András Váradi

Detection and identification of the new potential synthetic cannabinoids 1-pentyl-3-(2-iodobenzoyl)indole and 1-pentyl-3-(1-adamantoyl)indole in seized bulk powders in Hungary

3-Naphthoyl- and 3-phenylacetylindoles represent a group of substances of cannabimimetic activity with affinities - strongly influenced by their functional groups - to cannabinoid receptors CB1 and CB2. Some of them have been described as ingredients of herbal blends also known as smart products by several research groups. Recently further cannabimimetic substances possessing new chemical structures like benzoylindoles and adamantoylindoles have emerged. In Hungary, two powder samples were seized by the authorities and identified as 1-pentyl-3-(2-iodobenzoyl)indole (AM-679) and 1-pentyl-3-(1-adamantoyl)indole. Structure elucidation was carried out by LC-UV-MS/MS, LC-TOF-MS, GC-MS and NMR. The benzoylindole AM-679 is a known agonist of cannabinoid receptors while the adamantoylindole derivative also carries chemical features typical for cannabimimetics. It is thus assumed that both substances might be detected in smart products in the future.

Identification and characterization of the new designer drug 4′-methylethcathinone (4-MEC) and elaboration of a novel liquid chromatography–tandem mass spectrometry (LC–MS/MS) screening method for seven different methcathinone analogs

A fast and simple LC-MS/MS method was developed for screening mephedrone, butylone, methylenedioxypyrovalerone (MDPV), flephedrone, methylone and methedrone in bulk powder samples. Samples were separated on a reverse phase column using gradient elution with mixtures of water, acetonitrile and formic acid. After optimization a limit of detection of about 2ngmL(-1) was achieved using multiple reaction monitoring (MRM) mode. Total run time was less than 8min. Typical fragmentation characteristics of the studied compounds are discussed. The method was successfully applied to several unknown bulk powder samples seized by the Hungarian Customs and Finance Guard. One of the samples contained the new designer drug 4-methylethcathinone (4-MEC), which was identified and characterized by LC-MS/MS, NMR, FT-IR and LC-TOF-MS techniques. The method is also deemed to be applicable for the screening of simple dosage forms such as tablets and capsules.

Sulfate esters of morphine derivatives: Synthesis and characterization

Sixteen 3-O- and 6-O-sulfate esters of morphine, codeine and some of their N-methyl quaternary derivatives were synthesized by means of sulfation with pyridine-SO(3) complex and sulfuric acid/N,N-dicyclohexylcarbodiimide. Complete (1)H- and (13)C-NMR assignments are given for each of the synthesized compounds based on one- and two-dimensional homo- and heteronuclear measurements. Comparative analysis of chiral properties by circular dichroism and optical rotatory dispersion revealed characteristic differences in the spectra due to changes in charge, polarity and intramolecular association by strong hydrogen bonds in aqueous solution. The synthesized sulfate esters are prospective peripheral analgesics lacking central side effects and are also useful as reference substances for various analytical studies involving sulfate ester metabolites.

Bone marrow transplantation in accelerated chronic granulocytic leukaemia using dibromomannitol-preconditioning instead of total-body irradiation

In a preliminary study on five patients with accelerated CGL, transplantation of allogeneic matched bone marrow was shown to be feasible without whole-body irradiation. Animal experiments and studies with cells cultured in vitro suggest that the cytocastic drug used to kill leukaemic clones (Myelobromol-Chinoin) does not injure haemopoietic stroma. The administration of this protocol is cheap and easy. Our preconditioning does not, in itself, eradicate the malignant CGL clone immediately: 15-20% of marrow mitoses were Ph1+ one month after transplantation. For this reason, additional cytostatic therapy was given in the course of the 3rd to 6th post-transplant months. No Ph1+ cells were observed from the fourth post-transplant month onwards. Very few severe acute complications were seen and two out of three matched transplanted patients are disease-free 27 + and 13 + months later. On the basis of the developing normal spleen architecture and the changing pattern of circulating NAP score values, particularly the months-long persistence of distinctly low scores, and then the delayed emergence of normal levels, we put forward a hypothesis, emphasizing the role of environmental factors, including the formation of a normal haemopoietic stroma in the successfully transplanted CGL patient.

