Sie sind auf Seite 1von 7

C Pharmacology & Toxicology 2002, 91, 5763. Printed in Denmark .

All rights reserved

Copyright C ISSN 0901-9928

Afnities of Dihydrocodeine and its Metabolites to Opioid Receptors

Helmut Schmidt1, Stefan V Vormfelde2, Klaus Klinder3, Ursula Gundert-Remy4, Christoph H. Gleiter5, Gisela Skopp3, . Rolf Aderjan3 and Uwe Fuhr6
1 pharmazentrum frankfurt, Institute of Clinical Pharmacology, Johann-Wolfgang-Goethe-University, Theodor-SternKai 7, 60590 Frankfurt/Main; 2Department of Clinical Pharmacology, Georg-August-University, Robert-Koch-Str. 40, 37075 Gttingen; 3Institute of Legal Medicine, Ruprecht-Karls-University, Vostr. 2, 69115 Heidelberg; 4BgVV, Fachbereich Chemikalienbewertung, Postfach 330013, 14191 Berlin; 5Department of Clinical Pharmacology, EberhardKarls-University, Wilhelmstr. 56, 72074 Tbingen, and 6Institute of Pharmacology, Clinical Pharmacology, University of Cologne, Gleueler Str. 24, 50931 Kln, Germany

(Received November 6, 2001; Accepted February 11, 2002) Abstract: Dihydrocodeine is metabolized to dihydromorphine, dihydrocodeine-6-O-, dihydromorphine-3-O- and dihydromorphine-6-O-glucuronide, and nordihydrocodeine. The current study was conducted to evaluate the afnities of dihydrocodeine and its metabolites to m-, d- and k-opioid receptors. Codeine, morphine, d,l-methadone and levomethadone were used as controls. Displacement binding experiments were carried out at the respective opioid receptor types using preparations of guinea pig cerebral cortex and the specic opioid agonists [3H]DAMGO (m-opioid receptor), [3H]DPDPE (d-opioid receptor) and [3H]U69,593 (k-opioid receptor) as radioactive ligands at concentrations of 0.5, 1.0 and 1.0 nmol/l, respectively. All substances had their greatest afnity to the m-opioid receptor. The afnities of dihydromorphine and dihydromorphine-6-O-glucuronide were at least 70 times greater compared with dihydrocodeine (Ki 0.3 mmol/ l), whereas the other metabolites yielded lower afnities. For the d-opioid receptor, the order of afnities was similar with the exception that dihydrocodeine-6-O-glucuronide revealed a doubled afnity in relation to dihydrocodeine (Ki 5.9 mmol/ l). In contrast, for the k-opioid receptor, dihydrocodeine-6-O- and dihydromorphine-6-O-glucuronide had clearly lower afnities compared to the respective parent compounds. The afnity of nordihydrocodeine was almost identical to that of dihydrocodeine (Ki 14 mmol/l), whereas dihydromorphine had a 60 times higher afnity. These results suggest that dihydromorphine and its 6-O-glucuronide may provide a relevant contribution to the pharmacological effects of dihydrocodeine. The O-demethylation of dihydrocodeine to dihydromorphine is mediated by the polymorphic cytochrome P450 enzyme CYP2D6, resulting in different metabolic proles in extensive and poor metabolizers. About 7% of the caucasian population which are CYP2D6 poor metabolizers thus may experience therapeutic failure after standard doses.

The semisynthetic opioid dihydrocodeine is an analgesic and antitussive drug. It exerts its actions by interaction with opioid receptors of different types (m-, d- and k-opioid receptors). Since the beginning of the 1960s dihydrocodeine and dihydrocodeine-containing drugs were used by opioid addicts as a substitute drug to heroin. Recently this use increased by prescribing dihydrocodeine in maintenance treatment of addicts. Individual doses prescribed vary from 60 to 3000 mg dihydrocodeine daily (Frieem & Tschner 1991). Dihydrocodeine, the 7,8-dihydro analogue of codeine, has similar metabolic pathways as codeine (Fromm et al. 1995; Hufschmid et al. 1995). These include O-demethylation to dihydromorphine, formation of the corresponding 6- and/ or 3-O-glucuronides dihydrocodeine-6-O-, dihydromorPart of these results have been presented at the 7th Annual Conference of the German Society for Clinical Pharmacology and Therapeutics, May 9, 1997, Mannheim, Germany (Schmidt et al. 1997). Author for correspondence: Helmut Schmidt, pharmazentrum frankfurt, Institute of Clinical Pharmacology, Johann-WolfgangGoethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt/ Main, Germany (fax 496963017636, e-mail Helmut.Schmidt /

