Propranolol serum levels during twenty-four hours Propranolol serum levels during a 24-hr period were determined every 2 hr in 9 hospitalized patients with angina pectoris after oral administration of 40 mg of propranolol 3 times a day. After the first, second, and third tablets the mean maximum serum propranolol concentrations averaged 118 ± 71 ng Iml, 134 ± 97 ng Iml, and 118 ± 94 nglml and the mean minimum concentrations averaged 21 ± 18 nglml, 45 ± 25 nglml, and 54 ± 34 nglml (±SD), respectively. These data show a very wide inter- and intraindividual variation in serum propranolol levels. No relationship was found between serum level and blood pressure or dose (related to body weight).
E. Vervloet, Pharm.D., B. F. M. Pluym, M.D., J. Cilissen, K. Kahlen, and F. W. H. M. Merkus, Ph.D. Sittard and Amsterdam, The Netherlands Department of Cardiology and Clinical Pharmacology, R. C. Hospital, Sittard, and Department of Biopharmaceutics, University of Amsterdam
Propranolol has been used for ten years in the treatment of cardiovascular diseases and its pharmacologic properties have been investigated by many. 3, 7 From several studies it appears that there is a wide interindividual variation in plasma propranolol levels after oral administration of a single tablet. 1, 2. 9. 11 Since the common therapeutic regimen is one dose 3 times a day and the serum half-life (t 1h) is brief, the propranolol serum concentration has a tendency to fluctuate widely during a 24-hr period. Our study was undertaken to determine the fluctuations in propranolol serum concentrations and serum half-lives after the same oral dosage in hospitalized patients during a 24-hr period. Methods
This study was carried out in 9 hospitalized patients (aged 25 to 75 yr) who were admitted Received for publication Nov. 3, 1976. Accepted for publication May 3, 1977. Reprint requests to: Prof. F. W. H. M. Merkus, Ph.D., Department of Biopharmaceutics, University of Amsterdam, Plantage Muidergracht 14, Amsterdam, The Netherlands.
Table I. Mean coefficient of variation for different concentration ranges of propranolol Concentration range (nglml)
Mean coefficient of variation (%)
0- 50 51-100 101-150 151-200 201-350
9.0 5.7 8.6
4.9
9.4
to the Department of Cardiology for the treatment of angina pectoris. All patients had already received the therapy for 1 or 2 wk. One tablet of 40 mg propranolol was administered to each subject at 7:30 A.M., 1:30 P.M., and 7:30 P.M. Blood samples were taken every 2 hr, starting just before the first dose. This time interval was chosen because the maximum propranolol level developed at approximately 2 hr after oral administration. 1 At the time of each blood collection, resting heart rate and blood pressure were measured. The samples were centrifuged and the serum was frozen for at most 2 853
854
Vervioet et ai.
Clinical Pharmacology and Therapeutics
100
...:>. III
c:
... .!;; Q)
80
c:
.Q
III III 60
'E Q)
40
20
50
100
150
prOpranolol
200
ng I ml
Fig. 1. Linearity of the serum propranolol assay. Each point represents the mean of 4 determinations of known amounts of propranolol added to blank plasma (±SD). Equation of the standard curve: y = 0.45 x -0.25; correlation coefficient: r = 0.999.
wk. No change in serum propranolol concentration was observed in samples after as much as 2 wk storage in frozen condition. Propranolol concentrations were determined fluorimetrically by the method of Shand and associates. l l All patient samples were determined in duplicate. The mean coefficient of variation for the different concentration ranges is summarized in Table I. By this method only propranolol is extracted and measured but not its metabolite, 4-0Hpropranolol.
