Quantification of furosemide in camel plasma by high resolution mass spectrometry, application on pharmacokinetics.

We developed and validated a high-resolution liquid chromatography mass spectrometry method for the quantification of furosemide in camel plasma which was used for a pharmacokinetic study in camels. Plasma samples were extracted by supported liquid extraction and furosemide and internal standard (furosemide-D5) were separated on a an Agilent Zorbax XDB C18 column (50 × 2.1 mm i.d., 3.5 µm). Data was acquired in full-scan mode over a mass range of 200-400 Da in negative electrospray mode at a resolution of 70,000. Linear calibration curves were obtained over the concentration ranges of 1.0-10,000 ng/mL. The validated method was then successfully applied in evaluating the pharmacokinetics and metabolites of furosemide in six camels (Camelus dromedarus) and we were able to advice on a withdrawal time of furosemide treatment before racing.

Three-Dimensional Isotropic Fat-Suppressed Proton Density-Weighted MRI at 3 Tesla Using a T/R-Coil Can Replace Multiple Plane Two-Dimensional Sequences in Knee Imaging.

PURPOSE: To evaluate whether a 3 D proton density-weighted fat-suppressed sequence (PDwFS) of the knee is able to replace multiplanar 2D-PDwFS. MATERIALS AND METHODS: 52 patients (26 men, mean age: 41.9 ±â€Š14.5years) underwent magnetic resonance imaging (MRI) of the knee at 3.0 Tesla using a T/R-coil. The imaging protocol included 3 planes of 2D-PDwFS (acquisition time (AT): 6:40 min; voxel sizes: 0.40 - 0.63 × 0.44 - 0.89 × 3mm³) and a 3D-PDwFS (AT: 6:31 min; voxel size: 0.63 × 0.68 × 0.63mm³). Homogeneity of fat suppression (HFS), artifacts, and image sharpness (IS) were evaluated on a 5-point scale (5[excellent] - 1[non-diagnostic]). The sum served as a measure for the overall image quality (OIQ). Contrast ratios (CR) compared to popliteal muscle were calculated for the meniscus (MEN), anterior (ACL) and posterior cruciate ligaments (PCL). In 13 patients who underwent arthroscopic knee surgery, two radiologists evaluated the presence of meniscal, ligamental and cartilage lesions to estimate the sensitivity and specificity of lesion detection. RESULTS: The CR was higher in the ACL, PCL and MEN in 3D- PDwFS compared to 2D-PDwFS (p < 0.01 for ACL and PCL; p = 0.07 for MEN). Compared to 2 D images, the OIQ was rated higher in 3D-PDwFS images (p < 0.01) due to fewer artifacts and HFS despite the lower IS (p < 0.01). The sensitivity and specificity of lesion detection in 3D- and 2D-PDwFS were similar. CONCLUSION: Compared to standard multiplanar 2D-PDwFS knee imaging, isotropic high spatial resolution 3D-PDwFS of the knee at 3.0 T can be acquired with high image quality in a reasonable scan time. Multiplanar reformations in arbitrary planes may serve as an additional benefit of 3D-PDwFS. KEY POINTS: • 3D-PDwFS of the knee is acquired with high image quality• 3D-PDwFS can be achieved in only one measurement with a reasonable scan time• 3D-PDwFS with the advantage of multiplanar reformation may replace 2D-PD-weighted knee MRI Citation Format: • Homsi R, Gieseke J, Luetkens JA et al. Three-Dimensional Isotropic Fat-Suppressed Proton Density-Weighted MRI at 3 Tesla Using a T/R-Coil Can Replace Multiple Plane Two-Dimensional Sequences in Knee Imaging. Fortschr Röntgenstr 2016; 188: 949 - 956.

Pharmacokinetics and metabolism study of firocoxib in camels after intravenous administration by using high-resolution bench-top orbitrap mass spectrometry.

