J. vet. Pharmacol. Therap. 39, 54--61. doi: 10.1111/jvp.12235.

The pharmacokinetics of pimobendan enantiomers after oral and

intravenous administration of racemate pimobendan formulations in
healthy dogs
E. T. BELL*
J. L. DEVI*
S. CHIU* , 1
P. ZAHRA &
T. WHITTEM*
*Translational Research and Animal Clinical
Trial Study (TRACTS) Group, Faculty of
Veterinary and Agricultural Sciences,
University of Melbourne, Werribee, Vic.,
Australia; Racing Analytical Services Ltd,
Flemington, Vic., Australia

Bell, E. T., Devi, J. L., Chiu, S., Zahra, P., Whittem, T. The pharmacokinetics
of pimobendan enantiomers after oral and intravenous administration of
racemate pimobendan formulations in healthy dogs. J. vet. Pharmacol. Therap.
39, 5461.
Pimobendan is a benzimidazole-pyridazinone derivative, marketed as a racemic
mixture for the management of canine heart failure. Pharmacokinetics of the
enantiomers of pimobendan and its oral bioavailability have not been described
in dogs. The aim of this study was to describe pharmacokinetics of three formulations of pimobendan in healthy dogs: the licensed capsule product, and novel
liquid and intravenous formulations. A three-period, nested randomized twotreatment crossover design was used. Pimobendan was administered p.o. at
0.25 and i.v. at 0.125 mg/kg. Blood and plasma samples were analysed by
liquid chromatographymass spectrometry. Noncompartmental modelling was
used to describe the pharmacokinetics. Parameters were compared between formulations using a general linear model. Bioequivalence of the oral formulations
was tested using CI90 for AUC(0) and Cmax. Bioavailability of pimobendan
after oral dosing was 70%. Liquid and capsule formulations were bioequivalent
only for AUC. The positive enantiomer of pimobendan (PE) had a larger volume
of distribution than the negative enantiomer (NE) (281
48 vs. 215
68
mL/kg; P = 0.003) and a shorter half-life (21.7 vs. 29.9 min; P = 0.004). The
NE was distributed more quickly than the PE into blood cells. Enantiomers of
pimobendan have differing absorption, distribution and elimination. The pharmacokinetics of pimobendan in healthy dogs was described.
(Paper received 21 September 2014; accepted for publication 22 April 2015)
Dr Erin Bell, Faculty of Veterinary and Agricultural Sciences, University of
Melbourne, 250 Princes Highway, Werribee, VIC 3030, Australia.
E-mail: belle@unimelb.edu.au
1
Present address: Mississippi State University College of Veterinary Medicine, 240
Wise Center Drive, P.O. Box 6100, Mississippi State, MS 39762, USA

INTRODUCTION
Pimobendan is a benzimidazole-pyridazinone derivative, which
acts as both a positive inotrope and balanced vasodilator, and is
used in the management of congestive heart failure in dogs.
Positive inotropy is mediated by both calcium sensitization,
whereby pimobendan increases the affinity of calcium for
binding to troponin C on cardiac myocytes, and inhibition of
phosphodiesterase (PDE) III, which increases cyclic AMP concentration within myocytes and promotes the release of calcium
from the sarcoplasmic reticulum during systole (R
uegg, 1986;
Scholz & Meyer, 1986; Fujino et al., 1988a; Endoh et al., 1991).
The PDE III inhibition additionally causes both arterial and
venous vasodilation (Fujimoto & Matsuda, 1990). This combination of properties has led to use of pimobendan in the
54

management of congestive heart failure due to dilated

cardiomyopathy or valvular endocardiosis in dogs, and several
clinical trials in dogs with these conditions have reported
increased survival times and reduced clinical signs associated
with pimobendan treatment (Fuentes et al., 2002; Lombard
et al., 2006; Haggstrom et al., 2008; OGrady et al., 2008).
Pimobendan is a racemic mixture of chiral enantiomers (Chu
et al., 1995b), which have distinct pharmacokinetic properties
and pharmacodynamic effects (Williams & Lee, 1985; Drayer,
1986). The () enantiomer has a significantly greater calciumsensitizing and consequently positive inotropic effect than (+)
pimobendan (Fujino et al., 1988b). Stereoselective partitioning
of pimobendan into human red blood cells has also been
reported, with the concentration of () pimobendan within red
blood cells 1.8 times greater than that of (+) pimobendan in one
2015 John Wiley & Sons Ltd

