A submerged ceramic membrane reactor for the p-nitrophenol hydrogenation over nano-sized nickel catalysts.

The catalytic hydrogenation of p-nitrophenol to p-aminophenol over nano-sized nickel catalysts was carried out in a submerged ceramic membrane reactor. It has been demonstrated that the submerged ceramic membrane reactor is more suitable for the p-nitrophenol hydrogenation over nano-sized nickel catalysts compared with the side-stream ceramic membrane reactor, and the membrane module configuration has a great influence on the reaction rate of p-nitrophenol hydrogenation and the membrane treating capacity. The deactivation of nano-sized nickel is mainly caused by the adsorption of impurity on the surface of nickel and the increase of oxidation degree of nickel.

Stereoisomers of N-[1-hydroxy-(2-phenylethyl)-3-methyl-4-piperidyl]- N-phenylpropanamide: synthesis, stereochemistry, analgesic activity, and opioid receptor binding characteristics.

N-[1-(2-Hydroxy-2-phenylethyl)-3-methyl-4-piperidyl]-N-phenylpropanamide (ohmefentanyl,1) is an extremely potent analgesic agent with high affinity and selectivity for opioid mu receptors. There are three chiral carbons in 1, so eight optically active isomers are possible. Respective reaction of optically active 3-methyl-N-phenyl-4 -piperidinamines (5a-d) with (R)- or (S)-styrene oxide produced eight optically active intermediates which were subsequently converted to eight optically active isomers of 1 (1a-h). The absolute configurations of 1a-h were determined by X-ray analysis of (3R,4S,2'R)-(-)-cis-1a and (3R,4R,2'S)-(-)-trans-1g. The analgesic activity (mice, ip, hot plate) revealed their extreme stereodifferences; the ED50 values of (3R,4S,2'R)-(-)-cis-1a and (3R,4S,2'S)-(+)-cis-1b, which are the most potent isomers among eight isomers, were 0.004 65 (2990 times that of morphine) and 0.001 06 mg/kg (13 100 times that of morphine), respectively, while the corresponding antipodes 1d,c were the least potent compounds among the eight isomers. In agreement with pharmacological results, both 1a,b also had the highest receptor affinity and selectivity for the opioid mu receptor. The ratio of K(i)(DPDPE)&K(i)(DAMGO) was 22 800 for 1a and 22 500 for 1b. All isomers except 1c,d strongly inhibited the electrically evoked smooth muscle contraction of GPI and MVD but not that of RVD, and the inhibitory effects could be reversed by naloxone, which indicated that these compounds were potent mu agonists in GPI and MVD. There was a good linear correlation between the analgesic potencies (ED50) and the receptor affinities (K(i)(DAMGO)) or inhibitory effects (IC50) to GPI and MVD. These results suggested that the analgesic effects of ohmefentanyl are mediated by interaction between the agents and opioid mu receptors in the central nervous system and the 3R,4S configuration at the piperidine 3- and 4-carbon atoms and the S configuration at the phenylethyl 2-carbon atom are beneficial for analgesic potency and inhibitory effects in GPI and MVD and the same for an S or R configuration at the phenylethyl 2-carbon atom besides the 3R,4S configuration for receptor mu affinity and selectivity.

Electroacupuncture and morphine analgesia potentiated by bestatin and thiorphan administered to the nucleus accumbens of the rabbit.

The endogenous opioid peptide enkephalin (EK) is known to be degraded mainly by two enzymes, the dipeptidyl carboxypeptidase 'enkephalinase' and aminopeptidase. Microinjection of the enkephalinase inhibitor thiorphan or the aminopeptidase inhibitor bestatin into the nucleus accumbens of the rabbit produced a dose-dependent analgesic effect. This analgesic effect was totally reversed by the narcotic antagonist naloxone or by antibodies against [Met5]enkephalin (MEK) administered to the same site. Antibodies against [Leu5]enkephalin were not effective. Moreover, microinjection of thiorphan or bestatin into the nucleus accumbens resulted in a marked potentiation of the aftereffect of electroacupuncture (EA) produced analgesia, as well as the analgesia induced by a small dose of morphine. It is concluded that the analgesic effect elicited by EA and morphine is mediated, at least in part, by MEK-like immunoreactive substance(s) in the nucleus accumbens.

Analgesic activity and opioid receptor selectivity of stereoisomers of ohmefentanyl isothiocyanate.

