Liposome-based diclofenac for the treatment of inflammation in an acute synovitis model in horses.

Lameness as a result of joint disease is a major source of decreased athletic performance in the horse. Most treatment protocols include the administration of nonsteroidal anti-inflammatory drugs (NSAIDs). Phenylbutazone, alone or in combination with other treatments, is the most commonly and widely used NSAID, however it has the potential for serious side effects. The introduction of the liposome-based formulation of the NSAID diclofenac has shown promising effect as a safe and convenient treatment for lameness associated with osteoarthritis. The purpose of this study was to evaluate the effect of topical liposome-based diclofenac in an acute inflammation model using subjective lameness scores and objective lameness evaluation, carpal surface temperature and circumference, synovial fluid cell count and total protein content, and the biochemical markers interleukin-1 (IL-1), IL-6, and prostaglandin E(2) as determinants of inflammation. In this acute inflammation model, there was no overall difference between treatment and control groups.

Detection and identification of carprofen and its in vivo metabolites in greyhound urine by capillary gas chromatography-mass spectrometry.

Rimadyl (carprofen) was administered orally to the racing greyhound at a dose of 2.2 mg kg(-1). Following both alkaline and enzymatic hydrolysis, postadministration urine samples were extracted by mixed mode solid-phase extraction (SPE) cartridges to identify target analyte(s) for forensic screening and confirmatory analysis methods. The acidic isolates were derivatised as trimethylsilyl ethers (TMS) and analysed by gas chromatography-mass spectrometry (GC-MS). Carprofen and five phase I metabolites were identified. Positive ion electron ionisation (EI(+)) mass spectra of the TMS derivatives of carprofen and its metabolites show a diagnostic base peak at M(+)*. -117 corresponding to the loss of COO-Si-(CH(3))(3) group as a radical. GC-MS with positive ion ammonia chemical ionisation (CI(+)) of the compounds provided both derivatised molecular mass and some structural information. Deutromethylation-TMS derivatisation was used to distinguish between aromatic and aliphatic oxidations of carprofen. The drug is rapidly absorbed, extensively metabolised and excreted as phase II conjugates in urine. Carprofen, three aromatic hydroxy and a minor N-hydroxy metabolite were detected for up to 48 h. For samples collected between 2 and 8 h after administration, the concentration of total carprofen ranged between 200 and 490 ng ml(-1). The major metabolite, alpha-hydroxycarprofen was detected for over 72 h and therefore can also be used as a marker for the forensic screening of carprofen in greyhound urine.

Detection of fenspiride and identification of in vivo metabolites in horse body fluids by capillary gas chromatography-mass spectrometry: administration, biotransformation and urinary excretion after a single oral dose.

Studies related to the in vivo biotransforrmation and urinary excretion of fenspiride hydrochloride in the horse are described. After oral administration, the drug is metabolised by both phase I functionalisation and phase II conjugation pathways. Following enzymatic deconjugation, fenspiride and its phase I metabolites were isolated from post-administration biofluids using bonded co-polymeric mixed mode solid-phase extraction cartridges to isolate the basic compounds. Following trimethylsilylation (TMS), the parent drug and metabolites were identified by capillary gas chromatography-mass spectrometry (GC-MS). Fenspiride (A) and seven metabolites (B-->G) arising from oxidation on both the aromatic and heterocyclic substructures were detected in urine. The positive ion electron ionisation mass spectra of the TMS derivatives of fenspiride and its metabolites provided useful information on its metabolism. Positive ion methane chemical ionisation-GC-MS of the derivatives provided both derivatised molecular mass and structural information. Unchanged fenspiride can be detected in post-administration plasma and urine samples for up to 24 h. Maximum urinary levels of 100-200 ng ml(-1) were observed between 3 and 5 h after administration. After enzymatic deconjugation, the major phenolic metabolite (G) can be detected in urine for up to 72 h. This metabolite is the analyte of choice in the GC-MS screening of post-race equine urine samples for detection of fenspiride use. However, a distinct difference was observed in the urinary excretion of this metabolite between the thoroughbred horses used in UK study and the quarterbred and standardbred horses used for the USA administrations.

Characterization of urinary metabolites of testosterone, methyltestosterone, mibolerone and boldebone in greyhound dogs.

Androgenic steroids are used in female greyhound dogs to prevent the onset of estrus; moreover, these steroids also have potent anabolic activity. As anabolic steroids increase muscle mass and aggression in animals, the excessive use of these agents in racing greyhounds gives an unfair performance advantage to treated dogs. The biotransformation of most anabolic steroids has not been determined in greyhound dogs. The objective of the present study was to identify the urinary metabolites of testosterone, methyltestosterone, mibolerone, and boldenone in greyhound dogs. These steroids were administered orally (1 mg/kg) to either male or female greyhound dogs and urine samples were collected pre-administration and at 2, 4, 8, 12, 24, 72, and 96 h post-administration. Urine extracts were analyzed by high-performance liquid chromatography/mass spectrometry (HPLC/MS) to identify major metabolites and to determine their urinary excretion profiles. Major urinary metabolites, primarily glucuronide, conjugated and free, were detected for the selected steroids. Sulfate conjugation did not appear to be a major pathway for steroid metabolism and excretion in the greyhound dog. Phase I biotransformation was also evaluated using greyhound dog liver microsomes from untreated dogs. The identification of several in vivo steroid metabolites generated in this study will be useful in detecting these steroids in urine samples submitted for drug screening.