In this study, we developed a high-resolution liquid chromatography mass spectrometry method for the pharmacokinetic study of firocoxib followed by full method validation. Following a solid-phase extraction, the firocoxib and internal standard (celecoxib) were separated on an Agilent Zorbax ZDB C18 column (50 mm × 2.1 mm i.d., 3.5 µm) with a gradient elution using methanol and 0.1% aqueous formic acid. Data acquisition was performed at 25,000 resolution with the automatic gain set to 1,000,000 and the maximum injection time of 100 ms. Data were acquired in full-scan mode over a mass range of 100-550 Da in positive electrospray mode. Linear calibration curves were obtained over the concentration ranges of 0.5-200 ng/mL and no interfering peaks were detected at the retention time of firocoxib and internal standard in blank camel plasma samples. The mean extraction recoveries of firocoxib at three concentrations of 5, 25 and 75 ng/mL ranged from 92 to 104%. Coefficient of variation of intra-day and inter-day precision were both <10%. The accuracy of the method ranged from 95 to 107%. The validated method was then successfully applied in evaluating the pharmacokinetics and metabolism of firocoxib in camels (Camelus dromedarus) (n=5) following intravenous (i.v.) administration of a dose of 0.1 mgkg/body weight. The results obtained (mean ± SD) were as follows: the terminal elimination half-life (t1/2ß) was 5.75 ± 2.26 h, and total body clearance (ClT) was 354.1 ± 82.6 mL/kg/h. The volume of distribution at steady state (VSS) was 2344.4 ± 238.7 mL/kg. One metabolite of firocoxib was tentatively identified as desalkyl firocoxib (m/z 283). Firocoxib could be detected in plasma 3-5 days following i.v. administration in camels using a sensitive liquid chromatography high-resolution orbitrap mass spectrometry method.

Human neuroblastoma cells with acquired resistance to the p53 activator RITA retain functional p53 and sensitivity to other p53 activating agents.

Adaptation of wild-type p53 expressing UKF-NB-3 cancer cells to the murine double minute 2 inhibitor nutlin-3 causes de novo p53 mutations at high frequency (13/20) and multi-drug resistance. Here, we show that the same cells respond very differently when adapted to RITA, a drug that, like nutlin-3, also disrupts the p53/Mdm2 interaction. All of the 11 UKF-NB-3 sub-lines adapted to RITA that we established retained functional wild-type p53 although RITA induced a substantial p53 response. Moreover, all RITA-adapted cell lines remained sensitive to nutlin-3, whereas only five out of 10 nutlin-3-adapted cell lines retained their sensitivity to RITA. In addition, repeated adaptation of the RITA-adapted sub-line UKF-NB-3(r)RITA(10 µM) to nutlin-3 resulted in p53 mutations. The RITA-adapted UKF-NB-3 sub-lines displayed no or less pronounced resistance to vincristine, cisplatin, and irradiation than nutlin-3-adapted UKF-NB-3 sub-lines. Furthermore, adaptation to RITA was associated with fewer changes at the expression level of antiapoptotic factors than observed with adaptation to nutlin-3. Transcriptomic analyses indicated the RITA-adapted sub-lines to be more similar at the gene expression level to the parental UKF-NB-3 cells than nutlin-3-adapted UKF-NB-3 sub-lines, which correlates with the observed chemotherapy and irradiation sensitivity phenotypes. In conclusion, RITA-adapted cells retain functional p53, remain sensitive to nutlin-3, and display a less pronounced resistance phenotype than nutlin-3-adapted cells.

The pharmacokinetics and metabolism of meloxicam in camels after intravenous administration.

The pharmacokinetics and metabolism of meloxicam was studied in camels (Camelus dromedarus) (n = 6) following intravenous (i.v.) administration of a dose of 0.6 mg·kg/body weight. The results obtained (mean ± SD) were as follows: the terminal elimination half-life (t(1/2ß) ) was 40.2 ± 16.8 h and total body clearance (Cl(T) ) was 1.94 ± 0.66 mL·kg/h. The volume of distribution at steady state (V(SS)) was 92.8 ± 13.7 mL/kg. One metabolite of meloxicam was tentatively identified as methylhydroxy meloxicam. Meloxicam and metabolite were excreted unconjugated in urine. Meloxicam could be detected in plasma 10 days following i.v. administration in camels using a sensitive liquid chromatography tandem mass spectrometry (LC/MS/MS) method.

