Pharmacokinetics of ketamine and norketamine enantiomers after racemic or S-ketamine IV bolus administration in dogs during sevoflurane anaesthesia

Accepted Manuscript Pharmacokinetics of ketamine and norketamine enantiomers after racemic or S-ketamine IV bolus administration in dogs during sevoflurane anaesthesia

Noemi Romagnoli, Rima N. Bektas, Annette P. Kutter, Andrea Barbarossa, Paola Roncada, Sonja Hartnack, Regula BettschartWolfensberger PII: DOI: Reference:

S0034-5288(16)30722-6 doi: 10.1016/j.rvsc.2017.05.005 YRVSC 3321

To appear in:

Research in Veterinary Science

Received date: Revised date: Accepted date:

7 December 2016 3 May 2017 5 May 2017

Please cite this article as: Noemi Romagnoli, Rima N. Bektas, Annette P. Kutter, Andrea Barbarossa, Paola Roncada, Sonja Hartnack, Regula Bettschart-Wolfensberger , Pharmacokinetics of ketamine and norketamine enantiomers after racemic or S-ketamine IV bolus administration in dogs during sevoflurane anaesthesia, Research in Veterinary Science (2017), doi: 10.1016/j.rvsc.2017.05.005

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ACCEPTED MANUSCRIPT Pharmacokinetics of ketamine and norketamine enantiomers after racemic or Sketamine IV bolus administration in dogs during sevoflurane anaesthesia

Noemi Romagnolia, Rima N Bektasb, Annette P Kutterb, Andrea Barbarossaa,*, Paola

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Roncadaa, Sonja Hartnackb, Regula Bettschart-Wolfensbergerb

Department of Veterinary Medical Sciences, Alma Mater Studiorum – University of

Bologna, Via Tolara di Sopra 50, Ozzano Emilia (BO) 40064, Italy b

Anaesthesiology Section, Vetsuisse Faculty, University of Zurich, Winterthurerstr. 258 c,

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8057 Zurich, Switzerland

* Corresponding author: Andrea Barbarossa

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Via Tolara di Sopra 50, Ozzano Emilia (BO) 40064, Italy Tel. +39 051 2097500

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E-mail: [email protected]

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ACCEPTED MANUSCRIPT Abstract The aims of this study were to measure plasma levels of R- and S-ketamine and their major metabolites R- and S- norketamine following single intravenous bolus administration of racemic or S-ketamine in sevoflurane anaesthetised dogs and to calculate the relevant pharmacokinetic profiles. Six adult healthy beagle dogs were used in the study. An

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intravenous bolus of 4 mg/kg racemic ketamine (RS-KET) or 2 mg/kg S-ketamine (S-KET) was administered, with a three-weeks washout period between treatments. Venous blood samples were collected at fixed times until 900 minutes and R- and S-ketamine as well as Rand S -norketamine plasma levels determined by liquid chromatography coupled with tandem

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mass spectrometry. Cardiovascular parameters were recorded during the anaesthesia until 240 min. All dogs recovered well from anaesthesia. No statistical differences between groups

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were detected in any cardiovascular parameter. The pharmacokinetics of S-ketamine did not differ when injected intravenously alone or as part of the racemic mixture in dogs

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anaesthetised with sevoflurane. Following racemic ketamine, the area under the curve of R-

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norketamine was statistically higher than the one of S-norketamine.

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Keywords: dog; enantiomers; mass spectrometry; S-ketamine; S-norketamine

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ACCEPTED MANUSCRIPT Introduction Ketamine is a dissociative drug, commonly used in both human and veterinary anaesthesia. Commercially available ketamine is a 50:50% racemic mixture of S-ketamine and Rketamine. In several species, it has been shown, that the same anaesthetic effect can be achieved with S-ketamine at a dose of 50-67% of racemic ketamine (Deleforge et al., 1991;

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Adams and Werner, 1997; Schmidt et al., 2005; Duque et al., 2008; Larenza et al., 2008a). In humans and horses, S-ketamine at equipotent doses of the racemic mixture induces a slightly shorter action (Doenicke et al., 1992; Larenza et al., 2008b). Pharmacokinetic studies of racemic and S-ketamine in non premedicated humans and horses showed that kinetics

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differed between the enantiomers (Ihmsen et al., 2001; Larenza et al., 2009a). It was suggested that R-ketamine inhibited the elimination of S-ketamine and as a consequence S-

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ketamine elimination was faster following administration of S-ketamine alone in comparison to racemic ketamine. On the other hand, no difference in pharmacokinetics between the two

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enantiomers was detected, when ketamine was administered to isoflurane anaesthetised ponies (Larenza at al., 2007).

