In Vitro Ketamine CYP3A Mediated Metabolism Study using Mammalian Liver S9 Fractions, cDNA Expressed Enzymes and Liquid Chromatography Tandem Mass Spectrometry

Ketamine is widely used in medicine in combination with several benzodiazepines including midazolam. The objectives of this study were to develop a novel HPLC-MS/SRM method capable of quantifying ketamine and norketamine using an isotopic dilution strategy in biological matrices and study the formation of norketamine, the principal metabolite of ketamine with and without the presence of midazolam, a well-known CYP3A substrate. The chromatographic separation was achieved using a Thermo Betasil Phenyl 100 x 2 mm column combined with an isocratic mobile phase composed of acetonitrile, methanol, water and formic acid (60:20:20:0.4) at a flow rate of 300 μL/min. The mass spectrometer was operating in selected reaction monitoring mode and the analytical range was set at 0.05–50 μM. The precision (%CV) and accuracy (%NOM) observed were ranging from 3.9–7.8 and 95.9.2–111.1% respectively. The initial rate of formation of norketamine was determined using various ketamine concentration and Km values of 18.4 μM, 13.8 μM and 30.8 μM for rat, dog and human liver S9 fractions were observed respectively. The metabolic stability of ketamine on liver S9 fractions was significantly higher in human (T1/2 = 159.4 min) compared with rat (T1/2 = 12.6 min) and dog (T1/2 = 7.3 min) liver S9 fractions. Moreover significantly lower IC50 and Ki values observed in human compared with rat and dog liver S9 fractions. Experiments with cDNA expressed CYP3A enzymes showed the formation of norketamine is mediated by CYP3A but results suggest an important contribution from others isoenzymes, most likely CYP2C particularly in rat.

Pretreatment Of Agricultural Waste With Pleurotus Sp. For Ethanol Production

Bioethanol is a liquid fuel obtained from fermentation of sugar/starch crops. Lignocellulosic biomass being less expensive is considered a future alternative for the food crops. One of the main challenges for the use of lignocellulosics is the development of an efficient pre-treatment process. Pretreatments are classified into three - physical, chemical, and biological pretreatment. Chemical process has not been proven suitable so far, due to high costs and production of undesired by-products. Biologically, hydrolysis can be enhanced by microbial or enzymatic pretreatment. Studies show that the edible mushrooms of Pleurotus sp. produce several extracellular enzymes which reduce the structural and chemical complexity of fibre. In the present study, P. ostreatus and P. eous were cultivated on paddy straw. Spent substrate left after mushroom cultivation was powdered and used for ethanol production. Saccharomyces sp. was used for fermentation studies. Untreated paddy straw was used as control. Production of ethanol from P. ostreatus substrate was 5.5 times more when compared to untreated paddy straw, while the spent substrate of P. eous gave 5 times increase in ethanol yield. Assays showed the presence of several extracellular enzymes in the spent substrate of both species, which together contributed to the increase in ethanol yield

Impacto en la calidad de la Inducción anestésica con premedicación en pacientes pediatricos

La inducción anestésica en niños es uno de los mayores retos para los anestesiólogos ya que es la fase más estresante del período perioperatorio para el paciente pediátrico. Existen diferentes intervenciones para minimizar la ansiedad perioperatoria y aumentar la cooperación del paciente pediátrico con la inducción anestésica. Entre las intervenciones exitosas la premedicación farmacológica con midazolam ha mostrado grandes beneficios en pacientes pediátricos. Metodología: Se realizó un estudio de casos y controles en pacientes pediátricos llevados a cirugía en la Fundación Cardioinfantil entre 2011-2014. Por medio de muestreo aleatorio por conveniencia se tomaron como casos pacientes con premedicación y controles pacientes sin premedicación. El éxito en la inducción se midió por medio de la escala ICC, usada a nivel mundial. Resultados: El promedio de edad fue 4.9 σ 3.01 años para los casos y 5.02σ3.2 años para controles, presentaron la misma distribución por género, 40.6% femenino, 59.3% masculino. El éxito de la inducción anestésica con midazolam mostró resultados significativos (OR 7.3 IC95% 4.3 – 12.5 p0,000), en hombres (OR 9.44 IC95%4.5 – 19.8 p0,000), en menores de 5 años (OR 10.33 IC95% 5.07 – 21.04 p0,000), en pacientes con antecedentes quirúrgicos (OR 12.2 IC95% 5.28 – 27.8 p0.000) o anestesias previas (OR 7.9 IC95% 4.4 – 14.4 p0,000). Discusión: El midazolam como agente farmacológico usado para premedicación en pacientes pediátricos presenta resultados exitosos contundentes, por lo cual debe usarse en todos los casos.