phine-3-O- and dihydromorphine-6-O-glucuronide, and Ndemethylation to nordihydrocodeine (g. 1). It has been shown that O-demethylation of dihydrocodeine to dihydromorphine is mediated by the polymorphic cytochrome P-450 enzyme CYP2D6 (Fromm et al. 1995; Kirkwood et al. 1997). About 7% of the caucasian population are poor metabolizers that lack the functional CYP2D6 enzyme. They are therefore able to form only small amounts of dihydromorphine and its conjugates. In contrast, in at least 1%, additional copies of the gene are present, resulting in increased enzyme expression (Meyer & Zanger 1997). The contribution of the various metabolites to the therapeutic effects of dihydrocodeine is not known but may be important in the light of different metabolic proles between patients. Thus, the objective of the current study was to evaluate the afnities of dihydrocodeine and its metabolites to cerebral m-, d- and k-opioid receptors as the basis of their pharmacological action. Materials and Methods
Reagents. All chemicals were of analytical or research grade and were purchased from the indicated sources if not otherwise stated: [3H]DAMGO ([tyrosyl-3,5-3H(N)]Tyr-D-Ala-Gly-N-Methyl-Phe-



Fig. 1. Chemical structure of dihydrocodeine and its essential metabolic steps.

Gly-ol, 55.3 Ci/mmol), [3H]DPDPE ([tyrosyl-2,6-3H(N)]enkephalin (2-D-penicillamine, 5-D-penicillamine), 36.0 Ci/mmol) and [3H]U69,593 ([phenyl-3,4-3H]U-69,593, [phenyl-3,4-3H](5a,7a,8b)()-N-methyl-N-(7-(1-pyrrolidinyl)-1-oxaspiro(4,5)dec-8-yl)benzeneacetamide, 47.5 Ci/mmol), NEN Life Science Products GmbH, Kln, Germany; codeine, DAMGO, DPDPE and polyethylenimine, Sigma-Aldrich Chemie GmbH, Steinheim, Germany; U69,593, Alexis Deutschland GmbH, Grnberg, Germany; Tris and sucrose, Carl Roth GmbH Co, Karlsruhe, Germany; Quicksafe A scintillator, Zinsser Analytic GmbH, Frankfurt/Main, Germany; dihydrocodeine, Knoll Deutschland GmbH, Ludwigshafen, Germany; dihydromorphine was synthesized from hydromorphone by reduction with NaBH4 (Becker et al. 1986); dihydrocodeine-6-O-, dihydromorphine-3-O- and dihydromorphine-6-O-glucuronide, and nordihydrocodeine, Lipomed AG, Arlesheim, Switzerland; levomethadone, Aventis Pharma Deutschland GmbH, Frankfurt/Main, Germany; d,l-methadone, Synopharm GmbH, Barsbttel, Germany; dichlorodimethylsilane, morphine and n-heptane, Merck Eurolab GmbH, Darmstadt, Germany. Preparation of cerebral membranes. Preparations of cerebral membranes of Pirbright-white guinea pigs (both sexes, weight 450550 g) were carried out essentially as described (Schwanstecher et al. 1992). The time between death and brain homogenization was about 10 min. Prehomogenization of tissue was done for one min. in 10 volumes of ice-cold Tris buffer (10 mmol/l Tris-HCl, 290 mmol/l sucrose, pH 7.4) using an electric food blender. The prehomogenate was homogenized with ten strokes of a teon glass homogenizer at 300 rpm. The homogenate was centrifuged at 1,000g and 4 for 15 min. The supernatant was centrifuged for 30 min. at 100,000g and 4 . The resulting pellet was rehomogenized in Tris buffer, immediately frozen in liquid nitrogen and stored at 80 until binding studies were performed. Frozen material remained stable for at least several months and was used within 3 months for the experiments. Measurement of protein concentration. The protein content in the incubation volume of 500 ml (containing 100 ml of membrane preparation) was between 0.13 and 0.29 mg determined according to Bradford (1976). Measurement of binding of the selective opioid agonists. The tritiated and non-tritiated peptides DAMGO and DPDPE were stored and