Experimental. Extraction procedure. The serum sample (2 ml) was transferred to a 15-ml centrifuge tube and 0.25 ml 4 N NaOH and 10.0 ml n-heptane (containing 1.5% isoamyl alcohol) was added. The tube was shaken for 10 mins. After centrifugation 8.0 ml of the organic layer was transferred to a 15-ml centrifuge tube containing 4.0 ml 0.1 N HCl. The tube was shaken for another 10 min, and after centrifugation the organic layer was discarded. The fluorescence of the water phase was measured by fluorescence spectrophotometry. To make a standard curve, known amounts of propranolol were added to
blank plasma and determined by the described method. As shown in Fig. 1, the standard curve was linear up to 200 ng/ml. Reagents. The reagents used were: n-heptane (Merck); isoamyl alcohol (Merck); Sodium hydroxide, 4 N pa; hydrochloric acid, 0.1 N pa; plasma solution (GPO from the Blood Transfusion Service of the Red Cross AmsterdamNetherlands) . Apparatus. The following equipment was used: Perkin-Elmer fluorescence spectrophotometer 204, exciter wavelength 290 nm, and analyzer wavelength 345 nm. Results
After the first, second, and third tablets, the mean peak propranolol concentrations in the 9 patients during 24 hr averaged 118 ± 71 ng/ml, 134 ± 97 ng/ml, and 118 ± 94 ng/ml (±SD, respectively.) The minimum concentrations obtained 2 hr before each maximum propranolol concentration averaged 21 ± 18 ng/ml, 45 ± 25 ng/ml, 54 ± 34 ng/ml, and 25 ± 18 ng/ml (Table II). The mean serum propranolol concentration (±SE) of the 9 patients is shown in Fig. 2.
Volume 22 Number 6
Propranolol levels during twentylour hours
855
240
200
E .....
140
Ol
c:
120
'0 "0 c:
cu... a.
...0
80
a. 40
13.30
7.30
19.30
t
t
7.30
hours
t
40 mg
40 mg
1.30
40 mg
Fig. 2. Mean propranolol serum levels in 9 hospitalized patients during 24 hr (±SD).
Table II. Serum propranolol concentration in 9 patients (ng/ml serum) Time of blood collection
E. D.
7:30 9:30 II :30 13:30 15:30 17:30 19:30 21 :30 23:30 1:30 3:30 5:30 7:30
6 49 30 20 50 30 21 48 40 31 22 19 13
Patient
IM. C. I W. E. IA. D. I
J. D.
25 235 135 78 290 175 113 308 156 118 84 55 45
12 200 110 47 294 133 70 211 113 74 45 24 14
63 156 118 88 135 130 97 148 114 114 107 79 58
32 90 71 46 98 75 56 97 73 49 46 36 35
The serum propranolol concentration in the patients differed widely as is clearly shown in the curves of the maximum and minimum serum concentrations in 2 patients (Fig. 3) with a lO-fold difference after the same oral dose. After administration of each tablet the serum tYz was calculated by the method of least squares for a 4-hr period starting at the peak propranolol
I
J. K.
10 147 79 42 152 86 52 135 90
61 47 28 22
I
J. B.
20 68 52 40 88 58 27 58 49 29 22 18 10
I P. C. Ic. B. 15 97 58 37 65 67 41 37 75 55 36 27 20
5 22 13 10 32 20 10 23 22 23 16 11 5
Mean serum concentration
21 118 74 45 134 86 54 118 81 62 47 33 25
SE
18 71 41 25 97 51 34 94
42 35 30 21 18
concentration. The serum tV2 after the first 2 tablets was stable within a narrow range in all but one patient (Patient J. B.) (Table III). The serum tYz after the third tablet was longer in almost all patients. Because the 2-hr sampling time in our study does not allow precise tV2 calculation, the significance of these findings is limited. No linear relationship between the
856 Vervloet et al.
Clinical Pharmacology and Therapeutics
300
250
E
200
OJ
c:
'0 '0
150
c:(I)
... a. 0... a. 100 patient
M.e.
50
13.30
7.30
19.30
t40 mg
t40 mg
1.30
t40 mg
7.30
hours
Fig. 3. The minimum and maximum propranolol serum levels in 2 patients.
Table III. Half-lives of elimination of propranolol from serum in 9 patients after a dosage of 40 mg propranolol/day at 7 :30 A.M., 1 :30 P.M., and 7:30 P.M.
