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Vol. 27, Issue 6, 645-650, June 1999
Clinical Pharmacology Laboratory, Departments of Pharmacology and Psychiatry, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania
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Abstract |
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 Top
 Abstract
 Introduction
Materials and Methods
Results
Discussion
References
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A comparative study of the plasma pharmacokinetics and tissue distribution of the d-threo enantiomers of methylphenidate (MPH), para-bromomethylphenidate (p-Br MPH), and para-methoxymethylphenidate (p-OCH3 MPH) was conducted in rats after i.p. administration of a 37 µmol/kg dose. The plasma kinetic data was fit to a two-compartment model with absorption and lag time as well as evaluated by noncompartmental methods. All three compounds attained maximal concentration within 10 min of injection. Calculated mean residence time and elimination half-life values for d-p-Br MPH were significantly longer than those for d-MPH and d-p-OCH3 MPH, and clearance of the bromo derivative was substantially lower than the latter two compounds. Tissue distribution studies of the three d-threo enantiomers revealed that para-substitution of d-MPH had a profound effect on the distribution pattern of these drugs. The highest concentration of drug was found in the kidney and lung for d-MPH, lung and liver for d-p-Br MPH, and lung and brain for d-p-OCH3 MPH. The bromo derivative was found in the highest concentration in the central nervous system at 30, 120, and 180 min whereas levels of d-MPH were twice as high as d-p-OCH3 MPH at 30 min but slightly lower than the latter at 120 min. Related studies on the lipophilicity, plasma protein binding, and resistance to plasma degradation of these compounds were also conducted. The combined data from these experiments along with the pharmacokinetics and central nervous system distribution of these drugs provide explanations for discrepancies between the in vivo and in vitro activity of these compounds described in previous work.
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Introduction |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Chirality is a key factor for
the pharmacokinetics of methylphenidate (MPH; Ritalin, methyl
threo-dl-2-phenyl-2-(2'-piperidyl)acetate hydrochloride).1
(Fig. 1.) Racemic MPH is absorbed from the gut and metabolized with
substantial first pass effects. Rats selectively oxidize the
l-threo enantiomer of MPH quite extensively to
its p-hydroxy derivative, which is ultimately excreted in
the feces after glucuronidation (Faraj et al., 1974
; Patrick et al.,
1986a). In humans, the drug undergoes significant enantioselective
presystemic clearance favoring removal of the
l-threo isomer by deesterification to ritalinic acid (Srivinas et al., 1987). Despite the availability of kinetic data
on racemic and optically pure MPH, there are still no reports in the
literature on the pharmacokinetics, metabolism, and tissue distribution
of enantiomeric MPH analogs.
MPH appears to exert its pharmacological effects through potent
inhibition of dopamine and norepinephrine reuptake into the presynaptic
nerve terminus (Patrick et al., 1986b). Many of the investigations
of MPH and its derivatives have examined in vitro neurochemical
activity or in vivo pharmacodynamics in rats using racemic or
diastereomeric mixtures. A recent study in our group developed new
chemical syntheses and compared the neurochemical and locomotor
activity in rats of optically enriched threo MPH enantiomers
and their p-bromo and p-methoxy derivatives (Thai et al., 1998). Similar to the case with the parent compound, the pharmacological profile of the para-substituted MPH
derivatives was such that the d-isomers were more active
than the l-isomers. When comparing the d-threo
enantiomers of MPH, p-bromo MPH, and p-methoxy
MPH, we also noted a disparity between the in vitro and in vivo
activities of these drugs. In particular, the neurochemical potency of
d-methoxy MPH at dopaminergic and noradrenergic uptake sites
was similar to d-MPH, yet its locomotor-inducing effects were much less pronounced than the latter in terms of both amplitude and time duration. The d-p-bromo analog was also
shown to sustain locomotor activity for at least 180 min, which was
more than twice the duration of any other compound tested. To define
the role of metabolism in the overall in vitro/in vivo profile of these drugs, we investigated the pharmacokinetics and distribution of the
d-threo enantiomers of MPH and its
para-substituted derivatives in rats after i.p.
administration, the same route of drug delivery used in the locomotor
studies reported previously. The data from these studies would provide
valuable preliminary information on the potential of these derivatives
as alternative therapies for treatment of attention deficit
hyperactivity disorder as well as imaging probes for central nervous
system (CNS) dopaminergic activities.
