Experimental separation steps influence the protein content of corona around mesoporous silica nanoparticles.

In order to direct nanocarriers to their targets efficiently, we have to understand the interactions occurring at the nano-bio interface between nanocarriers and human proteins, which forms the layer called the corona. However, experiments aiming to identify and quantify the proteins in the corona, especially critical steps in the separation of nanoparticles from biological media may affect the corona composition. Here, we used nano-LC MS/MS to compare the protein corona contents obtained after using two different separation methods. We showed that applying centrifugation versus magnetization to isolate nanoparticles surrounded by a corona resulted in protein loss and a reshuffling of their respective abundances.

A pharmacokinetic study of 48-hour sevoflurane inhalation using a disposable delivery system (AnaConDa®) in ICU patients.

BACKGROUND: Little is known regarding sevoflurane kinetics and toxicity during long-term sedation of intensive care unit (ICU) patients using the AnaConDa® system. The objective of the present study was to establish a pharmacokinetic description of 48-h sevoflurane administration, and to estimate plasma concentrations of metabolites. METHODS: Forty-eight hour sedation with sevoflurane vaporized via an AnaConDa® device, with an end-tidal concentration objective of 1.5% (v/v), was initiated in 12 non-obese patients who did not have hepatic or renal failure but who required sedation for more than 48 h in our ICU. Plasma sevoflurane, hexafluoroisopropanol, and fluoride concentrations were determined over this time period and pharmacokinetic analysis was performed. RESULTS: The mean plasma concentration of sevoflurane was 76 mg/L at 24 h and 70 mg/L at 48 h. Wash-out of plasma sevoflurane correlated with a rapid decrease in the mean end-tidal sevoflurane level. The mean free plasma fraction of hexafluoroisopropanol never exceeded 8 mg/mL. The mean fluoride concentration was 0.8 µmol/L on day 0, 51.7 µmol/L on day 1, and 68.1 µmol/L on day 2 (P<0.0001). The distribution volume was 53 L, the elimination constant 2.9 h-1, the transfer constant from compartment 1 to compartment 2 (K1-2) 1.2 h-1, the K2-1 0.26 h-1, the half-life of elimination 3.78 h, and the total clearance 156 L/h. CONCLUSION: Following 48 hours of sedation using sevoflurane inhalation administered using an AnaConDa® delivery device, sevoflurane washout was rapid. Plasma fluoride levels accumulated over the study period without apparent nephrotoxicity.

[Stability and compatibility of acetaminophen, ketoprofen and amoxicillin in a fail-safe intravenous administration set]. / Stabilité et compatibilité du paracétamol, du kétoprofène et de l'amoxicilline administrés avec un perfuseur de sécurité

OBJECTIVES: Intravenous infusion takes an important place in the current therapy in hospitals and pharmaceutical firms keep improving their infusion medical devices, particularly with the development of an intravenous administration set with an automatic infusion stop. The aim of our study consists in an evaluation of the stability of acetaminophen, ketoprofen and amoxicillin during infusion and stasis of drugs for several hours in the dropper chamber and in the tube of this device. STUDY DESIGN: Analytical study. MATERIALS AND METHODS: Ten milligram per millilitre acetaminophen (acetaminophen 1 and acetaminophen 2), 1 mg/mL ketoprofen and 20 mg/mL amoxicillin were prepared and infused individually and successively with the intravenous administration set under ambient light and temperature. Collected samples of each drug during the infusion and after 24 hours of stasis in the medical device were monitored by a visual inspection, pH and osmolality assessment and chromatographic analysis of each drug with a validated stability-indicating method. RESULTS: Repeated individual perfusions of 10 mg/mL acetaminophen 1 or 1 mg/mL ketoprofen are possible through a fail-safe intravenous administration set, while repeated individual amoxicillin infusions are not because of the unstability of this drug. There is also no problem to infuse successively acetaminophen and ketoprofen through this medical device because these drugs are stable. However, we underlined an incompatibility between acetaminophen 2 and ketoprofen, which advises against the use of intravenous administration set for successive infusions of these two drugs. CONCLUSION: Despite the technical innovation of a fail-safe intravenous administration set, we have to stay aware of mixture consequences in intravenous infusion field.

Proton pump inhibitor administration via nasogastric tube in pediatric practice: comparative analysis with protocol optimization.

