Plasma and Peritoneal Ceftriaxone Concentrations After Intraperitoneal Administration in Horses With Septic Peritonitis.

Intraperitoneal ceftriaxone administration in healthy horses results in high and prolonged peritoneal concentrations. Recent findings suggest that intraperitoneal ceftriaxone might increase survival rates in horses affected by peritonitis. The present study aimed to evaluate plasma and peritoneal concentrations of ceftriaxone after intraperitoneal administration in horses with septic peritonitis. Twenty-six horses presenting clinical, laboratorial, and sonographic findings compatible with the disease were included. All horses received daily intraperitoneal ceftriaxone (25 mg/kg bwt) in addition or not with other antibiotics and support therapies. High-performance liquid chromatography was used to determine plasma and peritoneal ceftriaxone concentrations before and after 12 and 24 hours of ceftriaxone administration. Mean plasma concentrations 12 and 24 hours after administration were, respectively, 1.84 ± 0.43 and 0.37 ± 0.07 µg/mL, and mean peritoneal concentrations were 5.7 ± 2.84 and 0.42 ± 0.13 µg/mL. Ceftriaxone concentration was lower in comparison with previous studies in healthy horses and presented under the minimal inhibitory concentration for enterobacteria (≤1 µg/mL) and for gram-positive isolates (≤0.5 µg/mL) at 24 hours. The variation of the results obtained between healthy horses and with septic peritonitis demonstrated that pharmacokinetics/dynamics are different between these patients and suggests the use of an interval of dose of 12 hours.

Application of in vitro Drug Metabolism Studies in Chemical Structure Optimization for the Treatment of Fibrodysplasia Ossificans Progressiva (FOP).

Currently no approved treatment exists for fibrodysplasia ossificans progressiva (FOP) patients, and disease progression results in severe restriction of joint function and premature mortality. LDN-193189 has been demonstrated to be efficacious in a mouse FOP disease model after oral administration. To support species selection for drug safety evaluation and to guide structure optimization for back-up compounds, in vitro metabolism of LDN-193189 was investigated in liver microsome and cytosol fractions of mouse, rat, dog, rabbit, monkey and human. Metabolism studies included analysis of reactive intermediate formation using glutathione and potassium cyanide (KCN) and analysis of non-P450 mediated metabolites in cytosol fractions of various species. Metabolite profiles and metabolic soft spots of LDN-193189 were elucidated using LC/UV and mass spectral techniques. The in vitro metabolism of LDN-193189 was significantly dependent on aldehyde oxidase, with formation of the major NIH-Q55 metabolite. The piperazinyl moiety of LDN-193189 was liable to NADPH-dependent metabolism which generated reactive iminium intermediates, as confirmed through KCN trapping experiments, and aniline metabolites (M337 and M380), which brought up potential drug safety concerns. Subsequently, strategies were employed to avoid metabolic liabilities leading to the synthesis of Compounds 1, 2, and 3. This study demonstrated the importance of metabolite identification for the discovery of novel and safe drug candidates for the treatment of FOP and helped medicinal chemists steer away from potential metabolic liabilities.

Study of the in vitro metabolic profile of a new α2-adrenergic agonist in rat and human liver microsomes by using liquid chromatography-multiple-stage mass spectrometry and nuclear magnetic resonance.

