1Département de Pharmacie Clinique, de Pharmacocinétique et dEvaluation du Médicament, Institut des Sciences Pharmaceutiques et Biologiques de Lyon, 8 avenue Rockefeller, F-69373 Lyon Cedex 08, France. 2Service dAnesthésie et réanimation, Centre Hospitalier Universitaire Lyon-Sud, 165 chemin du grand Revoyet, F-69495 Pierre Bénite Cedex, France. 3Service Pharmaceutique, Hôpital Neuro-cardiologique, 59 Bd Pinel, F-69393 Lyon, France. 4Fédération de Biochimie, Laboratoire C, Hôpital E. Herriot, 3 place dArsonval, F-69437 Lyon Cedex 03, France. 5Laboratoire de Physiologie Rénale et Métabolique, INSERM U499, Faculté de Médecine Laennec, 8 rue G. Paradin, F-69008 Lyon, France*Corresponding author: Fédération de Biochimie, Laboratoire C, Hôpital E. Herriot, 3 place dArsonval, F-69437 Lyon Cedex 03, France
Accepted for publication: December 3, 2001
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Abstract |
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Methods. The extent of phase I and phase II metabolism of propofol was studied in 18 female and 17 male patients after an anaesthesia induced and maintained for at least 4 h with propofol. The infusion rates (mg kg1 h1) of propofol were (mean (SD)) 4.1 (1.0) and 4.5 (1.3) for males and females, respectively. Urine was collected from each patient for the periods 04, 48, 812, and 1224 h after the start of propofol administration. In a preliminary study, the three main glucuro-conjugated metabolites were isolated from urine and characterized by magnetic resonance spectroscopy. The quantification of these metabolites for the different collection periods was then performed by a HPLCUV assay.
Results. Total recovery of propofol in the metabolites studied amounts to 38%, of which 62% was via the PG metabolite and 38% via cytochrome P-450. This percentage is significantly higher than that previously reported from patients after a bolus dose of propofol. Extreme values for PG (024 h period) were included from 73 to 49%. There was no significant difference between female and male patients in the metabolite ratio.
Conclusions. We conclude that the extent of hydroxylation in propofol metabolism was higher than in previous findings after administration of anaesthetic doses of propofol. Moreover, the ratio between hydroxylation and glucuronidation of propofol is subject to an inter-patient variability but this does not correlate with the dose of propofol. However, the variation of the metabolite profile observed in the present report does not seem to indicate an extended role of metabolism in pharmacokinetic variability.
Br J Anaesth 2002; 88: 6538
Keywords: anaesthetics i.v., propofol; pharmokinetics, propofol
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Introduction |
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The aim of this study was to assess, first, the metabolite profile of propofol after an i.v. propofol anaesthesia which was maintained for at least 4 h in a larger Caucasian population, and secondly to compare female and male patient profiles.
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Patients and methods |
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All urine was collected from each patient for the periods 04, 48, 812, and 1224 h after the start of propofol administration. The volume of urine for each period was measured and after thorough mixing, a 40-ml aliquot was added to polypropylene haemolysis tubes and stored at 25°C before analysis.
Drugs and chemicals
Acetonitrile (HPLC grade) was purchased from MerckTM (Darmstadt, Germany). Ethyl acetate, 1-naphthyl-sulphate (1-NS) and acetic acid were obtained from SigmaTM (St Quentin Fallavier, France).
Analytical procedure
1-QG, 4-QG, and PG were extracted from human urine samples included in this study by liquid/liquid extraction using ethyl acetate. The mixture was mixed for 2 h, then the organic layer was removed and evaporated to dryness under nitrogen at 40°C. The residue was re-dissolved in water acidified with acetic acid and several fractions were injected onto the preparative chromatograph apparatus. Preparative chromatography was performed using a stainless steel column (250x20 mm, dp; 1025 µm, 120 Å) packed with ShiseidoTM Capcell Pak C18 from InterchimTM (Montluçon, France). The metabolites were separated with a linear gradient using wateracetic acid (pH 3.8) and acetonitrile as solvents. Aliquots corresponding to each metabolite were freeze-dried to obtain a lyophilized powder. Characterization of the glucuro-conjugate metabolite was performed using 1H-nuclear magnetic resonance spectroscopy (NMR) in a Spectrospin 500 MHz NMR spectrometer from BrukerTM (Wissembourg, France).
Chromatographic conditions for the quantification of glucuro-conjugated metabolites were described previously.10 Urine samples were thawed, mixed, and centrifuged at 4000g (20 min, 4°C). The sample was then diluted (1:5, v/v) with phosphate buffer (25 mM, pH 3.8), and 20 µl of 1-NS, used as internal standard, was added to a final concentration of 40 µM. This mixture was then vortexed and 20 µl was injected onto the HPLC column. Stock solutions of 1-QG, 4-QG, and PG were prepared in phosphate buffer (25 mM, pH 3.8), then serially diluted to concentrations of 10, 25, 50, 100 and 250 µg ml1 in blank urine. These concentrations were used to assess the accuracy and linearity of the method on 3 different days for the three metabolites.