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Materials and Methods |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Male Sprague-Dawley rats (225-250 g) were obtained from Hilltop
Lab Animals, Inc. (Scottdale, PA). Drug-free rat plasma was obtained
from Zivic-Miller Laboratories, Inc. (Zelienople, PA). The
d-threo enantiomers of MPH, p-bromo
MPH, and p-methoxy MPH were prepared by asymmetric synthesis
and recrystallized as white hydrochloride salts. The enantiomeric
purities of these optically active compounds were determined by a
previously described gas chromatographic derivatization technique and
found to be >98% (Thai et al., 1998). Racemic threo
ethylphenidate (EPH) was obtained by conventional esterification
methods from ritalinic acid. Derivatization grade pentafluoropropionic
anhydride (PFPA) was obtained from Acros Organics, Fisher Scientific,
Pittsburgh, PA. The medical-grade polysiloxane tubing used to
construct the cannula was obtained from Baxter Scientific (McGraw Park,
IL.).
Surgical Cannulation. Male Sprague-Dawley rats (225-250 g) were anesthetized by i.p. administration of a 200-mg/kg dose of chloral hydrate diluted to a concentration of 75 mg/ml in water. Supplemental methoxyflurane (Metofane) was administered as needed. The right jugular vein was exposed and cannulated with a polysiloxane cannula, which was then passed under the skin and fixed near the base of the neck. Daily flushing of the cannula with 0.028% heparinized saline was necessary to maintain patency. Rats were allowed to recover for at least 60 h from time of surgery before pharmacokinetic studies. All procedures involving handling of animals were approved by the Institutional Animal Care and Use Committee at the University of Pittsburgh.
Drug Treatment. Male Sprague-Dawley rats (225-250 g) were given food and water ad libitum. They were housed two to four per cage and maintained in a temperature-controlled colony room with a 12-h light/dark schedule. The animals were injected i.p. with a 37 µmol/kg dose of either d-threo MPH · HCl (10 mg/kg), d-threo para-bromomethylphenidate (p-Br MPH) · HCl (12.9 mg/kg), or d-threo para-methoxymethylphenidate (p-OCH3 MPH) · HCl (11.1 mg/kg) dissolved in 1.0 ml/kg saline using a 26-gauge 1/2-inch needle.
For plasma pharmacokinetic studies, blood samples (250 µl) were collected through the cannula at 2, 5, 10, 20, 30, 45, 60, 90, 120, and 180 min after injection. Heparinized (0.028%) saline (250 µl) was flushed through the cannula into the rat to purge the line and to replace lost fluid volume. The blood samples were transferred to a 500 µl polypropylene tube containing 7.5 µg of powder Na2EDTA, and the plasma was separated from whole cells by centrifugation at 3000g for 3 min and then stored at
80°C until analysis.
For the tissue distribution studies, rats were decapitated at 30, 60, and 180 min and trunk blood was collected into a Becton Dickinson & Co.
(Lincoln Park, NJ) purple top vacutainer tube containing powder
Na2EDTA. The whole blood was centrifuged at 3000g for 15 min and the plasma was transferred to a
polypropylene tube for storage at 80°C until analysis.
After blood collection, whole brain, heart, left lung, liver, and left
kidney were quickly dissected free and stored in polypropylene tubes
until analysis.
Analytical Methods.
Plasma samples were analyzed for MPH,
p-OCH3 MPH, or p-Br MPH by
a modification of a gas chromatography-mass spectrometry (GC-MS) method
described by Patrick et al. (1984). A 100 µl volume of plasma was
added to an 8-ml glass screw top tube containing 1.0 ml of 0.1 M HCl
and 100 ng of EPH as internal standard. A 4:1 v/v solution of
heptane/ethyl acetate (3.0 ml) was added, and the tube was agitated at
high speed on a mechanical shaker for 2.5 min and centrifuged for 10 min. The top organic layer was aspirated and discarded. The
acidic aqueous layer was alkalinized with 1.0 ml of 0.75 M aqueous
Na2CO3 and then extracted
with 2.0 ml of heptane/ethyl acetate (4:1 v/v). After shaking for 5 min and centrifuging for 10 min, the top organic layer was transferred to a
clear glass screw top tube and evaporated in a Savant Speed Vac
concentrator (Savant, Inc., Holbrook, NY). The resulting residue was treated with 50 µl of PFPA in 25 µl of ethyl acetate at 60°C for 45 min. Samples were dried in a Savant Speed Vac concentrator for
30 min, reconstituted in 50 µl of amyl acetate, transferred to
crimp-top vials with 100 µl volume V-type inserts, and injected into
a GC-MS by auto-injection.
