Chapter

Until recently, the only available method of treatment of opiate addicts in Germany was drug withdrawal and long-term behavioral therapy with abstinence from opiates as the goal. Since 1987 l-methadone (L-Polamidon©) has been used in methadone substitution programs (LMMT) but racemic methadone has not been legalised for this indication (German Drug Law; Art. 13 Abs. 1 BtmG) (Saß, 1991). The possibility of using racemic methadone instead of l-methadone in the treatment and maintenance of opiate addicts was first discussed in Frankfurt am main. The main reasons for this interest was the difference in costs of the two drugs, l-methadone being substantially more expensive than racemic methadone because of the more expensive production method and smaller production volume.

The analgesic activity of d,l-methadone is mainly due to the l-isomer. The analgesic potency ratio levo- : dextromethadone is 50 : 1 (Scott et al., 1948). In a study in ex-morphine addicts, opiate abstinence symptoms could be alleviated using l-methadone but not using d-methadone (Isbell and Eisenman, 1948). This research group also showed that d-methadone at high doses produced typical morphine-like responses but side effects were also seen (Fraser and Isbell, 1962). Sullivan et al. (1972) suggested that d-methadone metabolites may have an analgesic effect. In a study using 30 healthy, non-opiate tolerant volunteers, Olsen et al. (1976) found similar respiratory and pupillary effects using a 7.5 mg l-methadone dose and a 15 mg d,l-methadone dose. One subject showed slightly depressed respiration on high-dose d-methadone but the changes induced by l-methadone were of longer duration than those of d,l-methadone. In a double-blind study no significant difference was observed in methadone side effects in 66 subjects receiving methadone maintenance treatment (MMT) either as racemic methadone or as l-methadone (Judson et al., 1976). A recent pharmacodynamic study showed that l-methadone had a 10-fold higher binding affinity for µ₁ receptors than d-methadone (Kristensen et al., 1995). This is conclusive evidence that the l-methadone isomer is the main analgesic component in racemic methadone.

Comparisons of d-methadone and l-methadone pharmacokinetics in humans have shown ambiguous results. Kreek et al. (1979) obtained a longer elimination half-life for l-methadone in all three MMT subjects investigated. Beck et al. (1991) reported a single case in which the elimination half-life for l-methadone was shorter than that for d-methadone. These investigators also demonstrated that the l-methadone:d-methadone plasma concentration ratio differed not only between subjects but also within subjects. A between-subject difference in the l-methadone:d-methadone plasma concentrations ratio was also shown by Kristensen and Angelo (1992) who found a ratio of 0.8 (SD 0.2, range 0.5-1.1).

The plasma protein binding of methadone is stereoselective (Eap et al., 1990). In human plasma the unbound fraction of l-methadone is lower than that of d-methadone and this accounts for the lower bioavailability of l-methadone (Romach et al., 1981). The most important binding protein for methadone in plasma is α₁-acid glycoprotein suggesting that changes in the concentration of this binding protein e.g. due to acute or chronic inflammation, may alter the amount of unbound methadone. A recent study showed a significantly longer mean elimination half-life (37.5 h) and clearance (0.158 L/min) for l-methadone compared with d-methadone (Kristensen et al., 1996). Eap et al. (1996) found marked variability in the l-methadone:d-methadone plasma concentration ratio in opiate addicts who were changed from l-methadone maintenance treatment to racemic methadone treatment.

There are no published reports of qualitative differences in opiate craving between l-methadone and d,l-methadone treatment when the doses administered represent equivalent doses of l-methadone. The replacement of l-methadone by d,l-methadone in the clinical programs in Frankfurt am Main provided us with a unique opportunity to conduct a double-blind study of differences between l-methadone and d,l-methadone application during MMT. The specific aims of the study* were:

- to compare the doses of l-methadone and d,l-methadone required for control of opiate craving,

- to determine illicit use of opiates, cocaine and benzodiazepines,

- to measure plasma concentrations of methadone enantiomers and their main metabolite EDDP,

during l-methadone treatment and after the replacement of l-methadone by d,l-methadone.

