Phase I metabolism of ganstigmine. Rat, dog, monkey and human liver microsomal extracts investigated by liquid chromatography electrospray tandem mass spectrometry.

Ganstigmine, a new acetylcholinesterase inhibitor, was incubated with rat liver microsomes and the resulting metabolites were identified by high-performance liquid chromatographic/mass spectrometric (HPLC/MS) and HPLC/MS/MS analyses. The results showed the formation of eight main metabolites, among which geneseroline and molecules corresponding to mono-hydroxylated, demethylated and reduced ganstigmine. The metabolic profile drawn for humans, dog and monkey showed a pattern very similar to that of rat: only in the case of liver dog microsomes higher amounts of geneseroline and of a metabolite identified as demethylated and reduced drug were detected.

Polymorphism of rac-5,6-diisobutyryloxy-2-methylamino-1,2,3,4-tetrahydro-naphthalene hydrochloride (CHF 1035). I. Thermal, spectroscopic, and X-ray diffraction properties.

The polymorphism of rac-5,6-diisobutyryloxy-2-methylamino-1,2,3,4-tetrahydro-naphthalene hydrochloride (CHF 1035) was investigated. Three different crystal forms (Form I, Form II, and Form III) were obtained by recrystallization procedures from common organic solvents. The polymorphs were characterized by Raman and carbon-13 nuclear magnetic resonance ((13)C NMR) spectroscopy, in solution and in solid state (cross polarization-magic angle spinning), powder X-ray diffractometry, and thermal methods (differential scanning calorimetry, hot stage microscopy, and thermogravimetry). Moreover, the diffraction patterns of Form I, collected at controlled temperatures, gave evidence of the presence of two reversible structural rearrangements at approximately 60 and approximately 75 degrees C. These structural variations were confirmed by the results obtained by differential scanning calorimetry and hot stage microscopy techniques. The analysis of the Raman spectra allowed the identification of peculiar absorption bands for each polymorph. Form III was the stable crystal form at room temperature as determined by the basis of slurry conversion method.

Raman and solid state 13C-NMR investigation of the structure of the 1 : 1 amorphous piroxicam : beta-cyclodextrin inclusion compound.

The results of a Raman and solid state 13C-NMR spectroscopic investigation aimed at studying the conformation of piroxicam (P) and its interaction with beta-cyclodextrin (betaCD) in 1 : 1 amorphous PbetaCD inclusion compound are reported. The 1700-1200 cm(-1) FT-Raman and the 13C CP/MAS NMR spectra of 1 : 1 PbetaCD inclusion compound are discussed and assigned in comparison with those of the three main modifications of piroxicam (alpha, beta, and monohydrate). The FT-Raman and 13C-NMR results show that in 1 : 1 PbetaCD inclusion compound piroxicam mainly assumes the zwitterionic structure typical of monohydrate, even if the presence of a different structure, that is, beta form, is not excluded. Piroxicam monohydrate, differently from alpha and beta forms, is characterized by a zwitterionic structure with an internal proton transfer and an increased charge delocalization, as shown by our spectroscopic results. The charge delocalization characteristic of this zwitterionic structure gives rise to the interaction with betaCD via electrostatic and hydrogen bonds. The possibility of a host-guest interaction between piroxicam and betaCD is not excluded; the guest molecule can be accommodated in betaCD cavity by interaction via hydrophobic bonds.

Crystal forms of piroxicam pivalate: preparation and characterization of two polymorphs.

This study investigates the polymorphism of piroxicam ester with pivalic acid. Two crystal modifications were prepared by recrystallization from toluene (form 1) and ethyl acetate (form 2). Data regarding preparation conditions, solid state properties, and physicochemical characterization of two polymorphs by means of FT/IR spectroscopy, X-ray diffractometry on powder, and thermal analysis are reported. Heat of fusion rule and thermodynamic formulas consistently indicate an enantiotropic stability relationship of forms 1 and 2 with a calculated transition point (32 degrees C) near the ambient temperature. The phase diagrams of each polymorph with piroxicam were also investigated in order to gain information about the thermal behavior of their solid mixtures. Liquidus curves calculated by the Schröder-Van Laar equation from fusion enthalpies and temperatures were found to agree satisfactorily with experimental results obtained by first heating runs with differential scanning calorimetry.

Structural characterization of two polymorphic forms of piroxicam pivalate.