80°C until analysis. The extraction
procedure was carried out on 500 µl of supernatant in identical
fashion to the 100 µl aliquot of plasma.
Standard curves were constructed from spiked blank tissue prepared
under the conditions described above. Drug concentration in tissue is
expressed in terms of micrograms of drug per grams of tissue. Range of
concentrations for the standard curve was between 10 and 25,000 ng/g of
tissue. For both rat plasma and tissue assays, the regression
coefficient (r) > 0.99. Inter- and intra-assay variability were
acceptable with no more than 14% coefficient of variation. Recovery of
all compounds from tissue and plasma was >60%.
Partition Coefficients. The partition coefficients of d-MPH, d-OCH3 MPH, and d-Br MPH were determined between peanut oil and 0.067 M Na2HPO4 buffer (pH 7.4) at room temperature. Both phases were saturated with one another before the assay. Each compound was dissolved in buffer (5.0 ml) to a concentration of 1.0 µg/ml and shaken with peanut oil (5.0 ml) for 1 h. Layers were centrifuged and then separated. A 1.0-ml aliquot of the phosphate buffer phase was then withdrawn for analysis.
EPH (100 ng) was added to each 1.0-ml aliquot as internal standard. For analysis of the buffer layer, 1.0 ml of 0.75 M aqueous Na2CO3 was then added and extracted with 2.0 ml of heptane/ethyl acetate (4:1 v/v). The organic layer was dried and derivatized with PFPA according to methods described above for plasma samples. Standard curves were prepared in 0.067 M phosphate buffer. After determining the drug concentration in buffer, peanut oil concentration was calculated assuming mass balance. Partition coefficients were obtained by dividing concentration of compound in peanut oil by the concentration of compound in phosphate buffer and were performed in triplicate.Plasma Protein Binding. Binding experiments were performed using a Spectrum equilibrium dialyzer (Spectrum Medical Industries, Inc., Los Angeles, CA). Teflon cells with two chambers of 1.3 ml volume each were separated by Spectra-Por 1 regenerated cellulose membranes (Spectrum Medical Industries, Inc., Los Angeles, CA) with a molecular weight cut-off of 6000 to 8000 Da. The membranes were washed with water before use. After assembling the cell, one of the chambers was filled with 1.0 ml of NaCl/Na2HPO4 buffer (66 mM Na2HPO4 in 50 mM NaCl adjusted to pH 7.4). Rat plasma (1.0 ml) containing 0.15% Na2EDTA to inhibit plasma esterases along with 1 µg/ml of d-MPH, d-OCH3 MPH, or d-Br MPH was added carefully to the other chamber.
Equilibrium dialysis was conducted in a 37°C water bath with a rotational speed of 10 rpm for 2 h. The contents of each chamber were transferred to a polypropylene vial for storage at80°C before
analysis. Aliquots of 400 µl volume of plasma and buffer were
extracted, derivatized with PFPA, and analyzed by GC-MS. Standard
calibration curves were generated by spiking drug-free Na2EDTA-treated plasma and sodium phosphate
buffer. All experiments were performed in quadruplicate.
Rat Plasma Esterase Activity.
