Paracetamol pharmacokinetics and metabolism in young women

Abstract

Background:
There is relevant between individual variability in paracetamol clearance in young women. In this pooled study, we focused on the population pharmacokinetic profile of intravenous paracetamol metabolism and its covariates in young women.

Methods:
Population PK parameters using non-linear mixed effect modelling were estimated in a pooled dataset of plasma and urine PK studies in 69 young women [47 at delivery, 8/47 again 10-15 weeks after delivery (early postpartum), and 7/8 again 1 year after delivery (late postpartum), 22 healthy female volunteers with or without oral contraceptives].

Results:
Population PK parameters were estimated based on 815 plasma samples and 101 urine collections. Compared to healthy female volunteers (reference group) not on oral contraceptives, being at delivery was the most significant covariate for clearance to paracetamol glucuronide (Factor = 2.03), while women in early postpartum had decreased paracetamol glucuronidation clearance (Factor = 0.55). Women on contraceptives showed increased paracetamol glucuronidation clearance (Factor = 1.46). The oestradiol level did not further affect this model. Being at delivery did not prove significant for clearance to paracetamol sulphate, but was higher in pregnant women who delivered preterm (<37 weeks, Factor = 1.34) compared to term delivery and non-pregnant women. Finally, clearance of unchanged paracetamol was dependent on urine flow rate.

Conclusions:
Compared to healthy female volunteers not on oral contraceptives, urine paracetamol glucuronidation elimination in young women is affected by pregnancy (higher), early postpartum (lower) or exposure to oral contraceptives (higher), resulting in at least a two fold variability in paracetamol clearance in young women.

Full text

R E S E A R C H A R T I C L E Open Access
Paracetamol pharmacokinetics and
metabolism in young women
Karel Allegaert
1,2*
, Mariska Y. Peeters
3
, Bjorn Beleyn
2,4
, Anne Smits
1,2
, Aida Kulo
5,6
, Kristel van Calsteren
2,7
,
Jan Deprest
2,7
, Jan de Hoon
4,5
and Catherijne A. J. Knibbe
3,8
Abstract
Background: There is relevant between individual variability in paracetamol clearance in young women. In this
pooled study, we focused on the population pharmacokinetic profile of intravenous paracetamol metabolism and
its covariates in young women.
Methods: Population PK parameters using non-linear mixed effect modelling were estimated in a pooled
dataset of plasma and urine PK studies in 69 young women [47 at delivery, 8/47 again 10–15 weeks after
delivery (early postpartum), and 7/8 again 1 year after delivery (late postpartum), 22 healthy female volunteers
with or without oral contraceptives].
Results: Population PK parameters were estimated based on 815 plasma samples and 101 urine collections.
Compared to healthy female volunteers (reference group) not on oral contraceptives, being at delivery was
the most significant covariate for clearance to paracetamol glucuronide (Factor = 2.03), while women in early
postpartum had decreased paracetamol glucuronidation clearance (Factor = 0.55). Women on contraceptives
showed increased paracetamol glucuronidation clearance (Factor = 1.46). The oestradiol level did not further
affect this model. Being at delivery did not prove significant for clearance to paracetamol sulphate, but was
higher in pregnant women who delivered preterm (<37 weeks, Factor = 1.34) compared to term delivery and
non-pregnant women. Finally, clearance of unchanged paracetamol was dependent on urine flow rate.
Conclusions: Compared to healthy female volunteers not on oral contraceptives, urine paracetamol
glucuronidation elimination in young women is affected by pregnancy (higher), early postpartum (lower) or
exposure to oral contraceptives (higher), resulting in at least a two fold variability in paracetamol clearance in
young women.
Keywords: Acetaminophen, Glucuronidation, Oestradiol, Oral contraceptives, Paracetamol, Pregnancy,
Progesterone
Background
Characterizing pharmacokinetics (PK) and pharmaco-
dynamics (PD) in specific subpopulations is essential to
improve therapeutic effectiveness while minimizing ad-
verse events [1, 2]. Gender related differences in body
weight, physiology (e.g. pregnancy) or endocrinology
(e.g. menstrual cycle) may affect PK. This concern is
also reflected in the Food and Drug Administration
(FDA) guidance on bioavailability and bioequivalence
studies. This guidance recommends that in vivo bioequiv-
alence studies should be conducted in representative indi-
viduals, taking into account age, gender or race. If the
drug is intended for use in both sexes, one should attempt
to include similar proportions of male and female volun-
teers [3]. The same case can be built for specific set-
tings, like drugs intended to be used during pregnancy
(e.g. pruritus of pregnancy, tocolytics, gestational dia-
betes or hypertension) [4]. We aim to quantify the im-
pact of covariates on paracetamol metabolism in young
women, including pregnancy and postpartum [4–10].
