240
Br. J. clin. Pharmac.
LETTERS TO THE EDITORS
(I 9 7 7), 4
CEREBROSPINAL FLUID CONCENTRATION OF METOPROLOL IN A HYPERTENSIVE PATIENT Among the various theories of how ,B-adrenoceptor blocking drugs lower blood pressure, it has been suggested that these drugs act via a mechanism in the central nervous system (CNS) (Day & Roach, 1974; Reid, Lewis, Myers & Dollery, 1974). Animal studies have shown that after different equilibration times, the various ,B-adrenoceptor blockers distribute between brain tissue and plasma according to their lipid solubility (Johnsson & Regardh, 1976). The most lipophilic ,B-adrenoceptor blocker, propranolol, reaches a brain: plasma concentration ratio of about 15: 1 within 30 min of intravenous administration (Hayes & Cooper, 1971) whereas the least lipophilic drug of the series, practolol, reaches a distribution ratio of 0.8: 1 after 12-24 h (Scales & Cosgrove, 1970). Metoprolol, which is intermediate between the two extremes of lipid solubility, rapidly reaches a brain: plasma concentration ratio of 3: 1 (Bodin, Borg, Johansson, Ramsay & Skanberg, 1975). A study in man using postmortem samples showed that the brain: plasma distribution of propranolol was in agreement with that found in animals (Myers, Lewis, Reid & Dollery, 1975). In living subjects the only part of the human CNS that is readily accessible for drug assay is the cerebrospinal fluid (CSF). We have collected by lumbar puncture a sample of CSF from a patient who had been taking metoprolol 50 mg three times daily for 2 months. At the time of taking the CSF sample, prior to air encephalography, a blood sample was also taken for measurement of plasma metoprolol concentration. The patient was a 64-year-old woman who presented with blurred vision. She was found to have a casual blood pressure of 220/136 mmHg, impaired renal function, papilloedema, retinal haemorrhages and exudates and bilateral hemianopia. Her blood pressure was subsequently controlled on cyclopenthiazide-KCl and metoprolol and her papilloedema soon resolved, though the renal impairment and visual field defects remained. Plasma and CSF were assayed for metoprolol by electron capture gas-liquid chromatography (g.l.c.). The method used was a modification of that used by Jack & Reiss (1974) for the assay of oxprenolol. To 1 ml of plasma or CSF was added 100 ng propranolol (internal standard), 0.3 ml 0.75 mol/l NaOH and 5 ml ether/chloroform mixture (4: 1). After mixing and centrifuging, the organic layer was extracted with 3 ml 0.1 mol/l HCI. The aqueous layer was then made
alkaline by the addition of 0.5 ml 0.75 mol/l NaOH and re-extracted with 5 ml of the ether/ chloroform mixture. The organic extract was evaporated to dryness, redissolved in 25AI 1 dry ethyl acetate and 25 Mil trifluoroacetic anhydride,
sealed and heated at 500 for 30 min. The derivative thus formed was evaporated to dryness and redissolved in 100,l dry ethyl acetate for injection into a Varian 2100 gas chromatograph. The g.l.c. column (1.5 m x 3 mm) was packed with 4% OVIOI on Chromasorb W 100-120 mesh. The oven, injector port and 3H detector temperatures were 1950C, 2100C and 2550C respectively. The carrier gas was oxygen-free nitrogen at a flow rate of 25 ml/min. Under these conditions retention times were 2.7 min for metoprolol and 3.7 min for propranolol. The concentration of metoprolol was 341 ng/ml in plasma and 267 ng/ml in CSF. CSF metoprolol concentration thus represented 78% of the total drug concentration in the plasma. The degree of binding of the drug by plasma protein was not determined in this instance. However, Johansson, Appelgren, Borg & Elofsson (1974) have reported that protein-bound metoprolol accounts for 11 % of the total serum metoprolol concentration in vitro. This observation in one human apparently confirms that of Bodin et aL (1975), made in mice and rats, that metoprolol enters the CNS. Although it may be true in humans, as in mice and rats, that the concentration in brain tissue at equilibrium is about three times that in plasma, the concentration in CSF seems to be approximately equal to the concentration of nonprotein-bound drug in plasma. The author wishes to acknowledge the excellent technical assistance of Miss B.C. Galland and the co-operation of Mr D.G. Ferry, Toxicology Research Unit, Medical Research Council of New Zealand, for the use of GLC facilities. The work was carried out under the supervision of Professor F.O. Simpson, to whom the author is grateful for advice.
