Conformational analysis and structural comparisons of (1R,3S)-(+)- and (1S,3R)-(-)-tefludazine, (S)-(+)- and (R)-(-)-octoclothepin, and (+)-dexclamol in relation to dopamine receptor antagonism and amine-uptake inhibition

Conformational analysis with molecular mechanics (MM2(85] and molecular superimposition studies of (1R,3S)-(+)- and (1S,3R)-(-)-4-[3-(4-fluorophenyl)-6-(trifluoromethyl)indan-1-yl]-1- piperazineethanol (tefludazine) and (S)-(+)- and (R)-(-)-octoclothepin have been employed to identify biologically active conformations of these compounds with respect to dopamine receptor antagonism and amine-uptake inhibition. In contrast to what is commonly assumed, these studies indicate that the conformation of (S)-(+)-octoclothepin responsible for the dopamine receptor antagonism is different from the one observed in the crystal. From least-squares molecular superimpositions with the potent and stereoselective dopamine receptor antagonist (1R,3S)-tefludazine, biologically active conformations for the two compounds on the dopamine receptor have been deduced. This analysis also rationalizes the enantioselectivity of octoclothepin on the dopamine receptor. The X-ray structure of (S)-(+)-octoclothepin is shown to correspond structurally to the 1S,3R enantiomer of tefludazine, which is an amine-uptake inhibitor. This correspondence provides a structural basis for the norepinephrine (NE) uptake blocking properties of octoclothepin. It is predicted that the enantioselectivity of the NE-uptake inhibition of octoclothepin should be low with the S-(+) enantiomer as the more active optical isomer. A comparison of the deduced biologically active conformation of (S)-(+)-octoclothepin with (+)-dexclamol is also discussed on the basis of earlier derived superimposition studies with (+)-dexclamol.     

Aporphines. 48. Enantioselectivity of (R)-(-)- and (S)-(+)-N-n-propylnorapomorphine on dopamine receptors

The enantiomers [(S)-(+) and (R)-(-)] of N-n-propylnorapomorphine (NPA) were synthesized. (R)-NPA was obtained by the acid-catalyzed rearrangement of N-n-propylnormorphine. (R)-NPA also was converted to (RS)-N-n-propylnorapomorphine dimethyl ether by dehydrogenation of the 10,11-O,O'-dimethyl ether of (R)-NPA with 10% palladium on carbon in acetonitrile, followed by reduction with sodium cyanoborohydride under acidic conditions. Alternatively (RS)-NPA 10,11-O,O'-dimethyl ether was obtained via total synthesis. (+)-Dibenzoyl-D-tartaric acid was used to resolve (RS)-NPA dimethyl ether. Ether cleavage gave (S)-NPA isolated as the hydrochloride salt in greater than 99.9% enantiomeric purity, as determined by circular dichroism (CD) spectra. The pharmacological activities of (S)- and (R)-NPA were evaluated with subnanomolar concentrations of 3H-labeled apomorphine (APO), ADTN, and spiroperidol (SPR) for competition for binding to a membrane-rich subsynaptosomal fraction of calf caudate nucleus. IC50 (nM) values for (R)-NPA vs. (S)-NPA were as follows: [3H]APO, 2.5 vs. 66; [3H]ADTN, 2.0 vs. 60; [3H]SPR, 174 vs. 1400. The efficacy of (R)- and (S)-NPA in stimulating dopamine-sensitive adenylate cyclase from both homogenates of rat corpus striatum and pieces of intact carp retina was also evaluated. Three behavioral effects in the rat (stereotyped behavior, sedation, and catalepsy) were also examined. Only (R)-NPA induced stereotypy; (S)-NPA failed to antagonize this action of the R isomer. The effects of (R)- and (S)-NPA on adenylate cyclase agreed with the behavioral effects and radioreceptor binding assays in that the R isomer was the strongly preferred enantiomer at dopamine receptors. The S enantiomer of NPA was, however, the weakly preferred configuration for rat liver catechol O-methyltransferase. A dopamine-receptor model accommodates the configuration of NPA and related aporphines.     

The in vitro metabolic inversion of R(-) to S(+) indoprofen.