* Results of this work have been presented in part at the 36 ^th Spring Meeting of the Deutsche Gesellschaft für experimentelle und klinische Pharmakologie und Toxikologie, Mainz 1995 [16] and at the 1^stCongress of the European Association of Clinical Pharmacology and Therapeutics, Paris 1995 (Staib et al., 1996).

Methods

Subjects and study design

A consecutive series of 40 subjects, currently on l-methadone maintenance treatment (LMMT) and being treated as outpatients by the Malteser-Hilfsdienst in Frankfurt am Main were included in the study. Participation in the study required written informed consent. The study received approval from the Ethics Committee of the University of Frankfurt am Main. Inclusion criteria were: males or females older than 18 years who had been receiving LMMT for at least 1 month. Exclusion criteria were: confirmed AIDS-disease (HIV positive subjects were included in the study), pregnancy and non-controllable illicit drug use. Subjects were dropped from the study in the event of: serious adverse reactions, non-compliance, personal or medical reasons and withdrawal of consent. At the start of the study the 40 subjects were randomly divided into two groups of 20 subjects and thereafter all treatments were carried out under double-blind cinditions. One group remained on l-methadone treatment throughout the 3-week period (l-methadone group), whereas the other group changed from l-methadone to d,l-methadone treatment (d,l-methadone group) at twice the dose of l-methadone on Day 8 .

Drug administration and blood sampling

The daily doses of l-methadone-HCl (L-Polamidon®, Hoechst, Germany) or d,l-methadone-HCl (Symoron®, Brocades, The Netherlands) were prepared as a methadone linctus 5 mg·ml^-1and individualised depending on the previous l-methadone-HCl usage. They received the daily oral dose at about noon. During the study period the dose was adjusted according to the subjects needs.

Venous blood samples were collected in heparinized tubes one or two days prior to the change from l-methadone to racemic methadone on Day 8. Further samples were collected on Day 15 and between Day 20 and Day 22. The samples were immediately centrifuged for 10 min at 1500 g and the supernatant plasma stored at -25° C until required for further analysis.

Sample preparation

Plasma (0.5 ml) was mixed with 50 µl methanol and either 100 µl trihexyphenidyl-HCl (2 µg·ml^-1; Procedure A) or 50 µl dextropropoxyphene-HCl (1 µg·ml^-1; Procedure B) as internal standard (IS) in a stoppered glass tube and made alkaline with 0.5 ml potassium carbonate. This mixture was extracted into 3.5 ml n-hexane by gentle agitation for 90 min at room temperature. After centrifugation (3000 rpm for 5 min) the tube contents were cooled to -25° C and the unfrozen upper layer (n-hexane layer) decanted and evaporated to dryness under a gentle stream of dry nitrogen. The dry residue was redissolved in 100 µl mobile phase. In order to prepare calibration curves the same procedures were performed using plasma from healthy, drug-free volunteers spiked with d,l-methadone-HCl (racemic methadone-HCl, Symoron®, Brocades, The Netherlands), l-methadone-HCl (l-methadone-HCl, L-Polamidon®, Hoechst, Germany) or EDDP-perchlorate (Sigma Chemical Co., USA) over a concentration range of 5-400 ng·ml^-1.

Analytical equipment

The HPLC system consisted of an HP 1050 Series with quarternary pump and variable wavelength detector (Hewlett Packard, USA), a Model 7125 sample injector (Rheodyne Inc., USA) fitted with a 50 µl loop and an HP 3395 integrator (Hewlett Packard, USA) in combination with a BD41 Recorder (Kipp & Zonen, The Netherlands).

In Procedure A separation was performed on a Supelcosil LC-ABZ column (50x4.6 mm ID) packed with 5-µm-diameter particles and protected by a 20-mm Supelguard column (Supelco, USA). The mobile phase consisted of potassium dihydrogenphosphate buffer (25 mM, pH 2.5) mixed with acetonitrile (78.5:21.5, v/v). The flow rate was set at 1.5 ml·min^-1 with UV detection at 206 nm.