What GPs need to know about palliative-care drugs: ketamine

This is the first in a short series of articles that focus on what GPs should consider when monitoring and prescribing specialist-initiated palliative-care drugs. This first article summarises the key issues for patients receiving ketamine.

Ketamine, propofol, and the EEG: A neural field analysis of HCN1-mediated interactions

Ketamine and propofol are two well-known, powerful anesthetic agents, yet at first sight this appears to be their only commonality. Ketamine is a dissociative anesthetic agent, whose main mechanism of action is considered to be N-methyl-D-aspartate (NMDA) antagonism; whereas propofol is a general anesthetic agent, which is assumed to primarily potentiate currents gated by γ-aminobutyric acid type A (GABAA) receptors. However, several experimental observations suggest a closer relationship. First, the effect of ketamine on the electroencephalogram (EEG) is markedly changed in the presence of propofol: on its own ketamine increases θ (4–8 Hz) and decreases α (8–13 Hz) oscillations, whereas ketamine induces a significant shift to beta band frequencies (13–30 Hz) in the presence of propofol. Second, both ketamine and propofol cause inhibition of the inward pacemaker current Ih, by binding to the corresponding hyperpolarization-activated cyclic nucleotide-gated potassium channel 1 (HCN1) subunit. The resulting effect is a hyperpolarization of the neuron’s resting membrane potential. Third, the ability of both ketamine and propofol to induce hypnosis is reduced in HCN1-knockout mice. Here we show that one can theoretically understand the observed spectral changes of the EEG based on HCN1-mediated hyperpolarizations alone, without involving the supposed main mechanisms of action of these drugs through NMDA and GABAA, respectively. On the basis of our successful EEG model we conclude that ketamine and propofol should be antagonistic to each other in their interaction at HCN1 subunits. Such a prediction is in accord with the results of clinical experiment in which it is found that ketamine and propofol interact in an infra-additive manner with respect to the endpoints of hypnosis and immobility.

Effect of enzymatic pretreatment and increasing the organic loading rate of lipid-rich wastewater treated in a hybrid UASB reactor

This study aimed at evaluating the effect of increasing organic loading rates and of enzyme pretreatment on the stability and efficiency of a hybrid upflow anaerobic sludge blanket reactor (UASBh) treating dairy effluent. The UASBh was submitted to the following average organic loading rates (OLR) 0.98 Kg.m(-3).d(-1), 4.58 Kg.m(-3).d(-1), 8.89 Kg.m(-3).d(-1) and 15.73 Kg.m(-3).d(-1), and with the higher value, the reactor was fed with effluent with and without an enzymatic pretreatment to hydrolyze fats. The hydraulic detention time was 24 h, and the temperature was 30 +/- 2 degrees C. The reactor was equipped with a superior foam bed and showed good efficiency and stability until an OLR of 8.89 Kg.m(-3).d(-1). The foam bed was efficient for solid retention and residual volatile acid concentration consumption. The enzymatic pretreatment did not contribute to the process stability, propitiating loss in both biomass and system efficiency. Specific methanogenic activity tests indicated the presence of inhibition after the sludge had been submitted to the pretreated effluent It was concluded that continuous exposure to the hydrolysis products or to the enzyme caused a dramatic drop in the efficiency and stability of the process, and the single exposure of the biomass to this condition did not inhibit methane formation. (C) 2011 Elsevier B.V. All rights reserved.