For esterase activity, fresh plasma obtained by collecting trunk blood
from decapitated rats in heparinized tubes was used. Each of the
d-threo enantiomers of MPH, p-Br MPH,
and p-OCH3 MPH were incubated in
plasma (100 µl) as well as 0.067 M phosphate buffer (100 µl) at
37°C in a final concentration of 1.0 µg/ml. At 1, 2, 4, 7, and
9 h, samples were removed and EPH (100 ng) was added as internal
standard. After alkalinizing the solution with 1.0 ml of 0.75 M aqueous
Na2CO3, 2.0 ml of
heptane/ethyl acetate (4:1 v/v) was added and the compound was
extracted into the organic layer by vortexing and centrifugation. The
top organic layer was then transferred to a clean glass tube and stored
at 80°C until time of analysis. Samples were dried down and
derivatized according to methods described above for plasma samples to
determine levels of the esterified compounds. The study was conducted
in triplicate for each time point and initial pharmacokinetic
parameters were determined from plots of log concentration versus time
using a noncompartmental analysis function in the computer program
WinNonlin (Scientific Consulting, Inc, Apex, NC).
Pharmacokinetic Analysis. Plasma concentration versus time data from each individual experiment was evaluated by noncompartmental analysis as well as a two-compartment model with an absorption phase and lag time using the computer programs PKAnalyst (Micromath, Salt Lake City, UT) and WinNonlin (Scientific Consulting, Inc.). Initial estimates of exponential parameters for compartmental analysis were obtained with the computer program V-Fit (R. Brueckner, M.D., Walter Reed Army Institute of Research). These estimates were then used to fit the data by nonlinear least-squares methods. The goodness of fits were evaluated with informational criteria.
The pharmacokinetic parameters, Tmaxcalc (time at maximum drug concentration), Cmaxcalc (maximum drug concentration), and k
(distributional rate constant) were
obtained from compartmental analysis only and are expressed as the
means ± S.E. of four experiments. The remaining parameters are
the means ± S.E. of values calculated by both noncompartmental
and compartmental methods. The distribution and elimination half-lives
were calculated by dividing 0.693 by the rate constants,
k and k
,
respectively. The Tmaxobs (time at maximum
drug concentration) and Cmaxobs (maximum
drug concentration) are the means ± S.E. of actual observed
values for the four experiments.
The mean residence time (MRT) to the last sampling time was calculated
by the equation:
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lt) is the area under the
first moment curve to the last time and
AUC(0lt) is the area under the curve to the
last time.
Clearance (CL) was calculated as:
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is the elimination rate constant.
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Results |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Pharmacokinetics of d-MPH, d-Br MPH, and d-OCH3 MPH. The plasma pharmacokinetics of the d-threo isomers of MPH and its para-substituted analogs were investigated 3 days after surgery in rats surgically implanted with cannula. These animals were found not to be significantly affected by the surgical procedure or anesthesia after 24 h. They were active and possessed normal appetite on the day of drug administration.
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-elimination phase. At all time points after 20 min, concentrations of d-MPH and
d-OCH3 MPH were not significantly
different from one another, but both were significantly lower than
those of d-Br MPH.
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half-life of d-Br MPH (130 ± 25.8 min) was greater than twice the half-lives of the other two compounds
(51.6 ± 3.56 min for d-MPH; 62.5 ± 15.3 min for
d-OCH3 MPH). The MRT was also greater for bromine-substituted MPH (62.6 ± 4.58 min) relative to
d-MPH (45.4 ± 1.14 min) and
d-OCH3 MPH (43.7 ± 5.60 min). In
accordance with the trends for half-life and MRT, clearance of
d-MPH (318 ± 82.0 ml/min/kg) was greatest followed by
d-OCH3 MPH (238 ± 34.4 ml/min/kg) and d-Br MPH (65.6 ± 3.88 ml/min/kg). The
area under the concentration-time curve (AUC) to last time of
d-Br MPH (134 ± 12.6 µg · min/ml) was 2 to 3 times
greater than d-OCH3 MPH (52.0 ± 10.9 µg · min/ml) and d-MPH (40.6 ± 6.45 µg · min/ml). Although AUC to infinity of
d-OCH3 MPH (56.8 ± 15.3 µg · min/ml) and d-MPH (43.3 ± 6.78 µg · min/ml) were
similar to AUC to the last time for each corresponding drug, the AUC to
infinity of d-Br MPH (203 ± 12.5 µg · min/ml) was
nearly twice its value for AUC to last time. A significant proportion
of AUC to infinity for d-Br MPH came from extrapolation
based on the pharmacokinetic modeling. The apparent distribution
volumes for d-MPH (5.77 ± 1.39 liters) and
d-OCH3 MPH (6.38 ± 2.24 liters)
were similar whereas d-Br MPH (2.98 ± 0.61 liters)
appeared to be confined to a smaller volume. Statistical analyses of
these values by ANOVA and ad hoc methods revealed that there was a
difference for d-Br MPH versus both d-MPH and
d-OCH3 MPH with respect to half-life, AUC to infinity, AUC to last time, and MRT.
