J.vet. Pharmacol. Therap. 13,93-104,1990.

The influence of furosemide on plasma

elimination and urinary excretion of drugs
in standardbred horses
A. J. STEVENSON,* M. P. WEBER,* F. T O D I , ? M. MENDONCA,? J. D. FENWICK,?
E. KWONG,+ L. YOUNG,$ R. L E A V I T T , $ R. NESPOLO,$ P. B E A U M I E R ,
S. T I M M I N G S & S. K A C E W I
*Race Track Division, Agriculture Canada, Ottawa and Jerseyville, Ontario, ?Lynn &Johnston
Laboratories, Lachine, Quebec, +Can Test Ltd., Vancouver, British Columbia, QMannTesting
Laboratories, Mississauga, Ontario and TDepartment of Pharmacology, University of Ottawa,
Ontario, Canada
Stevenson, A.J., Weber, M.P., Todi, F., Mendonca, M., Fenwick, J.D., Kwong, E.,
Young, L., Leavitt, R.,Nespolo, R., Beaumier, P., Timmings, S. & Kacew, S. T h e
influence of furosemide on plasma elimination and urinary excretion of drugs in
standardbred horses. J. vet. Phannacol. Therap. 13, 93-104.
A study of the effects of intravenous administration of either 150 mg or 250 mg
of furosemide to standardbred mares pre-treated with other drugs was
undertaken to determine whether a unique pattern of drug elimination into
urine and from plasma for each compound occurred. Furosemide significantly
reduced the plasma concentrations of codeine compared to control 2 4 h after
furosemide administration. In contrast, the plasma concentrations of theophylline, phenylbutazone, pentazocine, guaifenesin and flunixin were not markedly
altered by furosemide. In the case of acepromazine, clenbuterol and fentanyl,
the data generated were insufficient to state with certainty whether or not
furosemide affected the plasma concentrations of these three drugs. A
significant reduction was noted in the urinary concentrations of guaifenesin,
acepromazine, clenbuterol, phenylbutazone, flunixin, fentanyl and pentazocine
within 1 4 h of furosemide administration. T h e urinary concentrations of
theophylline remained reduced as long as 8 h after furosemide injection.
Furosemide administration to horses pre-treated with codeine resulted in
depression of urinary morphine concentrations 2 4 h and 9-12 h after
furosemide injection. A lower furosemide dose (150 mg) produced changes in
drug urinary excretion and plasma elimination equivalent to the higher dose
(250 mg). It is evident that furosemide affects the urinary and plasma
concentrations of other co-administered drugs but not in a predictable fashion,
which limits the extrapolation of these results to as yet untested drugs.
A. J . Stevenson, Race Track Division, Agriculture Canada, PO Box 5904, Station F,
Ottawa, Ontario, C a d K2C 3x7,

INTRODUCTION
T h e occurrence of exercise-induced pulmonary hemorrhage (EIPH) is common in racing

horses (Cook, 1974; Pascoe et a / . , 1985). T h e

reported incidence of EIPH is variable and
dependent on factors such as length of fibreoptic endoscope, frequency of examination,
93

94 A. J. Stevenson et al.

degree of exercise, age as well as breed of

horse (Clarke, 1985). In general, there is now
a consensus that the prevalence of EIPH in
racing thoroughbred horses is between 40 and
75% (Sweeney & Soma, 1984; Pascoe et al.,
1985). This condition is considered to be a
serious problem in that the ability of the
affected horse to run may be jeopardized.
Some racing jurisdictions have permitted the
pre-race use of the diuretic furosemide to
control EIPH in the racing horse (Combie et
al., 1981); however, in a subsequent study
Sweeney et al. (1984) found that furosemide
administered prior to exercise failed to prevent EIPH. At present scientific studies have
shown that furosemide does not improve
performance of a horse beyond the innate
capability of the animal (Gabel ef at., 1977;
Tobin et al., 1978; Soma et al., 1985).
In recent years data have accumulated to
demonstrate that the racing horse has been
subjected to polypharmacy. Various investigators have reported that furosemide is extensively used in racing horses in conjunction
with analgesic, non-steroidal anti-inflammatory agents (NSAIDs) or anesthetics
(Roberts et al., 1976; Tobin et al., 1977). Since
furosemide exerts a diuretic action, this drug
may dilute out and interfere with the testing
of certain drugs in equine urine and plasma
(Tobin, 1978). Although there is a consensus
that furosemide affects the elimination of
other drugs, it would appear that the data
obtained are variable and dependent on the
drug testing protocol. Studies were thus
undertaken to examine the influence of furosemide on the plasma and urinary concentrations of nine other drugs. In this study the
effects of two different doses of furosemide
on the disposition of these nine drugs in
standardbred horses were determined by
three independent laboratories.
MATERIALS AND M E T H O D S
Standardbred mares weighing between 400
and 500 kg and stabled at a harness racing
training farm were used in this study. All
animals were fed a daily ration consisting of
hay, whole oats and sweet feed (commercial
grain-molasses mixture), and had free access
to water.