In Procedure B enantioselective separation was performed on a Chiral-AGP column (100x4 mm ID) protected by a Chiral-AGP guard column (10x3.0 mm ID, ChromTech AB, Sweden). The mobile phase consisted of sodium dihydrogen phosphate buffer (10 mM, pH 5.0) mixed with acetonitrile and dimethyloctylamine (873:127:0.5, v/v). The flow rate was set at 0.9 ml·min^-1 with UV detection at 206 nm. The analytical procedure was performed in an air-conditioned room at approximately 20 C°.

Analytical procedures

In order to determine the plasma levels of EDDP and the two enantiomers of methadone, two separate procedures were performed. In Procedure A the plasma concentrations of EDDP and methadone (as a mixture of d-methadone and l-methadone or exclusively l-methadone) and EDDP were determined simultaneously. Using the Supelcosil LC-ABZ column under the above mentioned conditions the determination was found to be both sensitive and specific for both methadone (as a mixture of d-methadone and l-methadone or exclusively l-methadone) and its metabolite EDDP. Typical chromatograms of a solution in buffer (a), an extracted plasma sample spiked with d,l-methadone, EDDP and IS (b) and an extracted plasma sample from one of the opiate addicts (c) are shown in Fig. 1. Retention times for EDDP, IS and d,l- methadone were 2.5, 3.9 and 5.5 min respectively. Detection limits in plasma (signal-to-noise ratio of at least 3) were approximately 4 ng·ml^-1 for EDDP and 6 ng·ml^-1 for methadone. The calibration curves for both EDDP and d,l-methadone in plasma showed linearity in the concentration range 5-200 ng·ml^-1 (r = 0.982) and 20-400 ng·ml^-1 (r = 0.991) respectively. The calculated recovery values were 66.6±5.1% (n=20) for EDDP, 79.3±6.5% (n=20) for d,l-methadone and 92.7±3.8% (n=20) for IS.

Since Procedure B provides the ratio of l-methadone to d-methadone the plasma concentrations of each enantiomer of methadone can be calculated. Ratios between l-methadone and d-methadone in each plasma sample were calculated by dividing the peak area of l-methadone by the peak area of d-methadone. The enantioselective separation was both sensitive and specific using the Chiral-AGP column under the above mentioned conditions. Typical chromatograms of a solution in buffer (a), an extracted plasma spiked with d,l-methadone and IS (b) and an extracted plasma sample from one of the opiate addicts (c) are shown in Fig. 2. Retention times for IS, l-methadone and d-methadone were 6.8, 8.1 and 10.0 min respectively. The calibration curves for d-methadone and l-methadone in plasma showed linearity (r = 0.931 and r = 0.988 respectively) in the concentration range 80-230 ng·ml^-1. The calculated recovery values were 68.7±12.0% (n=12) for IS.

Calculations

The concentration of l-methadone (C_l) in each plasma sample was calculated as:

C_l = C_d,l · r / (r + 1), where C_d,l is the plasma concentration of d,l-methadone determined using Procedure A and r is the l-methadone:d-methadone ratio with Procedure B. The concentration of d-methadone (C_d) is: C_d = C_d,l · r ^-1 / (r ^-1 + 1).

Illicit drug use

Additional drug use during the study period was assessed by analysis of urine. The samples were taken once weekly. The urine was analyzed for presence of the following drugs (metabolites); heroin, cocaine, barbiturates and benzodiazepines. Illicit drug use for each group was calculated as the sum of all drugs, except methadone, present in the urine sample of each subject.

