Vet. World, 2012, Vol.5(7):424-428

RESEARCH

Pharmacokinetics and bioavailability of tulathromycin
following intravenous, intramuscular and subcutaneousadministrations in healthy rabbits
K. Abo-El-Sooud, N. A. Afifi, A. M. Abd-El-Aty
Pharmacology Department,
Faculty of veterinary Medicine, Cairo University, Giza, Egypt
Corresponding author: Khaled Abo-EL-Sooud, E-mail: kasooud@yahoo.com
Fax: 0020235725240 – 002035710305.
Received:01-03-2012, Accepted: 06-03-2012, Published Online: 17-04-2012
doi: 10.5455/vetworld.2012.424-428

Abstract
Aim: This work was performed to investigate the pharmacokinetics of the triamilide antibiotic, tulathromycin in healthy
rabbits.
Materials and Methods: Ten rabbits in each group were given a single dose of 2.5 mg/kg body weight (bw) of tulathromycin
via intravenous (IV), intramuscular (IM) and subcutaneous (SC) administrations. The concentration of tulathromycin inplasma was determined by microbiological assay Bacillus subtilis ATCC 6633 as the test organism.
.
Results: Following IV administration, the total body clearance (Cltot) was 321.70 ml/kg h, the volume of distribution atsteady-state (Vdss) was 13.26 L/kg and the value of the elimination half-life (t1/2β) was 29.29 h. After SC administration, theelimination half-life (t1/2el), mean residence time (MRT) and maximum plasma concentration (Cmax) were significantly higher(36.22 h, 52.54 h and 882.19 ng/ml) than after IM route (31.69 h, 45.89 h and 714.72 ng/ml), respectively. Tulathromycin wasbound to the extent of 36% to plasma protein of healthy rabbits. The absolute bioavailabilities were 88.07 and 94.25% after
IM and SC injections.
Conclusion: Thus a single dose of tulathromycin is promising treatment for most respiratory disease in rabbits.
Key words: Pharmacokinetics, tulathromycin, bioavailability, rabbits.

To cite this article: Abo-El-Sooud K, Afifi NA, Abd-El-Aty MA (2012) Pharmacokinetics and bioavailability of
tulathromycin following intramuscular and subcutaneous administrations in healthy rabbits, Vet World, 5(7):424-428, doi: 10.5455/vetworld.2012.424-428
Introduction

Snuffles is considered the most common
infectious respiratory disease observed in pet rabbits[1]. Common clinical signs include nasal discharge,sneezing and conjunctivitis [2,3]. The most commonly
agent implicated with these symptoms is Pasteurella
multocida [4,5]. However, other pathogens are cited,such as Staphylococcus spp. and Bordetella
bronchiseptica, although apparently this is by nomeans an exhaustive list [1]. Macrolide antibiotics are
antibacterial agents that are widely used in thetreatment of infections of the respiratory system, soft
tissue and skin in humans and domestic animals [6].
Tulathromycin, like other macrolides, binds to the 50S
subunit of bacterial ribosomes and thereby inhibits
protein synthesis, leading to inhibition of cell divisionand cell death. Although macrolides are generally
regarded as bacteriostatic, tulathromycin actually exhibits
mixed bacteriostatic and bactericidal activity [7].
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Repeated administration of these products over several
days is typically required to achieve a therapeutic
effect. Prolonged exposure to antibiotics is important
for the treatment and/ or prevention of several diseases
in rabbits, however single administration therapy isdesirable for producers that wish to minimize animal
handling. Tulathromycin is a novel triamilide antibiotic
approved for the treatment of respiratory diseases inanimals. It may be retained in the lung for many days
after administration of a single dose [8-10]. The drug
has a long plasma elimination half-life in cattle of 90 h
[7,11], and therapeutic concentrations have been
detected in lung tissue for 10 to 15 days after a single
dose [11,12]. Therefore, tulathromycin possibly will
be recommended for use in rabbits for several
respiratory diseases including pneumonia. To ourknowledge there have not been any reports publishedon the pharmacokinetics of tulathromycin in rabbits.
Consequently, the aim of the present study was toinvestigate the pharmacokinetic profile of tulathro-