Furosemide (trade name Lasix) was purchased from Hoechst Canada Inc., Animal
Health Agriculture Division, Regina, Saskatchewan. Furosemide was administered by
the intravenous (i.v.) route at a dose of either
150 or 250 mg. These doses of furosemide
were selected based on the range of doses
currently permitted by certain racing jurisdictions in the United States for pre-race treatment of horses. In addition to furosemide,
there was a total of nine other test drugs
administered in this study. Under our experimental conditions, the administration of test
drug alone is labelled as the control group
(Type 1). The 150-mg group was test-drug
followed by 150 mg of furosemide (Type 2);
and the 250-mg group was test-drug followed
by 250 mg furosemide (Type 3). The same
four horses were used for a test drug. Since
these three types of experiments were carried
out for each individual test drug the total
number of groups in this study was 27. The
test drugs used, drug source, dose, route of
administration and duration between drugs
are listed in Table I.
In the mares receiving only the test drug,
plasma samples were collected immediately
prior to as well as hourly during a 12-h period
following test-drug administration. I n the
groups given test drug followed by either 150
or 250 mg of furosemide, samples of plasma
were collected immediately prior to test-drug
administration and hourly during a 12-h
period beginning 1 h after furosemide injection (Table I). A plasma sample was also
obtained 24 h after furosemide administration. Urine samples for all three group types
were collected immediately prior to and hourly following test-drug administration for a
12-h period. As in the case of plasma, a urine
sample was collected 24 h after test-drug
administration. Only in the case of fentanyl
was an additional urine sample collected 30
min after test-drug administration. Blood
samples were obtained by venipuncture (BD
6527 Vacutainer; Becton-Dickinson) and immediately centrifuged. Plasma was removed
and stored at -16C for shipment. Urine
samples were obtained via an indwelling Foley
catheter, over a 2 4 2 8 - h period, depending
on the experiment. Urine volume, pH and
specific gravity were measured before storage
at -16C and shipment.

4 h
3 h
2 h

I1

I1

oral or
I.V.

i.v.

I.V.
I.V.

320 clg
500 nig
1.5 g

2 g

2 g
300 mg
300 mg

Boeliringer Ingelheini Ltd.,

Burlington, Ontario
Schering Canada Inc.,
Pointe-Claire, Quebec
Squibb Canada Inc., Montreal,
Quebec
Armitage-Carroll Div. of
Langfortl Inc., Guelph, Ontario
Kougier lnc., Chanibly, Quebec
Winthrop Lab.. Div. of Sterling
Drug Inc., Aurora, Ontario
Allen k I 1anl)urys (Glaxo
Canada Ltd.), Toronto, Ontario

Ventipulmin
Banamine
Aminophylline
Bute Injectable
Cuaifenesin Syrup
Talwin V
Codeine Iliosphate
Injection

Flunixin
Theoph ylline
Phen ylbutazone
Guaifenesin
Pentazocine
Codeine

An example of a protocol using acepromazine as test drug is as follows: in the control group four mares received 10 mg of
acepromazine i.v. In the 150-mg treatment group, the same four horses received 10 mg of acepromazine i.v. and after 1 h these animals
were given 150 mg of furosemide i.v. In the 250-mg treatment group, the same four mares were administered 10 mg of acepromazine i.v.
followed 1 h later by 250 mg of furosemide i.v.

I h
oral

11

2 h

30 rnin

i.v.

I.V.

i.v.

Clenbuterol

400 clg

Janssen Pharmaceuticals Inc.,

Mississauga, Ontario

Sublimaze

Fenian yl

l h

I.V.