The paper reports a study on the metabolic inversion of indoprofen (2-[4-(2-isoindolinyl-1-one)-phenyl]-propionic acid) following incubation of the drug with liver microsomes from non-induced and phenobarbital-induced rats. The enantiomeric composition of the drug was determined after different incubation times of the racemate and the individual isomers. The S(+)/R(-) ratio was evaluated by densitometry following HPTLC separation of the R(+)-1-phenylethylamides. After incubation of the racemate and the individual isomers, no detectable amounts of indoprofen catabolites were extracted from the acidified incubation mixture. An appreciable enrichment in the S(+) enantiomer was observed after incubation of both racemate and R(-)-indoprofen; the S(+)/R(-) ratio reached a maximum after 1 h. Values were higher in the case of induction. After incubation of S(+)-indoprofen, a small but statistically significant decrease of the S(+)/R(-) ratio was observed. The increase of the S(+)-isomer concentration observed following incubation of R(-)-indoprofen can be ascribed to metabolic inversion by phenobarbital-inducible liver enzymes.     

Preparation and properties of (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl- (R)-(+)-alpha-hydroxy-alpha-(4-[125I]iodophenyl)-alpha-phenyl acetate and (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(S)-(-)-alpha-hydroxy-alpha- (4-[125I]iodophenyl)-alpha-phenyl acetate as potential radiopharmaceuticals

rac-4-Nitrobenzilic acid was synthesized and resolved with quinidine and quinine to give the corresponding (R)- and (S)-salts. The resolved diastereomeric salts were converted to (R)- and (S)-4-nitrobenzilic acids and subsequent esterification gave their corresponding ethyl esters. Transesterification with (R)-(-)-3-quinuclidinol afforded (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(R)-(+)-alpha-hydroxy-alpha- (4-nitrophenyl)-alpha-phenyl acetate and (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(S)-(-)-alpha-hydroxy- alpha-(4-nitrophenyl)-alpha-phenyl acetate. After hydrogenation, the (R,R)- and (R,S)-amines were converted to the respective triazene derivatives. The triazene derivatives reacted with sodium [125I]iodide to give (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(R)-(+)- alpha-hydroxy-alpha-(4-[125I]iodophenyl)-alpha-phenyl acetate and (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(S)-(-)-alpha-hydroxy- alpha-(4-[125I]iodophenyl)-alpha-phenyl acetate. The evaluation of their affinities to muscarinic acetylcholine receptors (MAcChR) shows that (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(S)-(-)-alpha-hydroxy-alpha-(4- [125I]iodophenyl)-alpha-phenyl acetate exhibits an affinity for the MAcChR from corpus striatum that is approximately threefold lower than that of (R)-(-)-1-azabicyclo[2.2.2]oct-3-yl-(R)-(+)-alpha-hydroxy-alpha-(4- [125I]iodophenyl)-alpha-phenyl acetate.     

The effect of food or sucralfate on the bioavailability of S(+) and R(-) enantiomers of ibuprofen.

This randomized, multiple cross-over pharmacokinetic study was undertaken to determine if food or sucralfate alter the bioavailability of the active S(+) enantiomer of ibuprofen. Eleven healthy adult male volunteers were given three single 600-mg doses of ibuprofen (separated by 1 week) administered either in a fasting state, after a standardized breakfast, or with sucralfate 1 g. The main outcome measures were area under the concentration (AUC), maximum peak plasma concentration (Cmax), and time to reach peak concentration (tmax) for total, S(+), and R(-) enantiomer serum ibuprofen levels, drawn up to 10 hours after dosing. The AUC for R(-) ibuprofen was significantly lower than S(+) ibuprofen in all three treatment groups. The treatments had no different effects on AUC for S(+), R(+), or total ibuprofen. There was no difference in the ratio of S(+):R(-) enantiomers across different treatment groups, but the intersubject variability was significant (P < .05). The S(+) ibuprofen Cmax was greater than the R(-) ibuprofen Cmax for all treatment groups (P < .05). Sucralfate reduced the peak concentration of both S(+) and R(-) enantiomers when compared with fasting (P < .05). There was a slight but nonsignificant increase in the mean time to achieve peak concentration of both S(+) and R(-) enantiomers. Neither food nor sucralfate has a significant effect on ibuprofen enantiomer pharmacokinetics, but interindividual variability contributes significantly to the variability of enantiomer bioavailability.     