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Tissue and Plasma Distribution of d-MPH,
d-Br MPH, and d-OCH3 MPH.
Table 2 shows levels of each compound
measured in rat plasma and tissue at 30, 120, and 180 min after i.p.
injection of the same dose used in the kinetic studies. Distribution
patterns of each drug were quite different from one another.
Nonetheless, levels of compound in tissue were higher in almost all
cases than in plasma. For d-MPH, the order of tissue
concentration at 30 min was kidney > lung > brain > heart 
liver. The d-Br derivative was distributed
with the order lung > liver > kidney > brain > heart. Levels of d-OCH3 MPH were
ranked as lung > brain > kidney > heart > liver.
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Rat Plasma Esterase Activity. The in vitro degradation of the d-threo isomers and their aromatic-substituted derivatives was investigated in rat plasma as well as in 0.067 M phosphate buffer (pH 7.4) at 37°C. The d-bromo derivative was most resistant to plasma esterases followed in order by d-OCH3 MPH and d-MPH. The calculated plasma half-lives shown in Table 4 were obtained from plots of time versus log concentration of each drug over a 9-h time span. The half-lives of d-OCH3 and d-Br MPH in rat plasma were 3- and 8-fold longer than that of d-MPH.
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Partition Coefficients and Plasma Protein Binding. The partitioning of these drugs between peanut oil and Sorenson's phosphate buffer (pH 7.4) was evaluated as a model of their body lipid solubility. The partition coefficients reported in Table 5 were obtained by dividing the drug concentration in peanut oil by its buffer concentration. Thus, d-Br MPH with a value of 14.8 was much more lipid-soluble than the other two compounds. The partition coefficient of d-OCH3 MPH was approximately 1.6 times greater than d-MPH. Plasma protein binding of these compounds was also determined by equilibrium dialysis and is reported in Table 5. Aromatic substitution increased percentage binding by 2.2- and 3.5-fold for methoxy and bromine derivatives, respectively.
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Discussion |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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This work evaluated the plasma pharmacokinetics and tissue
distribution of the d-threo enantiomers of MPH,
p-Br MPH, and p-OCH3 MPH
given i.p. at a dose of 37 µmol/kg. These drugs absorbed quickly and
reached peak levels within the first 10 min of i.p. dosing in similar
fashion to an oral route (Patrick et al., 1984). In addition, the
elimination half-life of d-MPH obtained in two separate studies by Aoyama (1994; 1996) were 46.2 min and 42.7 min. These values
are in agreement with the 51.6-min value obtained by us for
d-MPH as well as the 50-min half-life reported for racemic MPH by Patrick et al. (1984). Further comparisons of other parameters were cautiously avoided because of the differences in the route, amount, and form of dose in each study.
We previously reported a nonlinear relationship between the in vitro
neurochemical potency of the enantiomers of MPH, p-Br MPH,
and p-OCH3 MPH at dopaminergic and
noradrenergic sites and the in vivo locomotor-inducing activity of
these isomers (Thai et al., 1998). In particular,
d-OCH3 MPH possessed a potency of 2.1 and 2.3 for [3H]DA and
[3H]NE uptake inhibition relative to racemic
MPH, yet it did not act as strongly as racemic MPH in locomotion
assays. The d-Br analog was also shown to possess
approximately 8- to 14-fold greater in vitro activity in the same
assays as compared with d-MPH. The locomotor activity of
d-MPH was significantly shorter than d-Br MPH,
which possessed activity even at 180 min.
Upon examination of Fig. 2, it is immediately apparent that during
-phase elimination, decay of p-Br MPH is much slower
than the other two d-threo enantiomers as
evidenced by its more gradual slope on the concentration-time curve.