handled either in silanized glass or silanized polypropylene tubes with exception to the incubation periods. In these periods polystyrene tube-strips were used designed for the utilized cell harvester. Since polystyrene cannot be silanized we compared silanized polypropylen with polystyrene tubes but we found no signicant difference in receptor binding. Binding assays were performed for 120 min. at 37 with TrisHCl, 50 mmol/l, pH 7.4 in polystyrene tubes (Macrowell tube-strips, 12 tubes/strip, Zinsser Analytic GmbH, Frankfurt/Main, Germany). The incubation mixtures contained 0.5 nmol/l of the m-selective [3H]DAMGO (Kosterlitz & Paterson 1981), 1.0 nmol/l of the d-selective [3H]DPDPE (Cotton et al. 1985) or 1.0 nmol/l of the kselective [3H]U69,593 (Lahti et al. 1985) in inhibition binding experiments and 0.120 nmol/l of the three tritiated opioid agonists in saturation binding experiments. Unlabelled test opioids used to displace these ligands from the particular binding site included DAMGO, DPDPE, U69,593, dihydrocodeine, its metabolites dihydromorphine, dihydrocodeine-6-O-, dihydromorphine-3-O- and dihydromorphine-6-O-glucuronide, nordihydrocodeine and the control ligands morphine, codeine, d,l-methadone and levomethadone. The highest concentration of these substances was 1 mmol/l with exception of DHM3G and DHM6G (10 mmol/l) because of insufcient amounts of material. Non-specic binding was determined in the presence of 1 mmol/l up to 1 mmol/l of the different unlabelled ligands in each experiment. Incubations were terminated by rapid vacuum ltration through Whatman GF/B glass microbre lters, Merck Eurolab GmbH, Frankfurt/Main, Germany (25 mm diameter, wetted with incubation buffer containing 0.15% polyethylenimine for at least 1 hr at 4 ) using the Brandel Cell Harvester, Model M-12SP, Dunn Labortechnik GmbH, Asbach, Germany. Filters were washed twice with 5 ml incubation buffer at 4 . Filtration and washing took less than 15 sec. The lters were transferred to scintillation vials containing 4 ml of the scintillation cocktail. The [3H]-content on the lters was determined by a liquid scintillation counter (Canberra-Packard Liquid Scintillation Analyzer TRI-CARB 2100TR, Canberra-Packard GmbH, Dreieich, Germany). In saturation binding experiments non-specic binding was determined in the presence of 100 mmol/l of the particular unlabelled ligand. Data were transformed and tted by Scatchard Plot analysis and linear regression using the equation Bound/Free (BmaxBound)/Kd where Bound is the concentration of the speci-



binding) were converted into Ki values (equilibrium inhibition constants) according to the equation of Cheng & Prusoff (1973) Ki IC50/(1L/Kd) where L is the concentration of the tritiated opioid agonist. The equilibrium dissociation constants Kd were determined by displacement of the tritiated by the particular non-tritiated opioid agonists and were compared to the Kd values resulting from the saturation binding experiments.

Results Saturation binding experiments revealed Kd and Bmax values of 2.60.3 nmol/l and 8110.7 fmol/mg protein with [3H]DAMGO for the cerebral m-opioid receptor of the guinea pig (n4). With [3H]DPDPE at the cerebral d-opioid receptor the values were 3.10.4 nmol/l and 806.4 fmol/ mg protein (n4). At the cerebral k-opioid receptor the values were 3.50.3 nmol/l and 1428.2 fmol/mg protein with [3H]U69,593 (n3). Representative Scatchard Plots are shown in g. 2. Inhibition binding experiments in guinea pig cerebral membranes with the three tritiated radioactive ligands gave the displacement curves shown in g. 35. The Kd values for [3H]DAMGO, [3H]DPDPE and [3H]U69,593 were 2.20.2, 3.00.4 and 2.90.2 nmol/l, respectively. The Ki values for the displacing opioids are shown in table 1. The Hill coefcients for all compounds were in the range from 0.8 to 1.2 with the exception of d,l-methadone (1.6, m-opioid receptor), levomethadone (1.4, m- and d-opioid receptor) and DHM6G (0.6, d-opioid receptor). All active unlabelled substances had their greatest afnity to the m-opioid receptor and, with the exception of morphine, their lowest afnity to the k-opioid receptor. Binding of dihydromorphine, dihydromorphine-6-O-glucuronide and morphine at the m-opioid receptor exhibited the greatest afnities of all drug/receptor combinations tested. The afnities of dihydromorphine and its 6-O-glucuronide to the m-opioid receptor were at least 70 times higher,

