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CUN. CHEM.

39/6, 1033-1036 (1993)

Dipyrone Interference on Several Common Biochemical Tests


Neus Gasc#{243}n,13
Caries Otal, Cecilia Martmnez-Br#{252},1
Javier Merc#{233},
Mariano Cort#{233}s,
Rosa Arcelus,
Jos#{233}
M Queraltd, Juan M Sanchez,2 and Francisco GonzIiez-Sastre
We studied the effect in vitro and in vivo of dipyrone on the
determination of several biochemical tests in two analyzers, a Hitachi 747 and a Kodak Ektachem 700. From
studies in vitro, we found significant interference by dipyrone (P <0.05) in the determination of creatine kinase
(CK), lactate dehydrogenase (LO), uric acid, triglycerides,
cholesterol, aspartate aminotransferase,
alanine aminotransferase, and urea nitrogen with both instruments,and
in the determination of creatinine in the Ektachem ana-

lyzer. We also studied the effect of intravenouslyadministered dipyrone in 14 patients. Dipyrone interfered significantly (P <0.05) in the determinationof CK, LD, uric acid,
triglycerides, and cholesterol with both instruments, and
creatinine only with the Ektachem analyzer. Using highperformance liquidchromatography (HPLC), we measured
concentrations of dipyrone in the serum of patients who
had received the drug and observed a negative correlation
between the concentrations of dipyrone in the blood and
the percentage of each analyte concentration.
Indexing Terms: analytical error

drug y

multilayer

film

analysis
Dipyrone
(noramidopyrine
methanesulfonate)
is an
effective analgesic, antipyretic, and antiinflammatory
drug. The pharxnacokinetics
of dipyrone after oral administration
is well documented (1,2) but the pharmacokinetics after an intravenous dose of this drug has not
been described. In spite of its recognized undesirable
effect (agranulocytosis)
(3), dipyrone is one of the most
widely used analgesic drugs in our hospital.
We found that creatinine
was undetectable
when
patients serum samples were measured by an enzymatic method (4, 5) in the Kodak Ektachem 700 analyzer; when measured by a kinetic Jaffe method (6) in a
Hitachi
747 analyzer, the same samples showed the
presence of creatinine.
After examining the medical
history of the patients, we found that all of them had
received an intravenous
injection of 2 g of dipyrone
before blood collection.
We therefore studied in vitro the effect of dipyrone on

the analytical
determination of creatinine
and several
other analytes. We then studied the effect on the analytical results of administering
dipyrone to patients
before withdrawing
blood samples.
We also measured the concentrations of dipyrone in
the blood of these patients by HPLC. The procedures

followed were approved by the ethical committee


hospital.

of our

Material and Methods


For all measurements we used either a Hitachi 747
analyzer (Boehringer
Mannheim, Mannheim, Germany) with Boehringer Mannheim reagents or a Kodak
Ektachem analyzer (Eastman Kodak, Rochester, NY).
In both analyzers we measured creatinine, uric acid,
triglycerides, cholesterol, creatine kinase (CK), total
protein, aspartate aminotransferase
(AST), alanine
aminotransferase (ALT), urea nitrogen (BUN), calcium,
L-y-glutamyltransferase
(GGT), lactate dehydrogenase
(LD), phosphate, sodium, potassium, and chloride.4 For
measuring creatinine and calcium with the Hitachi 747
we used a kinetic Jaff#{233}
method and cresolphthaleincomplexone, respectively.
ChromatographicMeasurement
Dipyrone was provided by Sigma (St. Louis, MO; cat.
no. D-8890). HPLC analysis was performed with a 25 x
4 cm Lichrosorb RP-18 (5 jim) reversed phase column
(Merck,, Darmstadt, Germany). The mobile phase consisted of potassium phosphate 0.1 moIIL adjusted to pH
3 with phosphoric acid and mixed with acetonitrile (200
mJ.JL). The flow rate was 1 mJ.jmin. The standard
solution was prepared by dissolving 2 g of dipyrone in
distilled water to give a concentration of 200 mgIL (57
jimol/L). Perchloric acid (0.33 moIJL) was used to precipitate the samples, which were centrifuged at 6600 x
g for 10 mm. The supernate was injected onto the
column and measured at 258 nm.
Studies in Vitro