GABA agonists. Resolution, absolute stereochemistry, and enantioselectivity of (S)-(+)- and (R)-(-)-dihydromuscimol

(RS)-5-(Aminomethyl)-2-isoxazolin-3-ol (dihydromuscimol, DHM) is a potent 4-aminobutyric acid (GABA) agonist, the inhibitory effects of which on neurons are sensitive to the antagonist bicuculline methochloride (BMC), and it also interacts with the GABA uptake system in vitro. (S)-(+)-DHM (4) and (R)-(-)-DHM (5) were obtained in optically pure forms via resolution of tert-butyloxycarbonyl-protected DHM (1) using cinchonidine as the only resolving agent. The optical purity and absolute stereochemistry of 4 and 5 were established by chemical correlation to the (S)-(+) enantiomer of 3-hydroxy-4-aminobutyric acid (GABOB). While 4 was a specific and potent BMC-sensitive GABA agonist in vivo and in vitro, possibly the most potent GABA agonist so far described, the inhibition of GABA uptake by DHM proved to reside exclusively in the (R)-(-) enantiomer (5). The affinity of 5 for BMC-sensitive GABA receptor sites in vitro was some 50 times lower than that of 4. Compounds 4 and 5 can be considered semirigid isosteres of the conformationally flexible GABA analogues (S)-(+)- and (R)-(-)-GABOB, respectively, which show a very low degree of enantioselectivity with respect to GABA synaptic mechanisms. This correlation between the degree of enantioselectivity and conformational mobility of chiral GABA analogues might be of importance for the design of new drugs with specific actions at synapses at which GABA is the transmitter.     

Synthesis and chromatographic separation of the glucuronides of (R)- and (S)-propranolol

One of the major metabolites of propranolol (Inderal) is the O-glucuronide. In order to further study its disposition, possible metabolism, and contribution to the antihypertensive effect of propranolol, we have synthesized and separated the two diastereomeric propranolol O-beta-D-glucuronides (9a,b). These compounds were prepared by reaction of naphthol with epichlorohydrin and treatment of the resulting (2RS)-1'-(2,3-epoxypropoxy)naphthalene (2) with sodium azide to give (2RS)-1-(1'-naphthoxy)-3-azido-2-propanol (3). Alkylation of 3 with methyl (2,3,4-tri-O-acetyl-1-bromo-1-deoxy-alpha-D-glucopyranosid)uronate (4) gave methyl (2RS)-[1-(1'-naphthoxy)-3-azido-2-propyl-2",3",4"-tri-O-acetyl-beta-D- glucopyranosid]uronate (5a,b). Reductive alkylation, followed by HPLC separation of the diastereomers, gave methyl (2R)- and (2S)-[1-(1'-naphthoxy)-3-(isopropylamino)-2-propyl-2",3",4"-tri-O-acetyl- beta-D-glucopyranosid]uronate (6a,b). Hydrolytic removal of the acetyl and methyl protecting groups gave the free glucuronides, which were then converted to the sodium salts, 9a,b. The stereochemistry of the glycoside linkage was deduced from the 400-MHz 1H NMR spectra. The absolute configuration of the aglycon portion was determined after Glusulase hydrolysis by derivatization with (R)-(+)- or -(-)-alpha-methylbenzyl isocyanate and comparison of the HPLC retention volumes with those of derivatized reference (R)- and (S)-propranolols.     

The differential activities of R (+)- and S(-)-zacopride as 5-HT3 receptor antagonists

R(+)- and S(-)-zacopride were assessed as potential 5-HT3 receptor antagonists in behavioural and biochemical tests. The S(-)isomer was more potent than the R(+)isomer to antagonise the hyperactivity induced by the injection of amphetamine or the infusion of dopamine into the nucleus accumbens in the rat. In contrast, the R(+)isomer was more potent to reduce the aversive behaviour of mice to a brightly illuminated environment and in a marmoset human threat test, to facilitate social interaction in rats, to increase performance in a mouse habituation test and prevent a scopolamine-induced impairment, and to antagonise the inhibitory effect of 2-methyl-5-hydroxytryptamine to reduce [3H]acetylcholine release in slices of the rat entorhinal cortex. In binding assays, [3H]S(-)-zacopride and [3H]R(+)-zacopride labelled homogenous populations of high-affinity binding sites in the rat entorhinal cortex, R(+)-zacopride compete for a further 10 to 20% of the binding of [3H]R(+)/S(-)-zacopride or [3H]R(+)-zacopride in excess of that competed for by (S)(-)-zacopride. It is concluded that both isomers of zacopride have potent but different pharmacological activities, with the possibility of different recognition sites to mediate their effects.     