Human tolerability studies with D-Met2, Pro5-enkephalinamide

As reported previously D-Met2,Pro5-enkephalinamide (EA) is a highly active enkephalin analogue. To examine its human tolerability male volunteers were treated s.c. with increasing doses (0.1-30.0 mg). The observed autonomic effects were as follows: feeling of heaviness in the limbs, dry mouth, pallor of the face and conjunctival injection. There was no significant change in blood pressure, pulse and respiratory frequency. The autonomic effects appeared within 15-30 min. However, its effects on mood and wakefulness i.e. slight drowsiness, decrease in psychic tension and emotional detachment developed only later. The serum prolactin level increased dose-dependently, while the growth hormone (HGH) content showed biphasic dose-response pattern. The TSH content increased only at the highest doses applied (10.0-30.0 mg).

Effects of methysergide, bromocriptine and naloxone on prolactin, growth hormone and TSH release induced by d-Met2,Pro5-enkephalinamide in man

Abstractd-Met2,Pro5-enkephalinamide (EA) 10 mg, given SC, induced a dramatic rise in serum prolactin (PRL) and growth hormone (GH) levels in healthy male volunteers. The TSH content was also moderately elevated. Naloxone 0.8 mg administered IV abolished these effects. Bromocriptine 2.5 mg given per os also antagonized EA-induced PRL and TSH release but potentiated the GH surge. Methysergide 2.0 mg administered orally partially reversed EA-elicited PRL release, further augmented GH liberation and did not modify TSH output. The data indicate that inhibition of the dopaminergic tone and/or activation of certain serotonergic mechanisms play an important role in the EA-induced release of PRL and TSH. However, primarily other neurotransmitters might mediate the GH liberation elicited by this opioid peptide.

New morphine analogs produce peripheral antinociception within a certain dose range of their systemic administration

Growing data support peripheral opioid antinociceptive effects, particularly in inflammatory pain models. Here, we examined the antinociceptive effects of subcutaneously administered, recently synthesized 14-O-methylmorphine-6-O-sulfate (14-O-MeM6SU) compared with morphine-6-O-sulfate (M6SU) in a rat model of inflammatory pain induced by an injection of complete Freund’s adjuvant and in a mouse model of visceral pain evoked by acetic acid. Subcutaneous doses of 14-O-MeM6SU and M6SU up to 126 and 547 nmol/kg, respectively, produced significant and subcutaneous or intraplantar naloxone methiodide (NAL-M)–reversible antinociception in inflamed paws compared with noninflamed paws. Neither of these doses significantly affected thiobutabarbital-induced sleeping time or rat pulmonary parameters. However, the antinociceptive effects of higher doses were only partially reversed by NAL-M, indicating contribution of the central nervous system. In the mouse writhing test, 14-O-MeM6SU was more potent than M6SU after subcutaneous or intracerebroventricular injections. Both displayed high subcutaneous/intracerebroventricular ED50 ratios. The antinociceptive effects of subcutaneous 14-O-MeM6SU and M6SU up to 136 and 3043 nmol/kg, respectively, were fully antagonized by subcutaneous NAL-M. In addition, the test compounds inhibited mouse gastrointestinal transit in antinociceptive doses. Taken together, these findings suggest that systemic administration of the novel compound 14-O-MeM6SU similar to M6SU in specific dose ranges shows peripheral antinociception in rat and mouse inflammatory pain models without central adverse effects. These findings apply to male animals and must be confirmed in female animals. Therefore, titration of systemic doses of opioid compounds with limited access to the brain might offer peripheral antinociception of clinical importance.

New opioid receptor antagonist: naltrexone-14-O-sulfate synthesis and pharmacology.