Proton pump inhibitors (PPIs) are largely prescribed to children because their efficacy and tolerance are now well-established. One disadvantage resides in the absence of liquid form which causes problems for their administration in nasogastric tubes. Indeed, the absence of use recommendations involves many misuses responsible for inefficiency and/or tube obstruction. We tried to evaluate if PPIs can be administered through pediatric nasogastric tubes. We administered four PPIs (Omeprazole, esomeprazole, lansoprazole and lansoprazole orally disintegrating tablet) through nasogastric tubes. For each PPI a study plan was drawn up to assess the influence of different variables: the volume of water to dissolve or put in suspension the PPIs (2ml or 5ml), the volume of tube flush-through water post-PPI administration (2ml, 5ml or 10ml), the length (50cm or 125cm) and the diameter (6 or 8 French) of the polyurethane tubes. For each assay an analysis of each active ingredient at the tube outlet by UV spectrometry was carried out. All 6 F tubes were obstructed by PPIs. Through 8 F tubes, we observed a mean recovery of active ingredient of 86.2% for lansoprazole orally disintegrating tablet, 36.9% for esomeprazole but only 7.1% for lansoprazole and 3.9% for omeprazole. It is disadvised using omeprazole and lansoprazole through 8 F nasogastric tubes because no condition ensures the transit of an efficient concentration of active ingredient. For esomeprazole, the best conditions of administration were a water volume of 5ml and a rinse volume of 5ml but only a half of the microgranules administered were recovered. The most satisfactory results were obtained with lansoprazole orally disintegrating tablet. A 5ml volume of water diluent for suspension and a 10ml volume of flush-through water made it possible to deliver the full lansoprazole dose administered.

Comparative study and optimisation of the administration mode of three proton pump inhibitors by nasogastric tube.

Patients in intensive care often develop stress-induced ulcers. As a preventive measure, proton pump inhibitors (PPIs) are administered by nasogastric tube. However, some PPIs can block the tube. The aim of this study was to compare the behaviour of three PPIs (omeprazole, lanzoprazole and esomeprazole) during the transit of the granules through the tube and to optimise their modes of administration. For each IPP, the experiment was designed to study the influence of four variables: the tube material (silicone or polyurethane), the solvent used to dilute the granules (water or apple juice), the mode of administration (in two or three doses) and the rinse volume (10 or 20 ml). We counted the granules before transit and at the tube outlet, and assayed the active drug ingredient by UV spectrometry. The assay showed complete transit of esomeprazole through the tube, but average losses of omeprazole and lanzoprazole of 39 and 33%, respectively, were observed. No significant improvement was obtained by the variables 'diluent' and 'mode of administration'. The variable 'rinse' had a significant influence. For lanzoprazole, a polyurethane tube allowed recovery of on average 86% of the active ingredient. Esomeprazole is thus the choice PPI for the treatment of patients by nasogastric tube. Using a polyurethane tube and a rinse volume of 20 ml, the administration of lanzoprazole by tube can be considered. Use of omeprazole is not recommended because none of the modes of administration tested ensured that a sufficient concentration of active ingredient reached the stomach.

Impact of deep freezing on the stability of 25 mg/ml vancomycin ophthalmic solutions.

For the treatment of certain eye infections, ophthalmic solutions 'laced' with 25 mg/ml vancomycin are sometimes prepared. Their physical and chemical stability and the maintenance of their sterility were studied after deep freezing at -20 +/- 2 degrees C and thawing, followed or not by refrigeration for 48 h at 4 +/- 2 degrees C. Physical and chemical analysis comprised visual inspection turbidity, determination of pH and osmolality, and assay of vancomycin by high performance liquid chromatography with ultraviolet detection. For microbiological analysis a 25 mg/ml vancomycin ophthalmic solution was filtered through two membranes and cultured on trypticase-soy and Sabouraud-glucose solid media. Any colonies were then counted. These physical, chemical and microbiological analyses demonstrated the stability of 25 mg/ml vancomycin ophthalmic solutions in 5% glucose deep frozen at -20 +/- 2 degrees C for 3 months. The vancomycin concentration varied by no more than 5% of the initial concentration, and no breakdown product was evidenced. Neither pH (mean=3.8 +/- 0.1) nor osmolality (mean=318.3 +/- 5.6 mOsm/kg) varied significantly, and remained compatible with intraocular administration. No particle or bacterial combination was found in the course of the study. The thawing procedure (at ambient temperature or under warm running water from a tap) did not modify the stability of the eye drops. Likewise, storage in a refrigerator for 48 h after thawing did not modify stability. The advantage of storing vancomycin 25 mg/ml ophthalmic solutions for 3 months in deep freeze is that a stock of chemically and microbiologically controlled preparations can be held ready for administration to patients, thereby allowing prompter dispensing, as the eye drops are not made up extemporaneously, while the improved control over production ensures that patients receive solutions of constant quality, as every batch prepared is systematically inspected.