Ohmefentanyl is a very potent and highly selective agonist for mu-opioid receptors. We now study analgesia, in vitro activity and opioid receptor affinity of the stereoisomers of ohmefentanyl isothiocyanate. We found that some isomers of ohmefentanyl isothiocyanate had a potent analgesic effect and that all isomers except (3R,4S,2'S)-ohmefentanyl isothiocyanate had a more potent inhibitory action on the electrically evoked contractions of mouse vas deferens than of guinea pig ileum. The inhibitory actions could be antagonized by naloxone. However, compared with the activity of the corresponding stereoisomers of ohmefentanyl, these ohmefentanyl isothiocyanates had significantly reduced analgesia and in vitro activity. They also inhibited the binding of [3H]DPDPE ([D-Pen(2),D-Pen(5)]enkephalin) and [3H]DAGO ([D-Ala(2),Mephe(4),Gly-ol(5)]enkephalin) to opioid receptors in mouse brain membranes. The inhibitory effect of stereoisomers of ohmefentanyl isothiocyanate at mu-opioid receptors was markedly lower than that of their parent compounds. The affinity of stereoisomers of ohmefentanyl isothiocyanate for delta-opioid receptors was, however, greater than or equal to that of their corresponding stereoisomers of ohmefentanyl. The results showed that the introduction of an isothiocyanato group into the phenyl ring in position-1 of ohmefentanyl reduced bioactivity and affinity to mu-opioid receptors but that the selectivity of these compounds for delta-opioid receptors was enhanced. Isomer (3R,4S,2'R)-ohmefentanyl isothiocyanate showed highest selectivity for delta-opioid receptors (K(i)(mu)/K(i)(delta)=13.6) and potent analgesic activity (ED(50)=0.25 mg/kg).

Comparison of physical dependence of ohmefentanyl stereoisomers in mice.

Stereo-structural difference of ohmefentanyl stereoisomers on analgesic action and receptor affinity has been studied. To assess the difference of ohmefentanyl stereoisomers in physical dependence, the potency of physical dependence was quantified by estimating the ED50 value of ohmefentanyl stereoisomers in the naloxone-precipitated jumping test in mice. Morphine was used to assess the method and as a drug of comparison. The results indicate that the degree of physical dependence of morphine can been quantified by estimating the ED50 value of morphine withdrawal jumping induced by naloxone. A significant difference was observed in withdrawal jumping ED50 values among ohmefentanyl stereoisomers. Of these isomers, F9202 and F9204 had similarly potent analgesic action, but very significant difference in naloxone precipitated withdrawal response. Dependent potency index of F9204 was 618-fold weaker than that of F9202. It is concluded that a stereo-structural difference in physical dependence is found to exist among ohmefentanyl stereoisomers. Compound F9204 displayed a strong analgesic action and weak physical dependent potency.

Interactions of some 4-anilinopiperidines and 4-phenylpiperidines with the mu, delta- and kappa-binding sites.

The binding and pharmacological profiles of some 4-anilinopiperidine and 4-phenylpiperidine analogues were determined. The potency of the compounds to displace the binding of selective tritiated ligands was measured in homogenates of guinea-pig brain at 25 degrees C. All the compounds tested had high affinity for the mu-binding site, low affinity for the delta-site and were almost inactive at the kappa-site. The compounds were also tested for agonist activity in the guinea-pig ileum and the vasa deferentia of the mouse and rat. The pharmacological and binding profiles were compared.

Quantitative comparison of ohmefentanyl isomers induced conditioning place preference in mice.

Differences of analgesia and withdrawal response among ohmefentanyl stereoisomers have been studied. In the present study, Quantitative comparison of reinforcing effects of ohmefentanyl stereoisomers and morphine was performed by using a conditioned place preference design in mice. Results showed that morphine and ohmefentanyl stereoisomers were able to increase significantly the time spent in the drug-paired side with respect to vehicle treated animals. A good linear correlation between doses of drugs and number of mice with place preference was found within a given dose range. On the basis of the dose-response curve analysis, ohmefentanyl stereoisomers displayed a significant difference in place preference ED50. The addictive index (analgesic ED50/place preference ED50) was used to assess the addictive potential of drugs. It was demonstrated that the addictive potential of ohmefentanyl stereoisomers did not exhibit a large difference as addictive index. Among these stereoisomers, the addictive potential of compound F9208 was markedly lower than that of morphine.