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In dogs, the only study on stereoselective pharmacokinetics of racemic ketamine was

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performed under halothane anaesthesia. It demonstrated that following administration of racemic ketamine, S-ketamine showed a 35% higher elimination clearance than R-ketamine

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(Henthorn et al., 1999). Residual effects of ketamine or its metabolites can induce undesired psychomotor emergence reactions in recovery from anaesthesia. Those negative effects might be reduced following S-ketamine compared to racemic ketamine, as necessary dose rates are lower and elimination of the drug faster. Further cardiopulmonary effects might also differ between enantiomers. In dogs no comparative pharmacokinetic studies of racemic and Sketamine are available. The investigation of S-ketamine pharmacokinetics in detail is

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ACCEPTED MANUSCRIPT mandatory to allow its safe use and to elucidate all differences between S-ketamine and the current standard racemic ketamine. The authors supposed that the pharmacokinetics of S-ketamine and S-norketamine could be different if S-ketamine is administered alone or as part of the commonly used racemic mixture. In detail, based on their clinical experience, the authors expected that S-ketamine

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would be eliminated more rapidly when co-administered with the R- enantiomer.. The aims of this study were to measure plasma levels of R- and S-ketamine and their major metabolites Rand S- norketamine following single intravenous (IV) bolus administration of racemic or Sketamine in sevoflurane anaesthetised dogs and to calculate the relevant pharmacokinetic

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profiles.

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Materials and methods Animals

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The trial was approved by the committee for Animal Experimentation of the Canton of Zurich, Switzerland (67/2011). Six adult healthy beagle dogs, 3 females and 3 males (non-

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castrated), 38 (± 8) months of age and weighing 15.0 ±1.1 kg were used in the study. All the

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dogs were regularly checked for parasites and were vaccinated. Six days before the start of the experiments, blood cell counts were carried out and blood chemistry was checked. The weight

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of the individual dogs was evaluated the morning of the day the experiment was conducted. The dogs were fasted overnight before the experiment, but were given access to water ad libitum. The dogs were fed their standard diet once fully awake. Study design, instrumentation and monitoring The study was designed as a prospective, blinded, randomised crossover trial. General anaesthesia, necessary to allow the instrumentation of each animal, was induced via facemask using 8% sevoflurane in oxygen (4 L/min). Once unconscious, the tracheas of the dogs were

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ACCEPTED MANUSCRIPT intubated. Anaesthesia was maintained with 1.4 MAC = 3 % (+/-0.2 %) endtidal (Et) sevoflurane in oxygen and air (FiO2 0.35). The dogs were artificially ventilated to maintain EtCO2 35 +/- 5 mmHg, inspiratory peak pressure 12 +/- 2 cm H2O using a semi-closed circle system (Avance S 5 -Datex Ohmeda, Anandic Medical, Feuerthalen, Switzerland). All catheter sites were clipped and surgically prepared. Instrumentation consisted of the

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application of two peripheral venous 22 G catheters (Surflo IV Catheter 22G x 1”; Terumo. Leuven, Belgium), one used to inject the test drugs and carry out fluid therapy, and the second in case of problems during blood collection from the central venous catheter. A16 G x 16 cm long central venous catheter (Blue Flex Tip® Catheter, Arrow International, Teleflex Medical

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GmbH, Belp, Switzerland) was used to collect the blood samples for the pharmacokinetic analysis. For measurement of blood pressure and cardiac output using transpulmonary

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thermodilution (PiCCOplus, Software version 6.0, Pulsion Medical Systems, Munich, Germany) a 4-Fr 22 cm thermodilution catheter (PV2014L22N, Pulsion Medical Systems,

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Munich, Germany) was inserted to its full length into the femoral artery. As soon as the first peripheral venous catheter was inserted, 5 mL/kg/h lactated ringer solution were administered

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to maintain adequate blood volume. Heart rate (HR), intra-arterial blood pressures (systolic:

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SAP, mean: MAP, diastolic: DAP), cardiac output (CO), composition of in- and expired gases and core body temperature were all measured continuously using a multiparameter monitor

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(BN 850, GE Medical Systems, Anandic Medical Systems AG, Diessenhofen, Switzerland). Body temperature was maintained at 37.5-38.5 °C with a warm water blanket underneath the dog and a warm air device (Bair Hugger, 3M, Milan, Italy) to effect. The first recording of measured parameters took place before test drug administration and 5, 15, 30, 45, 60, 120, 180 and 240 min after it. Arterial blood was sampled anaerobically using specifically designed blood gas syringes (BD A-Line® Blutgasspritze, Becton Dickinson AG, Allschwil, Switzerland). Analysis of blood gases and pH as well as sodium (Na), potassium

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ACCEPTED MANUSCRIPT (K), chloride (Cl), calcium (Ca) and lactate concentrations were immediately performed using a blood gas analyser with co-oxymetry (Radiometer, ABL 700 Series, Radiometer GmbH, 8800 Thalwil, Switzerland). After 240 min all but the central venous catheter and the peripheral venous catheter not used for drug administration were removed and the dogs allowed to recover.