The crystal and molecular structures of two polymorphs of piroxicam pivalate are presented and discussed. A peculiarity of the high melting (154 degrees C) polymorph is the association of piroxicam pivalate molecules as centrosymmetric dimers by hydrogen bonding. Two centrosymmetrically related N-H...N hydrogen bonds maintain the dimer structure involving the amido nitrogen atom as donor and the pyridine nitrogen atom as acceptor. Molecular association of this type does not occur in the crystal structures of drugs belonging to the oxicam class of nonsteroidal antiinflammatory drugs. Two distinct conformations coexist in the crystal of the low melting polymorph (136 degrees C) with differing hydrogen bonding arrangements within domains of the crystallographically independent molecules. The occurrence of different molecular conformations (conformational polymorphism) associated with different hydrogen bonding schemes in discrete domains is an unusual structural feature. Structural data for the two polymorphs are also correlated with the relevant infrared spectra. Computer-generated X-ray powder diffraction patterns for the two polymorphs of piroxicam pivalate are in very good agreement with the experimental ones, thus confirming the validity of the single-crystal X-ray models.

HPLC method for the quantitation of cispentacin enantiomers in rat urine.

The two enantiomers of cispentacin, an antifungal antibiotic, are determined by reversed phase HPLC, after derivatization with Marfey's reagent, in 24 h urine samples collected from rats treated sc and iv with 20 mg/kg cispentacin racemate obtained by synthesis. The application range of the method is 25-250 mg/L for each enantiomer with a precision of 4.0-9.0%. Comparison with an authentic sample of natural origin (-)-cispentacin indicated that (-) enantiomer is excreted unchanged in smaller percentage than (+) enantiomer, which is almost completely eliminated as such.

Characterization of phospholipidic components of the natural pulmonary surfactant Curosurf.

The phospholipidic classes of the natural pulmonary surfactant Curosurf were separated by bidimensional TLC and isolated. Quantitative analysis of each class was made by colorimetric determination of phosphorus. FD/MS and NH3-CI/MS were used to identify individual components. GC was used for qualitative/quantitative analysis of fatty acid moieties as methyl esters, using internal standards, by comparison with authentic samples.

Nicotine potentiation of haloperidol-induced catalepsy: striatal mechanisms.

Nicotine potentiated the catalepsy produced by haloperidol. The excitotoxin quinolinic acid (QA) selectively destroys striatal neurons when injected directly into the striatum. Bilateral QA lesions of the rat striatum (150 nmol) significantly reduced the catalepsy produced by haloperidol as well as the ability of nicotine to potentiate haloperidol-induced catalepsy. A second experiment examined whether the ability of nicotine to potentiate haloperidol-induced catalepsy was associated with a potentiation of dopamine turnover following haloperidol. Nicotine alone produced a mild increase in dopamine turnover relative to saline treated controls while haloperidol produced a marked increase in dopamine turnover relative to saline- and nicotine-treated controls. However, the combined administration of haloperidol and nicotine did not further elevate dopamine turnover over that observed following haloperidol alone. The results indicated that: 1) nicotine could not potentiate haloperidol-induced catalepsy without an intact striatum and 2) the behavioral effect of nicotine and haloperidol cotreatment was not due to any change in dopamine turnover.

Teicoplanin metabolism in humans.

Teicoplanin, a lipoglycopeptide antibiotic, consists of five major components (A2-1 through A2-5), one hydrolysis component (A3-1), and four minor components (RS-1 through RS-4). All the major components contain an N-acyl-beta-D-glucosamine, but they differ in the lengths and branchings of their acyl-aliphatic chains. Previous studies with radiolabeled teicoplanin in rats and humans have shown that the drug is eliminated by the renal route and that metabolic transformation is very minor, about 5%. A possible metabolic transformation of teicoplanin into A3-1 was also suggested. In the present study in humans, two metabolites (metabolites 1 and 2; 2 to 3% of total teicoplanin) were isolated after intravenous administration of radiolabeled teicoplanin. After purification, their structures were determined by fast atom bombardment mass spectroscopy and 1H nuclear magnetic resonance spectroscopy on the basis of the well-known correlations established in this field, and they were found to be new teicoplaninlike molecules, bearing 8-hydroxydecanoic and 9-hydroxydecanoic acyl moieties. This metabolic transformation is likely due to hydroxylation in the omega-2 and omega-1 positions for metabolites 1 and 2, respectively, of the C-10 linear side chain of component A2-3. This might explain the low extent of metabolism of teicoplanin if we consider that only component A2-3 has a linear chain that is susceptible to such oxidation.

Microbial de-mannosylation and mannosylation of teicoplanin derivatives.