Pharmacokinetics of pimobendan in dogs 55

study (Chu et al., 1995a). Plasma and whole-blood pharmacokinetics of pimobendan (Przechera et al., 1991; Chu et al., 1995a,
b, 1999) and its enantiomers (Chu et al., 1995a) have been
described in humans; however, there is little available information about the pharmacokinetics of pimobendan in dogs.
The pioneer pimobendan formulation, Vetmedin (Boehringer
Ingelheim Vetmedica GmbH, Ingelheim, Germany), was registered with the Australian Pesticides and Veterinary Medicines
Authority (APVMA) in 2002. In Australia, this product was originally available as 1.25, 2.5 and 5 mg capsules; subsequently,
1.25 mg and 5 mg flavoured tablets have also become available.
The objective of this study was to describe the pharmacokinetics in dogs of two new preparations of pimobendan, an oral
solution formulation and an intravenous formulation, which
are planned for regulatory submission for approval. This study
aimed to describe the absolute and relative bioavailabilities of a
new oral pimobendan solution and the pioneer capsule formulations, with a primary hypothesis that the new oral solution
would be bioequivalent to the reference capsule formulation.
MATERIALS AND METHODS
Animals
The study was approved by the University of Melbourne
Animal Ethics Committee and was conducted according to the
Principles of Good Clinical Practice.
Ten adult cross-bred dogs (eight male and two female) were
enrolled in this study. The dogs were medium sized (mean
body weight 18.3 kg; range 12.221.6 kg). They were housed
individually or in pairs in a university animal research facility.
All dogs were determined to be healthy on the basis of physical
examination, haematology and serum biochemistry. The dogs
received no drugs other than the test drug during the study.
Study design and drug administration
The study comprised three periods, with a nested randomized
two-treatment crossover design and washout periods of no less
than 5 days. The dogs were allocated into two treatment groups
(A and B) by lottery without replacement. In period 1, group A
received the reference product (1.25, 2.5 or 5 mg racemate
pimobendan/capsule, Vetmedin capsule, Boehringer Ingelheim
Vetmedica GmbH) (REF) orally at a target dose of 0.25 mg/kg to
the nearest whole capsule, while group B received the investigational veterinary test product (1.0 mg racemate pimobendan/mL
solution; Apex Pimobendan Oral Solution; Apex Laboratories,
Somersby, NSW, Australia) (IVP) orally at the same absolute
dose as the REF product for each dog. In period 2, group A
received the IVP and group B received the REF, at the same dosages. The mean of the actual doses administered was 0.252
(range 0.2050.279) mg/kg. Oral dosing was immediately followed by administration of 10 mL water by syringe over the base
of the tongue. In period 3, all dogs received an experimental
aqueous solution formulation of racemate pimobendan
solubilized with 2-hydroxy-b-cyclodextrin (0.5 mg racemate
2015 John Wiley & Sons Ltd

pimobendan/mL solution, Pimobendan Intravenous Formulation; Apex Laboratories), at a dose equal in mg to one half of the
dose of the REF given to each animal, administered over 60 sec
through an intravenous catheter placed in a cephalic vein. Food
was withheld for at least 12 h prior to dosing in each period.
Sample collection
Blood was collected from preplaced intravenous cephalic catheters or by jugular venipuncture prior to drug administration
and then at the following times afterwards: 0.1, 0.25, 0.5,
0.75, 1, 1.25, 1.5, 1.75, 2, 2.5, 3, 4 and 6 h, for each of the
three treatment periods. Samples were collected into tubes containing lithium heparin and placed over wet ice until processing which was complete within 3 h.
Blood samples were divided into two aliquots of whole blood.
One aliquot was centrifuged at 4 C to separate plasma, which
was then aspirated and frozen at 70 C until analysis. The second aliquot was frozen as whole blood at 70 C until analysis.
Additionally, 0.1 mL of each baseline blood sample in each
treatment period was drawn up into a microhaematocrit tube
and centrifuged for 3 min to determine the packed cell volume
(PCV).
Analytical methods
The analytical method was based on solid phase extraction
from plasma or whole blood followed by normal-phase liquid
chromatographymass spectrometry (LCMS) analysis. Calibration curves for pimobendan were constructed in blank canine
plasma over a range 050 lg/L and blank canine whole blood
over a range 0100 lg/L. Quality control spiked plasma and
whole blood were also prepared at 5 and 25 lg/L. Mebendazole
was used as the internal standard.
Aliquots of plasma (200 lL) were diluted with formic acid
(2%, 400 lL). Each diluted sample was loaded onto a Bond Elut
Plexa PCX cartridge (60 mg, 3 mL, Agilent Technologies, Santa
Clara, CA, USA) which had been preconditioned with methanol
(600 lL) and formic acid (2%, 600 lL). The cartridge was then
washed successively with formic acid (2%, 600 lL) and methanol (600 lL). The sorbent was dried with nitrogen for 3 min
and finally eluted with ammonia in methanol (5% ammonium
hydroxide in methanol, 2 mL). The eluate was evaporated to
dryness under a stream of nitrogen at 60 C. Whole-blood samples (200 uL) were diluted with acetonitrile (800 uL) and vortexed, and the supernatant was decanted. Formic acid (2%,
2 mL) was then added and the sample extracted in the same
manner as plasma samples. The dried extracts were reconstituted in mobile phase (0.1% diethylamine in hexane/0.1%
diethylamine in ethanol, 90/10, 200 lL) for LCMS analysis.
The LCMS analysis was performed on an AB Sciex Q-Trap
mass spectrometer (AB Sciex, Framingham, MA, USA) coupled
to a Nexera LC 30-AD binary pump (Shimadzu, Kyoto, Japan).
Separation was achieved using a Chiracel OD-3 chiral column
(2.1 mm 9 150 mm/3 lm, Diacel Corporation, Osaka, Japan).
The mobile phase consisted of hexane containing 0.1%