Patient
E. D. M.C. W.E. A. D. 1. D. J. K. J. B. P. C. C. B. Mean
First half-life (hr)
Second half-life (hr)
Third half-life (hr)
3.1 2.5 l.9 4.9 4.2 2.2 5.3 2.9 3.5 3.4
3.2 3.0 2.0 4.7 5.7 2.6 2.4 2.7 2.4 3.2
6.3 3.6 2.7 4.6 6.1 3.6 3.3 3.9 3.8 4.2
Weight (kg)
73 74 80 74 73 82 91 76
serum propranolol concentration and the dose related to body weight (mg/kg/day) was detected, nor was there any correlation between resting heart rate or blood pressure and serum propranolol level. Discussion
These findings show a wide interindividual variation of propranolol serum levels in patients after oral administration of 40 mg propranolol 3 times during a 24-hr period (range, 23 to 308 ng/ml). In each patient there is also a great intraindividual variation in serum propranolol levels (Fig. 3 and Table II). There were 3- to 24-fold differences between minimum and maximum concentrations in Patients J. D. and W. E., respectively.
Volume 22 Number 6
The interpatient variability is partly due to a difference in excretion t1;2 but is more importantly caused by differences in rate and extent of metabolism. 1 , 10 From several studies 3 , 6 it appears that orally administered propranolol is extracted by the liver and for the greater part metabolized to an active metabolite, 4-0Hpropranolol. According to Paterson and coworkers 7 and Cleaveland and Shand 2 4-0Hpropranolol has a very short t1;2 (20 min). This metabolite, equally effective in beta-blocking properties, is rapidly eliminated and is detected only after oral administration. 4 Cleaveland and Shand 2 found no detectable effect of the additional beta-blocking properties of 4-0Hpropranolol after 5 days of continuous oral administration. It seems that in long-term angina pectoris therapy the propranolol level would be the most important parameter. In the continuous treatment of chronic angina pectoris the aim is to reduce the frequency of attacks as far as possible. This is done by reducing the cardiac work by means of blocking the catecholamines in order to reduce the cardiac oxygen consumption. According to Pine and colleagues B maximum therapeutic response in angina pectoris patients was observed between 64% and 98% of total blockade. This rather narrow range of beta blockade was related in their subjects to a very wide corresponding range of blood levels (14 to 90 ng/ml). In a report of McDevitt and Shand 5 with 3 healthy male subjects the propranolol plasma concentrations were a predictable function of the effectiveness in antagonizing the chronotropic effects of isoproterenol and exercise, according to the classical drug receptor theory for competitive antagonism. According to these studies every patient requires his own rather constant serum level for optimal protection in angina treatment, but from our data we can see that the propranolol concentration fluctuates widely over a 24-hr period. If these fluctuations are the same in the deeper compart-
Propranolol levels during twenty jour hours
857
ments, particularly in the heart, it must be concluded that the patients are not adequately protected by this therapy, especially not in the morning (Fig. 3). In actual practice propranolol therapy is withdrawn every evening and started the next day. In any case the full therapeutic significance of the remarkable intraindividual variations in propranolol serum levels is not clear.
References 1. Chidsey, C. A., Morselli, P., Bianchetti, G., Morganti, A., Leonetti, G., and Zanchetti, A.: Studies of the absorption and removal of propranolol in hypertensive patients during therapy, Circulation 52:313-318, 1975. 2. Cleaveland, C. R., and Shand, D. G.: Effect of route of administration on the relationship between beta adrenergic blockade and plasma propranolol level, CLIN. PHARMACOL. THER. 13: 18 1-185, 1972. 3. Hayes, A., and Cooper, R. G.: Studies on the absorption, distribution and excretion of propranolol in rat, dog and monkey, J. Pharmacol. Exp. Ther. 176:302-3II, 1971. 4. Johnson, G., and Regardh, C. G.: Clinical pharmacokinetics of ,B-andrenoceptor blocking drugs, Clin. Pharmacokinet. 1:233-263, 1976. 5. McDevitt, D. G., and Shand, D. G.: Plasma concentrations and the time course of beta blockade due to propranolol, CLIN. PHARMACOL. THER. 18:708-713, 1975. 6. Nies, A. S., and Shand, D. G.: Clinical pharmacology of propranolol, Circulation 52:6-13, 1975. 7. Paterson, J. W., Conolly, M. E., and Dollery, C. F.: The pharmacodynamics and metabolism of propranolol in man, Pharmacol. Clin. 2:127-133, 1970. 8. Pine, M., Favrot, L., Smith, S., McDonald, K., and Chidsey, C. A.: Correlation of plasma propranolol concentration with therapeutic response in patients with angina pectoris, Circulation 52:886-893, 1975. 9. Shand, D. G.: Individualization of propranolol therapy, Med. Clin. North. Am. 58:1063-1069, 1974. 10. Shand, D. G.: Propranolol, N. Eng!. J. Med. 293:280-285, 1975. 11. Shand, D. G., Nuckolls, E. M., and Oates, J. A.: Plasma propranolol levels in adults, CLIN. PHARMACOL. THER. 11:II2-120, 1970.