This observation is reinforced by the substantially longer elimination
half-life of the bromo derivative obtained by both noncompartmental and
compartmental analyses in relation to the parent compound and methoxy
derivative (Table 1). In addition, the MRT of d-Br MPH was
found to be longer and the clearance smaller for this drug. In light of
these results, we felt that perhaps the locomotor profiles of the three
drugs could be explained according to clearance arguments. That is, slower clearance may lead to a longer duration of activity. Although this was consistent with the findings for d-Br MPH, the
explanation did not hold for d-OCH3
MPH, which had similar clearance patterns to d-MPH despite
weaker locomotor stimulation. An interesting point concerning the
absolute values of clearance for these compounds is that clearance of
d-Br MPH was similar to hepatic blood flow in rats (~70
ml/min/kg). On the other hand, values for d-MPH and d-OCH3 MPH exceeded hepatic blood flow
by approximately 4-fold. This is consistent with the notion that both
d-MPH and d-OCH3 MPH are
cleared by extra-hepatic mechanisms such as plasma hydrolysis by
esterases. The half-lives of these compounds (in rat plasma as seen in
Table 4) were much less than d-Br MPH. In light of the above
information, pharmacokinetic parameters, such as clearance, MRT, and
elimination half-life, were consistent with the findings for
d-Br MPH but did not further elucidate potential mechanisms for the differences between d-OCH3 MPH
and d-MPH. We looked to tissue distribution with particular
emphasis on CNS penetrability for additional insight.
Much like the case with the plasma pharmacokinetic assessments, the
tissue distribution data was difficult to place within the context of
currently available literature because of the differences in route of
delivery, dose, and form (enantiomeric versus racemic) of drug.
Nonetheless, qualitative comparisons could be made between our results
and the available literature in terms of rank of tissue concentrations.
After a 1 mg/kg i.v. dose (Kotaki et al., 1988) as well as a 20 mg/kg
i.p. dose (Patrick et al., 1984), the distribution of racemic MPH was
in the order kidney > lung > brain > heart > liver. Our treatment with 10 mg/kg of d-MPH provided the
same outcome. The actual tissue levels reported by Patrick et al.
(1984) after the 20 mg/kg i.p. dose were found to be 4 to 7.5 times
higher than those in Table 2 obtained 30 min after a 10 mg/kg i.p.
dose. Drug levels at the 20 mg/kg dose may represent saturation
conditions such that the liver is unable to absorb and metabolize MPH
as efficiently from portal circulation.
Organ distribution patterns are complicated by several factors. The
most important determinants of organ distribution appear to be plasma
protein binding, lipid solubility, perfusion and extraction efficiency
of each tissue, and inherent tissue affinity for the drug. Metabolic
transformation will also alter the level of drug. MPH is known to bind
very weakly to 4% albumin with a total free fraction of 85% in human
plasma (Faraj et al., 1974). In accordance with these results, a free
fraction of 77.4% for MPH was obtained in our study using rat plasma.
The d-OCH3 and d-Br
derivatives were bound to plasma proteins to a greater degree. In
particular, the free fraction of d-Br MPH was 21.5%. This
represents an approximately 3.6-fold decrease in the amount of drug
available for distribution through lipid membrane layers in various
tissues relative to d-MPH when total plasma drug
concentrations are equivalent. Despite the increase in the plasma
protein binding of d-Br MPH, there were still sufficient
levels of free drug to produce significantly greater levels in all
organs, particularly the brain.