Ca2+-myristoyl switch and membrane binding of chemically acylated neurocalcins.

Neurocalcin is a member of a novel family of neuronal calcium sensors that belongs to the superfamily of EF-hand Ca(2+)-binding proteins. Neurocalcin is myristoylated on its N-terminus in vivo and can associate with biological membranes in a calcium and myristoyl-dependent manner. This process known as "Ca(2+)-myristoyl switch" has been best described for the photoreceptor specific protein, recoverin, as well as for several other neuronal calcium sensors. Here, we used reversed micelles to chemically acylate nonmyristoylated neurocalcin at its N-terminus with fatty acids of different lengths (from C12 to C16). This approach allowed us to prepare neurocalcin derivatives in which a single fatty acid is selectively linked to the N-terminal glycine of the polypeptide chain through an amide bond. The membrane binding properties of the monoacylated neurocalcins were then examined by cosedimentation with phospholipid vesicles and direct binding to lipid monolayers by surface plasmon resonance spectroscopy (Biacore). Our results show that neurocalcins monoacylated with lauric, myristic, or palmitic acid were able to associate with membrane in a calcium-dependent manner. This indicates that the Ca(2+)-myristoyl switch can function with different lipid moieties and is not strictly restricted to myristate. The ability to modify at will the fatty acid linked to the N-terminal glycine should be useful to analyze the contribution of the fatty acid moiety to the biological function of this family of neuronal calcium sensors.

Study of hydrophobic interactions between acylated proteins and phospholipid bilayers using BIACORE.

Intracellular proteins of eukaryotic cells are frequently covalently modified by the addition of long chain fatty acids. These modifications are thought to allow otherwise soluble proteins to associate with membranes by lipid-lipid based hydrophobic interactions. The purpose of this work was to quantify the effect of acyl chain length on hydrophobic interactions between acylated proteins and phospholipid monolayers. The binding of an artificially acylated model protein to electrically neutral phospholipids was studied by surface plasmon resonance, using BIACORE. Kinetic rates for the binding of bovine pancreatic ribonuclease A (RNase A), monoacylated on its N-terminal lysine with fatty acids of 10, 12, 14, 16 or 18 carbon atoms, to phospholipids on hydrophobic sensor chips, were measured. Unlike unmodified ribonuclease, acylated RNase A bound to the phospholipids, and the association level increased with the acyl chain length to reach a maximum for C16. Reproducible kinetics were obtained which did not fit a 1:1 Langmuir model but rather a two-step binding profile.

Induction of a catabolic state in rats by dexamethasone: dose or time dependency?

BACKGROUND: Daily injections of dexamethasone (DEX) given to adult rats are a recognized but nonstandardized model of stress. The aim of this work was to establish a reproducible and accurate model of stress in adult rats by chronic injection of DEX in order to standardize it. For this purpose, the effect of the duration of treatment and the effect of DEX dose were tested. To help understand the mechanisms of the catabolic effect of DEX, the study was extended to the metabolism of glutamine (GLN). In experiment 1, 60 male Sprague-Dawley rats (3 months old) were divided into 8 groups of 6 rats: groups G3, G5, G7, and G9 received 1.50 mg/kg/d of DEX by intraperitoneal (i.p.) injection for 3, 5, 7, or 9 days, respectively. Groups G3PF, G5PF, G7PF, or G9PF were pair-fed to groups G3, G5, G7, or G9, respectively. Group AL (n = 12) was healthy rats fed ad libitum. RESULTS: In treated rats, nitrogen balance reached its lowest value at day 5. After 9 days treatment by DEX, the catabolic state was reduced. An increase in GLN-synthetase activity and a decrease in muscle GLN content were related to DEX per se not to DEX-induced anorexia. In experiment 2, 25 rats were divided into 5 groups of 5 animals. Groups G0.75, G1.50, and G2.50 received 0.75, 1.50, and 2.50 mg/kg/d, respectively, of DEX by i.p. injection for 5 days. Group PF was pair-fed to group G2.50 and group AL was control rats. RESULTS: DEX induced a decrease in nitrogen balance that was dose-independent. GLN-synthetase activity was increased maximally in gastrocnemius by 0.75 mg/kg. CONCLUSIONS: Five days of treatment by DEX and a dose of 0.75 mg/kg/d induced a marked catabolic state.