A potent synthetic α2-adrenergic agonist called PT-31, (3-(2-chloro-6-fluorobenzyl)-imidazolidine-2,4-dione), was recently detected as a potential drug to be used as an adjuvant drug to treat chronic pain. The excellent pharmacological property of PT-31 highlights the importance in elucidating its metabolism, which could provide valuable information about its metabolite profile for further pharmacokinetics studies and additionally to estimate the impact of its metabolites on the efficacy, safety and elimination of PT-31. In this work, the study of the in vitro metabolism of PT-31 was initially carried out by using a liquid chromatography coupled to ion trap multiple-stage mass spectrometer (LC-IT-MSn) and a hybrid triple quadrupole/linear ion trap mass spectrometer (LC-QTrap). The production of at least three unknown oxidative metabolites was observed. Structural identification of the unknown metabolites was carried out by combination of LC-MS experiments, including selected reaction monitoring (SRM) and multi-stage full scan experiments. Further analysis of 1H-NMR led to the structural confirmation of the major metabolite. The results indicated that PT-31 was metabolized by a hydroxylation reaction in the imidazolidine-2,4-dione ring in rat and human liver microsomes, producing the metabolite 3-(2-chloro-6-fluorobenzyl)-5-hydroxyimidazolidine-2,4-dione in rat liver microsomes. A carbon hydroxylation onto the benzyl ring, produced two other minor metabolites of the PT-31 in rat liver microsomes.

Human topoisomerase inhibition and DNA/BSA binding of Ru(II)-SCAR complexes as potential anticancer candidates for oral application.

Three ruthenium(II) phosphine/diimine/picolinate complexes were selected aimed at investigating anticancer activity against several cancer cell lines and the capacity of inhibiting the supercoiled DNA relaxation mediated by human topoisomerase IB (Top 1). The structure-lipophilicity relationship in membrane permeability using the Caco-2 cells have also been evaluated in this study. SCAR 5 was found to present 45 times more cytotoxicity against breast cancer cell when compared to cisplatin. SCAR 4 and 5 were both found to be capable of inhibiting the supercoiled DNA relaxation mediated by Top 1. Interaction studies showed that SCAR 4 and 5 can bind to DNA through electrostatic interactions while SCAR 6 is able to bind covalently to DNA. The complexes SCAR were found to interact differently with bovine serum albumin (BSA) suggesting hydrophobic interactions with albumin. The permeability of all complexes was seen to be dependent on their lipophilicity. SCAR 4 and 5 exhibited high membrane permeability (P app  > 10 × 10-6 cm·s-1) in the presence of BSA. The complexes may pass through Caco-2 monolayer via passive diffusion mechanism and our results suggest that lipophilicity and interaction with BSA may influence the complexes permeation. In conclusion, we demonstrated that complexes have powerful pharmacological activity, with different results for each complex depending on the combination of their ligands.

Pharmacokinetic properties, in vitro metabolism and plasma protein binding of govaniadine an alkaloid isolated from Corydalis govaniana Wall.

Govaniadine (GOV) is an alkaloid isolated from Corydalis govaniana Wall. It has been reported to show a different number of biological activities including anti-urease, leishmanicidal and antinociceptive. The present study aims to characterize the GOV in vitro metabolism after incubation with rat and human liver microsomes (RLM and HLM, respectively) and to evaluate its pharmacokinetic properties. The identification of GOV metabolites was conducted by different mass analyzers: a micrOTOF II-ESI-ToF Bruker Daltonics® and an amaZon-SL ion trap (IT) Bruker Daltonics®. For the pharmacokinetic study of GOV in rats after intravenous administration, a LC-MS/MS method was developed and applied to. The analyses were performed using an Acquity UPLC® coupled to an Acquity TQD detector equipped with an ESI interface. The liver microsomal incubation resulted in new O-demethylated, di-hydroxylated and mono-hydroxylated compounds. Regarding the method validation, the calibration curve was linear over the concentration range of 2.5-3150.0ngmL-1, with a lower limit of quantitation (LLOQ) of 2.5ngmL-1. This method was successfully applied to a pharmacokinetic study. The profile was best fitted to a two-compartment model, the first phase with a high distribution rate constant (α) 0.139±0.086min-1, reflected by the short distribution half-life (t1/2α) 9.2±8.9min and the later one, with an elimination half-life (t1/2ß) 55.1±37.9min. The main plasma protein binding was 96.1%. This is a first report in this field and it will be useful for further development of govaniadine as a drug candidate.

Comparison of the effects of epidural or intravenous methadone on the minimum alveolar concentration of isoflurane in dogs.