Paracetamol is almost exclusively metabolized by the
liver. In adults, only 1–4 % is excreted in urine as
* Correspondence: karel.allegaert@uzleuven.be
1
NICU, University Hospitals Leuven, Herestraat 49, 3000 Leuven, Belgium
2
Department of Development and Regeneration, Cluster Organ Systems, KU
Leuven, Leuven, Belgium
Full list of author information is available at the end of the article
© 2015 Allegaert et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Allegaert et al. BMC Anesthesiology (2015) 15:163
DOI 10.1186/s12871-015-0144-3
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

unchanged paracetamol while the majority is excreted as
paracetamol-glucuronide (47–62 %) or paracetamol-
sulphate (25–36 %) [5–10]. A smaller part (8–10 %) is
oxidized by cytochrome P450 (including CYP2E1, but
also CYP1A2 and possibly CYP3A4) into 3-hydroxy-
paracetamol and the toxic metabolite N-acetyl-p-benzo-
quinone-imine (NAPQI) [5–9]. Compared to early post-
partum (10–15 weeks) observations, paracetamol clearance
was significantly higher (21.1 vs 11.7 l.h
−1
, + 80 %) at deliv-
ery. This higher clearance was due to a disproportional in-
crease in glucuronidation (11.6 vs 4.76 l.h
−1
, + 144 %), a
proportional increase in oxidation clearance (4.95 vs
2.77 l.h
−1
, 78 %) and primary renal clearance (1.15 vs
0.75 l.h
−1
,53%)[6].Thisincreaseinglucuronidation
clearance may in part be driven by oestradiol, and may
explain within and between individual differences in
paracetamol metabolism (e.g. oral contraceptives, fol-
licular vs luteal phase, postpartum, pregnancy, or dur-
ation of pregnancy) in young women [6, 8, 9, 11–14].
Based on a pooled analysis, we aimed to further explore
the impact of these covariates on paracetamol metabol-
ism based on plasma and urine collections in women at
delivery, in postpartum (early, or late) and healthy
volunteers, either or not on oral contraceptives (OC)
following intravenous (iv) paracetamol administration
[6, 11, 15].
Methods
Study populations and design
Young women at delivery, in early and late postpartum
This was an open-label, 3-period PK study (at delivery,
early, and late postpartum) conducted from August 2010
to March 2013 (EudraCT Number 2010-020164-37)
[6, 11, 16]. The study documents (study protocol, in-
formed consent, subsequent amendments) were reviewed
and approved by the local Ethics Committee of the
University Hospitals Leuven. This study followed GCP
(Good Clinical Practice) and local regulations. Written in-
formed consent of each woman (at least 18 years, adult-
hood according to the Belgian law) was obtained before
study initiation. The study was registered (www.clinical-
trials.gov, 19 October 2015, NCT02590900).
The administration of iv paracetamol (vial containing
1000 mg in 100 ml infusion solution, Perfusalgan®, Bristol
Myers Squibb Braine l’Alleud, Belgium) is part of routine
multimodal analgesia following caesarean delivery in the
University Hospitals Leuven [6, 11, 16]. Consequently,
patient consent was restricted to the collection of add-
itional blood samples, urine collection and the inclusion
in a database (demographic and clinical characteristics).
Pregnant women scheduled for elective or (semi)urgent
caesarean delivery and immediate postoperative iv para-
cetamol pain relief were considered. Women with known
paracetamol intolerance or who were already receiving
paracetamol in the period of 48 h prior to study were not
included [6, 11, 16].
In the first study period, an initial iv 2 g loading dose of
paracetamol (two vials) was administered to the patient by
the attending anesthesiologist within 5 min following
delivery of the newborn. Subsequent 1 g maintenance
doses were administered by the nurse at 6 h intervals for
maximal 24 h with a subsequent switch to oral paraceta-
mol. Paracetamol was administered either as 20 min
(loading dose) or 10 min (maintenance dose) infusion,
through a peripherally inserted venous catheter [6, 11, 16].
To further enrich the variability in clinical characteristics
at delivery compared to the earlier reported dataset [16],
an additional cohort of women undergoing preterm cae-
sarean delivery was recruited.
During the second study period, a subgroup of eight
women initially included in the first study period at deliv-
ery were admitted again for a single iv 2 g loading dose ad-
ministration and 6 h follow up, scheduled 10–15 weeks
after delivery of the newborn (early postpartum) [16].
Finally, the same subgroup of eight women were admitted
again about 1 year after delivery (late postpartum), using
the same study design.
For the duration of the first study period, subjects were
hospitalized at the maternity ward and for the second and
third study period at the Centre for Clinical Pharmacol-
ogy, University Hospitals Leuven, Leuven, Belgium. Only
cases with both plasma and urine observations were re-
cruited in this cohort. At the different time points, clinical
characteristics, including body weight and height, duration
of pregnancy, medical conditions and the use of oral con-
traceptives - when applicable - were registered.