10 mg

Ayerst Labs, Montreal, Quebec

Atravet

Acepromazine

Duration prior
to furosemide

Route of
administration

Dose

Drug source

Trade name

Test drug

T A B L E I. Test drugs used in this study

96 A . J. Stevenson et al.
Extraction and analysis

The extraction of drugs from plasma did

not require enzyme hydrolysis. In the case of
guaifenesin, pentazocine and codeine the
extraction of these drugs from urine was
initiated by enzyme hydrolysis utilizing figlucuronidase solid purchased from Sigma
Chemical Co., St Louis, M O (catalogue
number G-0751). At pH 5, this enzyme
preparation possessed 400 000 units of figlucuronidase activity per g solid and 22 900
units aryl sulfatase activity per g solid. To 1 ml
of sodium acetate buffer pH 5 was added 1.2
mg of 6-glucuronidase solid (approximately
500 unitdml) and this solution was subsequently added to 5 ml of urine which was then
incubated at 37C for 16 h. The p H of urine
hydrolysates or intact plasma (4 ml) was
adjusted to 14 in the case of guaifenesin with
5 - ~NaOH, to 9.5 for pentazocine with
aqueous sodium tetraborate and to pH 1 for
codeine with 50% HCI. Guaifenesin and
pentazocine were extracted from plasma or
urine hydrolysate with 2 X 5-ml dichloromethane and the solvent phase was evaporated to dryness under nitrogen. In the case
of codeine, there was an ether wash, readjustment of hydrolysate pH to 9.5 with
aqueous sodium tetraborate, a subsequent 2 x
5-ml dich1oromethane:isopropanol (90: 10)
extraction, and then evaporation of the organic extract with nitrogen. In all three cases
the residue was reconstituted with methanol
(200 PI for plasma) and aliquots were taken
for HPLC analysis. In the case of guaifenesin
the urine residue was reconstituted in 2 ml of
methanol while the methanol volume was 1 ml
for pentazocine and codeine. Aliquots were
then taken for HPLC analysis. T h e analytical
parameters used for HPLC are provided in
Table 11. The HPLC equipment was automated and furnished with a U.V. detector
(Waters Scientific, Ontario).
In the case of acepromazine, clenbuterol,
flunixin, theophylline, phenylbutazone and
furosemide, the volume of urine and plasma
used for extraction was 5 mi, except for
plasma flunixin, for which the volume was
2 ml. In the case of acepromazine or clenbuterol the urine or plasma was adjusted to
pH 11, with 2-N NaOH; for flunixin to pH 67 with phosphate buffer pH 6.6; for theophyl-

line to pH 9.5 with a solution consisting of

A:B in 1:l ratio where A is 25% aqueous
sodium sulfite and B is 460 ml of concentrated
NH40H plus 350 g of NH&I made u p to 1 1
with water; for phenylbutazone to pH 5.0 with
dilute HCI and sodium acetate buffer pH 5 ;
and for furosemide to pH 3.0 with glacial
acetic acid. Following pH adjustment, acepromazine and its metabolites were extracted
with 2 x 5-ml dich1oromethane:isopropanol
(90: 10); clenbuterol was extracted in 2 x 5-ml
hexane:isopropanol (90: 10) flunixin in 2 x 7ml ether; theophylline in 2 X 8-ml chloroform:isopropanol (90: 10); phenylbutazone in
2 x 8-ml ch1oroform:isopropanol (99: 1); and
furosemide in 2 X 6-ml hexane:ether:dichloromethane (1:1:1). After extraction the organic layers in all cases were evaporated to
dryness, reconstituted in methanol (200 pl
for plasma) and aliquots taken for HPLC
analysis (see Table 11). The volume of methanol used to reconstitute urinary residues of
flunixin and furosemide was 1 ml while 2 ml
was used for phenytbutazone and theophylline, and aliquots taken for HPLC analysis.
The concentration of fentanyl was measured using the radio-immunoassay (RIA)
technique of Michiels et al. (1977). Due to the
limited specificity of the RIA method, all
fentanyl concentrations are designated as
fentanyl equivalents.

Detection and identification

The urine from each horse was subjected to

an XAD-2 resin adsorption and extraction
procedure (Timmings et al., 1987). The urine
extracts obtained were spotted on pre-coated
Merck 6 0 - F ~ 5
chromatographic
~
plates with
silica gel with fluorescent indicator (British Drug House, Toronto, Ontario) and
developed in thin-layer chromatographic
chambers. The solvent system for developing
guaifenesin, pentazocine, codeine, morphine
and theophylline consisted of ethyl acetate:
methano1:ammonium hydroxide (85: 10:5).
The solvent system used for flunixin and
furosemide was dich1oromethane:acetic acid
(90: 10); for phenylbutazone and oxyphenbutazone was ch1oroform:methanol (928);
and for acepromazine metabolite and clenbuterol was methano1:ammonium hydroxide

Furosemide influence on drug elimination 97

T A B L E I I. High-pressure liquid chromatographic parameters used for each drug

Column packing
diameter (pm)$

Flow
rate
(mVmin)

0.02-M TEA:
METH (40:60)

10

1.5-2.5

110 (urine)
125 (plasma)