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Drug administration and sample collection

Three mL of blood were sampled into heparinised tubes before any treatment (T0) and were centrifuged immediately. Plasma was stored at -80 °C until the assay was carried out. Following instrumentation under sevoflurane anaesthesia, a single IV bolus of racemic

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ketamine 4 mg/kg (RS-KET group, 100mg/ml, Dr. E. Graeub AG, Bern) or S-ketamine 2 mg/kg (S-KET group, 50mg/ml, Dr. E. Graeub AG, Bern) was administered. After a three-

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week washout period, each dog underwent the same procedure but receiving the other dissociative agent. Three mL of venous blood was collected from the central venous catheter

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and transferred to labelled heparinised tubes (BD 3.5 mL Vacutainer, LH PST, REF 367374, Becton Dickinson, Temse, Belgium) at fixed times: 1, 2, 5, 10, 15, 20, 30, 45, 60, 75, 90, 105,

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120, 150, 180, 210, 240, 300, 360, 450, 540, 630, 720, 810 and 900 min after drug injection.

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To avoid contamination with the venous flushing solution, 5 mL of blood were collected before each sampling and re-injected thereafter. The venous catheter was flushed with 5 mL

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of physiologic 0.9% saline solution. Immediately after collection, the samples were kept at 4 °C until centrifugation (maximum 1 hour); plasma samples were frozen at -80 °C until the assay was carried out. At the end of the anaesthesia, all dogs received 22 mg/kg cefazoline (Kefzol®, Teva Pharma AG, Aesch, Switzerland) and 4 mg/kg carprofen (Rimadyl® ad us. vet., Pfizer, Zurich, Switzerland) IV. After the last blood sample was collected, the remaining catheters were removed. If the dogs were completely unremarkable (assessed by an experienced

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ACCEPTED MANUSCRIPT veterinarian), they were returned to their normal surroundings the following day and checked twice daily for abnormal behaviour and body temperature for three days. Analysis of the plasma concentrations of ketamine and norketamine enantiomers The plasma samples were analysed using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). A simple and quick extraction was performed using the technique

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described by Sergi et al. (2009), with slight modifications. After being thawed at room temperature, 150 μL of plasma were transferred to a microtube, then internal standards (Ketamine‐D4 and Norketamine‐D4, purchased from Sigma-Aldrich, St.Louis, MO, USA) and methanol were added, in order to obtain a total volume of 1.0 mL. The sample was then

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vortex-mixed for 30 sec, placed into an ultrasonic bath for 2 min and centrifuged at 8’000 ×g

Torrance, CA, USA) prior to analysis.

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for 5 min. The supernatant was finally filtered through a 0.2 μm PTFE filter (Phenomenex,

The LC system consisted of a Waters Aquity UPLC binary pump (Waters, Milford, MA,

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USA), equipped with a Phenomenex Lux 3u Cellulose‐3 (150 × 2.00 mm, 3.0 μm) column (Phenomenex, Torrance, CA, USA) kept at 45 °C. The mobile phase consisted of a mixture of

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acetonitrile and an aqueous solution containing 20 mM ammonium acetate and 0.1%

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ammonium formate, at a flow rate of 0.450 mL/min under programmed conditions, in order to obtain satisfactory separations of the two enantiomers of each compound. The LC was

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interfaced to a Waters Quattro Premier XE triple quadrupole mass spectrometer (Waters, Milford, MA, USA), operating in positive electrospray ionisation (ESI+) and in MRM (multiple reaction monitoring) mode. The specific transitions observed were: ketamine, m/z 238 → 125 and 179; ketamine‐D4, m/z 242 → 129 and 183; norketamine, 224 → m/z 125; norketamine‐D4, m/z 228 → 129. The capillary voltage was set at 3.00 kV, and the source temperature was 120 °C.