Compatibility of paclitaxel in 5% glucose solution with ECOFLAC low-density polyethylene containers-stability under different storage conditions.

The compatibility of paclitaxel with low-density polyethylene containers (ECOFLAC) was studied under different temperature and light conditions. Solutions of 0.4 and 1.2 mg/ml of paclitaxel in 5% glucose solution were prepared, put into ECOFLAC containers and stored: (i) at ambient temperature (20-25 degrees C) and in ambient light; (ii) at ambient temperature in the dark; and (iii) at +4 degrees C in the dark. Paclitaxel was assayed by high-performance liquid chromatography after visual inspection of the solutions. The results show that solutions of TAXOL in 5% glucose should not be stored for more than 5 days in glass or ECOFLAC containers because a whitish precipitate tends to form, lowering the paclitaxel concentration. The decrease in the paclitaxel concentration observed after chromatographic analysis ranged very widely (from 12 to 83% of the initial concentration). However solutions of TAXOL diluted in 5% glucose was stable for 5 days in ECOFLAC containers under all the storage conditions tested. These additive-free low-density polyethylene containers offer the advantage of not releasing DEHP into the paclitaxel solutions.

Kinetic impairment of nitrogen and muscle glutamine metabolisms in old glucocorticoid-treated rats.

Aged rats are more sensitive to injury, possibly through an impairment of nitrogen and glutamine (Gln) metabolisms mediated by glucocorticoids. We studied the metabolic kinetic response of adult and old rats during glucocorticoid treatment. The male Sprague-Dawley rats were 24 or 3 mo old. Both adult and old rats were divided into 7 groups. Groups labeled G3, G5, and G7 received, by intraperitoneal injection, 1.50 mg/kg of dexamethasone (Dex) for 3, 5, and 7 days, respectively. Groups labeled G3PF, G5PF, and G7PF were pair fed to the G3, G5, or G7 groups and were injected with an isovolumic solution of NaCl. One control group comprised healthy rats fed ad libitum. The response to aggression induced specifically by Dex (i.e., allowing for variations in pair-fed controls) appeared later in the aged rats (decrease in nitrogen balance from day 1 in adults but only from day 4 in old rats). The adult rats rapidly adapted to Dex treatment, whereas the catabolic state worsened until the end of treatment in the old rats. Gln homeostasis was not maintained in the aged rats; despite an early increase in muscular Gln synthetase activity, the Gln pool was depleted. These results suggest a kinetic impairment of both nitrogen and muscle Gln metabolisms in response to Dex with aging.

Monoacylation of ribonuclease A enables its transport across an in vitro model of the blood-brain barrier.

A major challenge in correcting disorders affecting the central nervous system is to induce blood-brain barrier (BBB) crossing of exogenous biological compounds such as proteins or specific nucleic acid sequences. Fatty acids, due to their high membrane affinity and low toxicity, are good potential candidates to promote this barrier crossing when covalently bound to proteins. In this paper, we report that regiospecific monoacylation of ribonuclease A (RNase A) enables its transport across an in vitro model of the BBB. Myristoylated, palmitoylated and stearoylated RNases A were prepared using reversed micelles as microreactors. All the purified acylated RNases A kept their original enzymatic activity. A single fatty acid moiety was linked to RNase A through the alpha-amino group of its N-terminal lysine as shown by powerful analytical techniques. The ability of monoacylated RNases A to cross an in vitro model of the BBB is strictly dependent on the acyl chain length, which must be at least 16 carbon atoms long.

Self-evolving microstructured systems upon enzymatic catalysis.

The consequences of cell microstructuration on enzyme functions is discussed in the framework of self-evolving microstructured systems. Molecular assemblies of amphiphiles or lipids are spontaneously formed by self-organisation. Among these different structures, reversed micelles, liquid crystalline mesophases and vesicles are hosts for enzymatic reaction studies. Inside a living cell, phospholipid metabolism is responsible for membrane structural modifications; the catalytic behaviour of lipolytic enzymes, mainly phospholipase (PL) A2, is described in relation with structural aspects of biological membranes. The implication in cellular regulation events of PLC and PLD is discussed in relation with the role of their reaction products as second messengers in membrane fusion processes. The in vitro synthesis of dialkyl phosphatidylcholines, via the enzymatic 'salvage pathway' which leads to the formation of vesicles upon phospholipid formation, is considered in relation with autopoiesis. More recent studies on self-evolving systems based on enzyme-surfactants reactions are detailed. The interactions between amphiphilic aggregates and enzymes allow to explore the OG/octanol/water phase diagram. Enzymatic formation of dipalmitoylphosphatidylcholine (DPPC) liposomes and non-ionic surfactant vesicles (NSV), starting from mixed micelles or open structures, finally sets an example of a biomimetic self-evolving system.