Elimination and phase II metabolism of ethanol in camels after intravenous administration.

Ethanol elimination was studied in camels (n=8) after a single bolus intravenous dose of 0.1g/kg bodyweight (BW). Blood samples were then collected at set intervals. Ethanol and ethyl glucuronide (EtG) in blood were analysed by validated static headspace gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry (LC-MS) methods, respectively. Blood-ethanol concentration-time profiles were plotted for each camel and these were evaluated. A simple linear regression model was fitted to the selected data points and the slope of the fitted line was used to estimate the elimination rate, the distribution factor and turnover rate, which were 5.15 mg/dL blood/h, 0.55 L/kg and 0.028 g/h/kg, respectively. Blood EtG concentration-time profiles were also plotted for each camel. The elimination half-life of EtG, estimated by linear regression (using the values obtained after ethanol was completely eliminated) was 2.18 h. The theoretical initial blood concentration of EtG (C(0)), obtained by extrapolation to time zero was 23.4 µg/dL. The results will be useful in monitoring alcohol doping in camels using either parent drug or metabolite.

The disposition of theophylline in camels after intravenous administration.

The pharmacokinetics of theophylline were determined after an intravenous (i.v.) dose of 2.36 mg/kg in six camels and 4.72 mg/kg body weight in three camels. The data obtained (median and range) for the low and high dose, respectively, were as follows: the distribution half-lives (t1/2 alpha) were 1.37 (0.64-3.25) and 2.66 (0.83-3.5) h, the elimination half-lives (t1/2 beta) were 11.8 (8.25-14.9) and 10.4 (10.0-13.5) h, the steady state volumes of distribution (Vss) were 0.88 (0.62-1.54) and 0.76 (0.63-0.76) L/kg, volumes of the central compartment (Vc) were 0.41 (0.35-0.63) and 0.51 (0.36-0.52) L/kg, total body clearances (Clt) were 62.3 (39.4-97.0) and 50.2 (47.7-67.4) mL/h.kg body weight and renal clearance (Vr) for the low dose was 0.6 (0.42-0.96) mL/h.kg body weight. There was no significant difference in the pharmacokinetic parameters between the two doses. Theophylline protein binding at a concentration of 5 micrograms/mL was 32.2 +/- 3.3%. Caffeine was identified as a theophylline metabolite but its concentration in serum and urine was small. Based on the pharmacokinetic values obtained in this study, a dosage of 7.5 mg/kg body weight administered by i.v. injection at 12 h intervals can be recommended. This dosing regimen should achieve an average steady state serum concentration of 10 micrograms/mL with peak serum concentration not exceeding 15 micrograms/mL.

The activity of mixed function oxidases, estimated by in vivo antipyrine clearance, is similar in horses and camels.

The activity of hepatic mixed function oxidases was compared in horses and camels (Camelus dromedarius) by studying the pharmacokinetics of antipyrine in seven camels and five horses following intravenous administration of a single dose of antipyrine (25 mg/kg). The data obtained (mean +/- SEM and median in brackets) in camels and horses, respectively, were as follows: the elimination half-lives were 3.25 +/- 0.23 (3.19) and 3.09 +/- 0.25 (2.90) hr; the apparent volumes of distribution (area method) were 0.691 +/- 0.045 (0.648) and 0.642 +/- 0.034 (0.676) l/kg; the volumes of distribution at steady state were 0.659 +/- 0.040 (0.607) and 0.620 +/- 0.030 (0.653) l/kg; the volume of the central compartment of the two-compartment pharmacokinetic model were 0.386 +/- 0.0523 (0.349) and 0.298 +/- 0.05 (0.308) l/kg; total body clearances were 0.148 +/- 0.008 (0.158) and 0.145 +/- 0.007 (0.147) l/kg/hr; the areas under the curves to infinity were 171.0 +/- 9 (165) and 175 +/- 8.0 (170) micrograms.ml.hr. There was no statistical significance in any parameter between camels and horses which suggests that the activity of hepatic mixed function oxidases is similar in horses and camels.