Fig. 2. Representative Scatchard Plots of [3H]DAMGO, [3H]DPDPE and [3H]U69,593 binding to guinea pig brain membranes. Concentrations of the tritiated agonists were in the range from 0.1 to 20 nmol/l. Non-specic binding was determined in the presence of 100 mmol/l of the particular unlabelled ligand.

cally bound [3H]-ligand, Free the concentration of the free [3H]ligand, Bmax the maximal number of binding sites and Kd the equilibrium dissociation constant. Calculated values for Bound/Free and Bound were normalized to protein content. Kd (1/Kdslope) and Bmax (intersection with the x-axis) values are presented as the arithmetic meanS.E.M. for at least 3 independent experiments. One representative experiment is plotted for each compound. In inhibition binding experiments the percentage of the bound radioactivity on the lters in relation to the total radioactivity in the incubations was between 0.9 and 2.2% for the three opioid receptor types. The fraction of the non-specic binding in relation to the specic binding was in the range from 13.7 to 32.5% for the m-, d- and k-opioid receptor. Results were analyzed using the SigmaPlot 5.0 scientic graphic software (Jandel Scientic Software GmbH, Erkrath, Germany). Results are presented as the arithmetic meanS.E.M. for at least 3 independent experiments. Relations to drug concentrations were analyzed by tting a logistic function to the experimental data by a non-linear least-squares routine assuming a single binding site. Data were weighted by the reciprocal of the corresponding S.E.M. If the Hill coefcients were outside the range from 0.8 to 1.2 additive models for two and more independent binding sites were tested but could not be tted to the data. Calculated IC50 values (values of 50% inhibition of specic

Table 1. Ki values (nmol/l) for displacement of specic opioid receptor agonists by dihydrocodeine, its metabolites and control substances. Values represent meansS.E.M. of 38 experiments as indicated in g. 35. [3H]DAMGO (m-opioid receptor) Dihydrocodeine and its metabolites Dihydrocodeine Dihydrocodeine-6-O-glucuronide Dihydromorphine Dihydromorphine-3-O-glucuronide Dihydromorphine-6-O-glucuronide Nordihydrocodeine Control substances Codeine Morphine d,l-Methadone Levomethadone DAMGO DPDPE U69,593 32512 54915 2.50.2 10000 4.50.5 43015 58917 4.90.3 24.82.1 9.31.1 2.20.2 [3H]DPDPE (d-opioid receptor) 5905359 3089144 13712 10000 11110 7074122 11442616 27312 54327 24015 3.00.4 2.90.2 [3H]U69,593 (k-opioid receptor) 14242466 39516520564 2237 10000 6743537 13707277 18061793 2063 167453 149245



Afnities of opioid moieties to m-opioid receptors depending on structural properties. Values represent Ki values in nmol/l. To compare the results the variability of the Ki values is presented as standard deviation. Literature data included for comparison are indicated by superscripts. Experimental conditions were not identical to those in the present study in all cases, for details see references. Dihydrocodeine 2.50.5 Codeine Reference opioid Dihydromorphine Morphine n. a.

Afnities of 3-O-demethyl derivatives 4.90.7 n. a. 1.20.3a 1.80.6c Afnities of 7,8-dihydro derivatives 32527 n. a. 24020c Afnities of N-demethyl derivatives 267108a no data Afnities of 6-O-glucuronides 4.50.9 23992a 790110c Afnities of 3-O-glucuronides n. a. 10000

n. a.