56 E. T. Bell et al.

diethylamine (solvent A) and ethanol containing 0.1% diethylamine (solvent B) run at 500 lL/min. The gradient started at
10% B, was held for half a minute and then ramped to 70% B
over 5 min. Solvent B was increased to 98% over the next minute and then held at 98% for a further 4 min. Solvent B was
returned to its starting point over a minute, and then, the column was allowed to equilibrate for 3 min.
The Turbo ion source operated with a spray voltage of
5500 V, temperature of 500 C, curtain gas at 30 units, ion
source gas 1 at 60 units and ion source gas 2 at 0 units. The
analytes were detected in positive ion selected reaction monitoring (SRM) mode. Two ms2 transitions for pimobendan and
one transition for mebendazole were chosen and used for the
quantification of (+) pimobendan and () pimobendan independently. Stereopure standards of the (+) and () forms of
pimobendan used for analysis were sourced from the Laboratory of the Government Chemist (Teddington, Middlesex, UK).
Validation was performed over 3 days by three different analysts. A set of six calibration standards was prepared and processed for each pimobendan enantiomer standard in plasma
(range: 050 lg/L) together with six low-quality controls
(5 lg/L) and six high-quality controls (25 lg/L). The (+)
pimobendan and () pimobendan were eluted from the column
at 4.43 and 5.37 min, respectively.
Data analysis and statistics
The cell-associated concentration of pimobendan was calculated using the following formula: red blood cell-associated
pimobendan concentration = whole-blood pimobendan concentration [plasma pimobendan concentration 9 (100  PCV)/
100]. The individual enantiomer concentrations were evaluated as well as their sum, allowing analysis of the pharmacokinetics of each enantiomer and of the racemate. The
concentration ratios of the enantiomers in whole blood at each
time point were calculated as the quotient of the positive over
the negative enantiomer concentrations.
The area under the whole-blood pimobendan concentration
time curve from time of dosing to the last quantifiable concentration time point postdosing plus the extrapolated portion of
the curve AUC(0) for periods 1, 2 and 3 were calculated
using the linear trapezoidal rule.
The absolute bioavailability for each oral product was calculated as
F

AUC01 oral  Dose IV

:
AUC01 IV  Dose oral

The relative bioavailability was calculated as

F

AUC01 IVP  Dose REF

:
AUC01 REF  Dose IVP

The maximum concentrations of pimobendan reached after

each dose (Cmax) in periods 1 and 2 were defined as the
observed maximum, taken directly from the pimobendan

concentration plots for each individual. The times taken after

oral dosing to achieve Cmax (Tmax) in periods 1 and 2 were
defined as the observed time of the Cmax, taken directly from
the pimobendan concentration plots for each individual.
The elimination rate constant (kel) was calculated by loglinear regression through the last four to eight quantified time
points in each period.
The terminal half-life (t1/2) was calculated as 0.693/kel. The
apparent volume of distribution (Varea) was estimated as
Varea

Dose
:
AUC01  kel

The clearance after intravenous administration was estimated as

Cl

Dose
:
AUC01

Other pharmacokinetic parameters were calculated using

standard formulae (Gibaldi & Perrier, 1982). Descriptive statistics for pharmacokinetic parameters were reported as their
means plus or minus their standard deviations. All pharmacokinetic calculations were achieved using validated spreadsheets
in Microsoft Excel v14.0.7 (Microsoft Corporation, Redman,
WA, USA). The pharmacokinetic parameters for each enantiomer and for racemate pimobendan in whole blood were compared between formulations using a general linear model with
factors for dog, period, sequence and formulation, using Stata
v11.2 for Windows (StataCorp, College Station, TX, USA).
Comparison of mean values between enantiomers within formulation used paired t-tests. P values 0.05 were considered
statistically significant. The bioequivalence of the two oral formulations was evaluated using the 90% confidence intervals
for the logarithm of the ratio of AUC(0) and Cmax, for a
range of plus or minus 20% (Westlake, 1988).

