CIRCADIAN TIME-EFFECT OF ORALLY ADMINISTERED

LORATADINE ON PLASMA PHARMACOKINETICS IN MICE

Dorra Dridi,1,2 Mossadok Ben-Attia,3 Mamane Sani,3 Nassim Djebli,2

Francois Ludovic Sauvage,4 and Naceur A. Boughattas1

1Laboratoire de Pharmacologie, Faculte de Medecine, Monastir Tunisie2Laboratoire de Pharmacologie, Faculte de Medecine, Limoges, France3Laboratoire de Biosurveillance de l’Environnement, Faculte des Sciences de Bizerte,Zarzouna, Tunisie4Service de Pharmacologie–Toxicologie, CHU Dupuytren, Limoges, France

Little is known about the chronopharmacokinetics of loratadine, a long-acting tricyclicantihistamine H1 widely used in the treatment of allergic diseases. Hence, the pharma-cokinetics of loratadine and its major metabolite, desloratadine, were investigated aftera 20 mg/kg dose of loratadine had been orally administered to comparable groups ofmice (n ¼ 33), synchronized for three weeks to 12 h light (rest span)/12 h dark (activityspan). The drug was administered at three different circadian times (1, 9, and 17 hafter light onset [HALO]). Multiple blood samples were collected over 48 h, andplasma concentrations of loratadine and desloratadine were determined by high per-formance liquid chromatography. There were no significant differences in Tmax of lor-atadine and desloratadine between treatment-time different groups. However, theelimination half-life (t1/2) of the parent compound and its metabolite was significantlylonger (p , 0.01) following administration at 9 HALO (t1/2 loratadine and desloratadine5.62 and 4.08 h at 9 HALO vs. 4.29 and 2.6 h at 17 HALO vs. 3.26 and 3.27 at 1 HALO).There were relevant (p , 0.05) differences in Cmax between the three treated groupsfor loratadine and desloratadine; 133.05 + 3.55 and 258.07 + 14.45 ng/mL at 9HALO vs. 104.5 + 2.61 and 188.62 + 7.20 ng/mL at 1 HALO vs. 94.33 + 20 and187.75 + 10.79 ng/mL at 17 HALO. Drug dosing at 17 HALO resulted in highestloratadine and desloratadine total apparent clearance values: 61.46 and 15.97 L/h/kg, respectively, whereas loratadine and desloratadine clearances (CL) were signifi-cantly slower (p , 0.05) at the other administration times (loratadine and deslorata-dine CL was 57.3 and 14.22 L/h/kg at 1 HALO vs. 43.79 and 12.89 L/h/kg at 9HALO, respectively). The area under the concentration-time curve (AUC) ofloratadine and desloratadine was significantly (p , 0.05) greater following drugadministration at 9 HALO (456.75 and 1550.57 (ng/mL) . h, respectively); it was

Submitted June 4, 2007, Returned for revision July 30, 2007, Accepted January 21, 2008The first two authors contributed equally to this work.Address correspondence to Naceur A. Boughattas, Laboratoire de Pharmacologie, Faculte de

Medecine, 5019 Monastir, Tunisie. Tel.: þ (216)73462200; Fax: þ(216)73460737; E-mail:naboughattas@yahoo.fr

Chronobiology International, 25(4): 533–547, (2008)Copyright # Informa HealthcareISSN 0742-0528 print/1525-6073 onlineDOI: 10.1080/07420520802257646

533

lowest following treatment at 17 HALO (325.39 and 1252.53 (ng/mL) . h,respectively). These pharmacokinetic data indicate that the administration time ofloratadine significantly affected its pharmacokinetics: the elimination of loratadineand its major metabolite desloratadine. (Author correspondence: naboughattas@yahoo.fr)

Keywords Loratadine, Desloratadine, Chronopharmacokinetics, Circadian rhythm,Mice

INTRODUCTION

Loratadine, chemically known as ethyl 4-(8-chloro-5,6-dihydro-11H-benzo[5,6]cyclohepta[1,2-b]pyridin-11-ylidene)-1-piperidinecarboxylic acidethyl ester, is a long-acting tricyclic antihistamine with selective peripheralhistamine H1-receptor antagonistic activity and fewer sedative effects thanconventional antihistamines (Barret, 1998; Kay & Harris, 1999).

Loratadine is indicated for the relief of nasal and non-nasal symptomsof seasonal allergic rhinitis. After oral administration, loratadine israpidly absorbed and undergoes substantial first-pass metabolism(Hilbert et al., 1987; Noehr-Jensen et al., 2006). It is metabolized todescarboethoxyloratadine (desloratadine), an active metabolite, mainlyby cytochrome P450 3A4 (CYP3A4) and by CYP2D6 in the liver(Yumibe et al., 1996).