Compatibility of tropisetron with glass and plastics. Stability under different storage conditions.

The compatibility of tropisetron (pure undiluted 1 mg mL(-1) and diluted with 5% glucose or 0.9% NaCl (saline)) with glass, poly(vinyl chloride), polypropylene or polyethylene containers has been studied over a period of two weeks. The drug solutions were exposed to different light and temperature conditions. Tropisetron was assayed by high-performance liquid chromatography. The results show that undiluted tropisetron is stable in polypropylene syringes for two weeks under all the storage conditions tested (daylight at room temperature, dark at room temperature, refrigerator at 4 degrees C). Some variations in concentration were observed after dilution of tropisetron but these remained within 10% of the initial concentration. Tropisetron can be stored undiluted at 1 mg mL(-1) in polypropylene syringes, although it is preferable to perform dilutions extemporaneously. Tropisetron diluted with 5% glucose or saline can be kept equally well in glass, poly(vinyl chloride) (Travenol bags) or polyethylene (ecoflac) containers.

Antioxidant activity of some ascorbic and cinnamic acids derivatives.

Some 4-benzoyl 3-hydroxy furan-2 (5H) ones (3a-d) and 2-amino 3-hydroxymethyl 4-aryl 4-oxo 2-butenoic acids (4a-h) have been synthesized. Compound 3c with an isobutyl substituent in the 5-position of the furan ring was the most effective (IC50 = 8.69 x 10(-4) M) in scavenging the superoxide anion. In vivo, 3c was also protective against reperfusion injury.

Crystallization of monoacylated proteins: influence of acyl chain length.

The crystallization of monoacylated proteins has been investigated using a model system. Acylated derivatives of bovine pancreatic ribonuclease A, differing in their acyl chain lengths (10 to 16 carbon atoms), have been prepared using reverse micelles as microreactors. With one fatty acid moiety per polypeptide chain, covalently attached to the NH2 terminus of the protein, all the modified proteins have similar enzymatic activity and hydrodynamic radius as the native protein. Only the caprylated derivative can give crystals which diffract to high resolution. The resolved structure indicates that: (i) the protein folding is not modified by the chemical modification, (ii) the capryl moiety is not buried within the molecule but available for external interactions. Dynamic light scattering experiments on concentrated solutions show that the protein-protein interactions are dependent on acyl chain length. Proteins with the longest attached chains (14 and 16 carbon atoms) tend to self-associate through acyl group interactions.

Synthesis of new serotonergic 2-substituted 4,6-diaryl pyridazin-3-ones.

A series of 4,6-diaryl pyridazin-3-ones substituted in the 2-position by [4-(4-aryl piperazin-1-yl]-but-2-ynyl moieties was synthesized and evaluated for antidepressant activity. The structures of these new pyridazine derivatives were confirmed by IR, 1H-NMR spectra and by elementary analysis. At 150 mg/kg i.p., they induced little or no reduction of the duration of immobility of mice in the forced swimming test. Head twitches produced by L-5-hydroxytryptophan in mice pretreated with pargyline were significantly potentiated by most of the tested compounds. In addition, pyridazine derivatives did not antagonize reserpine-induced palpebral ptosis or enhance the toxic effects of yohimbine and were almost devoid of anticholinergic properties in mice.

[4-Arylpiperazine derivatives: a seedbed of drugs for various therapeutic indications]. / Dérivés 4-arylpipéraziniques: une pépinière de médicaments aux indications thérapeutiques vari s?

The main focus of this review is the pharmacology and the therapeutic use of chemical related agents, the arylpiperazine derivatives. These compounds produce a variety of behavioural responses and pharmacological effects which directly and principally result from activation of serotonin systems. However, minor modifications in the chemical structure of these products involve important changes in affinity and selectivity for 5-HT receptors since it can also display significant affinity for dopaminergic, adrenergic or histaminergic receptors. The different arylpiperazine drugs therapeutically used are described as well as some compounds presently under investigation.