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ACCEPTED MANUSCRIPT The analytical method was validated in accordance with the EMEA/CHMP/EWP/192217/2009 guidelines at the beginning of the experiment. The linearity was satisfactory (R2 > 0.99) over a range from 5 to 8’000 ng/mL for both Sketamine and R-ketamine, and from 5 to 1’600 ng/mL for the norketamine enantiomers. The lower limit of quantification (LLOQ) was 5 ng/mL for all target compounds; interday and

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intraday accuracy and precision, expressed as relative standard deviation (SD) (n=9), were below 15%. Statistical analysis

Cardiopulmonary parameters and arterial blood gas data are reported as mean ± SD; for these

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parameters, a linear mixed effects model approach was performed using the software package R version 2.14.0 (R Development Core Team, 2011) and the package NLME (Pinheiro et al.,

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2011), to detect differences between the RS-KET and the S-KET treatment groups before and after test drug administration.

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The R-ketamine and S-ketamine concentration vs. time curves were analysed for each individual by XY plot using WinNonlin 6.3 software (Pharsight Corporation, Mountain View,

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CA, USA). The Wilcoxon rank test with Bonferroni correction was used to compare the

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concentrations of ketamine and norketamine enantiomers between the two different groups (the RS-KET group and the S-KET group) as well as to compare the calculated

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pharmacokinetic parameters between groups. The significance level was fixed at p<0.05. The statistical analysis was carried out using computer software (Med-Calc 12.7.5, MedCalc Software, Acacialaan 22, B-8400 Ostend, Belgium). Plasma concentrations obtained after IV administration were fitted using the following equation: C(t) = A e-αt + B e-β. All pharmacokinetic parameters were reported as mean ± SD and were determined using WinNonlin 6.3 (Pharsight Corporation, Mountain View, CA, USA). The individual plasma concentration-versus-time curves were fitted and the best

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ACCEPTED MANUSCRIPT compartment model was determined by application of the Akaike information criterion and Schwartz criterion (Schwartz 1978; Yamaoka et al. 1978). The following pharmacokinetic parameters were evaluated for ketamine enantiomers: AUC0→∞ (the area under the curve to infinity), T1/2el (the half-life of the elimination phase), Kel (the rate constants of the elimination phase); MRT (mean residence time), ClB (body clearance), Vc (volume of

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distribution for the central compartment); Cmax (peak metabolite concentration) and Tmax (time of peak metabolite concentration). For S-norketamine and R-norketamine, noncompartmental analysis was used to determine AUC, Cmax and Tmax. The pharmacokinetic

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parameters were calculated for each dog.

Results

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Animals and anaesthesia

The test drugs were administered 178 ± 59.5 min (RS-KET group) and 178 ± 39.1 min (S-

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KET group) after general anaesthesia induction (period required for dog instrumentation). The sevoflurane administration was stopped 415 ± 21.7 and 412 ± 19.8 min after the general

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anesthesia induction in the RS-KET group and in the S-KET group, respectively. All dogs

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recovered well from anaesthesia and were fully awake within 67.3 ± 13.5 min in the RS-KET group and in 87.8 ± 28.5 min in the S-KET group after the end of sevoflurane administration.

treatment.

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One dog in the RS-KET showed transitory laryngospasm at recovery, which resolved without

Cardiopulmonary parameters, pH, blood gases and electrolytes The cardiovascular parameters (HR, SAP, MAP, DAP and CO) as well as the results of blood gas analysis are reported in Table 1. No statistical differences between the groups were detected at any time point. [Table 1 here] Ketamine enantiomers plasma concentrations

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ACCEPTED MANUSCRIPT The plasma concentrations of the ketamine enantiomers and its metabolites R-norketamine and S-norketamine were plotted against the time points for the RS-KET and S-KET treatments (Figures 1 and 2, A and B). No statistical differences were detected between the concentrations of R-ketamine and S-ketamine as well as between S-ketamine concentrations following RS-KET and S-KET. After administration of racemic ketamine, the area under the

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curve to infinity of the metabolites R-norketamine was significantly higher than the one of Snorketamine, reported in Table 2. No R-ketamine or R-norketamine was detected after the administration of S-ketamine. [Figures 1 and 2 here] Pharmacokinetic parameters

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A 2-compartment model best described the S-ketamine and R-ketamine plasma concentration following IV administration of a bolus of the racemic ketamine and S-ketamine. The

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calculated parameters are reported in Table 2. The apparent volume of distribution in the central compartment was 0.75 ± 0.37 L/kg and 0.68 ± 0.37 L/kg for S-ketamine in the RS-

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KET group and the S-KET group, respectively (p = 0.75). The distribution half-life of Sketamine was 1.39 ± 0.37 min in the RS-KET group and 1.03 ± 0.60 min in the S-KET group

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(p = 0.06). The elimination half-life of S-ketamine was 15.56 ± 6.98 min in the RS-KET

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group and 13.07 ± 6.19 min in the S-KET group (p = 0.15). The elimination half-life of Rketamine was 15.42 ± 5.46 min. The total body clearance of S-ketamine was 71.82 ±12.80