Stereoselective determination of (beta S,gamma R and beta R,gamma S)-4-[3-(4-acetyl-3-hydroxy-2-propylphenoxy) propylthio]-gamma-hydroxy-beta-methylbenzenebutanoic acid in human and rat plasma by normal-phase high-performance liquid chromatography

A stereoselective high-performance liquid chromatographic method was developed for the determination of the enantiomeric composition comprising (beta S,gamma R and beta R,gamma S)-4-[3-(4-acetyl-3-hydroxy-2-propylphenoxy) propylthio]-gamma-hydroxy-beta-methylbenzenebutanoic acid in human and rat plasma. Both enantiomers, (beta S,gamma R)- and (beta R,gamma S)-1, were isolated from the plasma by liquid-liquid extraction, the organic phase was evaporated to dryness, and the residue was lactonized with p-toluenesulfonic acid followed by derivatization with R(+)-alpha-methylbenzylamine to form diastereomeric Schiff base adducts. Separation was performed on a silica column (Supelcosil) with a mobile phase composed of 97.8% hexane, 2% isopropyl alcohol, and 0.2% triethylamine. The column eluant was monitored with a variable wavelength UV detector set at 280 nm (0.005 AUFS). Based on the peak area ratios, the method shows excellent linear relationships with their corresponding concentrations and satisfactory intraday and interday assay precision and accuracy. The detection limit of this method is 25 ng/mL in plasma for each enantiomer. The method has been applied to the assay of plasma obtained from human subjects administered 1 in safety and tolerability studies and from rats administered 1 intravenously or per os.     

Lack of effect of cimetidine on the pharmacokinetics of R(-)- and S(+)-ibuprofen.

1. To investigate the effect of cimetidine on the pharmacokinetics of R(-)- and S(+)-ibuprofen, six healthy male volunteers received orally 800 mg racemic ibuprofen both in the drug-free state (control phase, C) and on the second day of a 3 day course of oral cimetidine, 1 g daily (treatment phase, T). The two phases (14 days apart) were randomised in a balanced cross-over manner. 2. The plasma concentrations of R(-)- and S(+)-ibuprofen were measured by high-performance liquid chromatography (h.p.l.c.). The protein binding of the enantiomers was assessed in a selection of plasma samples from each volunteer. Following alkaline hydrolysis of glucuronide conjugates, the urinary recoveries of ibuprofen and its major metabolites were measured by h.p.l.c. 3. There was no difference (P greater than 0.05, two-tailed Student's t-test; data expressed as mean +/- s.d.) between C and T phases in the total area under the plasma concentration-time curve of R(-)-ibuprofen (C 4514 +/- 1063 mg 1(-1) min vs T 4665 +/- 1435 mg 1(-1) min) and S(+)-ibuprofen (C 6460 +/- 1063 mg 1(-1) min vs T 6886 +/- 1207 mg 1(-1) min). Similarly, for each enantiomer, there was no difference between the two phases in the terminal half-life, the maximum plasma concentration or the time of its occurrence. 4. Cimetidine treatment had no effect (P greater than 0.05) on the time-averaged percent unbound in plasma of R(-)-ibuprofen (C 0.419 +/- 0.051% vs T 0.435 +/- 0.060%) and S(+)-ibuprofen (C 0.643 +/- 0.093% vs T 0.633 +/- 0.053%). (ABSTRACT TRUNCATED AT 250 WORDS)     

GABA agonists and uptake inhibitors. Synthesis, absolute stereochemistry, and enantioselectivity of (R)-(-)- and (S)-(+)-homo-beta-proline