Neoproterozoic 'snowball Earth' simulations with a coupled climate/ice-sheet model.

Ice sheets may have reached the Equator in the late Proterozoic era (600-800 Myr ago), according to geological and palaeomagnetic studies, possibly resulting in a 'snowball Earth'. But this period was a critical time in the evolution of multicellular animals, posing the question of how early life survived under such environmental stress. Here we present computer simulations of this unusual climate stage with a coupled climate/ice-sheet model. To simulate a snowball Earth, we use only a reduction in the solar constant compared to present-day conditions and we keep atmospheric CO2 concentrations near present levels. We find rapid transitions into and out of full glaciation that are consistent with the geological evidence. When we combine these results with a general circulation model, some of the simulations result in an equatorial belt of open water that may have provided a refugium for multicellular animals.

Development of analytical methods for the detection of metaraminol in the horse.

Aramine (metaraminol bitartrate) has been found in the possession of horse trainers and veterinarians who have been investigated for possible inappropriate drug administration to racing horses. Metaraminol (3-hydroxyphenylisopropanolamine) is a sympathomimetic amine that directly and indirectly affects adrenergic receptors, with alpha effects being predominant. Because it has the potential to affect the performance of a racing horse, its use is prohibited. In the present study, methods for the detection of metaraminol were developed. Metaraminol was found to be extracted with poor recovery (< 50%) from aqueous solutions by routine basic extraction or cation exchange/reversed-phase solid-phase extraction techniques. However, an extractive acetylation method gave good (> 90%) recovery of metaraminol from aqueous samples. Sequential urine samples collected from horses administered metaraminol intramuscularly at 0.02, 0.10, and 0.23 mg/kg were extracted by the developed extractive acetylation procedure and analyzed by gas chromatography-mass spectrometry (GC-MS) in full-scan and selected ion monitoring modes. Norphenylephrine was used as an internal standard for quantitative analysis. The maximum concentration of metaraminol occurred between 1 and 2 h postadministration. Metaraminol was detected in the 0.23 mg/kg administration urine for 24 h postadministration. Metaraminol was detected for the 0.10 and 0.02 mg/kg doses for approximately 8 h postadministration. No apparent biotransformation products were observed in a reaction mixture of metaraminol and horse liver microsomal reaction mixture. Comparison of gas chromatograms of the extracts of the postadministration urine samples with those of the pre-administration samples failed to reveal any exogenous compound other than metaraminol.

Effects of temperature, incubation period and substrate on production of fusaproliferin by Fusarium subglutinans ITEM 2404.

The kinetics of the production of fusaproliferin by Fusarium subglutinans ITEM 2404 in maize and rice cultures was investigated at various incubation temperatures. The growth rate of F. subglutinans was highest at 20 degrees C and 25 degrees C in maize cultures and at 15 degrees C in rice cultures. Although the growth rate was higher in rice than in maize, the maximal production of fusaproliferin was obtained in maize cultures, with a maximum yield (4309 microg g(-1)) at 20 degrees C for 6 weeks. In rice cultures the optimal incubation regimen was at 15 degrees C for 6 weeks, with a fusaproliferin level of 1557 microg g(-1). The production of fusaproliferin at 25 degrees C and 30 degrees C in both substrates was very low, with maximal yield at 25 degrees C of 979 microg g(-1) after 2 weeks and 143 microg g(-1) after 3 weeks in maize and rice cultures, respectively.

Isolation, purification, and structural characterization of flunixin glucuronide in the urine of greyhound dogs.

A urinary metabolite of flunixin in greyhound dogs was isolated and purified by a gradient-elution solid-phase extraction technique. The purified metabolite was shown to be hydrolyzed to free flunixin by strong base and by beta-glucuronidase, suggesting the presence of a C1-beta-glucuronide ester of flunixin. The metabolite was further characterized by positive-ion, tandem MS with electrospray ionization. Mass spectral data showed the presence of a protonated molecular ion (M+1) at m/z 473, which was consistent with the molecular weight of protonated flunixin glucuronide, and a product ion at m/z 297, which was consistent with the molecular weight of protonated flunixin. Collisionally induced dissociation of the m/z 297 product ion showed a fragmentation pattern consistent with that of standard flunixin. These data support the contention that this metabolite of flunixin in greyhound urine is the C1-beta-glucuronide of flunixin. Acyl glucuronide metabolites of some organic acid drugs have been shown to bind covalently to tissue proteins in vitro, in vivo, and ex vivo. The presence of this metabolite may, therefore, have pharmacokinetic and pharmacodynamic implications for flunixin in greyhound dogs, as well as in other animal species in which the acyl glucuronide of flunixin is a metabolite.

Environmental motivations for migration: population pressure, poverty, and deforestation in the Philippines.