E. Vervloet, Pharm.D., B. F. M. Pluym, M.D., J. Cilissen, K. Kahlen, and F. W. H. M. Merkus, Ph.D. Sittard and Amsterdam, The Netherlands Department of Cardiology and Clinical Pharmacology, R. C. Hospital, Sittard, and Department of Biopharmaceutics, University of Amsterdam
Propranolol has been used for ten years in the treatment of cardiovascular diseases and its pharmacologic properties have been investigated by many. 3, 7 From several studies it appears that there is a wide interindividual variation in plasma propranolol levels after oral administration of a single tablet. 1, 2. 9. 11 Since the common therapeutic regimen is one dose 3 times a day and the serum half-life (t 1h) is brief, the propranolol serum concentration has a tendency to fluctuate widely during a 24-hr period. Our study was undertaken to determine the fluctuations in propranolol serum concentrations and serum half-lives after the same oral dosage in hospitalized patients during a 24-hr period. Methods
This study was carried out in 9 hospitalized patients (aged 25 to 75 yr) who were admitted Received for publication Nov. 3, 1976. Accepted for publication May 3, 1977. Reprint requests to: Prof. F. W. H. M. Merkus, Ph.D., Department of Biopharmaceutics, University of Amsterdam, Plantage Muidergracht 14, Amsterdam, The Netherlands.
Table I. Mean coefficient of variation for different concentration ranges of propranolol Concentration range (nglml)
Mean coefficient of variation (%)
0- 50 51-100 101-150 151-200 201-350
9.0 5.7 8.6
4.9
9.4
to the Department of Cardiology for the treatment of angina pectoris. All patients had already received the therapy for 1 or 2 wk. One tablet of 40 mg propranolol was administered to each subject at 7:30 A.M., 1:30 P.M., and 7:30 P.M. Blood samples were taken every 2 hr, starting just before the first dose. This time interval was chosen because the maximum propranolol level developed at approximately 2 hr after oral administration. 1 At the time of each blood collection, resting heart rate and blood pressure were measured. The samples were centrifuged and the serum was frozen for at most 2 853
854
Vervioet et ai.
Clinical Pharmacology and Therapeutics
100
...:>. III
c:
... .!;; Q)
80
c:
.Q
III III 60
'E Q)
40
20
50
100
150
prOpranolol
200
ng I ml
Fig. 1. Linearity of the serum propranolol assay. Each point represents the mean of 4 determinations of known amounts of propranolol added to blank plasma (±SD). Equation of the standard curve: y = 0.45 x -0.25; correlation coefficient: r = 0.999.
wk. No change in serum propranolol concentration was observed in samples after as much as 2 wk storage in frozen condition. Propranolol concentrations were determined fluorimetrically by the method of Shand and associates. l l All patient samples were determined in duplicate. The mean coefficient of variation for the different concentration ranges is summarized in Table I. By this method only propranolol is extracted and measured but not its metabolite, 4-0Hpropranolol.
Experimental. Extraction procedure. The serum sample (2 ml) was transferred to a 15-ml centrifuge tube and 0.25 ml 4 N NaOH and 10.0 ml n-heptane (containing 1.5% isoamyl alcohol) was added. The tube was shaken for 10 mins. After centrifugation 8.0 ml of the organic layer was transferred to a 15-ml centrifuge tube containing 4.0 ml 0.1 N HCl. The tube was shaken for another 10 min, and after centrifugation the organic layer was discarded. The fluorescence of the water phase was measured by fluorescence spectrophotometry. To make a standard curve, known amounts of propranolol were added to
blank plasma and determined by the described method. As shown in Fig. 1, the standard curve was linear up to 200 ng/ml. Reagents. The reagents used were: n-heptane (Merck); isoamyl alcohol (Merck); Sodium hydroxide, 4 N pa; hydrochloric acid, 0.1 N pa; plasma solution (GPO from the Blood Transfusion Service of the Red Cross AmsterdamNetherlands) . Apparatus. The following equipment was used: Perkin-Elmer fluorescence spectrophotometer 204, exciter wavelength 290 nm, and analyzer wavelength 345 nm. Results
After the first, second, and third tablets, the mean peak propranolol concentrations in the 9 patients during 24 hr averaged 118 ± 71 ng/ml, 134 ± 97 ng/ml, and 118 ± 94 ng/ml (±SD, respectively.) The minimum concentrations obtained 2 hr before each maximum propranolol concentration averaged 21 ± 18 ng/ml, 45 ± 25 ng/ml, 54 ± 34 ng/ml, and 25 ± 18 ng/ml (Table II). The mean serum propranolol concentration (±SE) of the 9 patients is shown in Fig. 2.