Craving

The individual level of opiate craving was assessed subjectively using the Experience Sampling Method [Csikszentmihaly and Larson, 1987; de Vries, 1987). This method has been used previously in both ambulatory and clinical addiction research (Kaplan, 1992; de Vos et al., 1996). The subjects received the ESM questionnaire during their visit to the drug dispensary. The questionnaire contains 6 different questions to assess craving; (1) "Did you think about using?", (2) "Did you feel stoned?", (3) "Were you in control of yourself?", (4) "Did you feel restless?", (5) "Did you need dope quickly?", (6) "Did you feel the need to use dope?". The second and third questions were conceived as 'negative' indicators. The first, fourth, fifth and sixth questions are positive indicators. Items were scored yes or no. The 'yes' answers for the two negative items were counted each as -1, the 'yes' answers for the three positive items were each counted as +1. 'No' answers for the two negative questions were counted as 0 and for the four affirmative questions they were counted as -1. Total craving score was the arithmetic sum of the assigned value for all 6 questions. The individual craving scores ranged from -5 (absence of craving) to 3 (high craving) and were transformed on a scale of 0 to 8.

Statistics

Data were expressed as the mean ± SD. Students t-test was used to test the significance of the differences between the two groups. Pearson correlation was used to analyse the relationship between dose and plasma concentration.

Results

During the course of the study there were 2 drop-outs, a further 8 subjects did not fulfil the protocol conditions leaving and were excluded from the analysis leaving a total of 30 subjects, 18 males and 12 females, mean age 30 years (range 20-44) who completed the study.

Dose

Table 1 Mean dose and dose range by Day for the l-methadone and the d,l-methadone group

Day 8

l-methadone group (n=14)

d,l-methadone group (n=16)

mean dose (mg) ± SD

37.9 ± 10.5

38.0 ± 10.3

dose range (mg)

17.5 - 52.5

12.5 - 52.5

Day 15

mean dose (mg) ± SD

38.7 ± 11.0

38.7 ± 10.9

dose range (mg)

17.5 - 55.0

12.5 - 55.0

mean change in dose (mg)

+ 0.8 (= + 2.1 %)

+ 0.7 (= + 1.8 %)

number of dose changes (+/-)

7 / 1

10 / 0

Day 22

mean dose (mg) ± SD

40.5 ± 13.2

41.6 ± 12.2

dose range (mg)

15.0 - 65.0

12.5 - 57.5

mean change in dose (mg)

+ 2.6 (= + 6.9 %)

+ 3.6 (= + 9.5 %)

number of dose changes (+/-)

7 / 1

10 / 0

Table 1 shows the mean doses administered on Day 8, Day 15 and Day 22 to subjects in the l-methadone and the d,l-methadone groups (expressed in l-methadone equivalents). From Day 8 to Day 22 the doses administered increased on average by 6.9 % for the l-methadone group and 9.5 % for the d,l-methadone group. From Day 15 to Day 22 seven subjects in the l-methadone group requested a dose increase (range: 2.5 - 20 mg) and one subject requested a dose decrease (2.5 mg). In the d,l-methadone group 10 subjects requested a dose increase (2.5 - 10 mg). No significant differences with regard to the magnitude of doses and the number of changes were observed between the two groups.

Fig. 1

Fig. 1. Chromatograms of; (a) 50 µl buffer containing 20 ng EDDP-HClO₄, 40 ng d,l-methadone HCl and 100 ng trihexyphenidyl-HCl (IS); (b) 50 µl extracted plasma spiked with 20 ng HClO₄, 40 ng d,l-methadone-HCl and 100 ng trihexyphenidyl-HCl; (c) 50 µl extracted plasma from one of the opiate addicts spiked with 100 ng trihexyphenidyl-HCl.

E, T and M are the absorption peaks for EDDP, trihexyphenidyl and d,l-methadone respectively; the numbers at the top of the peaks represent retention times in minutes. The X-peaks are obtained from unidentified substances from plasma.

Fig. 2

Fig. 2. Chromatograms of: (a) 50 µl buffer containing 25 ng dextropropoxyphene-HCl (IS) and 20 ng d,l-methadone-HCl; (b) 50 µl extracted plasma spiked with 25 ng dextropropoxyphene-HCl and 40 ng d,l-methadone; (c) 50 µl extracted plasma from one of the opiate addicts spiked with 25 ng dextropropoxyphene.