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mL/min/kg in the RS-KET group and 72.68 ± 14.72 mL/min/kg in the S-KET group (p = 0.72). [Table 2 here]

Discussion In the present study in healthy dogs under steady state sevoflurane anaesthesia, resulting in stable cardiopulmonary function, the pharmacokinetic parameters of S-ketamine after a single IV injection of the racemic drug or the S-enantiomer did not significantly differ. There was

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ACCEPTED MANUSCRIPT also no difference in R-ketamine kinetics in comparison to the S-ketamine. The cardiopulmonary function and relevant electrolyte concentrations did not differ between the groups either. These results are in agreement with studies performed in ponies under isoflurane anaesthesia receiving a bolus of S-ketamine at 50% dose of racemic ketamine (Larenza et al., 2007). In this study, no differences in the distribution of S-ketamine following

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its administration as a sole drug or as part of the racemic mixture could be detected. Also the distribution half-lives of the two studies are remarkably similar suggesting that horses and dogs show similar ketamine metabolism under steady state inhalant agent anaesthesia. It is known that there are differences in plasma levels of peripheral venous and arterial blood

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samples (Chiou et al, 1981; Tuk et al., 1997). It has been suggested that pharmacokinetic analysis for potent short-acting drugs is more accurate based on arterial plasma levels (Chiou,

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1987). Nevertheless, our data gained with venous samples is in agreement with data of Larenza et al. (2007) gained with arterial samples. Probably frequent sampling and the use of

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a central venous catheter in the present study, allowed accurate reflection of the decline curve. The delay in comparison to arterial sampling was minimised by the use of a central venous

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catheter. As it was not attempted to link effect of ketamine to plasma levels, the venous

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sampling regime of the current study was adequate to discover differences between S- and RS-ketamine pharmacokinetics, especially as cardiopulmonary function was constant and

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identical between groups. In both treatment groups the plasma concentrations of ketamine enantiomers followed a bi-exponential decline, with a rapid initial distribution followed by a slower elimination, as previously reported for ketamine in dogs and other species (Kaka and Hayton, 1989; Delatour et al., 1991; Henthorn et al., 1999; Roncada et al., 2003; Pypendop and Ilkiw, 2005). The previous studies of ketamine pharmacokinetics in dogs were not stereospecific (Kaka and Hayton, 1989; Roncada et al., 2003; Pypendop et al., 2005) but also showed that a 2-compartment model best described the disposition of ketamine. Distribution

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ACCEPTED MANUSCRIPT half-lives and clearance rates were similar to ours, but elimination half-lives in our study were considerably shorter. This is probably a consequence of differences in co-medications, the number of samples and duration of sampling during the elimination phase in the different studies. In contrast to the present results, stereospecific studies in otherwise unpremedicated humans

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(Ihmsen et al., 1991) have reported a higher S-ketamine clearance in comparison to Rketamine, in particular when given as part of the racemate. Kharasch and Labroo (1992) hypothesized that an enzyme substrate competition between R-ketamine and S-ketamine could delay the N-demethylation of S-ketamine to S-norketamine, leading to a slower

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elimination of this enantiomer when given as part of the racemate. In vitro it was shown, that N-demethylation of S-ketamine by human liver microsomes is 20 % larger than that of R-

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ketamine (Karash et al., 1992). A study in Japanese volunteers (Yanagihara et al., 2003) on the other hand, showed no stereospecific difference in the pharmacokinetics of racemic

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ketamine. This difference to Ihmsen’s et al. study (1991) was attributed to variability between different human races.

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Looking at metabolite formation, a higher area under the curve of R-norketamine compared to

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S-norketamine (statistically significant) was observed after administration of the racemic mixture. This is not in agreement with the study in ponies (Larenza et al., 2007), where under

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isoflurane anaesthesia significantly lower AUC in R-norketamine compared to Snorketamine were found following racemic ketamine. The beagles in the present study were artificially ventilated, while Larenza et al. (2007) did not ventilate the ponies. Positivepressure ventilation interferes with cardiac function and venous return and has been shown to decrease systemic and hepato-splanchnic blood flow. During spontaneous breathing, the blood flow to the liver seems to be higher than during artificial ventilation (Jakob, 2010). We used sevoflurane, Larenza et al. (2007) isoflurane. The use of halogenated agents will have an

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ACCEPTED MANUSCRIPT influence on organ perfusion and as such also influences drug disposition and pharmacokinetics (Schwieger et al, 1991). Therefore, the three different inhalant anaesthetic agents might have further contributed to differences observed, but probably to similar extents. It seems likely that concerning formation of R- and S-norketamine there is a true speciesspecific difference between dogs and horses, but not between dogs and humans.