The mammalian circadian pacemaker resides in the paired suprachias-matic nuclei (SCN) and influences all biological processes. The behaviorand safety of many medications vary depending on dosing time relativeto 24 h rhythms of biochemical, physiological, and behavioral processesthat are under the direct control of the circadian clock (Burioka et al.,2005b; Holzberg & Albrecht, 2003; Ohdo, 2007; Takata et al., 2005).This circadian organization of living organisms is a key component of thepharmacokinetic variation (chronopharmacokinetics) reported in animalsand humans for many drugs (Bruguerolle et al., 2008; Labrecque &Beauchamp, 2003; Lemmer, 2005, 2006b; Milano & Chamorey, 2002;Reinberg, 1992; Reinberg & Smolensky, 1982; Reinberg et al., 1984;Widerhon et al., 2005).

Administration-time variation in therapeutic effects may be caused bycircadian rhythm-dependent differences in drug:

. resorption (Bruguerolle & Lemmer, 1993; Labrecque & Belanger,1991; Min et al., 1996);

. distribution (e.g., variation in membrane permeability; see Boulameryet al., 2007; Bruguerolle & Prat, 1989; Labrecque & Belanger, 1991);

. metabolism (e.g., variation of metabolizing enzyme activities or/andblow flow variation in affected organs); and/or

D. Dridi et al.534

. elimination (e.g., variation in renal blood flow, glomerular filtration,and pH; see Labrecque & Beauchamp, 2003; Labrecque & Belanger,1991).

Administration-time variation in therapeutic effects may also be dueto circadian differences in pharmacodynamic phenomena (e.g., alteredreceptor sensitivity or/and alteration in levels of agonists; see Buriokaet al., 2007; Han & Lee, 1998; Yao et al., 2006). Based on chronobiologi-cal studies, chronotherapy (time-dependent drug dosing) is especiallyrelevant when the symptoms of the disease vary predictably over time(Cui et al., 2005; Smolensky & Peppas, 2007; Yegnanarayan et al.,2006). Based on experimental chronobiology and chronopharmacologyinvestigations, chronotherapeutic interventions have been applied inthe treatment of a number of diseases, such as cancer (Boughattaset al., 1988, 1989, 1994, 2002), asthma (Burioka & Sasaki, 1996;Burioka et al., 2005a; Cloutier et al., 1993; Martin, 1993; Smolenskyet al., 2007b), cardiovascular disorders (Haus, 2007b; Hermida, 2007;Hermida et al., 2007; Lemmer, 1991, 2005, 2006a; Portaluppi &Lemmer, 2007; Smolensky & Peppas, 2007; Smolensky & Portaluppi,1999; Smolensky et al., 2007a; White & LaRocca, 2002), and endocrineand metabolic disorders (Haus, 2007a).

Although circadian changes in the pharmacokinetics of many drugs arewell-documented, little is known about the chronopharmacokinetics of lor-atadine in the mouse model. The objective of this study was to investigatethe circadian time-dependent pharmacokinetic changes of loratadine andits active metabolite after an oral administration of loratadine in a 20 mg/kg dose at three different circadian times (i.e., at 1, 9, and 17 h after lightonset [HALO]) using mice as an animal model.

MATERIALS AND METHODS

Animals and Synchronization

All of the animal experiments were performed according to theguidelines of care and use of laboratory animals and in accordancewith the standards of the journal (Touitou et al., 2006). A total of 99male Swiss albino mice aged 10 weeks (weight � 30 g) were purchasedfrom SIPHAT, Tunisia. All mice were synchronized for 15 days in aclean room especially designed for chronobiological studies (Reinberg &Smolensky, 1983) with an alternating 12 h light (L)/12 h dark (D) cycle(light: 07:00 h–19:00 h and dark: 19:00 h–07:00 h). The room tempera-ture was maintained at 228C and a relative humidity of 50%. The micewere housed three or five per cage with food and water ad libitum.

Chronopharmacokinetic Study of Loratadine in Mice 535

The animals were randomly assigned to one of three groups of 33 mice inorder to explore three circadian stages (1, 9, and 17 HALO). Thesynchronization of mice was checked by assessing the circadian rhythmin rectal temperature. Rectal temperature was measured with a digitalthermometer (YSI model 49 TA).

Chemicals

Loratadine and desloratadine were kindly provided by Ibn El BaytarLaboratories (Tunis, Tunisia). Acetone (Chromanorm for HPLC), propa-nol-2 (Normapur), ammonia (Normapur), and ammonium acetate werepurchased from Prolabo (Paris, France). Formic acid (99% pure) andmethyl clonazepam were purchased from Sigma, and acetonitrile wasobtained from Carlo Erba. Drug-free serum was taken from untreatedmice that were provided by SIPHAT (Tunisia).

