Kinetic disposition of lorazepam with focus on the glucuronidation capacity, transplacental transfer in parturients and racemization in biological samples

Abstract

The present study investigates the kinetic disposition with focus on the racemization, glucuronidation capacity and the transplacental transfer of lorazepam in term parturients during labor. The study was conducted on 10 healthy parturients aged 18–37 years with a gestational age of 36–40.1 weeks, treated with a single oral dose of 2 mg racemic lorazepam 2–9 h before delivery. Maternal venous blood and urine samples were obtained over a 0–48 h interval and the umbilical cord sample was obtained immediately after clamping. Lorazepam enantiomers were determined in plasma and urine samples by LC–MS/MS using a Chiralcel^® OD-R column. In vitro racemization of lorazepam required the calculation of the pharmacokinetic parameters as isomeric mixtures. The data were fitted to two-compartment model and the pharmacokinetic parameters are reported as means (95% CI): t_1/2a 3.2 h (2.6–3.7 h), K_a 0.23 h⁻¹ (0.19–0.28 h⁻¹), t_1/2 10.4 h (9.4–11.3 h), β 0.068 h⁻¹ (0.061–0.075 h⁻¹), AUC^0–∞ 175.3 (ng h)/ml (145.7–204.8 (ng h)/ml), Cl/F 2.6 ml/(min kg) (2.3–2.9 ml/(min kg)), Vd/F178.8 l (146.5–211.1 l), Fel 0.3% (0.1–0.5%), and Cl_R 0.010 ml/(min kg) (0.005–0.015 ml/(min kg)). Placental transfer of lorazepam evaluated as the ratio of vein umbilical/maternal vein plasma concentrations, obtained as an isomeric mixture, was 0.73 (0.52–0.94). Pregnancy changes the pharmacokinetics of lorazepam, with an increase in the apparent distribution volume, an increase in apparent oral clearance, and a reduction of elimination half-life. The increase in oral clearance may indicate an increase in glucuronidation capacity, with a possible reduction in the plasma concentrations of drugs depending on glucuronidation capacity as the major metabolic pathway.

Introduction

Lorazepam, a potent benzodiazepine, has been shown to have marked anxiolytic and sedative properties. Lorazepam can be administered as a premedicant for elective caesarean section or to women during labor [1], [2].

Lorazepam is marketed as a racemate of (+)-(S) and (−)-(R) enantiomers. The pharmacologically active (+)-(S) enantiomer has a 100- to 200-fold higher apparent affinity to the binding site of the receptor than (−)-(R)-lorazepam [3].

The pharmacokinetics of oral lorazepam has been well investigated in healthy male and female volunteers. Peak plasma concentrations of orally administered lorazepam are achieved within 2.5 h and mean values of absorption half-life generally are less than 30 min. The bioavailability of the 2 mg oral dose averages 89–93%. Lorazepam is less extensively bound to protein than are other benzodiazepines; its mean unbound fraction is 6.8%. The apparent volume of distribution of lorazepam (1.3–2.1 l/kg) indicates moderately extensive tissue uptake [4], [5], [6], [7], [8], [9], [10].

Lorazepam is mainly metabolized by glucuronidation (Fig. 1). At least 24 different UDP-glucuronosyltransferase (UGT) human genes have been identified and are classified in two families (UGT1 and UGT2) and three subfamilies (UTT1A, UGT2A and UGT2B) based on sequence homology. Lorazepam is not a substrate of UGT1A1, but appears to be a non-competitive inhibitor that may have a negative influence in patients with Gilberts's syndrome who already have an impaired glucuronidation of bilirubin [11]. It is not clear whether UGT2B7 metabolizes lorazepam and oxazepam although Patel et al. [12] suggested that 10% of Caucasians are poor glucuronidators of S-oxazepam. About 75% of an oral dose of lorazepam is excreted into human urine as lorazepam-glucuronide and 13.5% as oxidized metabolites and their glucuronides. Less than 1% is eliminated unchanged in urine [10], [13]. Chaudhary et al. [14] reported a diurnal variation in lorazepam elimination consistent with a fast-induced increase in hepatic glucuronidation during the night. Oral lorazepam clearance in male or female healthy volunteers ranges from 5 to 7 l/h [7], [9], [15] and elimination half-life from 9 to 16 h [4], [5], [6], [7], [8], [9].