Young healthy, non-pregnant female volunteers
To compare observations at delivery and in postpartum
with a reference group, eight healthy young non-pregnant,
female volunteers (18–40 years) were recruited. Using the
same sampling strategy, these women received a single iv
2 g loading dose and 6 h follow up. Clinical characteristics,
including body weight and height were collected. The
non-use of oral contraceptives was an explicit inclusion
criterion. This was to enable comparison with another
cohort of young women (n= 14) exposed to the same
loading dose (2 g iv paracetamol), followed by 1 g q6h for
48 h published by Gregoire et al. [15]. All these women
were on contraceptives, of whom 13 were on oral contra-
ceptives (ethinyloestradiol containing pill), one used a
levonorgestrel containing intrauterine device (this volun-
teer was classified as not exposed to ethinyloestradiol-
containing oral contraceptives).
Blood sampling and urine collection
Following delivery, seven blood samples (2 ml per sample)
were collected per subject. The first three samples were
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collected at 1, 2 and 4 h after initiation of the 2 g loading
dose. The next four samples were collected just before the
next maintenance doses (i.e. at 6, 12, 18 and 24 h). Blood
samples, drawn through a second, peripherally inserted
venous catheter dedicated for blood sampling only, were
collected into plastic lithium heparin tubes, immediately
centrifuged and plasma was stored at −20 °C until analysis.
In women undergoing a caesarean delivery, urine was
collected through a bladder catheter. Before the first dose,
the urine collection bag was emptied and a blank urine
sample was collected in order to exclude the possibility of
paracetamol being present in urine. Second and third
urine collections were harvested from 0 to 6 and 6–
24 h respectively, after the total urine volume was mea-
sured. After collection, urine samples were immediately
stored at −20 °C until analysis.
Inthesingledosestudies(earlypostpartum,latepostpar-
tum and healthy volunteers), a 2 g loading dose was admin-
istered to the subjects after they had voided. Four blood
samples at predetermined time points (1, 2, 4 and 6 h after
initiation of dosing) and one urine sample (extracted from
0 to 6 h urine collection) were collected following the same
principles described for the first study period [6, 11, 16]. In
the Gregoire et al. study, only plasma samples were
collected during repeated intravenous paracetamol admin-
istration [15].
Bioanalytical methods
Concentrations of unchanged paracetamol (plasma, urine)
and its metabolites paracetamol-glucuronide (urine) and
paracetamol-sulphate (urine) were determined by high
performance liquid chromatography (HPLC), according to
a previously validated and reported method [16]. The
lower limit of quantification for paracetamol in plasma
was 0.08 mg l
−1
, and for paracetamol and its metabolites
in urine 1 mg l
−1
. Coefficients of variation for intra- and
interday precision and accuracy were all below 15 % [16].
In the study of Gregoire et al., a HPLC method with
UV detection was used to quantify paracetamol concen-
trations in plasma, following a systematic dilution pro-
cedure (max 1/20). The analytical procedure in plasma
was shown to be linear from 0.020 to 10.0 mg/ml with
the limit of quantification at 0.020 mg/ml [15].
Oestradiol and progesterone levels were determined
for each patient at each study point via competitive
enzyme-linked immunosorbent assay (ELISA) with elec-
trochemiluminescence (MODULAR® ANALYTICS E-
170, Roche/Hitachi) by the clinical laboratory of the
University Hospitals Leuven [11].
Data analysis and population PK parameter estimates
The analysis was performed using non-linear mixed ef-
fect modeling (NONMEM, GloboMax LLC, Hanover,
MD, version VI) by use of the first-order conditional
estimation (Method 1) with η-εinteraction and
ADVAN6 TOL5. Parent drug and metabolites were
modelled simultaneously. For this purpose, the amounts
of unchanged paracetamol, paracetamol-glucuronide and
paracetamol-sulphate excreted in urine were calculated by
urinary concentration (mg l
−1
) multiplied by urine volume
and subsequently converted to milligram paracetamol
equivalents using a molecular weight of 151.2 mg mmol
−1
for paracetamol, 328.3 mg mmol
−1
for paracetamol-
glucuronide and 230.2 mg mmol
−1
for paracetamol-
sulphate.
S-plus (Insightful software, Seattle, WA, version 6.2) was
used to visualize the data. Model building was performed in
four different steps: (i) selection of the structural model
(one, two or three compartment model), (ii) choice of a
statistical sub-model, (iii) covariate analysis, and (iv) model
evaluation. Discrimination between different models was
made by comparison of the objective function. A value
of P< 0.01, representing a decrease of 6.63 points in the
objective function, was considered statistically signifi-
cant. In addition, goodness of fit plots including obser-
vations vs individual predictions, observations vs
population predictions, conditional weighted residuals
vs time and conditional weighted residuals vs popula-
tion predictions were used for diagnostic purposes. Fur-
thermore, the confidence interval of the parameter
estimates, the correlation matrix and visual improve-
ment of the individual plots were used to evaluate the
model.
