 days post inflammation in Trial 1 and -2-1 for Trial 2. Both CIM and AG also significantly reduced NO3-concentrations in plasma and TCF at 12 h postinitiation (Trials 1 and 2). Thus CIM, at the doses studied, was capable of reducing NO3- concentrations in this model as effectively as AG, a relatively specific inhibitor of iNOS activity.