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Pharmacokinetics and bioavailability of tulathromycin following I/V, I/M and S/C administrations in healthy rabbits

mycin in rabbits following a single IV, IM and SC
injections to estimate an appropriate dosage regimenof tulathromycin in rabbits.
Materials and Methods
Drug: Tulathromycin was used as 10% injectable
solution (Draxxin®, Pfizer Animal Health, New York,NY), Each ml of Draxxin® contains 100 mg oftulathromycin as the free base in a 50% propylene
glycol vehicle, monothioglycerol (5 mg/ml), withcitric and hydrochloric acids added to adjust pH.
Animals and Husbandry: Thirty healthy New
Zealand white rabbits of both sexes, 10-12 months old
and weighing 2.7-3.5 kg were used. Rabbits werehoused separately in individual cages under a 12-h
light/dark cycle and fed good quality hay (alfalfa)
and/or a pelleted feed concentrate (fiber 18%, protein
14%, calcium >1 and fat 2%) with free access to water.
The room temperature and relative humidity weremaintained at 20 and 220C, and between 30 and 60%,
respectively. The animals were allowed to acclimatize
and did not receive any drug treatment for at least 15
days preceding the study. The experiments werecarried out according to the National regulations on
animal welfare and institutional animal EthicalCommittee (IAEC).
Experimental design: The animals were allocated tothree groups of 10 rabbits each. Rabbits wereindividually weighed before drug administration anddoses were calculated precisely. Rabbits of all groupsof groups were injected a at a dose of 2.5 mg/kg bw of
10% solution of tulathromycin intravenously into theleft ear vein, intramuscularly into the left semimembranous muscles and subcutaneously in the scruff
using graduated 1 ml syringes, respectively.
Blood samples from the all groups (0.5 ml each)were taken via indwelling catheter into heparinized
Vacutainers (Becton Dickinson vacutainer Systems,
Rutherford, NJ, USA), from the right ear vein at 0
(blank sample), 0.25, 0.33, 0.5, 0.75, 1, 2, 4, 6, 8, 10,
12, 18, 24, 48 h and everyday for 10 days after all
routes of injections. Plasma was separated bycentrifugation at 2000g for 10 min and stored at -20 0C
until assayed.
Assay for Tulathromycin: Tulathromycin concentrations in plasma were determined by a microbiological agar plate assay [13] using Bacillus subtilis
ATCC 6633 as the test organism. Standard curves oftulathromycin (Pfizer Animal Health, New York, NY)
were prepared in pooled antibacterial-free plasma. All
samples were directly added to the culture plate. The
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limit of quantitation by this method was 20 ng/ml inplasma. The response of tulathromycin was linear over
the range of concentration between 20 and 2000 ng/ml
with a correlation coefficient (r2) of 0.998. The intraassay coefficient of variation rate of elimination (CV)was 8%. Negative control samples (non-treated)showed no bacterial inhibition, indicating no intrinsic
antibacterial activity of the samples.
In vitro plasma protein binding: The extent ofprotein-binding was determined in vitro using themethod of Craig & Suh [14] which is based on the
diffusion of the free antibiotic into the agar medium.
The drug was dissolved in phosphate buffer (pH 6.2)
and antibiotic-free rabbit's plasma at concentrations of200, 400, 600, 800 and 1000 ng/ml. The differences inthe diameter of the inhibition zone between thesolutions of the drugs in the buffer and plasma were
calculated. The percentage of protein bound fraction
was calculated according to the following equation:
Protein binding % = Zone of inhibition in buffer –Zone of inhibition in Plasma × 100 / Zone of inhibitionin buffer
Pharmacokinetic analysis: Plasma concentrationsof tulathromycin after IV, IM and SC administrations
were subjected to a compartmental analysis using anonlinear least-squares regression analysis with thehelp of a computerized curve-stripping software
package (R Strip; Version 5.0; Micromath Scientific
Software, Salt Lake City, UT, USA). Data were
examined by sequential weighted nonlinear regression.
Monoexponential, biexponential and triexponential
equations were fitted to individual plasma concentration–time data. Akaike's Information Criterion(AIC) [15], residual sum of squares (Rs) and analysisof residual's plots were used to discriminate between
models [16]. The distribution and elimination halflives (t1/2 and t1/2), the volume of distribution at steadystate (Vdss) were calculated according to standard equations[17]. The total body clearance was calculated as Cltot =Dose/ AUC. Statistical moments were used tocompute the non-compartmental model of area under
the concentration–time curves (AUC), area under the
first moment curve (AUMC); mean residence time(MRT) and bioavailability (F), where:F = [mean AUCIM or SC/ mean AUCIV] X 100.
Statistical analysis: The statistical analysis was
performed using the SPSS® 10.0 software package(SAS, Cary, NC, USA). Results are presented asarithmetic mean ± standard errors (SE). The
nonparametric Wilcoxon test was used to compare theVeterinary World, Vol.5 No.7 July 2012425