The possible interference of dipyrone on all of the


methods was studied in accordance with the
recommendations of the Sociedad Espanola de Qulmica
Clinica (7) for the detection and quantification
of drug
interference in vitro. To a serum pool prepared from
fresh serum samples we added dipyrone up to a final
concentration of 10 gIL (28 460 jimol/L), 10-fold the
expected maximum concentration obtained after a dose
of dipyrone, as suggested by Glick et al. (8); we also
prepared a drug-free pool of serum. Both pools were
analyzed 15 times for each analyte. The outliers were
eliminated
by applying Dixons criteria. We tested the
normality and homoscedasticity of the data by using the
Agostino normality and Snedecor F tests, respectively,
analytical

Servei de Bioquf mica i2 Unitat de Cures Intensives, Hospital


de Ia Santa Creu i Sant Pau, Avda. Pare Claret, 167, 08025
Barcelona, Spain.
Author for correspondence.
Received August 14, 1992; accepted November 25, 1992.

4Nonstandard abbreviati
. CK, creatine
kinase; LD, lactate
dehydrogenase; AST,aspartate aminotransferase; ALT, alanine aminotransferase; BUN, urea nitrogen; GGT,L-y.glutamyltransferase.

CUNICALCHEMISTRY, Vol.39, No. 6, 1993 1033

so that we could compare the means of both groups with


results from Students t-test or with the Mann-Whitney
U test.
If the differences between means were significant
(P
<0.01), the drug was considered to be an interferent.
To quantify the interference, we diluted the pooled sera
containing
10 g/L (28 460 moI/L)
dipyrone with serum from the drug-free
pool to obtain a series of
samples containing
dipyrone at 2846, 1423, 712, 356,
178, 89, 44, and 22 jimol/L. Each sample was analyzed
five times.
Differences of means between the drug-free pool and
the samples containing the interferent
were evaluated
for each method by using the Mann-Whitney
U test.
Studies In Vivo
The study group comprised

14 patients

from the

intensive care unit of our hospital who received 2 g of

Table 1. Minimum Dipyrone Concentration


Producing interference in Vitro

(amoIIL)

Kodak Ektacheai

HitachI

747

Nointerference

Creatinine
Uric acid
Triglycerides

22
44
44

22
4.4

Cholesterol

44
44

44
44

89
89

89
89

1423

1423

CK
AST
ALT
BUN

LD

dipyrone intravenously. Blood samples were obtained


before drug administration
(0) and 2, 5, 10, 30, 60, 120,
and 180 min after the dipyrone administration.
We
obtained blood samples at 360 mm from only 3 of the 14
patients. All analytes in all samples of serum were
measured in duplicate; the results were expressed as
percentages of the result at time 0.
One-way analysis of variance was used to analyze the
influence of the time of blood collection after drug
administration
on the determination
of each analyte.
Differences among individual
time points were tested
with Tukeys range test. The level of significance was P
<0.01.
To determine the duration of interference for each
quantity, we estimated differences of means between
time 0 results and the other time results by the MannWhitney U test. We used the Spearman correlation
coefficient
to determine the correlations
between the
percentages of the analyte concentrations showing interference and the blood concentrations of dipyrone.
Results
Studies in vitro. The minimum concentration of dipyrone that produced any interference with measurements
of the different analytes is shown in Table 1. The graphs
obtained by plotting the percentage of the original
concentration vs dipyrone concentration are shown in
Figures 1 and 2; only the analytes showing interference
from the indicated drug are represented.
Studies in vivo. The means and SD of concentrations
of each analyte at the different times of blood collection
are shown in Table 2 for the Kodak Ektachem and in