D, l-M and d-M are the absorption peaks for dextropropoxyphene, d-methadone and l-methadone respectively; the numbers at the top represent retention times in minutes.

Plasma concentrations of methadone enantiomers and EDDP

Fig. 1 and 2 show representative chromatograms for d,l-methadone and EDDP (Procedure A) and l-methadone and d-methadone (Procedure B). Table 2 shows the mean concentrations of l-methadone, d-methadone and EDDP in plasma in both groups.

The l-methadone dose:l-methadone plasma concentration ratio is an index of the bioavailability of l-methadone in individual subjects. The initial (on Day 8) value for this index in the l-methadone group was 4. 95 (SD: 2.50, range: 1.67 - 10.4) and in the d,l-methadone group 3.57 (SD: 1.07, range: 2.0 - 5.27). On Day 22 the ratios were 5.08 (SD: 2.71, range: 0.93 - 10.72) and 3.67 (SD: 1.98, range: 0.98 - 9.08) respectively. However, there was no significant differences in plasma concentrations and l-methadone dose:l-methadone plasma concentration ratio between Day 8 and Day 15 or 22. The correlation coefficient between the daily l-methadone dose (expressed in milligrams per kilogram body weight) and the l-methadone plasma concentration in the l-methadone group was 0.27 (P = .04). The correlation between the weight-corrected l-methadone dose and all measurements for the l-methadone plasma concentration was 0.34 (P = 0.001). The mean d-methadone:l-methadone plasma concentration ratio was 1.17 (SD = 0.28; range: 0.72 - 1.83). There was no significant difference between these ratios for Day 15: 1.20 (SD: 0.22, range: 0.72 - 1.61) and Day 22: 1.15 (SD: 0.33, range: 0.76 - 1.83). One subject in the l-methadone group had a detectable d-methadone concentration (21 ng·ml^-1) in her plasma on Day 15, apparently because of illicit racemic methadone use.

Table 2 Plasma concentrations of methadone enantiomers and EDDP by Day for the l-methadone and the d,l-methadone group

l-methadone group (n=14)

d.l-methadone group (n=16)

l-m

EDDP

l-m

d-m

EDDP

Day 8

mean concentration (ng.ml^-1) ± SD

185 ± 91

11 ± 8

135 ± 59

7 ± 4

concentration range (ng.ml^-1)

44 - 364

3 - 27

49 - 249

0 - 14

Day 15

mean concentration (ng.ml^-1) ± SD

184 ± 102

10 ± 6

147 ± 68

123 ± 54

18 ± 10

concentration range (ng.ml^-1)

41 - 426

3 - 22

34 - 346

47 - 270

2 - 34

Day 22

mean concentration (ng.ml^-1) ± SD

205 ± 110

12 ± 7

146 ± 82

135 ± 86

24 ± 14

concentration range (ng.ml^-1)

28 - 429

1 - 28

50 - 364

46 - 314

5 - 59

The l-methadone:EDDP plasma concentration ratio in the l-methadone group was 22.2 (range: 7.6 - 65.0; SD = 13.5) and the d,l-methadone:EDDP plasma concentration ration was 18.4 (range: 6.9 - 160.0; SD = 26.0). The plasma EDDP concentration in the d,l-methadone group increased 3-fold after starting treatment with d,l-methadone.

Illicit drug use

Differences between the two groups in illicit drug use were present on Day 8. In the d,l-methadone group, a total of 29 urines contained illicit drugs but in the l-methadone group only 20. On Day 22 the numbers were 30 (3.45 % increase) and 20 (no change) respectively. A decrease from 29 to 23 positive urines was observed on Day 15 in the d,l-methadone group. If only the additional use of heroin is taken into account, a significant increase was present in the l-methadone group on Day 22. The ratio changed from 1:14 on Day 8 to 6:14 on Day 22 (t = -2.33, P = .03). Apart from a small drop in the amount of heroin-positive urines in the d,l-methadone group on Day 15 (7:16), no change was seen between Day 8 and Day 22 (10:16). No significant difference between Day 8 and Day 22 was observed in the two groups when the total amount of illicit drugs in urine was taken into account (Table 3).