The paracetamol data were best described with a
three-compartment model, parameterized in terms of
the volume of the central compartment (V
1
), inter-com-
partmental clearances between central and peripheral vol-
umes (Q and Q
1
), peripheral volumes (V
2
and V
8
),
clearance to paracetamol-glucuronide (CL
P-G
=V
1
*k
13
),
clearance to paracetamol-sulphate (CL
P-S
=V
1
*k
14
),
clearance of unchanged paracetamol (CL
P-U
=V
1
*k
17
)
(Fig. 1). Clearance attributable to pathways other than
these measured in urine, the oxidative metabolites (CL
P-O
)
could not be significantly identified. With the current
study design, the metabolite volumes of distribution of
paracetamol-glucuronide and paracetamol-sulphate (V
3
and V
4
) cannot be identified, but were fixed to 18 % of the
central distribution volume of paracetamol in plasma [17].
Using this approach, the elimination rate of paracetamol-
glucuronide from plasma to urine (k
35
) equals the elimin-
ation rate of paracetamol-sulphate (k
46
). Relating the rate
of elimination of unchanged paracetamol (k
17
)tok
35
and
k
46
by estimation of a multiplication factor (MF) as k
35
=
MF*k
17
resulted in a significant decrease of objective func-
tion (ΔOF 40.9).
The uncertainty in the population parameters (coeffi-
cient of variation, CV) was estimated in NONMEM by
the covariance step. Individual estimates of the PK
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Fig. 1 Schematic representation of the pharmacokinetic model and its metabolites in plasma and urine [Abbreviations: P, paracetamol; P-G,
paracetamol-glucuronide; P-S, paracetamol-sulphate; P-U, unchanged paracetamol; V1, volume of the central compartment; V2 and V8, volumes of
the peripheral compartment; Q and Q1, inter-compartmental clearances between central and peripheral compartment; k, elimination coefficients]
Table 1 Clinical characteristics of the study population. Data are provided as by mean and standard deviation or incidence
Pregnancy and postpartum Healthy volunteers
at delivery postpartum, early postpartum, late no oral contraceptives contraceptives
Number of cases 47 8 (8 of 47) 7 (7 of 8) 8 14
Plasma samples, number and time 275, 0–24 h 32, 0–6 h 28, 0–6 h 32, 0–6 h 448, 0–24 h
Urine collections, number and time 78, 0–24 h 8, 0–6 h 7, 0–6 h 8, 0–6 h n.a.
Age (years) 30.9 (5.3) 32.1 (3.9) 32.9 (4.1) 31.1 (4.3) 23.5 (4.0)
Body weight (kg) 79.7 (12.9) 68.8 (11.2) 67.1 (13.5) 63.9 (6.6) 59.8 (8.9)
Body surface area (m
2
) 1.93 (0.19) 1.79 (0.17) 1.76 (0.2) 1.74 (0.1) 1.66 (0.14)
<37 weeks, at delivery 21/47 3/8 3/7 n.a. n.a.
37–41 weeks, at delivery 26/47 5/8 4/7 n.a. n.a.
Oestradiol (pg.ml
−1
) 4 833 (3 555) 86 (30) 75 (65) 79 (70) n.a.
Progesterone (ng.ml
−1
) 118 (95) 1.1 (0.55) 0.4 (0.2) 2.8 (3.9) n.a.
Follicular/luteal phase (number, each) n.a. 3/0 5/0 6/2 n.a.
Oral contraceptives (number/total) n.a. 4/8 2/7 0 13/14
Allegaert et al. BMC Anesthesiology (2015) 15:163 Page 4 of 11
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parameters were assumed to follow a log-normal
distribution. Therefore, an exponential distribution
model was used to account for between individual
variability. Residual errors were best described with a
proportional error model. The residual error for the
paracetamol data of Gregoire et al. [15] were best
described with a combined additive and proportional
error model.