RESULTS
The assay linearity was excellent with the coefficient of determination always in excess of 0.99. The lower limit of quantification for each pimobendan isomer was 2.5 lg/L with a %RSD
of 5.0% for the (+) isomer and 4.6% for the () isomer. The
recovery of both isomers from both dog plasma and whole
blood was greater than 85%.
A logarithmic concentrationtime plot of racemic pimobendan in whole blood, red blood cells and plasma after administration of the intravenous formulation of pimobendan is shown
in Fig. 1. Plasma pimobendan concentrations were found to be
comparatively low at all time points, often below the limit of
quantification, and were very variable both within and
between dogs. The mean plasma Cmax after oral dosing with
the oral solution, oral capsule and intravenous formulation
was 39.4
23.4, 38.1
18.3 and 51.1
28.5 lg/L, respectively. Because of this variability, these plasma concentration
2015 John Wiley & Sons Ltd

Pharmacokinetics of pimobendan in dogs 57

Comparison of oral formulations

The mean Cmax of racemic pimobendan in whole blood after
administration at a nominal dose of 0.25 mg/kg of the liquid
formulation was 407
94 lg/L, which was neither statisti10 000
Pimobendan concentration
(g/L)

Blood racemic
Cellular racemic

1000

Plasma racemic

100

10

1
0

60

120
Time (min)

180

240

Fig. 1. Concentrationtime plot of mean racemic pimobendan in whole

blood, red blood cells and plasma after intravenous administration of a
mean dose of 0.12 mg/kg of an experimental aqueous solution formulation of pimobendan to ten healthy dogs. Legend: X indicates mean
concentration of racemic pimobendan in whole blood; M indicates
mean concentration of racemic pimobendan in red blood cells; + indicates mean concentration of racemic pimobendan in plasma.

1000.0
Pimobendan blood
concentration (g/L)

data are not considered reliable. Therefore, the data analysis

focused on whole-blood pimobendan concentration.
Logarithmic concentrationtime plots of racemic, positive (+)
and negative () enantiomers of pimobendan in whole blood
after administration of an oral solution, oral capsule and intravenous formulation are shown in Figs 24.
Mean pharmacokinetic parameters of racemic pimobendan
in blood after administration of the oral solution, oral capsule
and intravenous formulation are shown in Table 1. The mean
extrapolated portion of AUC(0) was <0.5% of the total in
each individual dog for all formulations considered. Concentrations are reported as means
standard deviations.

100.0

10.0

1.0
0

30

60

90

120 150
Time (min)

180

210

240

Fig. 3. Concentrationtime plot of mean
SD racemic, (+) and

() pimobendan in blood after oral administration of a mean dose of
0.25 mg/kg of a capsule formulation of racemic pimobendan to ten
healthy dogs. Legend: indicates mean concentration of racemic
pimobendan in blood; indicates mean concentration of (+) pimobendan in blood; indicates mean concentration of () pimobendan in
blood.

cally different nor bioequivalent to that reached after administration of the capsule formulation (379
189 lg/L).
However, the extent of the mean difference was not clinically
relevant. The mean absorption time (MAT) was significantly
shorter for the liquid formulation (89
45 min) than for the
capsule formulation (131
59 min). Apart from this difference in the rate of absorption, the remainder of the concentrationtime plots was similar for the liquid formulation versus
the capsule formulation (Fig. 5). The t1/2 after liquid and capsule forms was similar (37.4
8.5 and 37.3
10.2 min,
respectively), as was bioavailability (F) of the two formulations
administered to fasted dogs: 69
16% for the liquid formulation and 67
14% for the capsule formulation.
The geometric mean of the ratios of AUC(0) for the IVP/
REF (the relative bioavailability) was 1.02. The 90% confidence
interval ranged from 0.874 to 1.199, which satisfied the
prechosen parameter for declaring bioequivalence on the basis
of AUC.

1000.0
Pimobendan blood
concentration (g/L)

Pimobendan blood
concentration (g/L)

1000.0

100.0

10.0

100.0

10.0

1.0
0

1.0
0

30

60

90

120 150
Time (min)

180

210

240

Fig. 2. Concentrationtime plot of mean
SD racemic, (+) and

() pimobendan in blood after oral administration of a mean dose of
0.25 mg/kg of a novel liquid formulation of racemic pimobendan to
ten healthy dogs. Legend: indicates mean concentration of racemic
pimobendan in blood; indicates mean concentration of (+) pimobendan in blood; indicates mean concentration of () pimobendan in
blood.
2015 John Wiley & Sons Ltd

30

60

90

120 150
Time (min)

180

210

240

Fig. 4. Concentrationtime plot of mean
SD racemic, (+) and ()

pimobendan in blood after intravenous administration of an experimental aqueous solution formulation of pimobendan, solubilized using
2-hydroxypropyl-b-cyclodextrin, at a mean dose of 0.12 mg/kg of
racemic pimobendan, to ten healthy dogs. Legend: indicates mean
concentration of racemic pimobendan in blood; indicates mean concentration of (+) pimobendan in blood; indicates mean concentration
of () pimobendan in blood.