Final concentration/original

concentration/original* 100

Final

100

100

100

CK

80
It

60\
KODAK

40
20

HIT*C141747

&

KODAX

500 1000 1500 2000 2500 3000

Dlpyroneconcentration
(pmol/L)

DipyroneconcentratIon(pmoIIL)

Finalconcentration/original
100

Finalconcentration/original
100

AST

80

500 1000 1500 2000 2500 3000

ALT

40

2:
500

1000 1500 2000 2500 3000 0

BUN

::

60

20

Finalconcentration/origInal
* 100

500 1000 1500 2000 2500 3000

500 1000 1500 2000 2500 3000

Dipyrone
concentration
(pmol/L)
Dlpyroneconcentration
(pmoi/L)
Dipyroneconcentration
(pmol/L)
Fig. 1. Effect of differentconcentrationsof dipyroneon the measurementof CK, LD, AST, ALT, and BUN
1034 CUNICAL CHEMISTRY, Vol. 39, No.6, 1993

Final concentratIon/original* 100

FinalconcentratIon/origInal
* 100
TRIGLYCERIDES

80
60
40
20-

\..

20

HITACHI 747

500

1000

Dipyrone

1500

2000

concentration

2500

3000

HITACHI 747

500

Final concentration/original

1000 1500

2000

2500

3000

Dipyroneconcentration(pmol/L)

(pmol/L)
* 100

Finalconcentration/original
* 100
1u9------

CHOLESTEROL

601-

CREATININE

HITACHI 747

4O\

40

KODAK

KODAK

2:!

20

500

1000

Dipyrone

1500

2000

concentration

2500

3000

500

1000

1500 2000

2500

3000

Dipyroneconcentration(pmoi/L)

(pmol/L)

Fig. 2. Effect of differentconcentrations of dipyroneon the measurementof uric acid,triglycerides,cholesterol,and creatinine(Ektachem)

Table 3. Percentages of Anaiyte Concentrations


Obtained with the HitachI 747 in the Studies
Conducted In Vivo

Table 2. Percentages of Analyte Concentrations


Obtained with the Kodak Ektachem in the Studies
Conducted in Vivo
Time of blood collection, mln
Analyt.
CK

10

30

60

120

180

86.1 89.7 93.0 94.0 92.4 91.6


19.9 17.2 12.5 8.5 6.2 7.9 9.2

84.3
0.001

Mean
SD
Uric acid
0.01
Mean
SD
W
0.01
Mean
SD
Triglycerides 0.001

Mean
SD
Cholesterol
Mean
SD

0.001

Mean
SD

Creatlnine

T1m of blood collection, mln

46.3 42.6 48.5 55.7 65.0 69.6 74.3


15.3 25.2 27.2 28.8 28.7 27.2 28.7
93.9 95.8 96.2 96.9 97.4 98.6
5.7 5.7 4.5 4.7 4.8 5.6 9.7

90.7

83.6

6.2

87.6 90.9 94.9 95.0 97.4 97.8


7.6 7.6 10.8 12.8 10.8 18.2

Anolyta

CK
Mean
SD
Uric acid
Mean
SD

0.001

LD

0.01

89.8

92.5

94.2

94.2

94.6

Mean

5.1

5.4

5.9

6.1

6.8

11.8

SD

30

60

120

180

0.001
45.8 62.2 68.2
10.0

SD
Cholesterol 0.001

88.4

10

91.2 92.2 95.4 97.1 94.1 91.8


9.2 10.0 8.6 5.3 2.6 5.1 6.9

Mean

4.5

90.4

Mean
SD
Triglycerides 0.001

84.9

13.0

10.1

77.8
6.0

83.0
7.4

85.6
7.2

87.1
9.2

89.4 89.0 91.5 92.7 91.7 95.2 92.5


7.6 8.2 7.8 6.9 7.8 7.0 11.6
79.4 86.8 89.9 93.1 96.5 93.0 95.7
10.2 5.6 4.5 5.3 5.0 5.4 10.7
88.6