Covariate analysis
The covariates body weight, body height, body surface
area, age, gestational age (GA), being at delivery, in early
Table 2 Parameter estimates (mean (CV%)) of the final population PK model for paracetamol and its metabolites in women at
delivery, early postpartum, late postpartum or healthy volunteers, with or without oral contraceptives (OC). For Cl
PG
,V
1
and
Q
1
both the final value and the equation is presented in which values in italic represent the value for the standard
population for that parameter
Parameter Mean Bootstrap mean
final model (CV%) (CV%)
Fixed effects
At delivery postpartum, early postpartum, late +
healthy volunteers
10 weeks later
CL
PG
(L/h) 7.33 (8.3) 2.02 (11.1)
0.55 (18.5) × 7.33 = 4.0 7.41 (9.7)
OC: 1.46 (12.5) × 4.0 = 5.8 OC:1.46 × 7.33 = 10.7 0.56 (19.5)
2.03(8.8) × 7.33 = 14.9 1.48 (10.8)
CL
PS
(L/h) 3.86 (5.5) 3.86 (5.5) 3.82 (5.6)
Preterm = 5.61 (7.9) 5.65 (8.4)
CL
PU
(L/h) 0.93 (6.3) + 0.0053 (28.2) × (UP-100) 0.94 (6.5)
0.0054 (29.8)
V
1
(L) 1.86 (6.3) × 18.5 =18.5 (7.9) 1.83 (6.4)
34.4 18.5 (7.4)
V
2
(L) 19.7 (33.6) 22.3 (37.9)
V
8
(L) 23.9 (5.4) 23.9 (5.0)
Q (L/h) 1.29 (15.0) × (BW/70) 1.34 (14.2)
Q
1
(L/h) 61.1 (6.8) 0.13 (17.9) × 61.1 = 7.9 61.1 (6.8) 61.6 (6.3)
0.13 (19.2)
MF 4.62 (11.8) 4.73 (10.7)
Interindividual variability
ω
CLpg
2
0.12 (23.0) 0.12 (23.2)
ω
V1
2
0.09 (24.1) 0.08 (24.4)
ω
CLpu
2
0.12 (61.6) 0.11 (58.7)
Residual error
σ
2(P plasma)
0.07 (12.8) 0.07 (13.8)
σ
2(P G)
0.29 (48.6) 0.29 (46.0)
σ
2(P S)
0.15 (26.1) 0.14 (23.5))
σ
2(P u)
0.15 (20.4) 0.15 (17.9)
σ
2(P plasma) Gregoire [
15
]
0.02 (21.6) 0.02 (19.4)
σ
2(P plasma), additive Gregoire [
15
]
0.016 (64.4) 0.016 (61.8)
Performance measures
−2LL 5286.743 5241.994 (3.8)
Values in parentheses are CV, coefficient of variation of the parameter values; OC; oral contraceptives; CL
PG
, clearance to paracetamol-glucuronide; CL
PS
, clearance
to paracetamol-sulphate, CL
u
, clearance to paracetamol unchanged; UP, urine production (urine volume (ml) divided by collection time (h)); V
1
, central volume; Q
and Q
1
, intercompartmental clearance between central and peripheral volumes; BW, body weight; V
2
and V
8
, peripheral volumes; MF multiplication factor for k17
compared with k35 and K46; ω
2
variance, the square root of the exponential variance of ηminus 1 is the percentage of interindividual variability in the parameters; σ
2
proportional within individual variance; −2LL, objective function
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postpartum (10–15 weeks after delivery), late postpar-
tum (1 year after delivery), healthy female volunteer,
term/preterm delivery (dichotomous), twin pregnancy,
maternal morbidity (pre-eclampsia, diabetes mellitus,
either type 1 or gestational), use of oral contraceptives,
oestradiol and progesterone levels and urine production
(ml h
−1
) were plotted subsequently against the individual
post-hoc parameter estimates and the weighted residuals
to visualize potential relationships. Based on these plots,
covariates were tested for their influence. Starting from
the basic model without covariates, the covariate model
was first built up using forward inclusion (p< 0.005,
representing a decrease of 7.88 points in objective func-
tion). The contribution of each covariate was
subsequently confirmed by stepwise backward deletion
(p< 0.001, representing a decrease of 10.82 points in
objective function). In the final model, all covariates as-
sociated with a significant increase in objective function
after elimination were maintained. The choice of the
model was further evaluated as described in the data
analysis.
Model validation
The internal validity of the population PK model was
assessed by the bootstrap re-sampling method (repeated
random sampling to produce another dataset of the
same size but with a different combination of individ-
uals) with stratification, taking into account the number
of individuals at delivery and postpartum. Parameters
obtained with the bootstrap replicates (250 times) were
Fig. 2 Diagnostics plots for the final PK model for awomen at delivery, bwomen in early postpartum and cwomen in late postpartum and d
healthy volunteers including observations vs individual predictions (left) and observations vs population predictions (right) for paracetamol
concentrations in plasma (circle, upper panels) and amount of paracetamol-glucuronide (diamond), paracetamol-sulphate (triangle), and unchanged
paracetamol (square) in urine (lower panels) as paracetamol equivalents with x = y identity line. The solid symbols indicate women on contraceptives,
the open symbols women with no contraceptives. In panel cthe group of healthy volunteers on contraceptives (n= 14, Gregoire) are distinguished
from women in late postpartum by a triangle down, the healthy volunteers with no contraceptives (n= 8) by symbols plus (circle plus; diamond plus,
triangle box and square plus)
Allegaert et al. BMC Anesthesiology (2015) 15:163 Page 6 of 11
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compared to the estimates obtained from the original
dataset.