[Effects of ohmefentanyl at anesthetic dose on plasma levels of corticosterone, cortisol and antidiuretic hormone in rats].

Ohmefentanyl (OMF) is a new mu opioid receptor agonist with high affinity and selectivity, and possesses anesthetic activity. With radioimmunoassay, the plasma levels of cortisol (C), corticosterone (CS) and antidiuretic hormone (ADH) in rats were measured. The results indicated that no significant differences in the plasma C, CS and ADH levels were observed between the saline control group and the OMF-treated group. Trauma (bone-crush injury) increased significantly the plasma CS level. However, pretreatment with OMF 4.0 micrograms.kg-1 reduced markedly the CS plasma levels in trauma-treated rats. The results suggest that OMF anesthesia itself showed no obvious effect on the plasma concentration of C, CS and ADH, but blocked the hormoral stress responses such as the increment of plasma CS level caused by trauma stimulus.

[Opioid dependence and tolerance in the guinea-pig ileum in vitro].

A rapid and simple method for the quantitative determination of opioid dependence and tolerance in the guinea pig ileum was introduced. Dependence, as indicated by a strong contraction of the ileum when challenged with naloxone, was produced by incubating ileum from native guinea pig with opioid at 37 degrees C for 1-6 h. The response of the ileum to naloxone was time-dependent and directly related to the normorphine concentration in the incubation fluid (10-1000 nmol/L) and to the challenge dose of naloxone (10-1000 nmol/L). There were lower naloxone-precipitated withdrawal responses in the guinea pig ileum incubated with pethidine, nalorphine and ohmefentanyl than with fentanyl, U50488H and morphine. Buprenorphine showed no withdrawal response. In spite of rapid development of dependence, segments of ileum incubated with normorphine at low concentration (10-300 nmol/L) showed no or slight tolerance to normorphine (3-fold). Only by incubating with high concentration of normorphine (1 mumol/L) was high tolerance detected. The results indicate that opioid dependence and tolerance do not develop in parallel.

The choice of opioid receptor subtype in isolated preparations by ohmefentanyl.

Ohmefentanyl has significant inhibitory actions on the electrically evoked contractions of guinea pig ileum and mouse vas deferens. Their IC50 values are 0.15 nM and 0.89 nM, respectively. In the guinea pig ileum and the mouse vas deferens, both mu-antagonist naloxone and (mu + kappa)-antagonist Mr. 2266 antagonize readily the agonist action of ohmefentanyl, indicating that ohmefentanyl acts on mu-receptors in the guinea pig ileum and the mouse vas deferens. Like other mu-agonists, ohmefentanyl has agonist activity but no antagonist action in the rat vas deferens. In the rabbit vas deferens, the activity of ohmefentanyl is very weak. The IC50 value is 149 nM. These results further indicate that ohmefentanyl is a potent and selective mu-agonist.

[Characterization of 9 slowly dissociated opioid ligands azabicyclononances compounds to mu, delta, and kappa receptors].

In the receptor binding assay, the relative affinity ratios of P-7548 [3-(beta-phenylethyl)-9 beta-methoxy-9 alpha-(m-propionoxyphenyl)-3- azabicyclo (3,3,1)nonane], P-7556 [3-(beta-phenylethyl)-9 beta-methoxy-9 alpha-(m-benzoyloxyphenyl)-3- azabicyclo(3,3,1)nonane] and P-7618 [3-(beta-phenylethyl)-9 beta-methoxy-9 alpha-[m-(1-cyclopentyl- propionoxyphenyl)]-3-azabicyclo(3,3,1)nonane] were 56:16:1, 1:4:1, and 6:0.2:1 at mu, delta, and kappa sites, respectively. These compounds possessed a tight binding to mu receptor. After washed 4 times, they still inhibited the [3H]ohmefentanyl binding by 70-80%. In the guinea pig ileum, they showed potent and persistent agonist activities, 607, 303, and 181 times respectively that of normorphine. These effects were readily antagonized by naloxone and Mr2266. In the mouse vas deferens (MVD), they also possessed long-lasting agonist activities. The effect of P-7556 on MVD was not antagonized by naloxone and Mr2266, indicating that P-7556 acted on delta receptor in MVD. In the rabbit vas deferens, P-7548, P-7556, and P-7618 antagonized the effect of U-50488H. We conclude that these azabicyclononanes are a series of opioid ligands with mu, delta agonist and kappa antagonist activities.