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Light sevoflurane anaesthesia was used in the present study in order to allow stress free instrumentation of the dogs and provide steady state conditions during the first hours of blood sampling. Henthorn et al. (1999) have also used a halogenated anaesthetic, namely halothane, to study stereoisomeric distribution of S- and R- ketamine to pulmonary tissue following

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administration of racemic ketamine. As there was a lack of stereoisomeric effect in pulmonary tissue distribution the authors concluded that distribution would be identical for other tissues

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as well – which is in agreement with the current study.

In conclusion, after racemic ketamine injection in dogs anesthetised with sevoflurane the

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pharmacokinetic profiles did not differ between S-ketamine and R-ketamine. Moreover, the PK parameters of S-ketamine were comparable when given alone or as part of racemic

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ketamine. Concerning metabolites, the only difference observed in the pharmacokinetic

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profiles was that the area under the curve of R-norketamine was statistically higher than the one of S-norketamine. Effects on cardiopulmonary function between S-ketamine (2 mg/kg)

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and racemic ketamine (4 mg/kg) were identical. The results of the present study suggest that, despite our initial hypothesis, the administration of S-ketamine alone at the dose rates tested might provide the same effects as the racemic mixture.

Conflict of interest

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ACCEPTED MANUSCRIPT The study was partly sponsored by Dr. E. Graeub AG, the manufacturer of the S- as well as racemic ketamine. The authors declare that there is no conflict of interest regarding the publication of this manuscript.

Acknowledgements

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The authors would like to thank Prof. W. Thorman, Regula Theurillat and Friederike Sandbaumhüter from the Clinical Pharmacology Laboratory Institute for Infectious Diseases of the University of Berne, Switzerland for the help with the analyses and the calculations of the pharmacokinetic data. This research did not receive any specific grant from funding

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agencies in the public, commercial, or not-for-profit sectors.

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Linear and Nonlinear Mixed Effects Models. R package version 3.1-102 [computer software].

Pypendop, B.H., Ilkiw, J.E., 2005. Pharmacokinetics of ketamine and its metabolite, norketamine, after intravenous administration of a bolus of ketamine to isoflurane-

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anesthetized dogs. Am. J. Vet. Res. 66, 2034–2038.

R Development Core Team, 2011. R: A language and environment for statistical computing.

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R Foundation for Statistical Computing, Vienna, Austria. ISBN 3-900051-07-0. URL

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http://www.R-project.org/.

Roncada, P., Zaghini, A., Riciputi, C., Romagnoli, N., Spadari, A., 2003. Kinetics of

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ketamine plasma and urine metabolite levels following intravenous administration in the dog.

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Vet. Res. Commun. 27(S1), 433–436.

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Schmidt, A., Oye, I., Akeson, J., 2005. Cerebral physiological responses to bolus injection of racemic, S(+)- or R(-)-ketamine in the pig. Acta Anaesth. Scand. 49, 1436–1442.

Schwartz, G., 1978. Estimating the dimension of a model. Ann. Stat. 6, 461–464.

Schwieger, I.M., Szlam, F., Hug, C.C., 1991. The pharmacokinetics and pharmacodynamics of ketamine in dogs anesthetized with enflurane. J. Pharmacokinet. Biop. 19, 149–156.

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Sergi, M., Bafile, E., Compagnone, D., Curini, R., D’Ascenzo, G., Romolo, F.S., 2009. Multiclass analysis of illicit drugs in plasma and oral fluids by LC-MS/MS. Anal. Bioanal. Chem. 393, 709–718.

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Tuk, B, Danhof, M., Mandema, J.W., 1997. The impact on arteriovenous concentration differences on pharmacodynamic parameter estimates. J. Pharmacokinet. Biop. 25, 39–62.

White, P.F., Marietta, M.P., Pudwill, C.R., Way, M.L., Trevor, A.J., 1976. Effects of

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halothane anesthesia on the biodisposition of ketamine in rats. J. Pharmacol. Exp. Ther. 196,

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Yamaoka, K., Nakagawa, T., Uno, T., 1978. Application of Akaike’s information criterion

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(AIC) in the evaluation of linear pharmacokinetic equations. J. Pharmacokinet. Biop. 6, 165–

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175.

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ACCEPTED MANUSCRIPT Table 1 Heart rate (HR), arterial blood pressures (SAP, MAP, DAP), cardiac output (CO), respiratory rate (RR), temperature (T), pH, arterial CO2 pressure (PaCO2), arterial oxygen pressure (PaO2), hemoglobin and electrolyte concentrations before (0 min) and after ketamine (RS-KET) and

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S-ketamine (S-KET) single bolus administration to six dogs. Data are reported as mean ± standard deviation over time (min).