The cyclic analogue of 4-aminobutyric acid (GABA), 3-pyrrolidineacetic acid (homo-beta-proline), is a potent agonist at GABAA receptors, it interacts effectively with GABA-uptake mechanisms, and it is a moderately potent inhibitor of GABAB receptor binding. (R)-(-)- (10) and (S)-(+)-homo-beta-proline (15) were synthesized via methyl (3S)-1-[(R)-1-phenylethyl]-5-oxo-3-pyrrolidinecarboxylate (5) and its 3R diastereomer (4), respectively. The mixture 3 consisting of 4 and 5 was synthesized via addition-cyclization reactions between (R)-1-phenylethylamine and itaconic acid (1). The diastereomers 5 and 4, which were separated chromatographically, were converted into (R)- (10) and (S)-homo-beta-proline (15), respectively. The absolute stereochemistry of 10 and 15 was established on the basis of an X-ray analysis of compound 5. The enantiomers 10 and 15 were shown to bind to GABAA and GABAB receptor sites with opposite stereoselectivity. Thus, (R)-homo-beta-proline (10) proved to be more than 1 order of magnitude more potent than the S enantiomer (15) as an inhibitor of GABAA receptor binding, whereas the GABAB receptor affinity of homo-beta-proline was shown to reside exclusively in (S)-homo-beta-proline (15). In contrast to the stereoselective receptor affinities of 10 and 15, these enantiomers were approximately equieffective as inhibitors of synaptosomal GABA uptake.     

Pharmacological differences between R(-)- and S(+)-ibuprofen

Ibuprofen (IBU) is a non-steroidal anti-inflammatory drug exhibiting optical isomerism. Only the racemate is in clinical use. In in vitro studies it has been demonstrated that only the S(+)-enantiomer inhibits the PG synthetase system. Nevertheless, it is widely believed that the sole use of the active isomer does not comprise any advantages since the inactive isomer is converted within the human body. In a triple cross-over study (300 mg S(+), 300 mg R(-), 600 mg racemic IBU; n = 8), we could show that the converted R(-)-IBU after racemate administration provides for only one third of the AUC of S(+)-IBU obtained after S(+)-application. Highest S(+)-peak plasma levels were reached after S(+)-IBU, lower ones after racemate. We, therefore, studied 4 patients with classical rheumatoid arthritis treated with 2-3 doses of 500 mg of S(+)-IBU/day over a two week period. Significant clinical recovery (Ritchie-index p less than 0.01; analogue scale pain p less than 0.05, motion p less than 0.01) was reached after one week. The results indicate that a reduction of dose and of metabolic load is possible if the S(+)-enantiomer is administrated.     

R(-) and S(+) stereoisomers of 11-hydroxy- and 11-methoxy-N-n-propylnoraporphine: central dopaminergic behavioral activity in the rat

R(-)11-Hydroxy-N-n-propylnoraporphine (11-OH-NPa) induced stereotyped behavior in the rat as potently (ED50 = 0.80 mg/kg, i.p.) as R(-)apomorphine (APO) and this effect was blocked by haloperidol; the 11-methoxy congener, R(-)11-MeO-NPa, had a weak effect (ED50 greater than 10 mg/kg) and the S(+) isomers had none. The isomer R(-)11-OH-NPa potentiated locomotion stimulated by apomorphine; S(+)11-OH-NPa inhibited it and the isomers of 11-MeO-NPa were inactive. Catecholaporphines usually are inactive orally, but both R(-) and S(+)11-OH-NPa were similarly potent after oral or parenteral administration. The isomer S(+)11-OH-NPa inhibited spontaneous and apomorphine-induced locomotion (ID50 = 1.8-2.7 mg/kg, p.o. and i.p.) and stereotyped behavior (ID50 = 3 mg/kg, p.o. or i.p.), all without inducing catalepsy. While apomorphine was short-acting (1-2 hr), the effects of R(-)11-OH-NPa persisted up to 6-7 hr and those of the S(+) isomer for at least 2.5 hr; moreover, the efficacy of R(-)11-OH-NPa increased markedly up to 3-4 hr, although its ED50 was independent of time (ED50 = 1.7-1.9 mg/kg, i.p. from 1-3 hr). The total effect of R(-)11-OH-NPa (p.o. or i.p.) over time was more than 10-times greater than that of injected apomorphine. These observations accord with the reported high (nM) affinity of 11-OH-NPa at cerebral DA receptor sites (D2 greater than D1) and weak interactions of the 11-methoxy congener. They support the conclusion that the R(-) and S(+) stereoisomers are neuropharmacologically active, respectively, as DA agonist and apparent antagonist, as was found with the enantiomers of N-n-propylnorapomorphine, perhaps due to the low intrinsic postsynaptic agonist activity of the S(+) isomers. Moreover, 11-OH-NPa was highly bioavailable orally and unusually long-acting; it may be absorbed slowly or have active metabolites. Hydroxy-substitution of aporphines at the 11-position, homologous to the 3-OH of DA, evidently is critical for affinity and activity at the DA receptor. These or other monohydroxyaporphines may represent leads to potentially useful DA agonist or antagonist drugs.     