Peptidyl carbamates as novel elastase inhibitors: structure-activity relationship studies.

Twenty-six novel peptidyl carbamates and thiocarbamates were synthesized and evaluated as elastase inhibitors. Eighteen compounds inhibited porcine pancreatic elastase, whereas only eleven of the newly synthesized compounds inhibited human leukocyte elastase. Neither of the other serine dependent proteases, trypsin or chymotrypsin, were affected by any of the active inhibitors. Structure-activity relationship studies indicated that inhibition was dependent on P1 and P'1 substitution as well as on the presence of the carbamate functionality. Placement of an isostere of valine at P1 and a 1-(phenyl mercaptotetrazole at P'1 resulted in the most active human leukocyte elastase inhibitor within this series of compounds (Ki - 3.0 x 10(-7) M).

Effects of structural variations on the rates of enzymatic and nonenzymatic hydrolysis of carbonate and carbamate esters.

The effect of structural variations on the rates of elastase-catalyzed hydrolysis of model carbonate and carbamate esters was studied using HPLC. It is shown that branching in the immediate vicinity of the carbonate or carbamate functionally results in decreased hydrolysis rates. Whereas aryl carbonates act as substrates for elastase, p-nitrophenyl butyl carbamate inhibits the enzyme. A novel method was developed for the entrapment and quantitation of 14CO2 produced upon hydrolysis of carbonyl-14C-labeled carbonate esters. This technique could be useful in studying the mechanism of enzymatic hydrolysis of this type of compound and has the potential of being adapted as a convenient method in the assessment of estrolytic activity of tissue homogenates.

Disposition of [14C]nonoxynol-9 after intravenous or vaginal administration to female Sprague-Dawley rats.

The disposition of nonoxynol-9 labeled with carbon-14 at the ethylene oxide units was studied following an iv or vaginal administration to female Sprague-Dawley rats. The results from the vaginal administration studies indicate 12.8% absorption of 14C radioactivity in 6.0 hr and 37.7% in 24.0 hr. Tissue distribution studies showed that the small and large intestines, including their contents, had the highest 14C activity by either route of administration. Radiomonitored HPLC of bile collected at 6.0 hr and urine at 6.0, 24.0, and 48.0 hr following an iv injection of [14C]nonoxynol-9 showed that the compound was completely metabolized in the body of the rat. The metabolites were primarily excreted in the feces and secondarily in the urine. Analysis of urinary metabolites containing the carbon-14 label, 6.0 hr following an iv dose, indicated the presence of highly polar neutral (53.27%) and acidic (39.23%) species.

Peptidyl carbamates incorporating amino acid isosteres as novel elastase inhibitors.

The design and synthesis of 13 novel peptidyl carbamates are described. When tested for inhibitory activity toward porcine pancreatic elastase, trypsin, and chymotrypsin, six compounds were found to specifically inhibit elastase without affecting the other two serine proteases. All the active inhibitors had an amino acid isostere at the P1 position. Kinetic studies indicated that the inhibition was competitive with Ki values ranging from 4.23 X 10(-5) to 2.4 X 10(-6) M. The degree of inhibition was found to be dependent on the specificity of the peptide chain for the extended subsites on the enzyme as well as on the nature of P1'. Preliminary work on one inhibitor indicates that the inhibition is reversible and proceeds via the rapid formation of a strong enzyme-inhibitor complex, followed by slow acylation of the serine residue on the active site of the enzyme. Peptidyl carbamates represent a novel class of elastase inhibitors.

Assessment of antiprotease activity of some carbamate derivatives.

Six carbamate derivatives were tested for inhibitory activity towards porcine pancreatic elastase, trypsin, and chymotrypsin. Only three esters that are isatoic anhydride derivatives were found to inhibit elastase competitively but nonspecifically. KI values for the best two inhibitors were determined from Dixon plots.