A.J. WOOD Wellcome Medical Research Institute, Department of Medicine, University of Otago Medical School, Dunedin, New Zealand. Received December 12, 1976
Br. J. clin. Pharmac. (1977), 4
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241
References BODIN, N-O., BORG, K.O., JOHANSSON, R., RAMSAY, C-H & SKANBERG, I. (1975). The distribution of metoprolol-(3 H) in the mouse and rat. Acta Pharmac. Tox., 36, SuppL V, 116-124. DAY, M.D. & ROACH, A.G. (1974). Central adrenoceptors and the control of arterial blood pressure.
Clin exp. Pharmac. Physiol, 1, 347-360. HAYES, A. & COOPER, R.G. (1971). Studies on the absorption, distribution and excretion of propranolol in rat, dog and monkey. J. Pharnac. exp. Ther., 176, 302-311. JACK, D.B. & REISS, W. (1974). Determination of low levels of oxprenolol in blood or plasma by gas-liquid chromatography. J. Chromat., 88, 173-176. JOHANSSON, K.A., APPELGREN, C., BORG, K.O. & ELOFSSON, R. (1974). Binding of two adrenergic
beta-receptor antagonists, alprenolol and H93/26 to human serum proteins. Acta Pharmn suec., 11, 333-346.
JOHNSSON, G. & REGARDH, C-G. (1976). Clinical pharmacokinetics of i3-adrenoceptor blocking drugs. Cin Pharmacokin, 1, 233-263. MYERS, M.G., LEWIS, P.J., REID, J.L. & DOLLERY,
C.T. (1975). Brain concentration of propranolol in relation to hypotensive effect in the rabbit with observations on brain propranolol levels in man. J. Pharmac. exp. Ther., 192, 327-335. REID, J.L., LEWIS, P.J., MYERS, M.G. & DOLLERY,
C.T. (1974). Cardiovascular effects of intracerebroventricular d-, 1- and dl- propranolol in the conscious rabbit. J. Pharmac. exp. Ther., 188, 394-399. SCALES, B. & COSGROVE, M.B. (1970). The metabolism and distribution of the selective adrenergic beta-blocking agent, practoloL J. Pharmac. exp. Ther., 175, 338-347.
PLASMA NITROGLYCERIN LEVELS AFTER SUBLINGUAL, ORAL AND TOPICAL ADMINISTRATION Nitroglycerin is administered clinically through any one of three routes: sublingual, oral or topical. While the therapeutic efficacy of the sublingual tablets is well established, the clinical utilities of nitroglycerin oral tablets and ointments are less well defined. The effectiveness of oral nitroglycerin is currently under serious debate. Metabolic studies (Needleman, Lang & Johnson, 1972) showed extensive first-pass metabolism of the drug in the liver, suggesting complete destruction of orally administered nitroglycerin. However, a recent clinical study (Winsor & Berger, 1975) has convincingly demonstrated significant clinical improvement of angina pectoris in patients given oral controlled release tablets. Nitroglycerin ointment appeared to elicit significant hemodynamic changes. After topical application of 15 mg of nitroglycerin, a significant decrease in left ventricular end-diastolic pressure started within 15 minutes and lasted until at least 60 minutes (Parker, Augustine, Burton, West & Armstrong, 1976). None of the studies, however, reported the plasma concentrations of nitroglycerin after drug administration. Our group has recently improved a gas chromotographic assay (Rosseel & Bogaert, 1973), extending quantitation of nitroglycerin in human plasma to levels as low as 0.1 ng/ml. The method involved stabilization of the drug in plasma with silver nitrate, followed by multiple extraction
using specially purified hexane. After addition of isosorbide dinitrate as the internal standard, the hexane extract was concentrated and injected into a glass column packed with 3% SP-240 1 on 100/120 supelcoport 01-1991 (Supelco, Inc., Bellefonte, Pa., U.S.A.). The column was maintained at 1400 C and quantitation was effected via a Nickel-63 electron capture detector. This improved technique allowed us to compare, apparently for the first time, the plasma nitroglycerin levels obtained from clinical doses of the drug through the three routes of administration. In a pilot study of a healthy volunteer, we have followed plasma nitroglycerin levels after administration of (i) a 0.3 mg sublingual tablet, (ii) a 2.5 mg and a 6.5 mg sustained-release oral capsule, and (iii) 1 of a 2% ointment (corresponding to 16 mg of nitroglycerin). The pulse rate and sphygmomanometric blood pressure of the sitting subject were concomitantly recorded. Figure 1 shows the plasma nitroglycerin levels obtained after administration of the various dosage forms. As expected, high drug levels were almost instantaneously achieved after the sublingual tablet, reaching about 1 ng/ml at the first measured data point (3 min). Thereafter, the plasma levels declined rapidly and no measurable nitroglycerin levels were detected after 16 min. In contrast, levels above 0.1 ng/ml of the drug were not observed until 20 min after application of the
Br. J. clin. Pharmac.