I R

SC

RS-KET Time (min)

0

5

15

30

45

60

120

180

240

HR [beats/min]

113±17.9

112 ±21.2

116±18.2

120±21.1

120±22.1

127±23.2

121±9.9

118±10.0

115±15.2

SAP [mmHg]

107±8.9

97±21.8

109±21.1

116±20.5

116±18.3

115±19.8

116±16.0

116±20.9

106±9.1

DAP [mmHg]

73±8.0

63±17.0

70±17.0

75±16.7

74±14.7

76±17.4

77±12.0

74±12.6

68±7.5

MAP [mmHg]

A M

U N

5

15

30

45

60

120

180

240

116±14.0

121±10.9

125±8.4

130±8.5

136±11.9

133±4.6

127±14.5

120±20.7

126±24.8

115±27.7

108±25.8

114±17.7

112±15.0

120±20.3

115±20.0

108±20.4

108±19.5

109±19.3

76±16.0

71±14.7

78±13.4

76±9.2

82±12.3

77±7.5

71±11.5

70±11.2

70±9.7

83±7.7

76±19.1

83±19.3

91±19.3

90±16.1

91±18.4

92±13.5

85±7.3

90±19.2

85±17.4

92±14.1

89±11.6

95±14.7

91±9.7

85±15.2

84±14.5

83±11.3

2.93±0.52

2.79±0.71

3.01±0.66

3.15±0.6

3.09±0.56

3.13±0.56

2.95±0.36

3.25±0.73

2.93±0.39

2.55±0.47

2.67±0.52

3.01±0.47

2.92±0.38

3.15±0.44

2.98±0.51

2.79±0.60

2.81±0.57

2.88±0.6

RR (breaths/min)

22±11.0

17±11.0

19±11.4

20±9.4

20±9.8

23±3.7

17±9.9

18±11.1

14±9.0

17±6.8

16±7.3

16±7.2

19±10.8

19±9.1

18±8.0

18±10.9

17±11.1

21±12.3

T (°C)

37.6±0.4

37.7±0.2

37.7±0.2

37.7±0.3

37.7±0.2

37.7±0.1

37.8±0.3

37.7±0.1

37.7±0.3

37.6±0.4

37.5±0.2

37.5±0.2

37.6±0.3

37.7±0.2

37.7±0.3

37.7±0.2

37.7±0.4

37.8±0.3

pH

7.37±0.01

7.35±0.03

7.34±0.03

7.36±0.03

7.35±0.03

7.37±0.1

7.38±0.03

7.38±0.03

7.41±0.1

7.37±0.04

7.35±0.03

7.34±0.02

7.34±0.03

7.33±0.04

7.34±0.1

7.37±0.03

7.36±0.03

7.39±0.03

33.7±2.7

34.4±3.7

36.0±4.6

33.2±2.5

31.9±3.0

32.7±2.4

32.1±2.0

30.7±4.4

31.7±3.7

33.6±2.3

34.2±1.9

34.6±2.4

34.8±4.3

34.2±5.2

32.9±2.6

30.4±3.00

191.0±16.4 175.8±20.4 185.5±8.1 189.1±11.1 167.8±24.3 193.5±25.1 189.6±7.1

185.6±25

CO [L/min]

PaCO2 [mmHg] PaO2 [mmHg]

9.1±1.2

8.4±1.6

8.6±1.5

8.9±1.4

K [mmol/L]

3.9±0.3

3.7±0.3

3.7±0.2

3.5±0.3

Na [mmol/L]

145.0±3.7

144.5±3.3

143.3±3.5

Ca [mmol/L]

1.3±0.02

1.4±0.03

1.3±0.03

Cl [mmol/L]

109.0±3.5

108.5±3.9

Lactate [mmol/L]

2.5±0.4

Glucose [mmol/L]

5.2±0.5

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Hemoglobin [g/dL]