Pharmacokinetics and bioavailability of tulathromycin following I/V, I/M and S/C administrations in healthy rabbits
10000

IM

Concentration of tulathrom ycin (ng/m L)

C o n c e n t ra t io n o f t u la t h ro m y c in ( n g /m L )

10000
SC

1000

100

10

1
0

12

24

36

48

60

72

84

96

108

120

132

144

156

1000

100

10

1
0

12

24

36

48

60

72

84

96

108

120

Time (hours)

Time (hours)

Figure-1. Mean ± SE of plasma concentrations oftulathromycin in rabbits after SC and IM administrations of
2.5 mg/kg b.w. (n=10)

Figure-2. Mean ± SE of plasma concentrations oftulathromycin in rabbits after IV injection of 2.5 mg/kg
b.w. (n=10)

Table-1: Mean ± SE plasma pharmacokinetic parameters of tulathromycin in normal rabbits (n=10)
following IV, IM and SC administrations at a dose rate of 2.5 mg/kg bw.
Parameterst

1/2 alphat1/2abt1/2beta
t1/2elVdss
Cltot
AUC0AUMC
MRT
Cmax
Tmax
F

Unit

IV

IM

SC

h
h
h
h
L/kg
ml/kg/hng/h/mlng/h2/ml
h
ng/ml
h
%

0.50 + 0.10--29.29 + 5.30--13.26 ± 2.10
321.70 + 15
7771.7 + 287
262855 + 723
37.25 + 2.45---------0.11 + 0.01--31.69 + 1.80----6844.7+ 252
231501+ 655
45.89 + 2.25
714.72 + 24
1.00 + 0.26
88.07 + 3.20--0.20 + 0.02**--36.22 + 2.26----7324.5 + 304
247729 + 578
52.54 + 2.85*
882.19 + 30**
1.55 + 0.26
94.25 + 4.10

t1/2alpha: distribution half-life; t½ab: absorption half-life; t1/2 (t½el): elimination half-life; Vdss: volume of distribution; Cltot: total bodyclearance; AUC: area under the curve by the trapezoidal integral; AUMC: area under moment curve by the trapezoidal integral;
MRT: mean residence time; Cmax: maximum plasma concentration; Tmax: time to peak concentration; F%: bioavailability. Valuesof diseased rabbits were significantly different from corresponding normal rabbits at * P<0.05, **P<0.01.

parameters obtained in healthy and diseased rabbits
following each route of administration. Means wereconsidered significantly different at p< 0.05 and P<0.01.
Results

All animals remained in good health throughout
the acclimatization and study periods. No adverse
effects were observed in the present studies when thetulathromycin formulation was administered torabbits at 2.5 mg/kg bw following IV, IM and SC
administrations. Mean (±SE) tulathromycin plasma
concentration–time curves in healthy rabbits
following IV, IM and SC injections are plotted on asemi-logarithmic graph in Figures 1&2. Values ofpharmacokinetic parameters are presented in Table 1.
Following a single IV route the plasma concentration
time curve of tulathromycin was best fitted to follow a
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two compartment open model. The total bodyclearance (Cltot) was 321.70 ml/kg.h, the volume of
distribution at steady-state (Vdss) was 13.26 L/kg andthe value of the elimination half-life (t1/2β) was 29.29 h.
After SC administration, the elimination half-life(t1/2el), mean residence time (MRT) and maximum
plasma concentration (Cmax) were significantly higher(36.22 h, 52.54 h and 882.19 ng/ml) than after IM route(31.69 h, 45.89 h and 714.72 ng/ml), respectively.
Tulathromycin was bound to the extent of 36%
to plasma protein of healthy rabbits. The absolutebioavailabilities were 88.07 and 94.25% after IM and
SC injections, respectively. The slow elimination andextensive distribution following a single dose oftulathromycin are fortunate pharmacokinetic
characteristics and promising for the treatment of
respiratory disease in rabbits.