LETTERS TO THE EDITORS
(I 9 7 7), 4
CEREBROSPINAL FLUID CONCENTRATION OF METOPROLOL IN A HYPERTENSIVE PATIENT Among the various theories of how ,B-adrenoceptor blocking drugs lower blood pressure, it has been suggested that these drugs act via a mechanism in the central nervous system (CNS) (Day & Roach, 1974; Reid, Lewis, Myers & Dollery, 1974). Animal studies have shown that after different equilibration times, the various ,B-adrenoceptor blockers distribute between brain tissue and plasma according to their lipid solubility (Johnsson & Regardh, 1976). The most lipophilic ,B-adrenoceptor blocker, propranolol, reaches a brain: plasma concentration ratio of about 15: 1 within 30 min of intravenous administration (Hayes & Cooper, 1971) whereas the least lipophilic drug of the series, practolol, reaches a distribution ratio of 0.8: 1 after 12-24 h (Scales & Cosgrove, 1970). Metoprolol, which is intermediate between the two extremes of lipid solubility, rapidly reaches a brain: plasma concentration ratio of 3: 1 (Bodin, Borg, Johansson, Ramsay & Skanberg, 1975). A study in man using postmortem samples showed that the brain: plasma distribution of propranolol was in agreement with that found in animals (Myers, Lewis, Reid & Dollery, 1975). In living subjects the only part of the human CNS that is readily accessible for drug assay is the cerebrospinal fluid (CSF). We have collected by lumbar puncture a sample of CSF from a patient who had been taking metoprolol 50 mg three times daily for 2 months. At the time of taking the CSF sample, prior to air encephalography, a blood sample was also taken for measurement of plasma metoprolol concentration. The patient was a 64-year-old woman who presented with blurred vision. She was found to have a casual blood pressure of 220/136 mmHg, impaired renal function, papilloedema, retinal haemorrhages and exudates and bilateral hemianopia. Her blood pressure was subsequently controlled on cyclopenthiazide-KCl and metoprolol and her papilloedema soon resolved, though the renal impairment and visual field defects remained. Plasma and CSF were assayed for metoprolol by electron capture gas-liquid chromatography (g.l.c.). The method used was a modification of that used by Jack & Reiss (1974) for the assay of oxprenolol. To 1 ml of plasma or CSF was added 100 ng propranolol (internal standard), 0.3 ml 0.75 mol/l NaOH and 5 ml ether/chloroform mixture (4: 1). After mixing and centrifuging, the organic layer was extracted with 3 ml 0.1 mol/l HCI. The aqueous layer was then made
alkaline by the addition of 0.5 ml 0.75 mol/l NaOH and re-extracted with 5 ml of the ether/ chloroform mixture. The organic extract was evaporated to dryness, redissolved in 25AI 1 dry ethyl acetate and 25 Mil trifluoroacetic anhydride,
sealed and heated at 500 for 30 min. The derivative thus formed was evaporated to dryness and redissolved in 100,l dry ethyl acetate for injection into a Varian 2100 gas chromatograph. The g.l.c. column (1.5 m x 3 mm) was packed with 4% OVIOI on Chromasorb W 100-120 mesh. The oven, injector port and 3H detector temperatures were 1950C, 2100C and 2550C respectively. The carrier gas was oxygen-free nitrogen at a flow rate of 25 ml/min. Under these conditions retention times were 2.7 min for metoprolol and 3.7 min for propranolol. The concentration of metoprolol was 341 ng/ml in plasma and 267 ng/ml in CSF. CSF metoprolol concentration thus represented 78% of the total drug concentration in the plasma. The degree of binding of the drug by plasma protein was not determined in this instance. However, Johansson, Appelgren, Borg & Elofsson (1974) have reported that protein-bound metoprolol accounts for 11 % of the total serum metoprolol concentration in vitro. This observation in one human apparently confirms that of Bodin et aL (1975), made in mice and rats, that metoprolol enters the CNS. Although it may be true in humans, as in mice and rats, that the concentration in brain tissue at equilibrium is about three times that in plasma, the concentration in CSF seems to be approximately equal to the concentration of nonprotein-bound drug in plasma. The author wishes to acknowledge the excellent technical assistance of Miss B.C. Galland and the co-operation of Mr D.G. Ferry, Toxicology Research Unit, Medical Research Council of New Zealand, for the use of GLC facilities. The work was carried out under the supervision of Professor F.O. Simpson, to whom the author is grateful for advice.