Drug Distribution and Biliary Excretion Pattern of a Cyclic Somatostatin Analog: A Comparison of (14)C Labeled Drug and a (131)I lodinated Drug Analog.

A cyclic somatostatin analog was compared to an iodinated analog of the same compound with respect to organ distribution and biliary excretion in the rat. The cyclic hexapeptide was radiolabeled with either (14)C or (131)I (tyrosine). Organ distribution of the iodinated compound as a function of time was nearly identical to that observed for the non-iodinated compound. Results indicated a rapid uptake by the liver and subsequent rapid excretion of the intact peptide in bile. Activity in other organs examined tended to fall off in a manner similar to the activity in blood with sequential samples. Because of the similarity in the in vivo behavior of the two compounds, the iodinated analog was deemed a suitable model for less invasive distribution studies, and was further examined in the dog using external gamma scintigraphy. In the unanesthetized dog the iodine activity was rapidly taken up by liver and collected in the gallbladder, thus exhibiting a similar rapid excretion pattern to that observed in the rat.

Peptidyl carbamate esters: a new class of specific elastase inhibitors.

A series of tripeptide carbamate esters were synthesized and tested for inhibitory activity towards porcine pancreatic elastase, trypsin and chymotrypsin. Two of these compounds specifically inhibited elastase [Ki = 4.25 x 10(-5)M and 3.00 x 10(-5)M], and exhibited no effect toward the other two serine proteases. These active tripeptidyl carbamates represent a novel class of specific elastase inhibitors.

[Trans-cerebral electric analgesia: reality, hypothesis or placebo? (author's transl)]. / Electro-analgésie transcérébrale: réalité, hypothèse ou placebo?

Sixty patients undergoing major digestive surgery received the same amount of drugs for induction of anesthesia; neuroleptic, morphinomimetic and pachycurares were used and calculated in mg/kg. A randomisation list permitted to define 45 patients who received in double blind (by means of a long electrical cord), an electrical current varying for each protocol; the control group (15 patients) did not receive an electrical current. Statistical study of the amount of drugs used for follow of anesthesia shows no real effect of electrical analgesia for digestive surgery.

[Very long duration anaesthesia (V. L. D. A.) considered by the surgeon (author's transl)]. / Les anesthésies de très longue durés (A. T. L. D.). Point de vue du chirurgien.

In the experience of the Universitary Surgical Clinic of Amiens, anaesthesia of more than 8 hours duration appears for 3 p. thousand of the total amount (9/3 000 on 1978). The authors distinguish between them "provided" very long duration operations and the "involuntary" ones (complex multitrauma, unexpected difficulties, etc.); that distinction is justified by many reasons among which one must consider mostly the team work setting up. The authors briefly report V. L. D. A. events in their own experience as in that reported in the medical literature. Also the practical difficulties resulting of V. L. D. A. in the intra-operative or post-operative periods are appraised. Then the authors propose some means able to reduce the number of V. L. D. A. and to assure a better proceeding of very long duration operations.

[Determination of alphaxalone in serum and urine by gas-liquid chromatography. Application in anaesthesiological pharmacology (author's transl)]. / Dosage de l'alphaxalone dans le sérum et les urines par chromatographie gaz-liquide. Application en pharmacologie anesthésiologique.

The authors propose a simple and reliable method for determination of alphaxalone [3 alpha-hydroxy(5 alpha)pregnane 11,20 dione] in serum and urine, during long-duration anaesthesia using Althesin, mixture of alphaxalone and alphadolone acetate [21 acetoxy, 3 alpha-hydroxy(5 alpha)pregnane 11,20 dione]. If assays of alphadolone appear difficult and without any precision, because of numerous interferences, the determination of alphaxalone in serum and urine seems interesting for pharmacokinetic and metabolic investigation. We confirm the quick rate of disappearance of alphaxalone from blood, associated with a very active metabolism in the liver. The results show that it exists a relation between the concentration of alphaxalone in blood and the value of anaesthesia judged on clinical and electrical criteria. We cannot however conclude that Althesin presents a cumulative effect, because we used important amounts of anaesthetic agent.