Synthesis and study of a molecularly imprinted polymer for the specific extraction of indole alkaloids from Catharanthus roseus extracts.

Two molecularly imprinted polymers (MIP) for catharanthine and vindoline have been synthesized in order to specifically extract these natural indole alkaloids from Catharanthus roseus by solid-phase extraction (SPE). Each MIP was prepared by thermal polymerisation using catharanthine (or vindoline) as template, methacrylic acid (or itaconic acid) as functional monomer, ethylene glycol dimethacrylate (EDMA) as cross-linking agent and acetonitrile (or acetone) as porogenic solvent. For catharanthine-MIP, a SPE protocol (ACN-AcOH 99/1 washing and MeOH-AcOH 90/10 elution) allows a good MIP/NIP selectivity (imprinting factor 12.6). The specificity of catharanthine-MIP versus related bisindole alkaloids was assessed by cross-reactivity study. The catharanthine-MIP specifically retained catharanthine and its N-oxide analogue but displayed a weak cross-reactivity for other Vinca alkaloids (vinorelbine, vincristine, vinblastine, vindoline, vinflunine). It appears that the catharanthine-like unit of these molecules are hardly trapped in catharanthine cavities located in the MIP, probably due to the sterical hindrance of the vindoline moiety. Finally, the MIP-SPE applied to C. roseus extract enabled quantitative recovery of catharanthine (101%) and the total removal of vindoline. Its capacity was determined and was equal to 2.43 µmol g(-1). Vindoline is a weaker base than catharanthine, so the vindoline-MIP was achieved with a strong acidic monomer (itaconic acid) to increase vindoline-monomer interactions and a modified washing solvent (ACN-HCOOH 99/1) to reduce non-specific interactions. The influence of the amount of HCOOH (protic modifier) percolated during the washing step upon the elution yield and the imprinting factor for vindoline was investigated. This preliminary optimisation of the washing step, and in particular the number of moles of acid percolated, seems useful to emphasize the use of MIP in conditions of high selectivity or high yield. A compromise was obtained with an imprinting factor equal to 7.6 and an elution recovery of 33%. However MIP-vindoline failed to achieve a specific extraction of vindoline since catharanthine was also extracted probably because of strong non-specific interactions occurring between catharanthine and the sorbent.

Conformational analysis and crystal structure of {[1-(3-chloro-4-fluorobenzoyl)-4-fluoropiperidin-4yl]methyl}[(5-methylpyridin-2-yl)methyl]amine, fumaric acid salt.

{[1-(3-Chloro-4-fluorobenzoyl)-4-fluoropiperidin-4yl]methyl}[(5-methylpyridin-2-yl)methyl]amine, fumaric acid salt (C(20)H(22)ClF(2)N(3)O, C(4)H(4)O(4)) (1) was synthesized and characterized by the complete (1)H, (13)C and (19)F NMR analyses. The conformation of the piperidin ring, in the solution state, was particularly studied from the coupling constants determined by recording a double-quantum filtered COSY experiment in phase-sensitive mode. (1)H NMR line-shape analysis was used, at temperatures varying between -5 and +60 degrees C, to determine the enthalpy of activation of the rotational barrier around the CN bond. Compound 1 crystallizes in the triclinic space group P1 with a=8.517(3) Angstrom, b=12.384(2) Angstrom, c=12.472(3) Angstrom, alpha=70.88(2) degrees, beta=82.04(2) degrees, gamma=83.58(2) degrees. The results strongly indicate that the solid and solution conformations are similar. Thermal stability and phases transitions were investigated by thermal gravimetric analysis (TGA) and differential scanning calorimetry (DSC). Furthermore polymorphism screening was studied from recrystallization of 1 performed in seven solvents and by slurry conversion in water. The X-ray powder diffraction (XRPD) and differential scanning calorimetry results suggested that 1 crystallizes into one crystalline form which melts at 157 degrees C (DeltaH=132 J g(-1)).

[Crystalline polymorphism of eflucimibe]. / Polymorphisme cristallin de l'éflucimibe.