34.6±1.4

90±15.6

0

S-KET

171.1±25.7 184.0±22.8 166.8±37.7

191.8±7.4 191.0±8.60 181.0±21.0 170.1±24.4

33.1±4.4 187.5±15

186.8±11.3 177.1±21.8

8.5±2.5

9.1±1.5

8.8±1.4

9.3±1.3

8.8±1.8

9.9±0.4

9.5±0.8

9.4±0.4

9.3±0.5

9.6±0.5

9.6±0.2

9.9±1.0

9.9±1.5

9.1±2.2

3.5±0.1

3.5±0.1

3.9±0.2

3.9±0.2

3.8±0.2

3.7±0.3

3.6±0.5

3.4±0.4

3.4±0.3

3.4±0.4

3.4±0.5

3.9±0.5

4.0±0.6

3.9±0.5

143.1±3.9

143.3±3.8

142.5±3.6

142.8±1.5

141.5±2.4

141.8±1.2

145.1±1.8

144.1±2.1

143.0±2.4

144.5±5.0

142.3±2.1

140.3±3.6

140.3±3.1

139.8±4.2

139.1±5.2

1.3±0.1

1.4±0.04

1.3±0.04

1.3±0.04

1.3±0.04

1.3±0.06

1.4±0.1

1.4±0.06

1.4±0.1

1.4±0.1

1.3±0.1

1.3±0.1

1.3±0.1

1.4±0.2

1.3±0.1

108.0±3.9

108.3±4.3

107.0±3.7

107.3±3.7

106.6±2.4

105.8±2.8

105.3±3.6

109.1±2.9

108.6±3.5

108.0±3.2

107.3±3.6

107.0±4.0

106.3±3.3

104.5±3.5

104.6±4.4

104.6±4.9

2.1±1.1

2.3±0.7

2.4±0.6

2.1±0.5

2.4±0.5

1.9±0.5

1.7±0.3

1.9±0.6

2.6±0.6

2.5±0.5

2.4±0.7

2.4±0.7

2.2±0.8

2.8±1.9

1.6±0.3

1.8±0.6

1.9±0.6

6.4±1.0

7.7±1.9

7.5±1.2

8.3±2.1

8.7±1.8

5.1±0.6

5.1±0.7

5.1±0.4

6.1±1.7

6.6±1.1

8.7±1.0

8.4±1.2

8.5±1.1

7.8±1.5

4.9±1.0

5.2±0.7

5.0±0.5

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ACCEPTED MANUSCRIPT Table 2 Mean ± standard deviation of pharmacokinetic variables for ketamine and norketamine enantiomers in plasma samples obtained from 6 dogs that received an intravenous bolus of Sketamine (S-KET) or racemic ketamine (RS-KET). AUC0→∞ (area under the curve to infinity); T1/2dis (half-life of the distribution phase); T1/2el (half-life of the elimination phase);

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Kel (rate constants of the elimination phase); MRT (mean residence time); ClB (body clearance); Vc (volume of distribution for the central compartment); Cmax (peak concentration); Tmax (time of peak concentration). a

P<0.05 between the two treatment groups; b P<0.05 between enantiomers within the RS-

S-KET

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Group

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KET group.

RS-KET

S-Ketamine

S-Ketamine

R-Ketamine

AUC0→∞ (µg min/mL)

28.48±5.74

28.64±5.41

30.27±4.84

T1/2dis (min)

1.03±0.60

1.39±0.37

1.25±0.15

T1/2el (min)

13.07±6.19

15.56±6.98

15.42±5.46

Kel (1/min)

0.06±0.02

0.05±0.03

0.05±0.05

MRT (min)

13.16±4.68

16.83±7.48

17.60±6.60

72.68±14.72

71.82±12.80

67.48±10.68

0.68±0.37

0.75±0.37

0.75±0.33

S-Norketamine

S-Norketamine

R-Norketamine

31.74±22.66a

30.11±14.12b

39.25±12.00a,b

0.37±0.14

0.40±0.13

0.37±0.12

9.17±2.04

11.67±4.08

11.67±4.08

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Compound

ClB (mL/min/kg)

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Vc (L/kg)

Cmax (µg/mL) Tmax (min)

AC

AUC0→∞ (µg min/mL)

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ACCEPTED MANUSCRIPT Figure legends

Figure 1. Mean plasma concentrations of R-ketamine and S-ketamine in the RS-KET group (A) and S-ketamine in the S-KET group (B), after administration of racemic ketamine 4 mg/kg or S-ketamine 2 mg/kg, respectively, to 6 dogs anaesthetized with sevoflurane in

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oxygen.

Figure 2. Mean plasma concentrations of R-norketamine and S-norketamine in the RS-KET group (A) and S-norketamine in the S-KET group (B), after administration of racemic

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ketamine 4 mg/kg or S-ketamine 2 mg/kg, respectively, to 6 dogs anaesthetized with

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sevoflurane in oxygen.

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Fig. 1

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Fig. 2

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ACCEPTED MANUSCRIPT Highlights  In human and horses the pharmacokinetic profiles of racemic and S-ketamine are different  PK data show no difference between racemic and S-ketamine in dog  Under 1.4 MAC sevoflurane anaesthesia in ventilated dogs following S-ketamine at 50%

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dose or racemate, cardiopulmonary parameters are identical

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