A.J. WOOD Wellcome Medical Research Institute, Department of Medicine, University of Otago Medical School, Dunedin, New Zealand. Received December 12, 1976
Br. J. clin. Pharmac. (1977), 4
LETTERS TO THE EDITORS
241
References BODIN, N-O., BORG, K.O., JOHANSSON, R., RAMSAY, C-H & SKANBERG, I. (1975). The distribution of metoprolol-(3 H) in the mouse and rat. Acta Pharmac. Tox., 36, SuppL V, 116-124. DAY, M.D. & ROACH, A.G. (1974). Central adrenoceptors and the control of arterial blood pressure.
Clin exp. Pharmac. Physiol, 1, 347-360. HAYES, A. & COOPER, R.G. (1971). Studies on the absorption, distribution and excretion of propranolol in rat, dog and monkey. J. Pharnac. exp. Ther., 176, 302-311. JACK, D.B. & REISS, W. (1974). Determination of low levels of oxprenolol in blood or plasma by gas-liquid chromatography. J. Chromat., 88, 173-176. JOHANSSON, K.A., APPELGREN, C., BORG, K.O. & ELOFSSON, R. (1974). Binding of two adrenergic
beta-receptor antagonists, alprenolol and H93/26 to human serum proteins. Acta Pharmn suec., 11, 333-346.
JOHNSSON, G. & REGARDH, C-G. (1976). Clinical pharmacokinetics of i3-adrenoceptor blocking drugs. Cin Pharmacokin, 1, 233-263. MYERS, M.G., LEWIS, P.J., REID, J.L. & DOLLERY,
C.T. (1975). Brain concentration of propranolol in relation to hypotensive effect in the rabbit with observations on brain propranolol levels in man. J. Pharmac. exp. Ther., 192, 327-335. REID, J.L., LEWIS, P.J., MYERS, M.G. & DOLLERY,
C.T. (1974). Cardiovascular effects of intracerebroventricular d-, 1- and dl- propranolol in the conscious rabbit. J. Pharmac. exp. Ther., 188, 394-399. SCALES, B. & COSGROVE, M.B. (1970). The metabolism and distribution of the selective adrenergic beta-blocking agent, practoloL J. Pharmac. exp. Ther., 175, 338-347.
PLASMA NITROGLYCERIN LEVELS AFTER SUBLINGUAL, ORAL AND TOPICAL ADMINISTRATION Nitroglycerin is administered clinically through any one of three routes: sublingual, oral or topical. While the therapeutic efficacy of the sublingual tablets is well established, the clinical utilities of nitroglycerin oral tablets and ointments are less well defined. The effectiveness of oral nitroglycerin is currently under serious debate. Metabolic studies (Needleman, Lang & Johnson, 1972) showed extensive first-pass metabolism of the drug in the liver, suggesting complete destruction of orally administered nitroglycerin. However, a recent clinical study (Winsor & Berger, 1975) has convincingly demonstrated significant clinical improvement of angina pectoris in patients given oral controlled release tablets. Nitroglycerin ointment appeared to elicit significant hemodynamic changes. After topical application of 15 mg of nitroglycerin, a significant decrease in left ventricular end-diastolic pressure started within 15 minutes and lasted until at least 60 minutes (Parker, Augustine, Burton, West & Armstrong, 1976). None of the studies, however, reported the plasma concentrations of nitroglycerin after drug administration. Our group has recently improved a gas chromotographic assay (Rosseel & Bogaert, 1973), extending quantitation of nitroglycerin in human plasma to levels as low as 0.1 ng/ml. The method involved stabilization of the drug in plasma with silver nitrate, followed by multiple extraction
using specially purified hexane. After addition of isosorbide dinitrate as the internal standard, the hexane extract was concentrated and injected into a glass column packed with 3% SP-240 1 on 100/120 supelcoport 01-1991 (Supelco, Inc., Bellefonte, Pa., U.S.A.). The column was maintained at 1400 C and quantitation was effected via a Nickel-63 electron capture detector. This improved technique allowed us to compare, apparently for the first time, the plasma nitroglycerin levels obtained from clinical doses of the drug through the three routes of administration. In a pilot study of a healthy volunteer, we have followed plasma nitroglycerin levels after administration of (i) a 0.3 mg sublingual tablet, (ii) a 2.5 mg and a 6.5 mg sustained-release oral capsule, and (iii) 1 of a 2% ointment (corresponding to 16 mg of nitroglycerin). The pulse rate and sphygmomanometric blood pressure of the sitting subject were concomitantly recorded. Figure 1 shows the plasma nitroglycerin levels obtained after administration of the various dosage forms. As expected, high drug levels were almost instantaneously achieved after the sublingual tablet, reaching about 1 ng/ml at the first measured data point (3 min). Thereafter, the plasma levels declined rapidly and no measurable nitroglycerin levels were detected after 16 min. In contrast, levels above 0.1 ng/ml of the drug were not observed until 20 min after application of the