58 E. T. Bell et al.
Table 1. Pharmacokinetic parameters of racemic pimobendan in blood
after oral administration at 0.25 mg racemate pimobendan/kg using a
novel oral liquid formulation, an oral capsule, and after intravenous
administration at 0.125 mg/kg using an aqueous solution formulation
of pimobendan, in ten healthy dogs. Values are expressed as
means
standard deviations

Dose (mg/kg)
t1/2 (min)
AUC(0inf)
(mgmin/L)
AUMC
(mgmin2/L)
MRT (min)
MAT (min)
Cl (Lh/kg)
Varea (L/kg)
F
Cmax (lg/L)
Tmax (min)

Oral (liquid)
formulation

Oral (capsule)
formulation

0.25
37.4
8.5
30.7
14.8

0.25
37.3
10.2
30.2
16.4

5643
5234

6050
3571

157

89

0.69

407

38.5

199

131

0.67

379

53.9

68
45*

0.16
94
15

63
59*

0.14
189
36.7

Intravenous
formulation
0.118
28.8
6.9
21.3
9.0
1634
1289
68
25

0.392
0.187
0.254
0.093

t1/2, terminal elimination half-life; AUC, area under the curve; AUMC,
area under the first moment curve; MRT, mean residence time; MAT,
mean absorption time; Cl, total body clearance; Varea, volume of distribution; F, bioavailability; Cmax, maximum blood concentration; Tmax,
time to maximum blood concentration.
*denotes statistical difference P 0.05 between oral formulations based
on racemate concentrations.

The bioavailability of the (+) and () enantiomers were

similar: 65
21% and 72
14%, respectively, for the liquid
formulation and 64
20% and 68
13%, respectively, for
the capsule formulation. There was no significant difference
between the enantiomers in time to reach maximum blood
concentration (Tmax) or t1/2. However, the Cmax of the ()
enantiomer was consistently higher than that of the (+) enantiomer, and the volume of distribution of the () enantiomer
was smaller than the (+) enantiomer for both the oral liquid
and capsule formulations.
After intravenous administration, the total body clearance (Cl)
of the (+) enantiomer was approximately double that of the ()
enantiomer, 0.561
0.335 vs. 0.304
0.129 Lh/kg, respectively (P = 0.004). This difference was only partly accounted for
by a larger Varea (0.281
48 vs. 0.215
68 L/kg). The harmonic mean for the t1/2 of the (+) enantiomer was also shorter
than its mirror being 21.7 vs. 29.9 min for the () enantiomer
(P = 0.004 Jacknife approach). Compared to the (+) enantiomer,
the () enantiomer moved much more quickly into association
with the cellular fraction of the blood; at 5 min after administration, the ratio of the (+) enantiomer to () enantiomer was 1.1
in plasma, but only 0.08 in red blood cells (Fig. 6).
After initial equilibration, the ratio of (+) to () pimobendan
gradually declined in whole blood and in red blood cells, but
was very variable in plasma (Fig. 6).
Pharmacokinetics of pimobendan in plasma versus blood

The geometric mean of the ratios of Cmax for the IVP/REF

was 1.18. The 90% confidence interval ranged from 0.852 to
1.644. The lower boundary of the confidence interval satisfied
the prechosen parameter for declaring bioequivalence on the
basis of Cmax. However, the upper boundary exceeded the 20%
interval, precluding a declaration of equivalent Cmax.
Pharmacokinetics of positive and negative enantiomers
Mean pharmacokinetic parameters of (+) and () enantiomers
of pimobendan in blood after administration of the oral solution, oral capsule and intravenous formulations of racemic
pimobendan are presented in Table 2.

There was a consistently greater concentration of (+) pimobendan than () pimobendan in plasma, the opposite to that
which was observed in blood. Also in contrast to the blood,
there was a second peak in plasma concentration of pimobendan at around 60 min postadministration of intravenous
pimobendan; this secondary peak was consistent between dogs.
The plasma concentrationtime curves also flattened at
between 60 and 90 min postadministration of the liquid formulation, and between 90 and 105 min postadministration of
the capsule formulation. These time points coincided with
peaks in the ratio of (+) to () enantiomers in plasma, so the
increase in plasma pimobendan concentrations at these times
may be attributable to an increase in the relative concentration
of the (+) enantiomer.

Pimobendan blood
concentration (g/L)

1000.0

DISCUSSION

100.0

10.0

1.0
0

30

60

90

120 150
Time (min)

180

210

240

Fig. 5. Concentrationtime plot of concentrations of racemic pimobendan in blood after oral administration of 0.25 mg/kg of pimobendan as
a capsule formulation (mean SD; open circles, dotted line) or as a
novel liquid formulation (mean + SD; crosses, solid line).