0.02-M TEA:
AN (85:15)

10

1.5 (0-4.8 min)

0.8 (4.9-7.4 min)

225

20

0 . 0 1 6 5 - ~TEA:
AN (13:87)

Flunixin

276

25

0.5% AAA:AN
(65:35)

Guaifenesin

225

20

U.V.detection
wavelength
(nm)

Injection
volume*
(PI)

Solventt

Acepromazine

238

20 (urine)
125 (plasma)

Clenbuterol

240

Codeine (plasma)

Drug

0.5% AAA:AN
(70:30)

Morphine (urine)

225

20

0 . 0 1 6 5 - ~TEA

Phenylbutazone

240

25

1% AAA:AN

1.5

Pentazocine

216

20

0 . 0 1 6 5 - ~TEA:
AN (87:13)

Theophylline

270

25

1% AAA:METH
(90:10)

(50:50)

*In the case of guaifenesin and pentazocine the injection volume for plasma and urine is equal.
t T h e abbreviations used were: TEA, triethylamine; METH. methanol; AN, acetonitrile; AAA. aqueous
acetic acid.
$The column was a C-18 Radial Pak attached to a Waters RCM-100 module.

(99:1). Visualization was accomplished by

examining plates under U.V. light and by
spraying the plates with specific solutions.
A Marquis spray consisting of sulphuric
acid:formaldehyde (9: 1 ) was used to visualize acepromazine, guaifenesin, pentazocine,
codeine and morphine. A Marquis, Mandelin
(containing 98.5 ml of sulphuric acid and
0.5 g of ammonium vanadate dissolved in
1.5 ml of water) and diazo [0.7% sodium
nitrite, 1% ammonium sulfamate, 0.5% N -(L
naphthy1)-ethylenediamine dihydrochloride]
spray sequence was used to visualize furosemide, clenbuterol, oxyphenbutazone and
phenylbutazone. In the case of theophylline a
modified Mandelin's spray (diluted 1:l with
methanol) was used to visualize the drug. A
Ludy Tenger (5 g of bismuth subcarbonate
dissolved in 15 ml of HCI, to which 30 g of
potassium iodide and then 85 ml of water
have been a d d e d w o p p e r chloride (150 g of
cupric chloride in 600 ml of water) 1:1 spray

mixture was used for flunixin. It should be

noted that gas-chromatography-mass-spectrometry was also utilized to confirm the
identity of all drugs and metabolites detected
in this study.
Data were statistically analyzed with Student's paired t-test. Significant differences
between the mean values are indicated when
the P value was (0.05.

RESULTS AND DISCUSSION

In horses given acepromazine alone, the

parent compound or its metabolite reduced
acepromazine (RAP)was detected in plasma
only u p to 1 h after d r u g administration (data
not shown). When furosemide at a dose o f
either 150 o r 250 mg was administered 1 h
after acepromazine, the plasma did not contain any acepromazine or its metabolite. T h e

98 A.

3.

Sleuenson et al.

10

10

15

20

25

Time ( h )

1.5

3.5

5.5

7.5

9.5

Collection midpoint ( h )

FIG. 1. Effect of furosemide on urinary acepromazine sulfoxide (a) concentrations and (b) excretion rate.
Each point represents the mean rt: SEM of four mares per group: control ( 0 ) ;150 mg ( 0 ) ;250 mg (0).For
experimental details, see legend to Table I. *Significant difference between control (test drug alone)
compared to acepromazine followed by 150 mg of furosemide ( P < 0.05). fsignificant difference between
control compared to acepromazine followed by 250 mg of furosemide ( P < 0.05).

concentration of the urinary metabolite, reduced acepromazine sulfoxide (RAPS), in the

absence or presence of diuretic is illustrated in
Fig. l(a). In all animals receiving acepromazine, only RAPS was detected in urine. T h e
concentration of RAPS was maximal at 1 h,
with the exception of one horse in which
RAPS concentrations reached a peak 9 h after
drug administration. In all cases RAPS concentrations decreased gradually from the
peak to a plateau after 12 h. When acepromazine administration was sequentially followed
by either furosemide dose, no RAPS was
detectable in urine 2 h after acepromazine. In
addition, 150 or 250 rng of furosemide significantly decreased urinary RAPS concentrations after 3 and 4 h. Data in Fig. l(a) show
that after 5-24 h the urinary RAPS concentrations were similar in all three groups. The
urinary excretion rate for acepromazine after
furosernide administration is presented in Fig.
l(b). Furosemide did not appear to affect the
urinary excretion rate of aceprornazine. Evidence thus indicates that furosemide markedly dilutes acepromazine metabolite con-