Volume 22 Number 6
Propranolol levels during twentylour hours
855
240
200
E .....
140
Ol
c:
120
'0 "0 c:
cu... a.
...0
80
a. 40
13.30
7.30
19.30
t
t
7.30
hours
t
40 mg
40 mg
1.30
40 mg
Fig. 2. Mean propranolol serum levels in 9 hospitalized patients during 24 hr (±SD).
Table II. Serum propranolol concentration in 9 patients (ng/ml serum) Time of blood collection
E. D.
7:30 9:30 II :30 13:30 15:30 17:30 19:30 21 :30 23:30 1:30 3:30 5:30 7:30
6 49 30 20 50 30 21 48 40 31 22 19 13
Patient
IM. C. I W. E. IA. D. I
J. D.
25 235 135 78 290 175 113 308 156 118 84 55 45
12 200 110 47 294 133 70 211 113 74 45 24 14
63 156 118 88 135 130 97 148 114 114 107 79 58
32 90 71 46 98 75 56 97 73 49 46 36 35
The serum propranolol concentration in the patients differed widely as is clearly shown in the curves of the maximum and minimum serum concentrations in 2 patients (Fig. 3) with a lO-fold difference after the same oral dose. After administration of each tablet the serum tYz was calculated by the method of least squares for a 4-hr period starting at the peak propranolol
I
J. K.
10 147 79 42 152 86 52 135 90
61 47 28 22
I
J. B.
20 68 52 40 88 58 27 58 49 29 22 18 10
I P. C. Ic. B. 15 97 58 37 65 67 41 37 75 55 36 27 20
5 22 13 10 32 20 10 23 22 23 16 11 5
Mean serum concentration
21 118 74 45 134 86 54 118 81 62 47 33 25
SE
18 71 41 25 97 51 34 94
42 35 30 21 18
concentration. The serum tV2 after the first 2 tablets was stable within a narrow range in all but one patient (Patient J. B.) (Table III). The serum tYz after the third tablet was longer in almost all patients. Because the 2-hr sampling time in our study does not allow precise tV2 calculation, the significance of these findings is limited. No linear relationship between the
856 Vervloet et al.
Clinical Pharmacology and Therapeutics
300
250
E
200
OJ
c:
'0 '0
150
c:(I)
... a. 0... a. 100 patient
M.e.
50
13.30
7.30
19.30
t40 mg
t40 mg
1.30
t40 mg
7.30
hours
Fig. 3. The minimum and maximum propranolol serum levels in 2 patients.
Table III. Half-lives of elimination of propranolol from serum in 9 patients after a dosage of 40 mg propranolol/day at 7 :30 A.M., 1 :30 P.M., and 7:30 P.M.
Patient
E. D. M.C. W.E. A. D. 1. D. J. K. J. B. P. C. C. B. Mean
First half-life (hr)
Second half-life (hr)
Third half-life (hr)
3.1 2.5 l.9 4.9 4.2 2.2 5.3 2.9 3.5 3.4
3.2 3.0 2.0 4.7 5.7 2.6 2.4 2.7 2.4 3.2
6.3 3.6 2.7 4.6 6.1 3.6 3.3 3.9 3.8 4.2
Weight (kg)
73 74 80 74 73 82 91 76
serum propranolol concentration and the dose related to body weight (mg/kg/day) was detected, nor was there any correlation between resting heart rate or blood pressure and serum propranolol level. Discussion
These findings show a wide interindividual variation of propranolol serum levels in patients after oral administration of 40 mg propranolol 3 times during a 24-hr period (range, 23 to 308 ng/ml). In each patient there is also a great intraindividual variation in serum propranolol levels (Fig. 3 and Table II). There were 3- to 24-fold differences between minimum and maximum concentrations in Patients J. D. and W. E., respectively.