91.1

93.4

94.8

95.1

94.2

94.0

6.7

5.3

5.1

4.7

4.7

5.5

9.2

0.01
81.7 84.0 87.0 90.1 93.4 97.3 96.3
13.0 10.0 9.8 7.3 6.9 7.9 10.1

Table 3 for the Hitachi 747. Table 4 shows the duration


of interference
after dipyrone administration.
The
Spearman
correlation coefficients (ra) relating dipyrone
blood concentrations and percentages of concentration
and their significance are shown in Table 5.
Chromatographic
results. The concentration of dipyrone vs the time of blood collection after drug administration is plotted in Figure 3, distributed in two graphs;
each line corresponds to one of the patients studied.

Discussion
In the studies in vitro, the results from the MannWhitney
U-test were consistent with the presence of
significant
negative interference
(P <0.05) by clipyrone
for the analytes listed in Table 1. The minimum
concentrations of dipyrone producing interference ranged from
22 to 1423 moI/L,
depending on the serum analyte

being measured.
No interference was observed for the measurement of
phosphate, GGT, total protein, sodium, potassium, or chloride in either instrument, and no interference was observed for creatimne in the Hitachi 747.
In the in vivo studies we found significant differences
calcium,

CLINICAL CHEMISTRY, Vol. 39, No. 6, 1993

1035

Dlpyrone(pmor/L)

Table 4. DuratIon of Interference after Drug


Administration In the Studies Conducted in Vivo

Dlpyrone (pmol/L)

Time, mm

Kodak Ektachem
Creatlnekinase
Uricacld
LD

>180
>180
60
60

Triglycerides
Cholesterol

>180
60

Creatinlne

HItachi 747
Creatinekinase
Uric acid

>180
>180
60
120
>180

LD

Triglycerides
Cholesterol

10

30 60

120180

0-

Time (mm)

10 30 60120180
Time (mm)

Fig. 3. Dipyroneconcentrationsin the serum of 14 patientswho


receIved 2 g of dipyroneintravenously
Abscissa: time of blood collection afterdipyrone administration

Table 5. Spearman Correlation Ranks (r) between


Dlpyrone Blood Concentrations and Percentages

centrations
centrations
p

Kodak Ektachem
Creatlnine
Creatine kinase
Uric acid
LD
Triglycerides
Cholesterol
HItachi 747
Creatine klnase
Uric acid
LD
Triglycerides

Cholesterol

67
35
82
82
81
81

-0.48

<o.oi

-0.32

<0.05
<0.01
<0.05
<0.01
<o.oi

-0.85
-0.22

-v.86
-0.39

14

0.41

82

0.80

82
82

-0.24
-0.50

82

-0.20

NS
<0.01
<0.05
<0.01
NS

NS, not signilicant.

between the concentration percentages at time 0 and


after different times of drug administration
for the
analytes listed in Tables 2 and 3. The interference
lasted more than 180 miii after drug administration for
CK in both instruments, for creatinine and triglycerides in the Ektachem analyzer, and for uric acid and
cholesterol in the Hitachi 747. From the results for the
three patients from whom we obtained blood samples
360 miii after dipyrone administration,
we conclude
that drug interference at 360 mm was not significant
(P = 0.001). The duration of drug interference for the
analytes listed in Table 4 was >180 miii.
We found a negative correlation between dipyrone con-

______

1036

CUNICAI.. CHEMISTRY, Vol. 39,

No.6,1993

in the blood and percentages

of analyte con-

(Table 5), with different levels of significance.


From our data we conclude that 2 g of dipyrone administered intravenously has a statistically
significant
negative effect on the measurement of CK, LD, uric acid,
triglycerides,
and cholesterol in both instruments
and on
creatinine

in the Ektachem

analyzer.

Because dipyrone

may negatively affect the measurement of analytes for


180 mm after drug administration,
we recommend that
blood samples should be collected at least 360 miii after
the intravenousinjection of dipyrone.
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