Cardiac and hemodynamic effects of S(-)- and (R(+)-DPI 201-106 and of racemic DPI 201-106 in conscious dogs

The cardiac and hemodynamic effects of (+/-)-DPI 201-106 (0.6 mg/kg), S(-)-DPI (0.3 mg/kg), R(+)-DPI (0.3 mg/kg), and their vehicle were compared in chronically implanted conscious dogs. (+/-)-DPI and S(-)-DPI induced qualitatively and quantitatively similar effects, increasing LV dP/dt, cardiac output and stroke volume and reducing total peripheral resistance. In contrast, R(+)-DPI decreased only stroke volume. It is concluded that (a) the positive inotropic effects of (+/-)-DPI in vivo are caused by its S(-) enantiomer, and (b) the peripheral vasodilating effects of (+/-)-DPI may not be linked to the drug's calcium antagonist properties.     

Probes for narcotic receptor mediated phenomena. 15. (3S,4S)-(+)-trans-3-methylfentanyl isothiocyanate, a potent site-directed acylating agent for the delta opioid receptors in vitro

Recently we reported the synthesis of the first enantiomeric pair of irreversible opioid ligands [(3S,4R)-(-)- and (3R,4S)-(+)-cis-4, SUPERFIT] and specific interaction of the latter with the delta receptor. Here we report another enantiomeric pair of irreversible opioid ligands, (+)-trans- and (-)-trans-3-methylfentanyl isothiocyanates [(3S,4S)-(+)-trans- and (3R,4R)-(-)-trans-4]. A single-crystal X-ray analysis of the 2,4,6-trinitrobenzenesulfonic acid salt of (+)-trans-3-methyl-N-phenyl-4-piperidinamine [(+)-trans-8] revealed it (and, therefore, 4) to have the trans configuration and the absolute configuration of (+)-trans-8 to be 3S,4S. The (+)-trans enantiomer of 4 was shown to be highly potent and about 10-fold more selective as an acylating agent than (-)-trans-4 for the higher affinity [3H]DADL (delta) binding site in rat brain membranes. In that assay, (+)-trans-4 and (+)-cis-4 were essentially equipotent as affinity ligands, and the levo enantiomers were considerably less potent. (+)-trans-4 was, thus, a potent, subtype-selective acylating agent for the delta opioid receptor in vitro. With membranes from NG108-15 neuroblastoma x glioma hybrid cells, containing only delta receptors, (+)-cis-4 was found to be a little more potent than (+)-trans-4. Similarly, (+)-cis-4 is the most effective inhibitor of adenylate cyclase in these membranes, (+)-trans-4 has weak activity, and the levo enantiomers are inactive. Only (+)-cis-4 was found to have antinociceptive activity in vivo.     