Simulations
Simulations were performed for women at delivery,
women 10–15 weeks postpartum (early postpartum) or
healthy volunteers/late postpartum upon a single iv
loading dose of 2 g of paracetamol, followed by 1 g q6h
for 24 h, with or without exposure to oral contraceptives
in the non-pregnant women.
Results
The pooled dataset was based on PK studies in 69 indi-
viduals. Forty-seven pregnant women were enrolled at
delivery, of whom eight were enrolled again 10–15 weeks
after delivery, and seven of these eight cases again 1 year
after delivery. Eight healthy female volunteers (not on
oral contraceptives) were recruited, and raw data on 14
healthy female volunteers on contraceptives (13 oral
contraceptives, one used a levonorgestrel covered intra-
uterine device) were provided by the sponsor of the
Gregoire study [15]. The clinical characteristics of the
different cohorts and the respective number of plasma
and urine observations collected are provided in Table 1.
In Table 2, the PK parameter estimates, the between and
within individual variability and the bootstrap analysis of
the final model are provided. Estimates in the specific
subgroups (at delivery, early postpartum, late postpartum,
on oral contraceptives) were provided as a Factor com-
pared to the estimates in volunteers and late postpartum
cases without oral contraceptives (reference group).
Figure 2 shows the observed versus individual predicted
concentrations/amounts and the observed versus popu-
lation predicted concentrations/amounts for plasma
and urine observations for the final model for (Fig. 2a)
women at delivery, (Fig. 2b) women in early postpartum
(10–15 weeks after delivery), and (Fig. 2c) late postpar-
tum (1 year) or healthy volunteers, with or without oral
contraceptives.
The systematic covariate analysis showed that being at
delivery was the most significant covariate for clearance
to paracetamol glucuronide (ΔOF 78.9, Factor = 2.03).
The influence of oestradiol levels or progesterone levels
on glucuronidation clearance - implemented as a power
function - resulted in decreases in objective function of
60.5 points and 68.9 points respectively. However, imple-
mentation of oestradiol or progesterone in addition to
being at delivery on glucuronidation did not further
improve the model. Women in early postpartum showed
a decreased paracetamol glucuronidation clearance
(Factor = 0.55) compared to healthy women (ΔOF = 26.4,
vs basic model; ΔOF 29.1 backward deletion vs final
model, p< 0.001). Women taking oral contraceptives
showed increased paracetamol glucuronidation clearance
Fig. 3 Model based simulation of plasma paracetamol disposition after 2 g loading dose, followed by 1 g paracetamol every 6 h in women with
different clinical characteristics [at delivery (a,circle), in early postpartum (b,triangle), in late postpartum or in healthy volunteers (c,cube)]. For the
band cpanel, simulations are provided with (white) or without (black) exposure to oral contraceptives
Allegaert et al. BMC Anesthesiology (2015) 15:163 Page 7 of 11
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

versus women without oral contraceptives (Factor = 1.46,
ΔOF 6.9, P< 0.01, vs basic model; ΔOF = 15.4 backward
deletion vs final model). Being at delivery did not prove
to be a significant covariate for clearance to paracetamol
sulphate. However, clearance to paracetamol sulphate
was higher in pregnant women who delivered preterm
(<37 weeks, Factor = 1.34) compared to term delivery
and non-pregnant women. Finally, clearance of un-
changed paracetamol was dependent on urine flow rate
(diuresis, mean urine flow 100 ml/h). The addition of
urine production (urine volume, ml divided by collection
time, h) as a linear equation on clearance of unchanged
paracetamol for the measured range 20–283 ml/h re-
sulted in a significant decrease of objective function
compared with the basic model (ΔOF 38.6, P< 0.001).
For missing values the urine production was assumed to
be 100 ml.h
−1
.
Central volume standardized for body weight signifi-
cantly improved the model. However, being at delivery as
a covariate for the central volume proved to be more
significant. Addition of body weight on central vol-
ume for the different groups did not further improve
the model. The inter-compartmental clearance (Q)
standardized for body weight(BW)provedsignifi-
cant. The inter-compartmental clearance Q1 was re-
duced in women in early postpartum (Factor = 0.13)
relative to the population mean of 61.1. (ΔOF = 46.6
backward deletion vs final model). The group late
postpartum could not be identified as significant co-
variate, which would suggest that the pharmacokinet-
ics 1 year postpartum equals the healthy volunteer group.
The impact of these covariates (pregnancy, early/late post-
partum, volunteers, with or without oral contraceptives)
on plasma paracetamol disposition is illustrated in Fig. 3.