Although pimobendan is commonly used in the management

of canine congestive heart failure, there has been little available information on the pharmacokinetics of pimobendan in
dogs. Schneider et al. (1997) described the pharmacokinetics of
pimobendan after oral and intravenous administration to
healthy dogs; however, this study used mostly supra-clinical
doses (0.256.75 mg/kg) and measured only plasma concentrations of pimobendan using an assay that was not enantiomer selective. While Schneider et al. (1997) did not present the
Cmax achieved after their 0.25 mg/kg intravenous dose, after
2015 John Wiley & Sons Ltd

Pharmacokinetics of pimobendan in dogs 59

Table 2. Pharmacokinetic parameters of (+)- and ()-enantiomers of pimobendan in blood after oral administration at 0.25 mg racemate pimobendan/kg using a novel oral liquid formulation, an oral capsule, and after intravenous administration at 0.125 mg/kg using an aqueous solution formulation of pimobendan, in ten healthy dogs. Values are expressed as means
standard deviations. Mean values for enantiomers within each
formulation were compared using 2-sample t-tests
Oral (liquid) formulation
+ enantiomer
38.6

10.5

1863

148

94

0.65

147

39.8

t1/2 (min)
AUC(0-inf) (mgmin/L)
AUMC (mgmin2/L)
MRT (min)
MAT (min)
Cl (Lh/kg)
Varea (L/kg)
F(relative)
Cmax (lg/L)
Tmax (min)

7.8
5.8*
1805*
65*
43*

Oral (capsule) formulation

 enantiomer
40.8
20.1
3674
159
84

9.2
8.6
3134
65
42

0.72
0.14
261
55
38.3
14.5

0.21*
41*
18.9

+ enantiomer
48

10.9

2168

195

141

0.64

139

54.2

40
7.5*
1580*
65
59*

0.20
82*
36.7

 enantiomer
40.5
19.2
3868
200
125

8.9
9.1
2058
63
59

0.68
0.13
241
110
53.9
36.7

Intravenous formulation
+ enantiomer
24.3

7.80

499

54

0.561

0.281

7.7
3.55*
446*
25*
0.335*
0.048*

 enantiomer
31.4

13.4

1125

75

0.304

0.215

7.7
5.50
853
25
0.129
0.068

*Statistical difference P 0.05 between enantiomers within formulation.

AUC01 LiquidDose Capsule
Relative bioavailability was calculated as F
: t , terminal elimination half-life; AUC, area under the curve; AUMC,
AUC01 CapsuleDose Liquid 1/2
area under the first moment curve; MRT, mean residence time; MAT, mean absorption time; Cl, total body clearance; Varea, volume of distribution;
F, bioavailability; Cmax, maximum blood concentration; Tmax, time to maximum blood concentration.

Pimobendan enantiomer ratio (+/)

2.0
1.8
1.6
1.4
1.2
1.0
0.8

30

60

90

120

150

180

210

240

0.6
0.4
0.2
0.0
Time (min)

Fig. 6. The ratio of mean concentrations of (+) to () enantiomers of

pimobendan in plasma (solid circles, dotted line), blood cells (open triangle, solid line) and blood (open circle, dotted line) after intravenous
administration of an experimental aqueous solution formulation of
pimobendan, solubilized using 2-hydroxypropyl-b-cyclodextrin, at a
mean dose of 0.12 mg/kg of racemic pimobendan, to ten healthy dogs.

the 2.25 mg/kg dose they achieved a Cmax of approximately

900 lg/L in plasma as estimated from their Fig. 1. In the current study after intravenous dosing at 0.125 mg/kg, the mean
Cmax in plasma approximated 50 lg/L. The Schneider study
showed that dose proportionality is likely acceptable up to
doses of about 2.25 mg/kg, and therefore, the results of their
study and the current study are not dissimilar.
However, the manufacturer of the licensed reference product
reports a maximal mean plasma concentration of pimobendan
of only 3.09
0.76 lg/L after a single oral administration of
0.25 mg/kg of the tablets, observed 14 h postdose (Summary
of Product Characteristics, Vetmedin capsule, Boehringer
2015 John Wiley & Sons Ltd