Probes for narcotic receptor mediated phenomena. 12. cis-(+)-3-Methylfentanyl isothiocyanate, a potent site-directed acylating agent for delta opioid receptors. Synthesis, absolute configuration, and receptor enantioselectivity

The first enantiomeric pair of irreversible opioid ligands [(+)- and (-)-4] were synthesized in greater than 99.6% optical purity as determined by HPLC analysis of diastereoisomeric derivatives of the intermediate 3-methyl-N-phenyl-4-piperidinamine enantiomers. Single-crystal X-ray analysis of the (R,R)-L-(+)-tartaric acid salt of (-)-9 revealed the absolute configuration to be 3S,4R. The absolute configuration of (-)-3 [cis-(-)-3-methylfentanyl] and (-)-4 derived from (-)-9 is thus 3S,4R and that of (+)-3 and (+)-4 is 3R,4S. The (+) enantiomer of 4 (SUPERFIT) was shown to be highly potent and specific for acylation of delta opioid receptors (to the exclusion of mu) in rat brain membranes like its achiral prototype FIT and was about 10 times as potent as the latter in this assay. The (+)-4 was about 5 times as potent as FIT in acylation of delta receptors in NG108-15 neuroblastoma X glioma hybrid cells and about 50 times as potent as its enantiomer. Both FIT and (+)-4 behaved as partial agonists in inhibition of delta receptor coupled adenylate cyclase in NG108-15 membranes and (+)-4 was 5-10 times more potent than FIT and about 100 times more potent than its enantiomer in this assay. Dibromination of amine 12, catalytic exchange of bromine with tritium gas, and reaction of the labeled amine with thiophosgene afforded [3H]-(+)-4 with a specific activity of 13 Ci/mmol. Previous experiments indicated (+)-4 acylates the same 58 000-dalton glycoprotein previously shown to be acylated by FIT but with less nonspecific labeling. In view of the high potency and specificity of (+)-4 and the availability of its enantiomer, it seems likely that these compounds will prove to be valuable tools for study of the opioid receptor complex.     

Excitatory amino acid agonists. Enzymic resolution, X-ray structure, and enantioselective activities of (R)- and (S)-bromohomoibotenic acid

The enantiomers of alpha-amino-4-bromo-3-hydroxy-5-isoxazolepropionic acid (4-bromohomoibotenic acid, Br-HIBO, 1) a selective and potent agonist at one class of the central (S)-glutamic acid receptors, were prepared with an enantiomeric excess higher than 98.8% via stereoselective enzymic hydrolysis of (RS)-alpha-(acetylamino)-4-bromo-3-methoxy-5-isoxazolepropionic acid (4) using immobilized aminoacylase. The absolute configuration of the enantiomers of Br-HIBO was established by X-ray crystallographic analysis, which confirmed the expected preference of the enzyme for the S form of the substrate 4. (S)- and (RS)-Br-HIBO were potent neuroexcitants on cat spinal neurones in vivo, while (R)-Br-HIBO was a very weak excitant. Correspondingly, the S enantiomer of Br-HIBO (IC50 = 0.34 microM) was considerably more potent than the R form (IC50 = 32 microM) as an inhibitor of [3H]-(RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid ([ 3H]AMPA) binding to rat brain synaptic membranes in vitro. In contrast, (S)- and (R)-Br-HIBO were approximately equipotent (IC50 values of 0.22 and 0.15 microM, respectively) as inhibitors of [3H]-(S)-glutamic acid binding in the presence of CaCl2. The enantiomers of Br-HIBO showed no significant affinity for those binding sites on rat brain membranes which are labeled by [3H]kainic acid or [3H]-(R)-aspartic acid.     

Synthesis and dopamine agonist and antagonist effects of (R)-(-)- and (S)-(+)-11-hydroxy-N-n-propylnoraporphine

The R-(-) and S-(+) enantiomers of 11-hydroxy-N-n-propylnoraporphine, (R)-3 and (S)-3, were synthesized in six steps from 1-(3-methoxy-2-nitrobenzyl)isoquinoline. Neuropharmacological evaluation of the R and S isomers (by affinity to dopamine receptor sites in rat brain tissues, induction of stereotyped behavior, and interaction with motor arousal induced by (R)-apomorphine in the rat) indicated that, similar to the 10,11-dihydroxy congener 2, both enantiomers can bind to dopamine receptors but that only (R)-3 activates them, whereas (S)-3 shows activity as a dopaminergic antagonist.     

Incorporation and retention of radiolabeled S-(+)-and R-(-)-methamphetamine and S(+)- and R(-)-N-(n-butyl)-amphetamine in mouse hair after systemic administration.