&NA; Opioid antagonists, naloxone and naltrexone have long been used in clinical practice and research. In addition to their low selectivity, they easily pass through the blood‐brain barrier. Quaternization of the amine group in these molecules, (e.g. methylnaltrexone) results in negligible CNS penetration. In addition, zwitterionic compounds have been reported to have limited CNS access. The current study, for the first time gives report on the synthesis and the in vitro [competition binding, G‐protein activation, isolated mouse vas deferens (MVD) and mouse colon assay] pharmacology of the zwitterionic compound, naltrexone‐14‐O‐sulfate. Naltrexone, naloxone, and its 14‐O‐sulfate analogue were used as reference compounds. In competition binding assays, naltrexone‐14‐O‐sulfate showed lower affinity for &mgr;, &dgr; or &kgr; opioid receptor than the parent molecule, naltrexone. However, the &mgr;/&kgr; opioid receptor selectivity ratio significantly improved, indicating better selectivity. Similar tendency was observed for naloxone‐14‐O‐sulfate when compared to naloxone. Naltrexone‐14‐O‐sulfate failed to activate [35S]GTP&ggr;S‐binding but inhibit the activation evoked by opioid agonists (DAMGO, Ile5,6deltorphin II and U69593), similarly to the reference compounds. Schild plot constructed in MVD revealed that naltrexone‐14‐O‐sulfate acts as a competitive antagonist. In mouse colon, naltrexone‐14‐O‐sulfate antagonized the inhibitory effect of morphine with lower affinity compared to naltrexone and higher affinity when compared to naloxone or naloxone‐14‐O‐sulfate. In vivo (mouse tail‐flick test), subcutaneously injected naltrexone‐14‐O‐sulfate antagonized morphines antinociception in a dose‐dependent manner, indicating its CNS penetration, which was unexpected from such zwitter ionic structure. Future studies are needed to evaluate its pharmacokinetic profile.

Glucosides of morphine derivatives: synthesis and characterization

Six 3-O- and 6-O-glucosides of morphine and codeine derivatives were synthesized by means of glucosylation with acetobromo-α-d-glucose. O-Glucosylation at C6 was carried out by the Koenigs-Knorr method, whereas the 3-O-glycoside of morphine was synthesized directly upon stirring morphine with acetobromo-α-d-glucose and aqueous sodium hydroxide in acetone. Complete 1H and 13C NMR assignments are presented for each synthesized compound based on one- and two-dimensional homo- and heteronuclear NMR techniques. Circular dichroism, ultraviolet absorbance, and high-resolution mass spectroscopy data ensure identification and structural characterization of the O-glucoside conjugates. The synthesized glucoside conjugates are potential analgesics; the presented spectral and chromatographic data are useful references for various analytical and metabolic studies including samples of biological origin.Graphical Abstract

Synthetic and quantum chemical study on the regioselective addition of amines to methyl maleamate

AbstractSynthetic and theoretical studies were performed to gain insight into the regioselectivity in the mechanism of aspartyl-isoaspartyl formation, modeled by additions of ammonia and primary amines to methyl maleamate. Reactions between maleamate and aliphatic, araliphatic amines or O-methyl acetimidate lead to the formation of N-substituted isoasparaginates. The size of the amine and the activating effect of the amide and ester group on the double bond are the determining factors of the site of addition. The formation of both isomers was observed only in the case of ammonia addition. The regioselectivity was predicted on the basis of the charge distribution for low-energy methyl maleamate conformers, calculated at the B3LYP/6-311++G(2df,2pd)//B3LYP/6-31+G(d) level, both in gas phase and in methanol. The methyl isoasparaginate over methyl asparaginate product ratio was computed based on the free energy Boltzmann distribution of their conformers. The calculated 2 : 1 ratio is in agreement with the experimental regioselectivity of the addition of nitrogen nucleophiles.n FigureRegioselective formation of asparaginates