43033 54934 270100c

4.70.5a 0.60.5a 8.911.11.21.4b 3.51.1c 3720a 36050c

n. a.

n. a., not applicable; aChen et al. (1991); bLoser et al. (1996); cMignat et al. (1995).

whereas the other metabolites had lower afnities than dihydrocodeine. In detail, 6-O-glucuronidation of dihydromorphine only slightly affected the afnity to m-receptors, while the 3-O-glucuronide had a much lower afnity. Dihydrocodeine and dihydromorphine revealed two-fold greater afnities than their 7,8-unsaturated analogues. Dihydrocodeine also had a two-fold greater afnity than its 6-O-glucuronide. A comparison between the effects of glucuronidation, 7,8-dihydrogenation, N-dealkylation and 3-O-dealky-

lation on m-receptor afnities of the opioid moieties is shown in table 2. With respect to the d-opioid receptor, afnities were about 5 to 50 times lower than those for binding at the mopioid receptor, but the order of afnities was similar with the exception that dihydrocodeine-6-O-glucuronide revealed a two-fold higher afnity compared with dihydrocodeine. Displacement of [3H]U69,593 from k-opioid receptors resulted in Ki value relations different to the other opioid

Fig. 3. Displacement curves of the m-opioid receptor ligand [3H]DAMGO (0.5 nmol/l) by DAMGO, dihydrocodeine, its metabolites and control substances as indicated. Points represent meansS.E.M. of 48 experiments.



Fig. 4. Displacement curves of the d-opioid receptor ligand [3H]DPDPE (1.0 nmol/l) by DPDPE, dihydrocodeine, its metabolites and control substances as indicated. Points represent meansS.E.M. of 47 experiments.

receptor types. The 6-O-glucuronides of dihydrocodeine and dihydromorphine revealed clearly lower afnities compared to the respective parent compounds whereas morphine/dihydromorphine (Ki values 206 and 223 nmol/l, respectively), codeine/dihydrocodeine (Ki values 18061 and 14242 nmol/l, respectively) and d,l-methadone/levomethadone (Ki values 1674 and 1492 nmol/l, respectively) had almost identical afnities at the k-opioid receptor. At the m- and d-opioid receptor d,l-methadone yielded only half of the afnity of levomethadone leading to the

conclusion that d-methadone has no afnity at these receptor types. However, at the k-opioid receptor the afnities of d,l-methadone and levomethadone were almost identical with the result that d-methadone has an afnity comparable to that of levomethadone. Discussion In the present study, the afnity proles of dihydrocodeine, of ve of its most important metabolites and of control

Fig. 5. Displacement curves of the k-opioid receptor ligand [3H]U69,593 (1.0 nmol/l) by U69,593, dihydrocodeine, its metabolites and control substances as indicated. Points represent meansS.E.M. of 38 experiments.



opioids at the three opioid receptor types were determined in guinea pig cerebral cortex membranes. The binding properties of opioids in this tissue have been shown to closely reect the binding to cloned human receptors (Knapp et al. 1995). Afnities of control substances presented were similar to those reported in the literature. Modications of the different opioid molecules at corresponding positions appear to have similar effects on the afnity to the m-receptor, the main site of opioid action (table 2). While glucuronidation at C-6 or N-demethylation did not change afnity or were related to a small decrease in afnity only, O-demethylation brought about a pronounced increase in afnity. 7,8-Dihydro derivatives had slightly higher afnities when compared to their unsaturated counterparts (Chen et al. 1991; Mignat et al. 1995; Loser et al. 1996). 3-O-Glucuronides had essentially no relevant afnity to opioid receptors, and the apparent displacement of the radioactive ligands observed may well result from contamination with the more active parent substances (Bartlett & Smith 1995; Lser et al. 1996). Thus, it seems that 6-glucuronide metabolites may provide some contribution to the effects of any opioid, depending on the actual concentration at the binding site. E.g. morphine-6-O-glucuronide was found to exert analgesic effects in several clinical studies (Ltsch & Geisslinger 2001). Upon administration of 3-O-methyl derivatives it appears that the 3-O-demethylated metabolites, because of their much greater afnities, may mediate an important part of the pharmacological effects even when formed only in small amounts. Individual diversity in expression and activity is present for any of the enzymes involved in the metabolism of opiates. As a consequence, there are individual patterns of metabolite formation of dihydrocodeine. This appears to be most important for the formation of dihydromorphine and its 6-O-glucuronide having the greatest afnity to the mopioid receptor. This metabolic step in man depends on the activity of CYP2D6, which shows a genetic polymorphism (Kroemer & Eichelbaum 1995; Meyer & Zanger 1997). Indeed, after a single oral dose of 60 mg dihydrocodeine hydrogentartrate the percentage of urinary recovery of dihydromorphine and its glucuronides was 8.9% in CYP2D6 extensive metabolizers, in contrast to the 1.3% observed in poor CYP2D6 metabolizers (Fromm et al. 1995). In addition, there is no saturation of CYP2D6-dependent O-demethylation of dihydrocodeine in extensive metabolizers (Ammon et al. 1999), even when administered in daily doses up to 1350 mg (Mikus et al. 1998). Thus, the pharmacological effects of dihydrocodeine may be lower in poor metabolizers and more pronounced in ultrarapid metabolizers. Corresponding results have been reported for the effects of codeine depending on the formation of its metabolite morphine via CYP2D6 (Poulsen et al. 1996; Eckhardt et al. 1998) but these are not unequivocal (Vree et al. 2000). Results concerning dihydrocodeine were also contradictory (Fromm et al. 1995; Jurna et al. 1997; Wilder-Smith et al.