Fig. 4 Clearance to paracetamol-glucuronide (CL
P-G
,grey, l.h
−1
), clearance to paracetamol-sulphate (CL
P-S
,transparent, l.h
−1
) and clearance of
unchanged paracetamol (CL
P-U
,striped, l.h
−1
) as estimated at delivery, in early postpartum, or in late postpartum/healthy volunteers are provided
with the impact of the other covariates (preterm on CL
P-S
at delivery, oral contraceptives (OC) on CL
P-G
in non-pregnant women). The sum reflects
the total paracetamol clearance, while the coefficients of variation can be retrieved in Table 2
Allegaert et al. BMC Anesthesiology (2015) 15:163 Page 8 of 11
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Discussion
The current study explored the variability in the differ-
ent metabolic and elimination clearance estimates in
young women following iv paracetamol administration.
To allow for an analysis of the different metabolic path-
ways, we applied an earlier described model, based on
simultaneous collection of plasma and urine [16]. Using
this approach, we clearly confirmed the significantly
higher (Factor = 2.03, 15.8 l.h
−1
) clearance to paracetamol
glucuronide at delivery and significantly lower (Factor =
0.55, 4.66 l.h
−1
) clearance in early postpartum when com-
pared to healthy female volunteers (7.33 l.h
−1
) [11, 16]. In
addition, the use of oral contraceptives (Factor = 1.46) –
obviously limited to non-pregnant women - was also
found to affect clearance to paracetamol glucuronide. Be-
sides these major effects on paracetamol metabolic clear-
ance, there was a minor impact of preterm (Factor =1.34),
but not for term delivery on clearance to paracetamol
sulphate and of the urine flow on elimination of un-
changed paracetamol in urine (Fig. 4). Finally, these clin-
ical covariates performed better as predictors of altered
paracetamol glucuronidation clearance when compared to
oestradiol or progesterone levels.
Both the impact of pregnancy and oral contraceptives
on intravenous paracetamol clearance have been reported
earlier in literature (Table 3) [9, 10, 14, 16, 18, 19]. In the
current pooled analysis, we clearly linked this raised
clearance with a raised paracetamol glucuronidation activ-
ity and initially hypothesized that this was associated with
oestradiol as biomarker. This hypothesis was based on the
fact that endogenous estrogens are both a substrate as well
as an inducer of glucuronidation enzymes [12, 13, 20, 21],
and similar observations have been described for ethiny-
loestradiol [22]. However, the use of oestradiol as bio-
marker in itself was not superior to the use of the more
readily available clinical characteristics (pregnancy, post-
partum, exposure to oral contraceptives) in our model.
Besides the impact of pregnancy and oral contracep-
tives on paracetamol glucuronidation, the clearance of
unchanged paracetamol was dependent on the urine
output. This confirms earlier observations of Miners et
al., who quantified the effects of high and low urine flow
rates on the urinary metabolic ratios for paracetamol
glucuronidation, sulphation and oxidation at steady-state
in seven (four female, three male) healthy young adults
[7]. Metabolic partial clearances were unaffected by
urine flow rate, but individual paracetamol metabolic ra-
tios varied 2.5- to 3.2-fold over a 7.4-fold range of urine
flow rates (48–360 ml.h
−1
).
Beyond changes in paracetamol disposition, we
hypothesize that this pattern of raised phenotypic
glucuronidation driven by pregnancy or oral contracep-
tives is of relevance to explain and predict within and be-
tween individual variability in disposition of drugs that
mainly undergo UDP-glucuronosyltransferase (UGT)1A6,
1A1, 1A9 or 2B15 driven glucuronidation. Consequently,
we anticipate a similar pattern for other drugs that
undergo glucuronidation, including lamotrigine (UGT1A4,
plasma concentrations increase in postpartum, range + 75–
351 %, reflecting decreased clearance), propofol (UGT1A9,
Table 3 Overview of the pharmacokinetics of intravenous (iv) paracetamol in cohorts of women as retrieved in literature
Author Study characteristics Clearance (l/h) Distribution volume (l/kg)
Ochs et al. [18] single iv, 650 mg, young women, age matched study design
Controls (n= 10), 21–30 year, 54 (SE 2.1) kg 16.8 (SE 0.6) 0.98 (SE 0.08)
Oral contraceptives (n= 10), 62 (SE 2.5) kg 22.7 (SE 2.3) 0.98 (SE 0.06)
Sonne et al. [19] single iv, 1 000 mg, 2 episodes in each individual 16.6 1.01
3 women 54–56 kg, 29–33 years
Scaveno et al. [14] single iv, 650 mg, 30 post-menopausal women
controls (n= 18): 45 (SE 3.9) years, 64.9 (SE 3.3) kg 16.6 (SE 0.69) 0.85 (SE 0.04)