The importance, in therapeutics, of the concept of bioavailability and on-going quality research in the formulation of a drug has prompted us to examine the crystalline polymorphism of eflucimibe as from the research phase. This study has been carried out by re-crystallization of the product in organic solvents having a different polarity in a variety of experimental temperature and pressure conditions, then, subsequently, by re-cooling the previously dissolved substance. The analytical methods applied to identify and then describe the polymorphic forms are thermogravimetry analysis (TGA), differential scanning calorimetry (DSC), X-ray powder diffraction from synchrotron radiation (XRPD), infrared spectrometry (IR), solid-state nuclear magnetic resonance spectrometry (SSNMR) and lastly maximum solubility measurements. By means of XRPD, two polymorphic forms called A and B have been clearly identified at ambient temperature. These two crystalline forms were obtained in a reproducible way, then described by DSC, XRPD, IR and SSNMR. Differential scanning calorimetry analysis has shown for polymorphic form A two endothermic phenomena with low energy at about 35 masculine and 118 degrees C attributed by XRDP to conformational polymorphism. The complex endothermic event that extends between 75 masculine and 105 degrees C appears to correspond to successive alterations of a lamellar structure. The solid-solid transition observed at 110 degrees C is due to a change of crystalline phase, then the melting point occurring at about 130 degrees C. For form B, two changes of crystalline phase are clearly observed at about 80 masculine and 120 degrees C. The reversibility of these changes was observed by thermomicroscopy in polarized light. Form A, which is less soluble in absolute ethanol than form B, is the more stable thermodynamically in the temperature range from 25 masculine to 50 degrees C where the data have been obtained. The Van't Hoff diagram layout for each polymorphic form appears to reveal an A<-->B transition temperature in a temperature range lower than 25 degrees C. This study, undertaken as from the research phase, has enabled us to highlight the polymorphism of this new chemical entity by means of XRDP by explaining the nature of the endothermic phenomena observed by DSC, and lastly identify the thermodynamically more stable polymorphic form, thus contributing to a better knowledge of this future drug.

Experimental conditions for the continuous subcutaneous infusion of four central analgesics in rats.

For the analysis of pharmacotherapeutic regimens for chronic pain in animals, it is important to establish delivery methods in which analgesics can be administered continuously and at a constant rate for a prolonged period of time. This allows for the assessment of how drug effects may vary over time in the presence of ongoing pain. The present study determined, for four analgesic compounds, the maximal doses that met all of the following criteria: (i) water-soluble, (ii) stable over 14 days at 38 degrees C, and (iii) devoid of undesirable side-effects in normal rats, as assessed by evolution of body weight and temperature after the subcutaneous implantation of an osmotic mini-pump that continuously infused the compounds over a 14-day period. The results showed the maximal doses to be 5 mg/rat/day for morphine hydrochloride, 2.5 mg/rat/day for imipramine hydrochloride, 20 mg/rat/day for ketamine hydrochloride, and 10 mg/rat/day for gabapentin. These doses were further found to be sufficient to express each compound's representative pharmacological activity. The conditions identified here appear appropriate for future studies of these four compounds in rat models of chronic pain and neuropathic allodynia.

Efficient applications of capillary electrophoresis-tandem mass spectrometry to the analysis of adrenoreceptor antagonist enantiomers using a partial filling technique.

Throughout the separation of chiral basic drugs by capillary electrophoresis (CE) with neutral hydroxypropyl-beta-cyclodextrin (HP-beta-CD) as chiral selector, the sensitivity of detection can be improved by using tandem mass spectrometric (MS-MS) detection with a partial filling technique rather than with UV spectrometric detection. Prior to sample injection. the capillary was partly filled with HP-beta-CD dissolved in volatile ammonium formate buffer (pH 4, ionic strength 50 mM). The effects of modifying the HP-beta-CD concentration in the selector zone and the length of the separation zone on the enantioresolution and the signal-to-noise ratio of the pseudo-molecular MH+ ion were investigated. For a given selector zone length, as the concentration of the neutral cyclodextrin increases, the resolution between enantiomers becomes higher (the opposite of the behavior of the signal-to-noise ratio) and then reaches an optimum value. The decrease of the selector zone length lowered the resolution between the enantiomers but increased peak efficiencies and signal-to-noise ratio values. Accordingly, partial capillary filling at 80% (v/v) and 10 mM concentration of HP-beta-CD was selected as a suitable compromise between resolution and sensitivity of MS detection. Limits of detection for each adrenoreceptor antagonist enantiomer were 5 ng/ml (0.02 microM) in CE-MS-MS instead of 150 ng/ml (0.60 microM) in CE-UV, which enhances sensitivity by a factor of 30.

Absolute configuration of (+)-(S)-2-(5-fluoro-2-methoxy-1,4-benzodioxan-2-yl)imidazolinium bromide.

The title compound, C(12)H(14)FN(2)O(3)(+).Br(-), crystallizes in the non-centrosymmetric P2(1)2(1)2(1) space group. The absolute configuration of the pharmacologically active molecule could be resolved in the hydrobromide salt, the structure of which is reported. The molecule of the title compound has the S configuration. The molecular packing in the crystal is stabilized by weak N-H.Br [N.Br = 3.240 (4) and 3.302 (4) A] hydrogen bonding.