1998). Therefore, clinical studies addressing efcacy and safety of dihydrocodeine in man depending on the CYP2D6 phenotype are required, especially in addicts receiving dihydrocodeine in maintenance treatment in widely different doses up to 3000 mg daily (Frieem & Tschner 1991). Because the relative afnity of dihydrocodeine and dihydromorphine was the same at all opioid receptors tested, clinical effects like analgesia and clinically relevant common side effects of opioids irrespective of the receptor type involved may be used in such studies to assess the role of the metabolite pattern in dihydrocodeine therapy. Acknowledgements The study was supported by the Bundesministerium fr Gesundheit, Germany (grant 3221720/8). We are grateful to Dr. Linz, Dr. Hock and their coworkers from Aventis Pharma Deutschland GmbH, Frankfurt/Main, Germany, for providing access to guinea pig brain membrane preparations.

Ammon, S., U. Hofmann, E. U. Griese, N. Gugeler & G. Mikus: Pharmacokinetics of dihydrocodeine and its active metabolite after single and multiple oral dosing. Brit. J. Clin. Pharmacol. 1999, 48, 317322. Bartlett, S. E. & M. T. Smith: The apparent afnity of morphine3-glucuronide at mu1-opioid receptors results from morphine contamination: demonstration using HPLC and radioligand binding. Life Sci. 1995, 57, 609615. Becker, H. G. O., G. Domschke, E. Fanghnel, M. Fischer, K. Gewald, R. Mayer, D. Pavel, H. Schmidt & K. Schwetlick: Organikum. VEB Deutscher Verlag der Wissenschaften, Berlin, Germany, 1986. Bradford, M. M.: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 1976, 72, 248254. Chen, Z. R., R. J. Irvine, A. A. Somogyi & F. Bochner: Mu receptor binding of some commonly used opioids and their metabolites. Life Sci. 1991, 48, 21652171. Cheng, Y. & W. H. Prusoff: Relationship between the inhibition constant (K1) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. Biochem. Pharmacol. 1973, 22, 30993108. Cotton, R., H. W. Kosterlitz, S. J. Paterson, M. J. Rance & J. R. Traynor: The use of [3H]-[D-Pen2,D-Pen5]enkephalin as a highly selective ligand for the delta-binding site. Brit. J. Pharmacol. 1985, 84, 927932. Eckhardt, K., S. Li, S. Ammon, G. Schanzle, G. Mikus & M. Eichelbaum: Same incidence of adverse drug events after codeine administration irrespective of the genetically determined differences in morphine formation. Pain 1998, 76, 2733. Frieem, D. H. & K. L. Tschner: Codein und Dihydrocodein als Ausweich-und Ersatzdrogen. Fortschr. Neurol. Psychiatr. 1991, 59, 164169. Fromm, M. F., U. Hofmann, E. U. Griese & G. Mikus: Dihydrocodeine: a new opioid substrate for the polymorphic CYP2D6 in humans. Clin. Pharmacol. Therap. 1995, 58, 374382. Hufschmid, E., R. Theurillat, U. Martin & W. Thormann: Exploration of the metabolism of dihydrocodeine via determination of its metabolites in human urine using micellar electrokinetic capillary chromatography. J. Chromatogr. B Biomed. Appl. 1995, 668, 159170. Jurna, I., W. Komen, J. Baldauf & W. Fleischer: Analgesia by dihy-