centration in urine for only 2 h and that this

effect does not appear to be d u e to a change in
urinary drug excretion rate. T h e 150-mg dose
of furosemide is as effective as the 250-mg
dose in producing a dilution of acepromazine.
Clenbuterol was not detected in any of the
plasma samples in the absence or presence of
furosemide. Clenbuterol, when administered
alone, was found in urine after 2 h and a
similar concentration was still detected 11 h
after administration (data not shown). By 12 h
following drug administration, clenbuterol
was not detected in the urine. In horses given
clenbuterol followed by either furosemide
dose, clenbuterol was not detected in urine
obtained 1 h after furosemide. In some horses
clenbuterol appeared in the urine at 2 - 8 h
post-furosemide, despite the fact that clenbuterol concentrations were falling prior to
furosemide administration. In contrast to
aceprornazine where there was a return to
control values, the urinary concentration of
clenbuterol was still lower than control 12 h
after furosemide treatment. As in the case of
acepromazine, the urinary excretion rate of

Furosemide influence on drug elimination 99

0.01

1
0

15

20

25

Time (h)

II
0

1.5

3.5

5.5

7.5

9.5

11.5

Collection midpoint ( h )

FIG. 2. Effect of furosemide on (a) plasma and urinary flunixin concentrations and (b) urinary fluxinin
excretion rate. Each point represents the mean +. SEM of four mares per group. In the control group ( 0 )
horses received 500 mg of flunixin i.v. In the 150-mggroup (0)the same four mares were given 500 mg of
flunixin i.v. followed 2 h later by furosemide at 150 mg i.v. In the 250-mg group (0)the same four horses
were injected with 500 mg of flunixin i.v. and after 2 h 250 mg of furosemide was given. *Significant
difference between control compared to flunixin followed by 150 mg of furosemide (P < 0.05). tsignificant
difference between control compared to flunixin followed by 250 mg of furosemide (P < 0.05).

clenbuterol was not markedly altered by furosemide suggesting that the decrease in urinary
concentration was associated with dilution of
d r u g concentration d u e to a rise in urine
volume.
Figure 2(a) illustrates the elimination of
flunixin from.plasma and its excretion into
urine in the presence or absence of diuretic.
I n horses administered only flunixin, the
maximal concentration in plasma or urine was
noted after 1 h. Regardless of the dose of
furosemide, there was a significant reduction
in the urinary concentration of flunixin u p to
3 h after furosemide administration. However, furosemide did not significantly affect
plasma flunixin concentrations. Figure 2(b)
skows that furosemide in general did not
markedly alter the urinary excretion rate of
flunixin indicating that the observed reduction in urinary flunixin concentration was d u e
to the dilution effect of furosemide.
T h e influence of furosemide on theophylline elimination is shown in Fig. 3(a). In horses
given theophylline alone, its plasma and urine

concentrations were maximal after 1 h, decreased steadily with time and were still
present after 24 ,h. A similar elimination
pattern of theophylline from horse plasma
was reported by Errecalde el af. (1984). Furosemide did not significantly enhance the
elimination of theophylline from plasma.
However, Fig. 3(a) clearly shows that both
furosemide doses significantly reduced the
urinary concentration of theophylline from I
to 7 h after diuretic treatment. T h e degree of
reduction of urinary theophylline concentrations was independent of furosemide dose.
T h e effect of furosemide o n urinary
theophylline concentrations was prolonged
compared to its effects on the other drugs. In
general, the urinary excretion rate of
theophylline was not significantly altered by
furosemide except for its reduction between
5-6 and 7-8 h after diuretic treatment (Fig.
3b). T h e decrease in urinary theophylline
concentration observed after furosemide is
thus d u e to the diluting effect of the diuretic.
As in the case of theophylline, the maximal

100 A. J . Stevenson et al.

10

I
0
Time ( h )

1.5

3.5

5.5

7.5

9.5

11.5

Collection midpoint ( h )

FIG. 3. Effect of furosemide on (a) plasma and urinary theophylline concentrations and (b) urinary
theophylline excretion rate. Each point represents the mean 2 SEM of four horses per group. T h e control
group ( 0 )received 1.5 g of theophylline i.v. In the 150-mg group ( 0 )the same four mares were injected with
1.5 g of theophylline i.v. and 2 h later with 150 mg of furosemide i.v. In the 250-mg group (0)the same four
horses received 1.5 g of theophylline followed 2 h later with 250 mg of furosemide i.v. *Significant difference
between control cornpared to theophylline followed by 150 mg of furosemide (P < 0.05). ?Significant
difference between control compared to theophylline followed by 250 mg of furosemide (P < 0.05).