Relation of pretreatment sequence diversity in NS5A region of HCV genotype 1 with immune response between pegylated-INF/ribavirin therapy outcomes

Hepatitis C virus (HCV) infection frequently persists despite substantial virus-specific immune responses and the combination of pegylated interferon (INF)-alpha and ribavirin therapy. Major histocompatibility complex class I restricted CD8+ T cells are responsible for the control of viraemia in HCV infection, and several studies suggest protection against viral infection associated with specific HLAs. The reason for low rates of sustained viral response (SVR) in HCV patients remains unknown. Escape mutations in response to cytotoxic T lymphocyte are widely described; however, its influence in the treatment outcome is ill understood. Here, we investigate the differences in CD8 epitopes frequencies from the Los Alamos database between groups of patients that showed distinct response to pegylated alpha-INF with ribavirin therapy and test evidence of natural selection on the virus in those who failed treatment, using five maximum likelihood evolutionary models from PAML package. The group of sustained virological responders showed three epitopes with frequencies higher than Non-responders group, all had statistical support, and we observed evidence of selection pressure in the last group. No escape mutation was observed. Interestingly, the epitope VLSDFKTWL was 100% conserved in SVR group. These results suggest that the response to treatment can be explained by the increase in immune pressure, induced by interferon therapy, and the presence of those epitopes may represent an important factor in determining the outcome of therapy.

Ketamine alters oscillatory coupling in the hoppocampus

Recent studies show that higher order oscillatory interactions such as cross-frequency coupling are important for brain functions that are impaired in schizophrenia, including perception, attention and memory. Here we investigated the dynamics of oscillatory coupling in the hippocampus of awake rats upon NMDA receptor blockade by ketamine, a pharmacological model of schizophrenia. Ketamine (25, 50 and 75 mg/kg i.p.) increased gamma and high-frequency oscillations (HFO) in all depths of the CA1-dentate axis, while theta power changes depended on anatomical location and were independent of a transient increase of delta oscillations. Phase coherence of gamma and HFO increased across hippocampal layers. Phase-amplitude coupling between theta and fast oscillations was markedly altered in a dose-dependent manner: ketamine increased hippocampal theta-HFO coupling at all doses, while theta-gamma coupling increased at the lowest dose and was disrupted at the highest dose. Our results demonstrate that ketamine alters network interactions that underlie cognitively relevant theta-gamma coupling.

Pre-emptive epidural ketamine or S(+)-ketamine in post-incisional pain in dogs: A comparative study

Objective-To compare the pre-emptive analgesic effects of epidural ketamine or S(+)-ketamine on post-incisional hyperalgesia.Study Design-Prospective randomized study.Animals-Twenty-four mongrel dogs (1-5 years, weighing 11.9 +/- 1.8 kg).Methods-Dogs were anesthetized with propofol (5 mg/kg intravenously) and a lumbosacral epidural catheter was placed. Dogs were randomly allocated to 3 groups, each with 8 dogs. The control group (CG) was administered saline solution (0.3 mL/kg); the ketamine group (KG) ketamine (0.6 mg/kg); and the S(+)-ketamine group (SG) S(+)-ketamine (0.6 mg/kg). The final volume was adjusted to 0.3 mL/kg in all groups. Five minutes after the epidural injection a surgical incision was made in the common pad of the right hind limb and was immediately closed with simple interrupted nylon suture. Respiratory (RR) and heart (HR) rates, rectal temperature (7, sedation (S), lameness score, and mechanical nociceptive threshold by von Frey filaments were evaluated before the propofol anesthesia and at 15, 30, 45, 60, 75, and 90 minutes and then at 2, 4, 6, 8, 12, and 24 hours after epidural injection.Results-There were no differences in RR, HR, T, or S between groups. Motor blockade of the hind limbs was observed during 20 +/- 3.6 minutes in KG and during 30.6 +/- 7.5 minutes in SG (mean SD). Mechanical force applied to obtain an aversive response was higher from 45 minutes to 12 hours in KG and from 60 to 90 minutes in SG, when compared with CG.Conclusions-Pre-emptive epidural ketamine induced no alterations in RR and FIR, and reduced post-incisional hyperalgesia for a longer time than did S(+) ketamine.Clinical Relevance-Although anesthetic and analgesic potency of S(+) ketamine is twice that of ketamine, the racemic form is seemingly better for post-incisional hyperalgesia. (C) Copyright 2004 by the American College of Veterinary Surgeons.