DIHYDROCODEINE BINDING TO OPIOID RECEPTORS drocodeine is not due to formation of dihydromorphine: evidence from nociceptive activity in rat thalamus. J. Pharmacol. Exp. Therap. 1997, 281, 11641170. Kirkwood, L. C., R. L. Nation & A. A. Somogyi: Characterization of the human cytochrome P450 enzymes involved in the metabolism of dihydrocodeine. Brit. J. Clin. Pharmacol. 1997, 44, 549555. Knapp, R. J., E. Malatynska, N. Collins, L. Fang, J. Y. Wang, V. J. Hruby, W. R. Roeske & H. I. Yamamura: Molecular biology and pharmacology of cloned opioid receptors. Faseb J. 1995, 9, 516 525. Kosterlitz, H. W. & S. J. Paterson: Tyr-D-Ala-Gly-MePheNH(CH2)2OH is a selective ligand for the m-opiate binding site. Brit. J. Pharmacol. 1981, 73, 299P. Kroemer, H. K. & M. Eichelbaum: Its the genes, stupid. Molecular bases and clinical consequences of genetic cytochrome P450 2D6 polymorphism. Life Sci. 1995, 56, 22852298. Lahti, R. A., M. M. Mickelson, J. M. McCall & P. F. Von Voigtlander: [3H]U-69593 a highly selective ligand for the opioid kappa receptor. Eur. J. Pharmacol. 1985, 109, 281284. Lser, S. V., J. Meyer, S. Freudenthaler, M. Sattler, C. Desel, I. Meineke & U. Gundert-Remy: Morphine-6-O-beta-D-glucuronide but not morphine-3-O-beta-D-glucuronide binds to mu-, delta- and kappa- specic opioid binding sites in cerebral membranes. Naunyn Schmiedebergs Arch. Pharmacol. 1996, 354, 192 197. Ltsch, J. & G. Geisslinger: Morphine-6-glucuronide: an analgesic of the future? Clin. Pharmacokinet. 2001, 40, 485499. Meyer, U. A. & U. M. Zanger: Molecular mechanisms of genetic


polymorphisms of drug metabolism. Annu. Rev. Pharmacol. Toxicol. 1997, 37, 269296. Mignat, C., U. Wille & A. Ziegler: Afnity proles of morphine, codeine, dihydrocodeine and their glucuronides at opioid receptor subtypes. Life Sci. 1995, 56, 793799. Mikus, G., A. Ulmer, K. Mrike & U. Hofmann: Heroin substitution therapy: Pharmacokinetics of dihydrocodeine. Naunyn Schmiedebergs Arch. Pharmacol. 1998, 358 (Suppl. 2), P47.28. Poulsen, L., K. Brsen, L. Arendt-Nielsen, L. F. Gram, K. Elbk & S. H. Sindrup: Codeine and morphine in extensive and poor metabolizers of sparteine: pharmacokinetics, analgesic effect and side effects. Eur. J. Clin. Pharmacol. 1996, 51, 289295. Schmidt, H., S. V. Loser, K. Klinder, U. Gundert-Remy, N. Riet brock, G. Skopp, R. Aderjan & U. Fuhr: Afnity of dihydrocodeine-6-b-D-glucuronide and nordihydrocodeine to the cerebral m-opioid receptor of guinea pig. Eur. J. Clin. Pharmacol. 1997, 52, A7 (abstract). Schwanstecher, M., U. Schaupp, S. Lser & U. Panten: The binding properties of the particulate and solubilized sulfonylurea receptor from cerebral cortex are modulated by the Mg2 complex of ATP. J. Neurochem. 1992, 59, 13251335. Vree, T. B., R. T. van Dongen & P. M. Koopman-Kimenai: Codeine analgesia is due to codeine-6-glucuronide, not morphine. Int. J. Clin. Pract. 2000, 54, 395398. Wilder-Smith, C. H., E. Hufschmid & W. Thormann: The visceral and somatic antinociceptive effects of dihydrocodeine and its metabolite, dihydromorphine. A cross-over study with extensive and quinidine-induced poor metabolizers. Brit. J. Clin. Pharmacol. 1998, 45, 575581.