concentration of guaifenesin in plasma and

urine was detected 1 h after drug administration (Fig. 4a). While guaifenesin could not be
detected in plasma beyond 8 h after treatment, it was still present in urine after 24 h.
When furosemide was administered to horses
pre-treated with guaifenesin, the urine excretion pattern was similar to .that noted for
guaifenesin alone. The only exception occurred 1 h after furosemide administration when
urinary guaifenesin was significantly reduced
compared to horses receiving guaifenesin
alone; however, by 2 h post-diuretic there was
no difference between control and treatment
group values. In contrast to other drugs,
furosemide increased the urinary excretion
rate of guaifenesin between 1 and 3 h postdiuretic (Fig. 4b). The reduction in urinary
guaifenesin concentrations seen after furosemide appears to be related to the predominance of dilution over excretion.
Data in Fig. 5(a) illustrate that in horses
given only codeine a peak plasma concentration was found 1 h after drug administration

with drug still present 8 h post-treatment.

Administration of furosemide at either dose
to codeine-pretreated horses resulted in a
significant reduction in plasma codeine concentrations 4-8 h after diuretic treatment.
Unlike the case of the other drugs for which
plasma drug concentrations were determined
in this study, the plasma concentration of codeine after the administration of furosemide
failed to return to the concentration found
when this drug was given alone. At present, it
is not clear why furosemide exerted a prolonged action on codeine plasma concentrations as compared to a transient effect on the
urinary concentration of its metabolite, morphine. Both codeine and morphine were
detected in urine after codeine administration, but since the concentration of morphine
was tenfold higher, only the morphine was
quantified. In horses administered codeine
alone, the maximal urinary morphine concentration was found after 2 h, declined gradually and was still detectable after 24 h. The
influence of furosemide on urinary morphine

Furosemide influence on drug elimination

0.01

10

15

20

25

Time ( h )

0 1.5

3.5

5.5

7.5

9.5

101

11.5

Collection midpoint ( h 1

FIG. 4. Effect of furosemide on (a) plasma and urinary guaifenesin concentrations and (b) urinary
guaifenesin excretion rate. Each point represent the mean k SEM of four mares per group. In the control
group ( 0 )2 g of guaifenesin was administered p.0. In the 150-mg group (0)the same four horses received
2 g of guaifenesin p.0. followed 1 h later by 150 mg of furosemide i.v. In the 250-mg group (0)the same four
mares were given 2 g of guaifenesin p.0. followed 1 h later with 250 mg of furosemide i.v. *Significant
difference between control compared to guaifenesin followed by 150 mg of furosemide (P < 0.05).
TSignificant difference between control compared to guaifenesin followed by 250 mg of furosemide (P <
0.05).

concentrations appeared to be biphasic. Furosemide significantly decreased the urinary

morphine concentration 1-2 h post-diuretic.
This was followed by a return to urine
morphine values similar to those noted in
the control group. However, 9-10 h postfurosemide the morphine concentrations
were again significantly lowered with a return
to test-drug values after 24 h. These results
are similar to those of Combie et al. (1981)
who reported that 250 mg of furosemide
produced a significant but transient (3-h)
decrease in urine morphine concentrations in
horses pre-treated with morphine. In contrast, a biphasic excretion phase was not noted
in their study. It is conceivable that the administration of morphine i.v. by Combie et al.
(I98 1) differs from the i.v. codeine injection
in our study and may account for this difference in urinary morphine concentrations. It is
of interest that furosemide significantly increased the urinary excretion rate of morphine I 4 h after diuretic treatment (Fig. 5b).

It would appear that the changes in urinary

morphine concentrations produced by furosemide are associated not only with dilution of
drug via an increase in urine volume but also
with enhanced d r u g excretion.
T h e plasma elimination and urinary excretion patterns for fentanyl, pentazocine and
phenylbutazone in the presence of furosemide have previously been reported (Roberts
et al., 1976; Miller et al., 1977; Tobin el af.,
1977; Combie et a f . , 1981; Soma et al., 1984).
Consequently, o u r results will be compared
without the use of figures. Fentanyl was only
measurable in plasma for 2 h after its administration alone. When furosemide was given
to fentanyl-pretreated horses. the plasma
fentanyl concentration was reduced and could
be detected only after 1.5 h. It is thus evident
that measurement of plasma fentanyl concentration after a single i.v. injection is not a
reliable index of concentrations of this d r u g
during or following a period of diuresis.
However, Soma et al. (1984) found that dur-

102 A. J. Stevenson et al.

Time ( h 1

Collection midpoint ( h )