Statistical analysis
Patient characteristics were compared using a MannWhitney U-test. The differences in metabolite profile between males and females or between the four collection periods for the same metabolite, were evaluated using analysis variance followed by the Scheffe post-hoc test. Comparison of metabolite profiles between the study of Sneyd and colleagues and the present study were by MannWhitney U-test.4 For analytical validation, the significance of the slope and the validity of the linear calibration curves were confirmed using FisherSnedecors F-test. For each metabolite, the homocedasticity for the calibration curve was tested using Cochrans test. In all cases P<0.05 was taken as the minimum level of statistical significance.
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Results |
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Metabolite profile
The proportion of the main urinary conjugated metabolites for the 024 h period is shown in Table 2. There was no significant difference between female and male patients for 1-QG, 4-QS, and PG. For 4-QG a small but significant difference was observed (P=0.04). However, this small difference can be considered, from a metabolic and clinical viewpoint, as negligible. PG is the main metabolite and accounts for approximately 62% of the total metabolite profile for the 024 h period. Extreme values for PG (024 h period) of 7073% for two females and one male and 4952% in one male were recorded. The production of 1-QG was greater than 4-QG, and this difference was statistically significant.
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Finally, the percentage of the propofol dose eliminated in the urine (over 024 h) from female and male patients was 39.8 (2.9) and 36.1 (2.9)%, respectively, equivalent to 38.0 (2.0)% in all patients (mean (SEM)). Unquantified amounts of unchanged propofol were detected in urine samples.
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Discussion |
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Our anaesthetic procedure included the administration of remifentanil. This new opioid is metabolized by several non-specific esterases in blood and in tissues, conferring a metabolism independent of liver function.14 In the present study, no correlation was found between the proportion of any metabolite and the total dose of remifentanil. In addition, acetaminophen and ketoprofen, two drugs administered in the postoperative period, undergo glucuronidation as their main metabolic pathways. UGT1A6 and UGT2B7 are the most effective metabolizers of acetaminophen and ketoprofen via glucuronidation, respectively.15 Although one cannot rule out, definitively, on this basis a metabolic interaction between propofol and these drugs, a modification of the metabolite ratio of propofol was unlikely under these conditions. From a methodological point of view, the quantification of 4-QS via the 4-QG equation has introduced an error in the amount of 4-QS. However, the UV spectra of both metabolites were very close, particularly in the region of 220 nm.10 Moreover, the recovery of 4-QS in urine was found to be relatively low, so a small deviation from the actual value would not modify markedly the relative proportions of the metabolites.
Differences between the present data, obtained from a larger population (n=35), and that obtained from Sneyds study in female patients could be explained by the limited size of their population (n=6), which may not have been representative, rather than by the different quantification methods that were used. Furthermore, the pharmacokinetics of propofol are well fitted by a triexponential model, with a short first elimination half-life assessed to 3040 min after an infusion from 3 to 9 mg kg1 h1.2 16 Thus, our anaesthetic procedure favours metabolic pathways present during steady-state conditions (mean 320 min for the duration of propofol infusion) and they do not correspond to the single dose data. Total recovery of the administered dose, during the 024 h period, for all patients was lower (38.0 (2.0)%) than that observed in female Caucasian patients (50.9 (4.0)%).4 Several recent studies reported that remifentanil could decrease the clearance of propofol as a result of decreased hepatic perfusion.17 18 This point is consistent with the fact that propofol has a high hepatic extraction (over 0.7) and thus, hepatic blood flow is a major determinant of its hepatic elimination.19 It has been established that modifications of drug disposition occur during surgery and anaesthesia, and the period of anaesthesia was markedly longer in our study (165540 min) than in Sneyd and colleagues study (50150 min).2 4 20 Consequently, this consideration explains the lower percentage of recovery of the propofol dose in the present study.
In the present study, the percentage of recovered conjugate was comparable with female and male patients in agreement with a previous study concluding that there were no gender differences in the pharmacokinetics of propofol in humans.21 None of the patients who smoked exhibited a different metabolite profile although an influence on acetaminophen glucuronidation was reported in male heavy smokers.22 However, the number of smoker patients in our study (n=4) was too small to draw any conclusions. Similarly, six elderly patients (over 70 yr old) were included. Their metabolite profile was in the same range as younger patients although a lower total body clearance has been described in the elderly.23
In summary, the present study has documented that the extent of hydroxylation in propofol metabolism was higher than reported previously after administration of an anaesthetic dose of propofol. For the whole study group, the ratio between hydroxylation and glucuronidation is subject to inter-patient variability. The relative abundance of CYP2B6, CYP2C9, and UGT1A8/9 may perhaps explain this inter-individual variation. However, first, the metabolite profile does not correlate with propofol dose and secondly, no unusual pattern was observed in a particular patient.4 However, inter-patient variability was not great compared with other anaesthetics such as alfentanil which exhibits an important inter-individual variability in hepatic metabolism with clinical consequences.24 25
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Acknowledgements |
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