Multiplicity of kappa opioid receptor binding in the rat cardiac sarcolemma.

In order to determine the opioid receptor binding in the rat cardiac sarcolemma, direct and displacement binding assays of tritiated bremazocine, a nonselective opioid, agonist and U69593, a specific kappa-receptor agonist, were performed in a sarcolemmal membrane preparation. Saturation studies revealed that there were substantial binding sites for both radioligands. The [3H]bremazocine had a binding capacity of 100 fmol/mg protein with a Kd of 3.4 nM while the binding of [3H]U69593 showed a Kd of 7.4 nM and a Bmax of 139 fmol/mg protein. Displacement binding assay showed that [D-Ala2, MePhe4, Gly-(ol)5]enkephalin (DAGO), a specific mu-agonist, exhibited no inhibitory effect on the binding of either [3H]bremazocine or [3H]U69593. [D-Ala2,D-Leu5]enkephalin (DADLE), a specific delta-agonist, had no inhibition on the binding of [3H]U69593, but displaced the binding of [3H]bremazocine at higher concentrations (10(-5)-10(-6)M). U50488, a specific kappa-agonist, completed with the binding of [3H]U69593 23 times more potently than with the binding of [3H]bremazocine. A residual binding sites of [3H]bremazocine was found with a Kd of 6.4 nM and a Bmax of 76 fmol/mg protein under conditions in which mu, delta and kappa 1 opioid bindings were suppressed by DAGO and DADLE at 200 nM and U50488 at 1 microM. DADLE at 5 microM, a concentration known to block kappa 2 binding sites, abolished the residual U50488-insensitive specific binding of [3H]bremazocine. Met5-Enkephalin-Arg6-Phe7, a specific kappa 2 ligand, displaced the binding of [3H]bremazocine much more readily than the binding of [3H]U69593. The present study provided evidence for the presence of two kappa-receptor subtypes in the cardiac sarcolemma: one U50488-sensitive and DADLE-insensitive, and the other U50488-insensitive and DADLE-sensitive. These two kappa-receptor binding subtypes may correspond to kappa 1 and kappa 2 receptor subtypes. Computer assisted non-linear regression analysis in the competition experiments of U50488 with [3H]U69593 afforded a best fit of the inhibition curves for two displacing components of kappa 1 binding sites: one with high affinity and low Bmax, another with low affinity and high Bmax.

Characterization of [3H]U69593 binding sites in the rat heart by receptor binding assays.

The binding properties of [3H]U69593, a selective k-ligand, in rat heart homogenates were characterized by direct and displacement receptor binding assays. It was found that there are substantial specific [3H]U69593 binding sites in the rat heart. They were saturable, reversible and stereospecific. Both association and dissociation rates were monophasic and the Scatchard plot was linear, indicating a homogeneity of binding sites. The Hill coefficient was close to 1, indicating an absence of cooperativity. The Bmax and Kd were 7.91 fmol/mg and 2.92 nM, respectively. The binding sites were most abundant in the right atrium, followed by right ventricle, left ventricle and left atrium in descending order. The study provides information on the properties and distribution of k-binding sites in the rat heart.

Effect of Tris, HEPES, and TES buffers on binding at mu-, delta-, and kappa-opioid sites in guinea pig brain.

In homogenates of guinea-pig brain minus cerebellum, the delta-binding of 1.5 nM [3H]-[D-Pen2,D-Pen5]enkephalin is little affected by the type and concentration of the three buffers, Tris-HCl, HEPES-KOH, or TES-KOH (10-75 mM). However, the mu-binding of 1 nM [3H]-[D]Ala2,MePhe4,Gly-ol5]enkephalin or the kappa-binding of 1.5 nM [3H]-U-69,593 is influenced by the choice of buffer. A suitable concentration of buffer for further analyses of opioid binding has been found to be 10 mM. At each site, the effects of MgCl2 on binding are the same whether 10 mM Tris-HCl, HEPES-KOH, or TES-KOH are used but variations of the effects of NaCl confirm the view that mu- and kappa-sites, but not delta-sites, are affected by choice of buffer. Furthermore, under some assay conditions the effects of NaCl and MgCl2 at the kappa-sites of guinea-pig cerebellum differ from their effects in brain minus cerebellum, indicating that these are differences of binding characteristics at the kappa-sites of these tissues.