UGT-catalyzed glucuronidation reactions are responsible for ∼35% of all drugs metabolized by phase II enzymes. Many of the individual UGT enzymes are expressed not only in liver but also in extrahepatic tissues, where the extension of glucuronidation can be substantial [16]. UGTs have not received substantial attention in the pharmacogenetic literature due to their overlapping activity and lack of selective probes [11]. Crom et al. [17] and Kearns et al. [18] assessed conjugation metabolism in healthy male volunteers or patients with cystic fibrosis using lorazepam administered by intravenous route. Herman et al. [10] reported that lorazepam cannot properly be used as a marker of conjugative metabolism due to the fact that it undergoes significant enterohepatic recirculation in humans. Herman et al. [19] associated neomycin and cholestyramine in an attempt to block the enterohepatic circulation of lorazepam and to permit an in vivo estimate of hepatic glucuronidation.

Pregnancy causes various physiologic changes that can lead to important variations in the pharmacokinetic process of absorption, distribution and elimination of drugs. Low concentrations of many drugs during pregnancy are consistent with increased hepatic blood flow, as well as increased volume of distribution and decreased binding to plasma proteins. Pregnant women are subject to an increased level of estrogens and progesterones resulting in changes in hepatic drug metabolism [20]. Previous studies in humans have demonstrated that pregnancy increases CYP2D6 and decreases CYP1A2 and N-acetyltransferase activities [21], [22]. Luquita et al. [23] reported a decrease in liver UGTs (UDP-glucuronosyltransferases) activity in pregnant rats affecting family 1 isoforms and UGT2B1. In postpartum animals, protein level recovered (UGT1A5 and UGT2B1) or even increased (UGT1A1 and UGT1A6) with respect to control rats.

Lorazepam crosses the placenta and spreads through the tissues of the fetus. Mc Bride et al. [1] reported that fetal concentrations rarely exceeded that in the mother following administration of 2.5 mg i.v. Kanto et al. [2] found that the levels of lorazepam were equal in the maternal and umbilical circulation 0.8–7.7 h after administration of 2 mg i.m. or 11–15.5 h after administration of a 2.5 mg oral dose. The authors [2] reported a serum protein unbound fraction of 14.0 ± 4.8% in the maternal circulation and 20.8 ± 3.1% in the umbilical circulation. The metabolism of lorazepam in neonates differs considerably from that occurring in children and adults as a function of the low capacity of neonates to conjugate lorazepam with UDP-glucuronic acid. Cappielo et al. [24] reported that uridine 5′-diphosphoglucuronic acid, the endogenous substrate of UGTs, is present in the human fetal liver at a concentration five-fold lower than in the adult liver, indicating a potential limiting factor for glucuronidation in the human fetus.

Kanazawa et al. [25] reported the stereoselective analysis of lorazepam in plasma samples collected from a patient who was being treated with racemic lorazepam and found only concentrations of (+)-(S)-lorazepam. These data were not confirmed by independent studies. Actually Pham-Huy et al. [3] reported the spontaneous racemization of lorazepam in polar medium, and thus the impossibility of a quantitative determination of the single enantiomers in plasma samples of rabbits.

Since lorazepam may be considered to be a marker of glucuronidation capacity and in view of the absence of clinical data about stereoselectivity and influence of pregnancy in the pharmacokinetics of lorazepam, the objective of the present study was to assess the pharmacokinetics of lorazepam and its conjugate as an isomeric mixture in parturients treated with a single dose of racemic lorazepam. LC–MS/MS analysis was performed on a chiral stationary phase in order to prove the in vitro racemization of lorazepam.