FIG. 5. Effect of furosemide on (a) plasma codeine and urinary morphine concentrations and (b) urinary
morphine excretion rate. Each point represents the mean f SEM of four mares per group. The control
group ( 0 )receiqd 300 mg of codeine i.v. T h e 150-mg group (0)(same f o u r mares) w er e given 300 mg of
codeine i.v. and after 2 h 150 mg of furosemide i.v. T h e 250-mg group (0)(same four horses) received
300 mg of codeine i.v. followed 2 h later by 250 mg of furosemide i.v. *Significantdifference between control
compared to codeine followed by 150 mg of furosemide (P < 0.05). ?Significant difference between control
compared to codeine followed by 250 mg of furosemide (P < 0.05).

ing continuous i.v. fentanyl infusion and

maintenance of plasma fentanyl concentrations, there was a n increased urinary excretion of fentanyl equivalents induced by furosemide. Under these experimental conditions
the plasma concentration was considered a
reliable index of d r u g concentrations during
diuresis. In urine the peak fentanyl concentration was attained at 30 min with this drug
being detectable 12 h after d r u g administration. Furosemide administration to fentanylpretreated horses resulted in a significant decrease in urinary fentanyl concentration 1-4 h
after furosemide treatment with a return to
concentrations seen with animals given only
fentanyl after 5-12 h. An identical urinary
fentanyl excretion pattern in the absence and
presence of furosernide was reported previously (Conibie ef al., 1981; Soma et al., 1984).
It is of interest that a dose of 150 mg of
furosemide in o u r study was less than that
used by other investigators yet this dose was
equally effective in reducing urinary fentanyl
concentrations as the 250-mg dose.

In horses treated with pentazocine, maximal plasma concentrations were found after
1 h and were detectable for 4 h. Furosemide
did not markedly affect plasma pentazocine
concentrations. In contrast, furosemide significantly reduced urinary pentazocine concentrations at 1-3 h after diuretic administration
with a n increase in urinary d r u g excretion
rate after 1-2 h. However, 4-22 h after
furosemide the urinary pentazocine concentration was similar to that noted in horses
given the analgesic alone. In separate reports,
Miller ef al. (1977) and Tobin et al. (1977)
found that in horses administered 0.33 rng/kg
of pentazocine followed 30 min later by 1 mgl
kg of furosemide there was a dramatic fall in
urinary pentazocine concentrations 1 h after
furosemide with a return to values as
observed in animals treated with pentazocine
alone by 4 h. It is worth noting that the
urinary elimination pattern for pentazocine is
similar between previous findings and o u r
data. Evidence thus suggests that furosemide
exerts a selective action o n urinary pentazo-

Furosemide influence on drug elimination

cine concentrations without an apparent

effect on its plasma concentrations and that
the fall in drug concentrations is probably due
to dilution via an increase in urine volume.
In agreement with reports by other investigators (Roberts et al., 1976; Gabel et al., 1977;
Tobin et al., 1977) data in the present study
showed that furosemide did not significantly
affect plasma phenylbutazone concentrations.
It is of interest that the plasma elimination of
the metabolite, oxyphenbutazone was also not
affected by either furosemide dose. Furosemide produces a rapid, dramatic fall in
urinary phenylbutazone concentrations i n the
horse (Roberts et al., 1976; Combie et al.,
1981; Gronwall, 1985). This effect was found
to be of short duration, with a return of
urinary phenylbutazone concentrations to
control values. In this study, a similar pattern
was observed. Furosemide significantly decreased urinary phenylbutazone concentrations 1-3 h after diuretic administration without an associated change in urinary drug
excretion rate. This was followed by a return
to values noted for horses given only phenylbutazone. In the case of urinary oxyphenbutazone, the effect of furosemide was more
prolonged with reduced concentrations
observed as long as 5 h after furosernide
treatment. In agreement with previous findings (Gabel et al., 1977) the effect of furosemide on urinary phenylbutazone concentration is not prolonged and a low (150-mg)
diuretic dose is equally effective as a larger
dose.
Our data show that furosemide administration to horses pre-treated with other drugs
results in a variable dilution of each drug in
urine. It is thus clearly evident that the ability
of furosemide to dilute the urinary concentrations of other drugs is not always equal, and
thus the elimination patterns of test drugs in
combination with furosemide can not be
reliably predicted.
ACKNOWLEDGMENTS
This study was funded wholly by the Race
Track Division of Agriculture Canada. T h e
authors gratefully acknowledge the assistance
of the staff of the Equine Drug Evaluation
Centre, Jerseyville, Ontario, F. Chen and P.