The effects of epidural and intravenous (IV) methadone (0.5mg/kg) on the minimum alveolar concentration of isoflurane (ISO(MAC)) were compared in dogs. Six dogs (16.5 ± 2.5 kg bodyweight) received three treatments in random order during isoflurane anaesthesia, with a 7 day washout interval between each study. Methadone was injected via a lumbosacral epidural catheter introduced 10 cm cranially into the epidural canal and the electrical stimulation for ISO(MAC) determination was applied either to the thoracic (EP(T) treatment) or to the pelvic limb (EP(P) treatment) during separate study days. In the IV treatment, ISO(MAC) was determined via electrical stimulation of the pelvic limb. Variables were recorded before (baseline), 2.5 and 5h after drug injection. The ISO(MAC) decreased significantly (P<0.05) from baseline at 2.5 and 5h after methadone in all treatments. At 2.5h, the magnitude of ISO(MAC) reduction did not differ between treatments (mean decreases from baseline: 30-33%). The ISO(MAC) reduction lasted longer following epidural methadone in the thoracic limb (decreases from baseline: 30% at 5h in the EP(T) treatment vs. 19% and 16% in the EP(P) and IV treatments, respectively). Although the isoflurane sparing effect provided by epidural methadone was not significantly greater than IV methadone during the initial stage (2.5h), it was more prolonged than the IV route in specific dermatomes (5h in the thoracic limb) with the epidural technique employed. Methadone may therefore provide a greater isoflurane sparing effect when administered epidurally, compared to IV, when noxious stimulation occurs in specific dermatomes.

Organophosphorus-induced delayed neuropathy: a simple and efficient therapeutic strategy.

Organophosphorus (OP) used as pesticides and hydraulic fluids can produce acute poisoning known as OP-induced delayed neuropathy (OPIDN), whose effects take long time to recover. Thus a secure therapeutic strategy to prevent the most serious effects of this poisoning would be welcome. In this study, tri-o-cresyl phosphate (TOCP, 500 mg/kg p.o.) was given to hens, followed or not by nimodipine (1mg/kg i.m.) and calcium gluconate (Ca-glu 5mg/kg i.v.). Six hours after TOCP intoxication, neuropathy target esterase (NTE) activity inhibition was observed, peaking after 24h exceeding 80% inhibition. A fall in the plasmatic calcium levels was noted 12h after TOCP was given and, in the sciatic nerve, Ca(2+) fell 56.4% 24h later; at the same time calcium activated neutral protease (CANP) activity increased 308.7%, an effect that lasted 14 days. Any bird that received therapeutic treatment after TOCP intoxication presented significant signs of OPIDN. These results suggest that NTE may be implicated in the regulation of calcium entrance into cells being responsible for the maintenance of normal function of calcium channels, and that increasing CANP activity is responsible to triggering OPIDN. Thus, with one suitably adjusted dose of nimodipine as well as Ca-glu, we believe that this treatment strategy may be used in humans with acute poisoning by neuropathic OP.

Stereoselective analysis of carvedilol in human plasma and urine using HPLC after chiral derivatization.

An enantioselective high-performance liquid chromatographic method for the analysis of carvedilol in plasma and urine was developed and validated using (-)-menthyl chloroformate (MCF) as a derivatizing reagent. Chloroform was used for extraction, and analysis was performed by HPLC on a C18 column with a fluorescence detector. The quantitation limit was 0.25 ng/ml for S(-)-carvedilol in plasma and 0.5 ng/ml for R(+)-carvedilol in plasma and for both enantiomers in urine. The method was applied to the study of enantioselectivity in the pharmacokinetics of carvedilol administered in a multiple dose regimen (25 mg/12 h) to a hypertensive elderly female patient. The data obtained demonstrated highest plasma levels for the R(+)-carvedilol (AUC(SS) (0-12) 75.64 vs 37.29 ng/ml). The enantiomeric ratio R(+)/S(-) was 2.03 for plasma and 1.49 for urine (Ae(0-12) 17.4 vs 11.7 microg).