Table 3 Number of illicit drugs detected in urine by Day for the l-methadone and the d,l-methadone group

l-methadone group (n=14)

d,l-methadone group (n=16)

Day 8

heroin

cocaine

barbiturates

benzodiazepines

total

Day 15

heroin

cocaine

barbiturates

benzodiazepines

total

Day 22

heroin

cocaine

barbiturates

benzodiazepines

total

Craving

Differences in the mean level of craving between the two groups were present on Day 8. Although both groups still received l-methadone on this day, the d,l-methadone group showed a higher mean level of craving. On Day 15 the mean craving level for both groups was decreased. On Day 22 the mean level of craving of the d,l-methadone group had returned to initial values whereas the mean level of craving for the group that remained on l-methadone increased slightly (5.4 %) up to Day 22. No significant differences in craving were seen between Day 8 and Day 22 for both groups. When the range of craving level is considered, a slight decrease was observed in the l-methadone group whereas the range of craving levels in the d,l-methadone group increased slightly (Table 4).

Table 4 Mean craving and craving range by Day for the l-methadone and the d,l-methadone group

Day 8

l-methadone group (n = 14)

d,l-methadone group (n = 16)

mean craving ± (SD)

3.7 ± (1.7)

4.7 ± (2.4)

craving range (lower-upper limit)

6 (0 - 6)

7 (1 - 8)

Day 15

mean craving ± (SD)

2.4 ± (1.0)

3.4 ± (2.1)

craving range (lower-upper limit)

4 (0 - 4)

7 (0 - 7)

Day 22

mean craving ± (SD)

3.9 ± (1.6)

4.7 ± (2.8)

craving range (lower-upper limit)

5 (1 - 6)

8 (0 - 8)

Discussion

The three clinical response parameters measured in this study are a) number of requests for dosage change, b) use of illicit drugs and c) degree of opiate craving. No significant differences in these 3 parameters was found between subjects on l-methadone and those on racemic methadone over the whole study but there was an increase in heroin use in the l-methadone group.

In addition to subjective and objective assessment of clinical performance, a newly developed HPLC stereospecific separation technique enabled us to determine plasma concentrations of the methadone enantiomers during methadone maintenance treatment.

The correlation coefficient for the weight-corrected l-methadone dose and the l-methadone plasma concentration was low in agreement with our previous findings using d,l-methadone (de Vos et al., 1995). The l-methadone:d-methadone ratio indicated higher mean l-methadone plasma concentration than d-methadone. Other studies, although not as extensive as the present study, have also shown higher l-methadone plasma concentrations in subjects receiving racemic methadone (Kreek et al., 79; Nakamura et al., 1982). A study in 12 non-opiate tolerant volunteers (Olsen et al., 1976) showed consistently lower l-methadone plasma concentrations in comparison with d-methadone. After changing from l-methadone to d,l-methadone on Day 15, no significant changes in the l-methadone:d-methadone ratio were observed in the d,l-methadone group between day 15 and day 22. The variability in this ratio between individuals was less marked (0.72 - 1.83) and this agrees with results of a recent study carried out in 22 opiate addicts, where the l-methadone:d-methadone ratio ranged from 0.63 to 2.40 (Eap et al., 1996). The sharp increase in EDDP plasma concentration from 7 to 24 ng·ml^-1 occurring in the d,l-methadone group after changing to d,l-methadone confirms suggestions that racemic methadone induces its own metabolism (Verebely et al., 1975; Nilsson et al., 1982; Eap et al., 1996). The non-significant decrease in the mean l-methadone:d-methadone plasma concentration ratio from 1.20 on Day 15 to 1.15 on Day 22 is in accord with Eap et al. (1996) who suggested a preferential metabolism of l-methadone compared to d-methadone when changing to d,l-methadone.