We examined the incorporation of unlabeled and tritiated enantiomers of methamphetamine (MA) and a more lipophilic analog N-(n-butyl)-amphetamine (BA) into the hair of pigmented (C57) and nonpigmented (Balb/C) mice after systemic administration. We also compared the ability of extraction methods to remove unlabeled and tritiated MA and BA enantiomers from the hair. R(-)-MA, S(+)-MA, [(3)H]R(-)-MA, [(3)H]S(+)-MA, R(-)-BA, S(+)-BA, [(3)H]R-(-)-BA, and [(3)H]S-(+)-BA were each administered to C57 and Balb/C mice (23 days of age) by i.p. injection at 8.8 mg/kg daily for 3 days. At 44 days of age, hair samples from the animals were treated with a brief methanol wash, a 24-h extraction with pH 6 phosphate buffer, and a final digestion in 1 N NaOH to free residual drugs from the hair. Labeled drugs in the extracts were quantitated by liquid scintillation counting. Unlabeled drugs were quantitated by gas chromatography/mass spectrometry (GC/MS). GC/MS analysis demonstrated MA and BA to be the major (>90%) species present in the blood during the 24 h after administration. Less than 10% of the MA was N-demethylated. No p-hydroxylated metabolites were found. Blood concentrations of tritiated MA and BA enantiomers measured by liquid scintillation counting agreed well with blood concentrations of unlabeled enantiomers measured by GC/MS. Hair concentrations of S(+)-MA were greater than those of R(-)-MA in both mouse strains, paralleling blood concentrations. There were no enantiomeric differences seen with BA hair accumulation in either strain of mouse. Significantly more MA and BA enantiomers were deposited in pigmented than in nonpigmented hair. With labeled and unlabeled compounds, approximately 30% of S(+)-MA and 60% of R(-)-MA in pigmented hair could be removed by a phosphate extraction. A significant amount of drug could not be removed from the hair by extraction. Greater amounts of drug could be extracted from nonpigmented hair than pigmented. Extracted and residual MA and BA concentrations in pigmented hair were significantly greater when labeled compounds were quantitated by liquid scintillation counting than when unlabeled compounds were quantitated by GC/MS. However, radiotracer and unlabeled drug concentrations were the same in nonpigmented hair. The results demonstrate that hair pigmentation is an important determinant in MA and BA deposition, and that MA and BA deposition is not enantioselective. The data demonstrate a significant amount of MA and BA accumulated is not easily amenable to exhaustive aqueous extraction from the hair. The use of tritiated MA and BA enantiomers demonstrates that a significant amount of MA and BA stored in pigmented hair is structurally different from parent MA and BA, perhaps associated with melanin components of hair.     

(S)-(+)-methadone is more immunosuppressive than the potent analgesic (R)-(--)-methadone

Methadone is a widely used synthetic opioid which is administered as a racemic mixture of (R)-(--)- and (S)-(+)-enantiomers, with only (R)-(--)-methadone possessing mu opioid receptor agonist activity. Methadone inhibits numerous immune functions in vitro at concentrations above 10 microM in a nonstereoselective and naloxone-insensitive fashion, suggesting the presence of nonclassical opioid receptors on immune cells. No in vivo data on the effects of methadone's enantiomers on immune function are available. Therefore, the stereoselectivity of methadone's analgesia (hot plate latency) in vivo and immune suppression ex vivo (splenocyte proliferation) was investigated in groups of Balb/c mice. Significant analgesia was observed in animals that received racemic methadone (P=0.0012, 52% MPE) and (R)-(--)-methadone (P=0.0002, 70% MPE) when compared to saline-treated controls, while (S)-(+)-methadone was devoid of any such effect (-4% MPE). In vivo (R)-(--)- and racemic methadone caused significant inhibition (P<0.001, greater than -70%) of basal proliferation compared to saline control. In stark contrast to analgesia, in vivo (S)-(+)-methadone caused significantly greater inhibition of basal proliferation (P<0.001, -130%) than (R)-(--)- and racemic methadone. The immune suppression caused by methadone is not purely a classical opioid response but involves nonclassical opioid receptors located at the central level, which have yet to be characterised. Moreover, the dose at which immune suppression occurred could be achieved clinically.