Volume 22 Number 6
The interpatient variability is partly due to a difference in excretion t1;2 but is more importantly caused by differences in rate and extent of metabolism. 1 , 10 From several studies 3 , 6 it appears that orally administered propranolol is extracted by the liver and for the greater part metabolized to an active metabolite, 4-0Hpropranolol. According to Paterson and coworkers 7 and Cleaveland and Shand 2 4-0Hpropranolol has a very short t1;2 (20 min). This metabolite, equally effective in beta-blocking properties, is rapidly eliminated and is detected only after oral administration. 4 Cleaveland and Shand 2 found no detectable effect of the additional beta-blocking properties of 4-0Hpropranolol after 5 days of continuous oral administration. It seems that in long-term angina pectoris therapy the propranolol level would be the most important parameter. In the continuous treatment of chronic angina pectoris the aim is to reduce the frequency of attacks as far as possible. This is done by reducing the cardiac work by means of blocking the catecholamines in order to reduce the cardiac oxygen consumption. According to Pine and colleagues B maximum therapeutic response in angina pectoris patients was observed between 64% and 98% of total blockade. This rather narrow range of beta blockade was related in their subjects to a very wide corresponding range of blood levels (14 to 90 ng/ml). In a report of McDevitt and Shand 5 with 3 healthy male subjects the propranolol plasma concentrations were a predictable function of the effectiveness in antagonizing the chronotropic effects of isoproterenol and exercise, according to the classical drug receptor theory for competitive antagonism. According to these studies every patient requires his own rather constant serum level for optimal protection in angina treatment, but from our data we can see that the propranolol concentration fluctuates widely over a 24-hr period. If these fluctuations are the same in the deeper compart-
Propranolol levels during twenty jour hours
857
ments, particularly in the heart, it must be concluded that the patients are not adequately protected by this therapy, especially not in the morning (Fig. 3). In actual practice propranolol therapy is withdrawn every evening and started the next day. In any case the full therapeutic significance of the remarkable intraindividual variations in propranolol serum levels is not clear.
References 1. Chidsey, C. A., Morselli, P., Bianchetti, G., Morganti, A., Leonetti, G., and Zanchetti, A.: Studies of the absorption and removal of propranolol in hypertensive patients during therapy, Circulation 52:313-318, 1975. 2. Cleaveland, C. R., and Shand, D. G.: Effect of route of administration on the relationship between beta adrenergic blockade and plasma propranolol level, CLIN. PHARMACOL. THER. 13: 18 1-185, 1972. 3. Hayes, A., and Cooper, R. G.: Studies on the absorption, distribution and excretion of propranolol in rat, dog and monkey, J. Pharmacol. Exp. Ther. 176:302-3II, 1971. 4. Johnson, G., and Regardh, C. G.: Clinical pharmacokinetics of ,B-andrenoceptor blocking drugs, Clin. Pharmacokinet. 1:233-263, 1976. 5. McDevitt, D. G., and Shand, D. G.: Plasma concentrations and the time course of beta blockade due to propranolol, CLIN. PHARMACOL. THER. 18:708-713, 1975. 6. Nies, A. S., and Shand, D. G.: Clinical pharmacology of propranolol, Circulation 52:6-13, 1975. 7. Paterson, J. W., Conolly, M. E., and Dollery, C. F.: The pharmacodynamics and metabolism of propranolol in man, Pharmacol. Clin. 2:127-133, 1970. 8. Pine, M., Favrot, L., Smith, S., McDonald, K., and Chidsey, C. A.: Correlation of plasma propranolol concentration with therapeutic response in patients with angina pectoris, Circulation 52:886-893, 1975. 9. Shand, D. G.: Individualization of propranolol therapy, Med. Clin. North. Am. 58:1063-1069, 1974. 10. Shand, D. G.: Propranolol, N. Eng!. J. Med. 293:280-285, 1975. 11. Shand, D. G., Nuckolls, E. M., and Oates, J. A.: Plasma propranolol levels in adults, CLIN. PHARMACOL. THER. 11:II2-120, 1970.