The single components of the teicoplanin complex, glycopeptide antibiotics active against Gram-positive bacteria, can be converted in the corresponding de-mannosyl derivatives by cultures of Nocardia orientalis NRRL 2450 or Streptomyces candidus NRRL 3218. Conversely, teicoplanin aglycone and other teicoplanin de-mannosyl derivatives can be converted in the corresponding teicoplanin mannosyl derivatives by cultures of Actinoplanes teichomyceticus ATCC 31121. The biological transformation yields are approximately 40% for de-mannosylation and 90% for mannosylation. The processes allow for the preparation of gram quantities of the de-mannosyl derivatives of teicoplanin and of teicoplanin mannosyl derivatives. De-mannosyl teicoplanin and teicoplanin mannosyl-pseudoaglycone were not amenable to preparation by either acidic or basic chemical hydrolysis.

Nicotine potentiates haloperidol-induced catalepsy and locomotor hypoactivity.

Nicotine was found to potentiate the catalepsy and reduced locomotion following the administration of haloperidol. The ability of various doses of nicotine (0.1, 0.2, or 0.3 mg/kg) to potentiate the catalepsy produced by haloperidol (0.1, 0.2 or 0.4 mg/kg) was investigated. Nicotine potentiated the cataleptic effects of both the 0.2 and 0.4 mg/kg doses of haloperidol, but had no effect following the lowest (0.1 mg/kg) dose of haloperidol. The nicotine potentiation of catalepsy produced by the highest dose of haloperidol was independent of the dose of nicotine used. Nicotine alone did not produce catalepsy. A second experiment evaluated the ability of nicotine to potentiate the decreases in spontaneous locomotor activity produced by haloperidol. Animals received nicotine (0.1 mg/kg) alone or in conjunction with haloperidol (0.1 or 0.4 mg/kg) and were tested in Digiscan Animal Monitors. Haloperidol produced a dose-related decrease in locomotion. Nicotine significantly potentiated the hypoactivity produced by both doses of haloperidol. These results indicated that: 1) nicotine produces a significant potentiation of both the catalepsy and locomotor decreases following haloperidol and 2) the Digiscam Animal Activity Monitors may provide a more sensitive assessment of the interaction between nicotine and haloperidol than the catalepsy bat test. These data suggest that adjunct treatment with nicotine may prove useful for treating neuroleptic responsive disorders such as Tourette Syndrome, schizophrenia and Huntington's disease.

Isolation and structure determination of two new analogs of teicoplanin, a glycopeptide antibiotic.

Teicoplanin is an antibiotic produced by fermentation of Actinoplanes teichomyceticus as a complex formed by five closely related glycopeptides characterized by different fatty acid chains of ten and eleven carbon atoms. In addition, minor quantities of related substances are present. Two of them, named RS-1 and RS-2, were shown to be teicoplanins having as fatty acid chains 10-methylundecanoic acid and n-dodecanoic acid, respectively. Other two related substances, named RS-3 and RS-4, have now been isolated and purified starting from fermentation broths of a mutant of the same microorganism producing them in substantial amounts. This was achieved by semipreparative reversed-phase liquid chromatography carried out on high-pressure scale. The structures were assigned on the basis of 1H NMR spectra and homonuclear COSY 2D experiments and fast atom bombardment MS spectrometry, in comparison with the large mass of data till now accumulated on teicoplanin. RS-3 and RS-4 are teicoplanins having as fatty acid chains 6-methyloctanoic acid and n-nonanoic acid, respectively.

Quantitative determination of propildazine in rat plasma by gas-liquid chromatography.

A gas-liquid chromatographic method for the evaluation of the new anti-hypertensive drug propildazine (ISF 2123) in rat plasma is described. The procedure involves separation of the drug from plasma by cation-exchange chromatography, subsequent acylation of the dried eluate with heptafluorobutyric anhydride and quantitation with electron-capture detection. Propildazine can be determined in concentrations down to ca. 0.4 microgram/ml.

Epoxide-diol metabolic pathway of cytenamide in the rat.

Cytenamide administered intraperitoneally to rats is biotransformed to cytenamide-10,11-epoxide and 10,11-dihydro-10,11-dihydroxycytenamide. These metabolites were separated by chromatographic methods and their structures elucidated by mass spectrometry. The structure of the epoxide was confirmed by direct comparison with an authentic sample. The formation of a chemical artifact, cytenamide-syn-11-hydroxylactone, in the urine was observed. The formation of these metabolites in vitro was demonstrated by incubation of cytenamide with rat liver microsomes.