conjugated oestrogens (n= 12): 46 (3.4) years, 60.2 (1.7) kg 16.6 (SE 0.25) 0.82 (SE 0.05)
Abernethy et al. [9] single iv, 650 mg, 16 women
controls (n= 8): 23–32 years, 48–66 kg 13.7 (SD 1.26) 0.96 (SD 0.08)
oral contraceptives (n= 8): 21–36 years, 48–77 kg 20.0 (SD 1.68) 1.04 (SD 0.08)
Wynne et al.[10] single iv, 500 mg, 42 female/5 male volunteers, .all results pooled
healthy, young: 25 (SE 1) years, 59 (SE 2) kg 16.6 (SE 0.71) 1.00 (SE 0.04)
healthy, elderly: 73 (SE 1) years, 66 (SE 2) kg 14.6 (SE 0.79) 1.07 (SE 0.03)
frail, elderly: 82 (SE 2) years, 53 (SE 4) kg 7.9 (SE 0.32) 0.81 (SE 0.03)
Kulo et al. [16] single iv, 2 000 mg
28 cases, at delivery 31.5 (20–42) years, 79 (57–110) kg 20.3 (11.8–62.8) 0.72 (0.52–1.56)
SE standard error, SD standard deviation
Allegaert et al. BMC Anesthesiology (2015) 15:163 Page 9 of 11
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

clearance 35 % higher during pregnancy) or benzodiaze-
pines (UGT2B7/15, clearance 75 % higher during preg-
nancy) [4, 12, 20–26]. Similar to the development and
validation of model-based approaches in the field of mat-
uration based on system specific information [27, 28], the
quantitative functions described can be used to quantify
the impact of pregnancy or oral contraceptives on pheno-
typic UGT1A1 or UGT1A6 glucuronidation.
Conclusions
Variability in paracetamol glucuronidation elimination in
young women was in part explained by pregnancy, early
postpartum or exposure to oral contraceptives. Oestradiol
or progesterone plasma levels also explained increased
paracetamol glucuronidation elimination. However, imple-
mentation of oestradiol or progesterone in addition to
being at delivery did not further improve the model. We
hypothesize that the pattern of raised phenotypic glucuro-
nidation and its variability in young women is of relevance
to predict within and between individual variability in
disposition of any drug that is subject to glucuronidation.
Abbreviations
BW: Body weight; CL: Clearance; CYP: Cytochrome p450; CV: Coefficient of
variation; ELISA: Enzyme-linked immunosorbent assay; FDA: Food and drug
agency; GA: Gestational age; GCP: Good clinical practice; HPLC: High
performance liquid chromatography; iv: Intravenous; k: Elimination rate;
MF: Multiplication factor; NAPQI: N-acetyl-p-benzoquinone-imine; OC: Oral
contraceptives; PD: Pharmacodynamics; P-G: Paracetamol glucuronide;
PK: Pharmacokinetics; P-O: Paracetamol, oxidative metabolites; P-
S: Paracetamol sulphate; P-U: Unchanged paracetamol;
Q: Intercompartmental clearance; V: Distribution volume.
Competing interests
Besides the funding from agencies and academic research organizations
mentioned below, the authors declare that they have no other competing
interests.
Authors’contributions
KA was the principal investigator of the studies on patients and pooled the
available data and built the dataset. MYP performed the population PK
analysis, supported and verified by CK. All other authors contributed to the
study design (BB, AS, AK, KvC, JdP, JdH), recruitment of patients and sample
collection (AS, AK, BB) or bio-analysis (BB, AK, KvC, JdH). All authors were
involved in interpretation of the data, the drafting the manuscript and the
subsequent revisions. All authors have read and approved the final
manuscript.
Acknowledgements
Karel Allegaert is supported by the Fund for Scientific Research, Flanders
(Fundamental Clinical Investigatorship 1800214 N), Aida Kulo by a Join EU-
SEE scholarship. K van Calsteren is supported by a KOOR clinical research
grant of the University Hospitals, Leuven. The clinical research was in part
supported by an unrestricted academic clinical research grant provided by
the Belgian Society for Anesthesia and Resuscitation.
Author details
1
NICU, University Hospitals Leuven, Herestraat 49, 3000 Leuven, Belgium.
2
Department of Development and Regeneration, Cluster Organ Systems, KU
Leuven, Leuven, Belgium.
3
Department of Clinical Pharmacy, St Antonius
hospital, Nieuwegein, The Netherlands.
4
Department of Pharmaceutical and
Pharmacological Sciences, KU Leuven, Leuven, Belgium.
5
Center for Clinical
Pharmacology, University Hospitals Leuven, Leuven, Belgium.
6
Institute of
Pharmacology, Clinical Pharmacology and Toxicology, Faculty of Medicine,
University of Sarajevo, Sarajevo, Bosnia Herzegovina.
7
Obstetrics and
Gynecology, University Hospitals Leuven, Leuven, Belgium.
8
Leiden Academic
Centre for Drug Research, Leiden University, Leiden, The Netherlands.
Received: 26 April 2015 Accepted: 11 November 2015
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