Ingelheim Vetmedica GmbH, 2012), with plasma levels of pimobendan below quantifiable levels by 4 h after oral administration. This contrasts to a mean plasma Cmax of 38.1
18.3 lg/L
achieved in the current study. This ten-fold disparity may be due
to differences in experimental design (e.g. analytical technique)
or may reflect a real difference in the absolute bioavailability of
the current studys reference capsule product and the tablet formulation used for production of the labelled information.
The pharmacokinetic parameters for the novel liquid formulation were similar to the capsule formulation, with the exception of an increased rate of absorption of the liquid
formulation. The two oral formulations are bioequivalent based
on AUC but not when considering Cmax. The suitability of
using Cmax as a bioequivalence parameter has been debated,
but in this case the difference in Cmax between formulations is
likely to be correctly reflecting the difference in absorption
rates (MAT), identifiable because of the availability from the
current study of intravenous data. The more rapid absorption
has potential to be advantageous as it reduces the lag time
from treatment to effect. The absolute mean increase in blood
pimobendan Cmax after the liquid oral formulations compared
to the reference formulation capsules was only 28 lg/L or 7%
of the reference product, and so although the difference was
statistically significant, it may be of little clinical relevance,
because the interindividual variance in volume of distribution
between the ten dogs was 37%.
In addition to more rapid drug absorption, a liquid formulation of pimobendan may have advantages over capsule or tablet
formulations, such as easier administration in some dogs, more
accurate dosing by body weight, avoidance of lodging of the
product in the oesophagus delaying passage into the stomach
(Westfall et al., 2001) and reduced cost of manufacture (Allen
et al., 2005). These may make an orally administered solution of
pimobendan an attractive alternative.

60 E. T. Bell et al.

The intravenous pharmacokinetics of pimobendan in dogs has

now been described. A single intravenous dose of 0.125 mg/kg
resulted in a maximum mean blood concentration greater than
that obtained with either oral formulations. An intravenous formulation of pimobendan is likely to be more useful than oral formulations for the emergency treatment of dogs with congestive
heart failure, due to a significantly more rapid onset of action
and avoidance of the need to orally medicate dyspnoeic dogs,
which can be associated with significant distress.
Measurement of positive and negative enantiomers of pimobendan did not identify any apparent differences in the elimination half-lives of the enantiomers. However, as previously
described for humans (Chu et al., 1995a), there appeared to be
stereoselective partitioning of enantiomers in dogs, with a
higher concentration of () pimobendan associated with red
blood cells; whether this pimobendan was within red blood
cells or adhered to their cell membranes was not determined.
Increased binding of () pimobendan within cardiac myocytes
has been suggested to be the mechanism by which () pimobendan elicited a greater contractile force than (+) pimobendan
in vitro (Fujino et al., 1988a, 1988b; Chu et al., 1995a). Also,
the () pimobendan isomer has been shown to be associated
with cardiotoxicity at distinctly lower doses than the (+) isomer
(Schneider et al., 1997). Although the relevance of stereoselective partitioning in vivo remains unknown, and may or may
not be clinically important, these findings would become clinically relevant to dose determination if pure-isomeric dose forms
become available.
The absolute bioavailability of orally administered pimobendan in the dog is approximately 70%. The current study did
not, however, determine dose proportionality, so this finding
should be constrained to the dose range investigated.
In conclusion, the pharmacokinetics of the two oral formulations after a single dose of 0.25 mg/kg was similar, with the
exception of a more rapid rate of absorption after administration of the liquid formulation. This faster absorption is likely to
result in a more rapid onset of action. The novel liquid formulation represents an alternative formulation which may have
practical advantages over the existing capsule and tablet formulations. An intravenous dose of 0.125 mg/kg of pimobendan in aqueous solution resulted in a maximal blood
concentration of pimobendan greater than that obtained with
the 0.25 mg/kg orally. Intravenous pimobendan may be a useful adjunctive therapy for the emergency treatment of dogs
with congestive heart failure due to dilated cardiomyopathy or
valvular endocardiosis, but the dosage suitable for such use
should take in to account the difference in bioavailability
between oral and intravenous formulations.

ACKNOWLEDGMENTS
The Authors would like to thank Mr. Garry Anderson for his
assistance with conduct of the statistical analyses. This project
was funded by Apex Laboratories Pty. Ltd., Somersby New
South Wales, Australia.