Continuous infusion of ketamine in hypovolemic dogs anesthetized with desflurane

Objective: To evaluate the cardiorespiratory effects of continuous infusion of ketamine in hypovolemic dogs anesthetized with desflurane.Design: A prospective experimental study.Animals: Twelve mixed breed dogs allocated into 2 groups: saline (n=6) and ketamine (n=6).Interventions: After obtaining baseline measurements (time [T] 0) in awake dogs, hypovolemia was induced by the removal of 40 mL of blood/kg over 30 minutes. Anesthesia was induced and maintained with desflurane (1.5 minimal alveolar concentration) and 30 minutes later (T75) a continuous intravenous (IV) infusion of saline or ketamine (100 mu g/kg/min) was initiated. Cardiorespiratory evaluations were obtained 15 minutes after hemorrhage (T45), 30 minutes after desflurane anesthesia, and immediately before initiating the infusion (T75), and 5 (T80), 15 (T90), 30 (T105) and 45 (T120) minutes after beginning the infusion.Measurements and main results: Hypovolemia (T45) reduced the arterial blood pressures (systolic arterial pressure, diastolic arterial pressure [DAP] and mean arterial pressure [MAP]), cardiac (CI) and systolic (SI) indexes, and mean pulmonary arterial pressure (PAP) in both groups. After 30 minutes of desflurane anesthesia (T75), an additional decrease of MAP in both groups was observed, heart rate was higher than T0 at T75, T80, T90 and T105 in saline-treated dogs only, and the CI was higher in the ketamine group than in the saline group at T75. Five minutes after starting the infusion (T80), respiratory rate (RR) was lower and the end-tidal CO(2) (ETCO(2)) was higher compared with values at T45 in ketamine-treated dogs. Mean values of ETCO(2) were higher in ketamine than in saline dogs between T75 and T120. The systemic vascular resistance index (SVRI) was decreased between T80 and T120 in ketamine when compared with T45.Conclusions: Continuous IV infusion of ketamine in hypovolemic dogs anesthetized with desflurane induced an increase in ETCO(2), but other cardiorespiratory alterations did not differ from those observed when the same concentration of desflurane was used as the sole anesthetic agent. However, this study did not evaluate the effectiveness of ketamine infusion in reducing desflurane dose requirements in hypovolemic dogs or the cardiorespiratory effects of ketamine-desflurane balanced anesthesia.

Relative potency of ketamine and S(+)-ketamine in dogs

The aim of this study was to determine the relative potency of racemic ketamine and S(+)-ketamine for the hypnotic effect and to evaluate the clinical anesthesia produced by equianesthetic doses of these two substances in dogs. One hundred and eight dogs were allocated in groups R2, R2.5, R3, R6, R9, R12, S2, S2.5, S3, S6, S9, and S12, to receive by intravenous route 2, 2.5, 3, 6, 9, and 12 mg/kg of ketamine or S(+)-ketamine, respectively. A dose-effect curve was drawn with the dose logarithm and the percentage of dogs that presented hypnosis in each group. The curve was used to obtain a linear regression, to determine the effective doses 100 and the potency relationship. In another experimental phase, eight groups of five dogs received 3, 6, 9 and 12 mg/kg of ketamine or S(+)-ketamine to evaluate the periods of latency, hypnosis, and total recovery. The times in which the dogs reached the sternal position, attempted to stand up for the first time, recovered the standing position, and started to walk were also recorded. The hypnotic dose for ketamine was 9.82 +/- 3.02 (6.86-16.5) mg/kg and for S(+)-ketamine was 7.76 +/- 2.17 (5.86-11.5) mg/kg. The time of hypnosis was longer in R3 and the first attempt to stand up occurred early in R6 when compared with S3 and S6 respectively. When R9 (100% of hypnosis with ketamine) and S6 [100% of hypnosis with S(+)-ketamine] were compared (1:1.5 ratio), the time to sternal position (12 +/- 2.5 and 20.2 +/- 5.6 min respectively) and the total recovery time (45 +/- 5.5 and 60.2 +/- 5.2 min respectively) were significantly shorter with S(+)-ketamine. It was concluded that the potency ratio between ketamine and S(+)-ketamine in dogs is smaller than the one reported in other species, and that the dose obtained after a reduction of 50%, as usually performed in humans, would not be enough to obtain equianesthetic effects in dogs.