Enhancement of second-migrating enantiomer peak symmetry of basic drugs by using dual-cyclodextrin system in capillary electrophoresis.

Today, chiral separations of cationic drugs by capillary electrophoresis are generally carried out by adding negatively charged cyclodextrins (CDs) to the running buffer while anionic or neutral drug separations require the use of dual-CD systems (mixtures of neutral and charged CDs). Chiral separation of some basic drugs (idazoxan, efaroxan, milnacipran) has been studied by using mixtures of sulfated-beta-CD (S-beta-CD) and hydroxypropyl-gamma-CD (HP-gamma-CD). The influence of the following parameters (nature and concentration of neutral CD, concentration of S-gamma-CD) on many separation factors (electrophoretic mobility, selectivity, efficiency, asymmetry factor, resolution) demonstrated that dual-CD systems are useful for chiral separation of basic drugs in order to improve the symmetry of the second-migrating enantiomer. Indeed, the neutral CD reduces the extent of electromigration dispersion by mobility tuning. Finally, the 0.5 mg/mL S-beta-CD/5 mg/mL HP-gamma-CD dual system has allowed the chiral separation of idazoxan, efaroxan and milnacipran enantiomers in less than 9 min.

[Structural analysis of vinorelbine in solution determined by nuclear magnetic resonance spectrometry]. / Analyse structurale de la vinorelbine en solution déterminée par spectrométrie de résonance magnétique nucléaire.

An active partnership between the National Centre for Scientific Research (CNRS) and the laboratories Pierre Fabre is underpinning the development of a new molecule, vinorelbine, whose tartrate received marketing authorization in France in 1989, under the name of Navelbine. This medicine was first recommended for the treatment of bronchial cancer "not small cell", then, in 1991, for the treatment of metastatic breast cancer. In 1994, its registration in United States was granted for the treatment of bronchial cancer "not small cell". Vinorelbine is obtained by hemisynthesis using two antecedent monomeric alkaloids, catharanthine and vindoline, followed by a modification of the catharanthine nucleus, so as to produce the first 5' nor vinca-alkaloid. The chemical structure of vinorelbine has been examined in our laboratory using nuclear magnetic resonance spectrometry. Bearing in mind their complexity, the total attribution of the proton spectrum and the carbon-13 spectrum has required experiments for homonuclear (1H-1H) and heteronuclear (1H-13C and 1H-15N) correlation. These experiments have been carried out using a BRUKER spectrometer operating at the nominal proton frequency of 200 MHz in direct detection mode, then with a 400 MHz spectrometer equipped with the reverse detection mode. The chemical structure has thus been analyzed with no ambiguity. The results of this structural study will be presented in due course. We have also undertaken a comparative conformational study between base vinorelbine in chloroform solution and ditartrate vinorelbine (Navelbine) in methanolic solution. The conformation of the vinorelbine molecule in solution in these different solvents have been studied with NOESY (Nuclear Overhauser Effect Spectroscopy) experiments. The results of these experiments have been confirmed by data stemming from molecular modelization.

Characterization of polymorphs and solvates of 3-amino-1-(m-trifluoromethylphenyl)-6-methyl-1H-pyridazin-4-one.

The characterization of two polymorphs of the title compound (F2692; 1) by differential scanning calorimetry (DSC), microanalysis, proton nuclear magnetic resonance spectroscopy, thermogravimetry, thermomicroscopy, infrared spectroscopy, and X-ray diffractometry is described. Both polymorphs are crystalline, with form II being more stable at temperatures less than 160 degrees C. The thermal behavior was studied at different rates of heating, and the enthalpies of transition were calculated from DSC data. The transformation of aqueous suspensions of form I to the water-stable form II is described, and the heats of solution and intrinsic aqueous dissolution rates of both polymorphs were determined. 1 also formed solvates with dimethyl sulfoxide and 1-methyl-2-pyrrolidinone. The solvates were studied by thermogravimetry, DSC, and infrared spectroscopy.

GLC determination of itanoxone in biological fluids.

A sensitive and reliable method for the quantitative determination of itanoxone, 4-[4'-(2-chlorophenyl)phenyl]-4-oxo-2-methylenebutanoic acid, in biological fluids is described. A quantitative ethyl acetate extraction of the plasma samples is followed by reduction and methylation of itanoxone. Quantification is achieved by GLC using electron-capture detection and an internal standard. The minimum concentration of itanoxone detected in plasma is 0.1 microgram/ml. Recovery of the titrated compound added to human plasma averaged 100.9 +/- 2.92% (RSD).