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Pharmacokinetics and bioavailability of tulathromycin following I/V, I/M and S/C administrations in healthy rabbits

Discussion

This study used the microbiological assay for
determination of tulathromycin in plasma which does
not separate the parent compound from the active
metabolites. Tulathromycin is metabolized slowly,
and the majority of drug is excreted unchanged infeces and urine [7]. Nevertheless, it measures the total
activity which could be more useful for pharmacodynamic evaluations than high performance liquid
chromatography (HPLC) methods [18]. In the present
study, pharmacokinetics of tulathromycin is characterized in normal healthy rabbits after three routes ofadministrations. Following IV administration oftulathromycin (2.5 mg/kg bw), the average volume of
distribution at steady-state (Vdss) is 13.26 L/kg which istypically similar to that reported in swine and larger
than that reported in cattle, 11 L/kg [12]. The calculated
half-life of elimination following IV administration in
rabbits was (29.29 h) shorter than the values reported
in swine (67.5 h) by Benchaoui et al., [19].
Consequently the systemic clearance of tulathromycin
in our study (321.70 L/h/kg) was faster than reported
in swine (181 L/h/kg) [19]. Differences in the kinetic
parameters are relatively common and frequently
related to interspecies variation, age, breed, healthstatus of the animals and/or the assay method used[20]. A basic nature, limited degree of ionization, andlipophilicity are characteristics of macrolides ingeneral, enabling extensive drug penetration into
tissues and fluids, and resulting in large volumes of
distribution [8,21,22]. Triamilides are basic
compounds and include in their structure three amine
sites with pKa1 = 8.49, pKa2 = 9.28 and pKa3 = 9.80.
These properties allow tulathromycin to achieve hightissue penetration. In this respect, Benchaoui et al.,[19] stated that tulathromycin is extensively
distributed to lung tissue following IM administrationand the peak concentrations in the lung were markedly
higher (lung Cmax = 3470 ng/g) than in plasma (Cmax =616 ng/mL), and were achieved by 24 h (Tmax) after
injection. Over the time course following administration, drug concentrations in the lung were 24.9 to
181 times higher than those measured in plasma. Also,
Evans, [7] stated that tulathromycin is slowly releasefrom cells, are thought to account for the prolonged
drug levels in lung and other tissue. Actual lung halflife values for cattle and swine were calculated as 184
and 142 hours, respectively [11,19]. Consequently, it
has been suggested that tulathromycin may be safe andeffective in treating P. multocida and other pulmonary
pathogens in rabbits.
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After SC administration, the elimination halflife (t1/2el), mean residence time (MRT) and maximum
plasma concentration (Cmax) were significantly higher(36.22 h, 52.54 h and 882.19 ng/ml) than after IM
route (31.69 h, 45.89 h and 714.72 ng/ml),
respectively. These values are lower than reported indomestic goats after SC injection by Clothier et al.,2011. The mean time of maximum plasma
concentration (Tmax) following SC injection (1.55 h) is
longer than estimated in swine [19] and in cattle[11&23]. Tulathromycin was bound to the extent of
36% to plasma protein of healthy rabbits. The value
was similar to the value reported in cattle (40%) by
Nowakowski et al., [11] and lower than reported ingoats (50%) by Clothier et al., [24].
The absolute bioavailabilities were 88.07 and94.25% after IM and SC injections, respectively
indicating excellent absorption of tulathromycin fromparenteral routes. Similar high values have beenobtained for tulathromycin after IM route in swine(87.7%) by Benchaoui et al., [19] and after SC route incattle (91%) by Nowakowski et al., [11].
Conclusion

We conclude that a single dose of tulathromycin
at 2.5 mg/kg bw is promising for treatment of
respiratory disease in rabbits owing to its fortunate
pharmacokinetic characteristics which are slow
elimination and extensive distribution.
Acknowledgements

The authors acknowledge Pharmacology
Department, Faculty of veterinary Medicine, Cairo
University for the great help in rearing and handling of
rabbits. This work has been performed by the authorsof their own interest.
Competing interest

Authors declare that they have no competing interest.
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