103

Price of Can Test Ltd. and J. Nikolajev of

Mann Testing Laboratories. T h e manuscript
was typed by L. Lefebvre.

REFERENCES
Clarke, A.F. (1985) Review of exercise induced
pulmonary haemorrhage and its possible relationship with mechanical stress. Equine Veiennaty
Journal, 17, 166172.
Combie, J.. Nugent. T. & Tobin, T. (1981) The
pharmacology of furosemide in the horse. V. The
duration of reduction of urinary concentration of
drugs. Journal of Equine Veterinary Science, 1 , 203207.
Cook, W.R. (1974) Epistaxis in the racehorse.
Equine Vetm'mly Journal, 6, 45-58.
Errecalde, J.O., Button, C., Baggot, J.D. & Mulders,
M.S.G. (1984) Pharmacokinetics and bioavailability of theophylline in horses. Jounzal of Veterinaty
Ph-ology
and Therapeutics, 7, 255-263.
Gabel, A.A., Tobin, T., Ray, R.S. & Maylin, G.A.
(1 977) Furosernide in horses: a review. Journal of
Equine Medicine and Surgety, 1, 215-218.
Gronwall, R. (1985) Effect of diuresis on urinary
excretion and creatinine clearance of the horse.
Amrican Journal of Veterinaly Research, 46. I6 161618.
Michiels, M., Hendricks, R. & Heykants, J. (1977) A
sensitive radioimmunoassay for fentanyl. European Journal of Clinical Pharmacology, 12, 153-158.
Miller, J.R., Roberts, B.L., Blake, J.W.. Valentine,
R.W. & Tobin, T. (1977) Drug interactions in the
horse. 111. Effects of furosemide on urinary
concentrations of glucuronide metabolites of
pentazocine. Research Communications in Chemical
Pathology and P h a m c o l o g y , 17, 447-456.
Pascoe, J.R., McCabe, A.E., Franti, C.E. & Arthur,
R.M. (1985) Efficacy of furosemide in the treatment of exercise-induced pulmonary hemorrhage in Thoroughbred racehorses. American
Journal of Irefennary Research, 46. 2000-2003.
Roberts, B.L.. Blake. J.W. & Tobin. T . (1976) Drug
interactions in the horse: effect of furosemide on
plasma and urinary levels of phenylbutazone.
Research Communicatioru in Chemical Patholog?i and
Pharmacology. 15, 257-265.
Soma, L.R., Korber. K.. Anderson, T. & Hopkins, J.
(1984) Effects of furosemide on the plasma and
urinary concentrations and the excretion of fentanyl: model for the study of d r u g interaction in
the horse. American Journal of Veterinav Research,
45, 1743-1749.
Soma. L.R., Laster. L.. Oppenlander. F. & BarrAlderfer. V. (1985) Effects of furosemide on the
racing times of horses with exercise-induced
pulmonary hemorrhage. American Journal of
Vetm'nav Research. 46. 763-768.
Sweeney, C.R. & Soma. L.R. (1984) Exerciseinduced pulmonary hemorrhage in Thoroughbred horses: response to furosemide or
hesperidin-citrus bioflavinoids. Journal of the

104 A . J . Stevenson et al.

American Veterinary Medical Association, 185, 195197.
Sweeney, C.R., Soma, L.R., Bucan, C.A. &Ray, S.G.
( 1984) Exercise-induced pulmonary hemorrhage
in exercising Thoroughbreds: preliminarv results
with pre-exercise medication. kornell Vetdinarian,
74, 263-268.
Timmings, S., Beaumier, P., Sutherland, D. &
Nikolajev, J. (1987) Metabolism and elimination
of mazindol by the horse after oral administration. In Proceedings of the Sixth Intemtional Cunference of Racing Analysts and Velennariam. Ed.
Crone, D . pp. 247-254. Royal Hong Kong Jockey
Club, Hong Kong.

Tobin, T. (1978) Pharmacology review: a review of

recent research on furosemide in the horse.
Journal of Equine Meduine and Surgety, 2 , 3 14-32 1.
Tobin, T., Roberts, B.L. & Miller, J.R. (1977) The
pharmacology of furosemide in the horse. I.
Effects on the disposition of procaine, methylphenidate, phenylbutazone, and pentazocine.
J o u m l of Equine Medicine and Surgeq, 1,402-409.
Tobin, T., Roberts, B.L., Swerczek, T.W. & Crisman, M. (1978) T h e pharmacology of furosemide
in the horse. 111. Dose and time response relationships, effects of repeated dosing and performance effects. Journal of Equine Medicine and
Surgery, 2 , 216-226.