REFERENCES
Allen, A.D., Anderson, D.P. & Jeffcott, L.B. (2005) Oral dosage forms
and delivery systems. In: The Merck Veterinary Manual. 9th edn. Ed.
Kan, C.M., Merck Sharp & Dohme Corp., New Jersey.
Chu, K.M., Shieh, S.M. & Hu, Y.P. (1995a) Plasma and red blood cells
pharmacokinetics of pimobendan enantiomers in healthy Chinese.
European Journal of Clinical Pharmacology, 47, 537542.
Chu, K.M., Shieh, S.M. & Hu, Y.P. (1995b) Pharmacokinetics and
pharmacodynamics of pimobendan in patients with dilated cardiomyopathy and congestive heart failure after single and repeated oral
dosing. Clinical Pharmacology and Therapeutics, 57, 610621.
Chu, K.M., Hu, Y.P. & Shieh, S.M. (1999) Cardiovascular effect and
simultaneous pharmacokinetic and pharmacodynamic modeling of
pimobendan in healthy normal subjects. Drug Metabolism and Disposition, 27, 701709.
Drayer, D.E. (1986) Pharmacodynamic and pharmacokinetic difference
between drug enantiomers in humans: an overview. Clinical Pharmacology and Therapeutics, 40, 125133.
Endoh, M., Shibasaki, T., Satoh, H., Norota, I. & Ishihata, A. (1991)
Different mechanisms involved in the positive inotropic effects of
benzimidazole derivative UD-CG 115 BS (pimobendan) and its demethylated metabolite UD-CG 212 Cl in canine ventricular myocardium. Journal of Cardiovascular Pharmacology, 17, 365375.
Fuentes, V.L., Corcoran, B., French, A., Schober, K.E., Kleemann, R. &
Justus, C. (2002) A double-blind, randomised, placebo-controlled
study of pimobendan in dogs with dilated cardiomyopathy. Journal of
Veterinary Internal Medicine, 16, 255261.
Fujimoto, S. & Matsuda, T. (1990) Effects of pimobendan, a cardiotonic
and vasodilating agent with phosphodiesterase inhibiting properties,
on isolated arteries and veins of rats. Journal of Pharmacology and
Experimental Therapeutics, 252, 13041311.
Fujino, K., Sperelakis, N. & Solaro, R.J. (1988a) Sensitization of dog
and guinea pig heart myofilaments to Ca2 + activation and the inotropic effect of pimobendan: comparison with milrinone. Circulation
Research, 63, 911922.
Fujino, K., Sperelakis, N. & Solaro, R.J. (1988b) Differential effects of
d- and l- pimobendan on cardiac myofilament calcium sensitivity.
Journal of Pharmacology and Experimental Therapeutics, 247, 519
523.
Gibaldi, M. & Perrier, D. (1982) Pharmacokinetics, 2nd edn. Marcel
Dekker, Inc., New York.
Haggstrom, J., Boswood, A., OGrady, M., Jons, O., Smith, S., Swift, S.,
Borgarelli, M., Gavaghan, B., Kresken, J.G., Patteson, M., Ablad, B.,
Bussadori, C.M., Glaus, T., Kovacevic, A., Rapp, M., Santilli, R.A.,
Tidholm, A., Eriksson, A., Belanger, M.C., Deinert, M., Little, C.J.,
Kvart, C., French, A., Ronn-Landbo, M., Wess, G., Eggertsdottir,
A.V., OSullivan, M.L., Schneider, M., Lombard, C.W., DukesMcEwan, J., Willis, R., Louvet, A. & DiFruscia, R. (2008) Effect of pimobendan or benazepril hydrochloride on survival times in dogs
with congestive heart failure caused by naturally occurring myxomatous mitral valve disease: the QUEST study. Journal of Veterinary
Internal Medicine, 22, 11241135.
Lombard, C.W., Joens, O. & Bussadori, C.M. (2006) Clinical efficacy of
pimobendan versus benazepril for the treatment of acquired atrioventricular valvular disease in dogs. Journal of the American Animal Hospital Association, 42, 249261.
OGrady, M.R., Minors, S.L., OSullivan, M.L. & Horne, R. (2008) Effect
of pimobendan on case fatality rate in Doberman Pinschers with
congestive heart failure caused by dilated cardiomyopathy. Journal of
Veterinary Internal Medicine, 22, 897904.
Przechera, M., Roth, W., Kuehlkamp, V., Risler, T. & Haehl, M. (1991)
Pharmacokinetic profile and tolerability of pimobendan in patients

2015 John Wiley & Sons Ltd

Pharmacokinetics of pimobendan in dogs 61

with terminal renal insufficiency. European Journal of Clinical Pharmacology, 40, 107111.
R
uegg, J.C. (1986) Effects of new inotropic agents on Ca ++ sensitivity
of contractile proteins. Circulation, 73, III78III84.
Schneider, P., Guettner, J., Eckenfels, A., Heinzel, G., von Nicolai, H.,
Trieb, G. & Lehmann, H. (1997) Comparative cardiac toxicity of the
IV administered benzimidazole pyridazinon derivative Pimobendan
and its enantiomers in female Beagle dogs. Experimental and Toxicologic Pathology, 49, 217224.
Scholz, H. & Meyer, W. (1986) Phosphodiesterase-inhibiting properties
of newer inotropic agents. Circulation, 73, III99III108.

2015 John Wiley & Sons Ltd

Westfall, D.S., Twedt, D.C., Steyn, P.F., Oberhauser, E.B. & VanCleave,
J.W. (2001) Evaluation of esophageal transit of tablets and capsules
in 30 cats. Journal of Veterinary Internal Medicine, 15, 467470.
Westlake, W.J. (1988) Bioavailability and bioequivalence of pharmaceutical formulations. In: Biopharmaceutical Statistics for Drug Development, Ed. Peace, K.E., pp. 329352. Marcel Dekker, Inc., New York.
Williams, K. & Lee, E. (1985) Importance of drug enantiomers in clinical pharmacology. Drugs, 30, 333354.