The improvement of the anti-hyperalgesic effect of ketamine and of its isomers by the administration of ifenprodil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

The effects of anesthesia with a combination of intramuscular xylazine-diazepam-ketamine on heart rate, respiratory rate and cloacal temperature in roosters

Objective To examine the anesthetic effects of a xylazine-diazepam-ketamine (XDK) combination in roosters.Study design Prospective experimental trial.Animals Six healthy white Leghorn roosters weighing 2.03 +/- 0.08 kg.Methods Each rooster was pre-medicated with xylazine (3 mg kg(-1), IM) and after 15 minutes anesthesia was induced with a diazepam (4 mg kg(-1)) and ketamine (25 mg kg(-1)) combination injected into the pectoral muscles. Heart and respiratory rates were recorded before anesthesia and every 15 minutes after induction for 165 minutes. Cloacal temperature was measured before and 15 minutes after pre-medication and every 75 minutes thereafter during anesthesia. Quality of induction and recovery were scored subjectively; duration of loss of righting reflex, abolition of response to a painful stimulus and palpebral reflex were also recorded.Results Intramuscular injection of xylazine smoothly induced loss of the righting reflex within 3-4 minutes. Loss of response to a painful stimulus occurred at 13.1 +/- 2.9 minutes (mean +/- SD) after the administration of the D-K combination, and lasted for 63.0 +/- 5.3 minutes. Roosters anesthetized with this combination had a significant decrease in heart and respiratory rates and cloacal temperature. The recovery period lasted for up to 4 hours (227.5 +/- 15.4 minutes). Quality of recovery was satisfactory for four roosters but excitation was noted in two birds.Conclusions and clinical relevance The XDK combination was a useful anesthetic technique for typhlectomy in roosters. Nevertheless this drug combination should be used with caution and cardiopulmonary parameters monitored carefully. Under the conditions of this experiment it was associated with a decreased cloacal temperature and prolonged recoveries.

Comparison between S-ketamine and clonidine in caudal analgesia

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influence of S(+)-ketamine analgesia in renal intraoperative ischemia: histological study in rats

OBJETIVO: Investigar, em ratos, o efeito da S(+)cetamina na histologia renal após hemorragia intra-operatória. MÉTODOS: Vinte ratos Wistar machos, anestesiados com pentobarbital sódico, foram divididos, aleatoriamente, em 2 grupos: G1 - controle (n=10) e G2 - S(+)cetamina (n=10), submetidos a hemorragia de 30% da volemia em 3 momentos (10% a cada 10 min) 60 min após anestesia. G2 recebeu S(+)cetamina, 15 mg. kg-1, i.m., 5 min após anestesia e 55 min antes do 1.º momento de hemorragia (M1). Foram monitorizadas a pressão arterial média (PAM), temperatura retal (T) e freqüência cardíaca. Os animais foram sacrificados (M4) 30 min após o 3.º momento de hemorragia (M3). Os rins e o sangue das hemorragias foram utilizados para estudo histológico e do hematócrito (Ht). RESULTADOS: Houve redução significativa da PAM, T e Ht. Na histologia, G1=G2 na dilatação tubular, congestão e necrose. A soma total dos escores foi significativamente diferente e G2>G1. CONCLUSÃO: Hemorragia e hipotensão determinaram alterações na histologia renal. O aumento da concentração sangüínea de catecolaminas provavelmente determinou escores mais altos de alterações histológicas com o uso de S(+)cetamina.