One reason for these observations may be found in the relative lipid
solubilities of these drugs. We chose peanut oil/pH 7.4 buffer as
the organic/aqueous partition because it seems to closely mimic
partitioning between lipid and serum in the body (Dayton and Perel,
1971). We found that the partition coefficient of d-Br MPH
between peanut oil and phosphate buffer at pH 7.4 was several-fold greater than d-MPH and
d-OCH3 MPH indicating greater lipid
solubility of d-Br MPH. In accordance with the partition
coefficient data, d-Br MPH's CNS levels at 30, 120, and 180 min were at least 2-fold higher than d-MPH and
d-OCH3 MPH. Trends in lipid solubility
were paralleled by a high brain/plasma ratio for d-Br MPH at
all time points shown in Table 3. The lipid solubility data for
d-MPH and d-OCH3 MPH, on
the other hand, were not in agreement with their relative brain/plasma
ratios. The ratio for the methoxy derivative was nearly twice as low
despite a slightly greater partition coefficient in peanut oil as
compared to d-MPH. Nonetheless, the CNS results help to shed
light on the locomotor activity profiles obtained in previous
experiments. The ability of these drugs to penetrate the blood-brain
barrier and to remain within the brain seems to be a crucial factor in
maintaining in vivo activity. The high levels of d-Br MPH in
the CNS throughout the 180-min period provide evidence that its longer
duration of locomotor inducing activity are the result of a maintenance
of above-threshold levels at the site of action. Differences in maximal
locomotor activity at 30 min for d-MPH and
d-OCH3 MPH coincide with the higher
brain concentration of the former. At 120 min, the locomotor activity
of rats treated with d-MPH and
d-OCH3 MPH returned to baseline; this
was coincident with a 4- to 14-fold decrease in the CNS levels of both drugs.
Metabolism of MPH by cleavage of the methyl ester group abolishes the psychostimulant properties of the drug. In an attempt to understand the role of deesterification in the organ distribution patterns of the three compounds, a study was undertaken to assess their relative resistance to plasma esterases. The half-life of d-Br MPH in rat plasma relative to the other two drugs was significantly longer, and this relative resistance to metabolism may have been responsible for its higher circulating plasma and tissue concentrations. The results for d-OCH3 MPH as compared to d-MPH were not consistent with the notion that resistance to esterases leads to higher overall drug levels. Although the methoxy derivative had a longer half-life in plasma in vitro, its in vivo plasma pharmacokinetic profile was not significantly different from that of d-MPH. Other metabolic factors, including liver oxidation, may be contributing. One final note on the results of the degradation study was the surprisingly longer half-life of d-Br MPH in plasma relative to buffer. In light of this compound's higher plasma protein binding, it was felt that the amount of drug available to plasma esterases was limited. The presence of plasma proteins may have provided an additional storage site for d-Br MPH and thus imparted a certain level of resistance to degradation in rat plasma.
In conclusion, we report the first comparative pharmacokinetic and tissue distribution study of the d-threo enantiomers of MPH and its p-bromine and p-methoxy analogs in rats. This study was initiated to obtain preliminary pharmacokinetic data that might aid further clinical exploration of these drugs. In addition, we hoped that it would provide possible explanations for the discrepancies between in vitro and in vivo potencies of these drugs. For d-Br MPH, the cumulative consequence of a slower elimination from plasma and favorable CNS penetrability and retention along with high intrinsic potency at the dopamine and norepinephrine transporter systems seems to lead to an extended duration of in vivo locomotor activity. Discrepant in vitro and in vivo activities of MPH and its methoxy derivative may be the result of lower brain levels of d-OCH3 MPH relative to d-MPH. Related studies including lipid solubility, plasma protein binding, and plasma stability lend further support to some of the pharmacokinetic and tissue distribution results. The data above would strongly support further exploration of d-Br MPH as a clinical candidate for a once daily dose treatment of attention deficit hyperactivity disorder and as an in vivo probe for imaging of CNS dopaminergic activity.
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Footnotes |
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Received September 23, 1998; accepted February 10, 1999.
This work was supported in part by National Institutes of Health/National Institute of General Medical Sciences Grant 5T32GM08208.
Send reprint requests to: Dr. James M. Perel, Ph.D., Director, Clinical Pharmacology Program, E-1203 Western Psychiatric Institute and Clinic, 3811 O'Hara Street, Pittsburgh, PA 15213-2593. E-mail: pereljm{at}msx.upmc.edu
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Abbreviations |
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Abbreviations used are:
MPH, methylphenidate;
p-Br MPH, para-bromomethylphenidate;
p-OCH3 MPH, para-methoxymethylphenidate;
GC-MS, gas
chromatography-mass spectrometry;
, elimination constant;
MRT, mean
residence time;
AUC, area under the concentration-time curve;
CNS, central nervous system;
EPH, racemic threo
ethylphenidate;
PFPA, pentafluoropropionic anhydride;
MSD, mass
selective detector.
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References |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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