"This paper uses a multinomial discrete choice model and data from the Philippines to examine migrant choice between alternative destinations. Travel costs and perceived opportunities at the upland frontier are more important than general (upland plus lowland) destination attributes that indicate more developed social infrastructure or greater expected welfare. For example, migration streams are larger to destinations where the public share of forestland and the road system are larger. These features also characterize regions of more rapid deforestation. Therefore, emigration policies must recognize their effects on deforestation at the frontier--and their anticipated indirect effects on downstream environments."

Detection of flunixin in greyhound urine by a kinetic enzyme-linked immunosorbent assay.

A two-step kinetic enzyme-linked immunosorbent assay was developed to detect the presence of flunixin in the urine of greyhound dogs. The assay system was developed using polyclonal antiflunixin antisera, a rabbit albumin-flunixin conjugate adsorbed onto polystyrene microtiter strips, and flunixin reference standards for calibration. The assay parameters were optimized and the performance characteristics were determined. The quantitative intra- and inter-run precisions (%CV) of the analysis of replicate (n = 10) flunixin-spiked urine samples were 9.9-12.5% and 10.2-13.6%, respectively. The linear dynamic range was 1-100 ng/mL, and the quantitative accuracy, as determined by calculation of percent error of measured flunixin in flunixin-spiked drug-free greyhound urine, was -16% to +14% over this range. The I50 of the ELISA was 17.3 ng/mL. The limit of detection was 25 ng/mL in greyhound urine. The reactivity in the assay system relative to flunixin (100%) was 147% for flunixin glucuronide, 25% for clonixin, and 5% for niflumic acid. The ELISA was capable of detecting total flunixin for up to 72 h in dogs administered flunixin at 0.55 mg/kg orally and up to 96 h in a dog that was administered flunixin at 1.0 mg/kg orally.

Effect of divalent cations in bacteriological media on the susceptibility of Xanthomonas maltophilia to imipenem, with special reference to zinc ions.

The susceptibility of Xanthomonas maltophilia strains to beta-lactams is known to vary according to the type of growth media used. We have assayed the divalent cation content of various susceptibility testing media and correlated this with the susceptibility to imipenem of 30 strains of X. maltophilia, by calculating the IC50. No correlation was found with Ca++ and Mg++ content (r = 0.005, P = 0.99), but a highly significant correlation with the Zn++ content (7.7-42.7 mumol/L) of the medium was found (r = 0.93, P = 0.003). The effect of Zn++ on the susceptibility of X. maltophilia to imipenem was further investigated by adding varying amounts of zinc sulphate to Oxoid Mueller-Hinton agar which has a low Zn++ content (14.2 mumol/L). A highly significant correlation between the Zn++ content and the IC50 was observed (r = 0.95, P = 0.001). Some variability was seen from one series of IC50 determinations to another and samples of ultra-pure water were processed in exactly the same fashion as the agar media and then assayed for cation content. No significant increase in Ca++ or Mg++ content of the water were observed but water autoclaved in universal containers with metal caps and rubber washers acquired up to 40 mumol/L of Zn++. Studies of the correlation between in-vitro sensitivity tests and the clinical performance of beta-lactams used against X. maltophilia will need to take account of the Zn++ content of the susceptibility testing media used.

More rapid identification of bacteraemia by manual rather than radiometric method.

Results of blood culture examination using the radiometric (Bactec-460) system for one year showed no overall improvement compared with those of the previous three years when a manual system with early blind subculture was used. The isolates from the manual system were available more often on solid media, 24 hours earlier, than when the radiometric system was used. In a further study of 1100 blood cultures the radiometric medium was tested for growth index as well as being subcultured blindly, irrespective of growth index, on the first day. Thirty six out of 54 (67%) of the blood cultures were positive on subculture but negative for growth index at this time. The overall cost of the radiometric system is also considerably more than that of the manual system.

Levamisole toxicosis in swine.

Levamisole toxicosis occurred in swine in a commercial swine operation after the animals were mistakenly injected intramuscularly with levamisole. Clinical signs consisted of vomition, salivation, ataxia, recumbency and death. Surviving animals recovered completely within 24 to 48 hr.

Rapid detection of bacteraemia by early subculture.

Inspection and blind subculture was carried out on 7031 consecutive blood cultures at 10 p.m. on the day they were received. Analysis of the results after a 2-year period showed that 119 out of 237 (50%) bacteraemic patients were detected at this stage. Preliminary sensitivity tests were done at this time and their results were available within 24 h of the blood cultures being received. Thus earlier specific therapy was possible in 50% of the cases of bacteraemia.

Determination of extractable mirex in whole blood.

A method for the determination of free mirex in blood has been developed in which whole blood is extracted with acetone-hexane (9+91), passed through Na2SO4, concentrated, and analyzed by electron capture gas-liquid chromatograpohy (GLC). Results were highest for fresh blood. Recoveries were verified by determining 14C-labeled mirex before and after extraction. Little mirex was detected in the samples after extraction. Hydrolysis of residual blood indicates that a metabolite of mirex is released. The possible metabolite has a GLC retention time and mass spectrum which resemble hydroxy metabolites of mirex.