Study Design

Loratadine was freshly prepared by adding sterile distilled water toachieve a 2 mg/mL concentration. A single dose of 20 mg/kg loratadinewas administered orally to each mouse in a fixed fluid volume (10 mL/kg b.w.) and at one of three circadian times (1, 9, and 17 HALO). Bloodsamples were withdrawn from the orbital sinus predose (0 h) and at0.16, 0.33, 0.5, 1, 2, 4, 6, 12, 24, and 48 h after oral administration of lor-atadine. All blood samples were collected in lithium heparin tubes andwere centrifuged immediately at 3,000 rpm for 15 min. Plasma sampleswere stored in a 2808C freezer until analysis.

Preparation of Standards

Stock solutions of loratadine and desloratadine were prepared separ-ately at 1 g/L in methanol. The internal standard (methyl clonazepam)was provided as 100 ng/mL methanolic solutions.

Preparation of Calibration Curves

A pool of stock solutions (loratadine and desloratadine) at 10 mg/L inmethanol was prepared by mixing appropriate amounts of individual stocksolutions at 1 g/L. Calibration standards were prepared by mixing 100 mLof mouse plasma, 100 mL of acetonitrile, 100 mL internal standard, andstandard solutions at concentration ranging from 1 to 500 ng/mL lorata-dine and desloratadine. Calibrators and samples were centrifuged at13,000 rpm for 10 min. The supernatant was transferred to 200 mL vialsand then loaded in the autosampler.

D. Dridi et al.536

Plasma Loratadine and Desloratadine Determination

Instrumentation and Conditions

Loratadine and desloratadine determination was performed using theturbulent flow chromatography-tandem mass spectrometry system (TFC-MS/MS). The system consisted of a Cohesive 2300 system (Cohesive technol-ogies, Milton Keynes, Road Map, Buckinghamshire, UK) equipped with anautosampler kept at 68C, two binary high-pressure Agilent 1100 pumps,and three six-port switching valves controlled by the Aria OS softwarepackage. The system configuration, the parameters, and the analyticalprocess have been previously described in detail (Sauvage et al., 2006).Briefly, online extraction was performed at a high flow-rate (1.25 mL/min)on a Cyclonew 50-mm particle size (50 � 0.5 mm I.D.) column (Cohesive tech-nologies) in alkaline conditions (phase A1: 20 mM ammonium acetate inwater with 0.1% ammonia; phase B1: mixture of acetone/acetonitrile/propa-nol-2 [50:30:20; vol/vol/vol]). The chromatographic separation was per-formed in acidic conditions (phase A2: 0.1% formic acid in water; phase B2:0.1% formic acid in acetonitrile) using an Xterra MS C18, 5 mm(50 � 2.1 mm I.D.) column (Waters, Milford, Massachusetts, USA) kept at458C, with a constant flow rate of 400 mL/min.

Detection was performed with a TSQ Quantum Discovery MS/MSsystem (Thermo-Electron, Les Ulis, France) equipped with an orthogonalelectrospray ionization source and controlled by the Xcalibur computerprogram. MS/MS detection was performed in the positive ion, multiple-reaction, monitoring (MRM) mode following two transitions for loratadine(m/z 383 . 337 m/z; 383 . 267), desloratadine (m/z 311 . 259 m/z;311 . 243), and internal standard (IS) methylclonazepan (m/z 329 .

279 m/z; 329 . 221).

Validation

Linearity, interassay precision, and accuracy of the method werestudied by preparing and analyzing a set of calibrating standards at 1,2.5, 5, 10, 50, 100, 250, and 500 ng/mL. The lower limit of detection(LOD) and the lower limit of quantification (LOQ) of the technique were0.5 ng/mL and 1 ng/mL, respectively. Linearity was checked up to500 ng/mL (r ¼ 0.999). The within-day coefficients of variation (CV)were less than 15%, except at the LOQ (,20%). Accuracy was good, withmean relative errors ,15% over the linearity range.

Pharmacokinetic Analysis

Pharmacokinetic parameters of loratadine were estimated using thenon-compartmental method (linear pharmacokinetics), and data were

Chronopharmacokinetic Study of Loratadine in Mice 537

derived directly from the plasma concentration-time profiles (Gabrielsson &Weiner, 2000). Peak plasma concentrations (Cmax) and time of Cmax (Tmax)of loratadine and desloratadine were obtained directly from the observedconcentration-time data. The area under the plasma concentration-timecurve (AUC) was calculated by the trapezoidal rule. The terminal elimin-ation constant k was estimated by linear regression of the terminal portionof the concentration-time curve, and the elimination half-life (t1/2) was calcu-lated as 0.693/K. The apparent oral clearance (CL/F) was calculated asDose/AUC, and its metabolic ratio (MR) was calculated as the ratio ofAUCdesloratadine to AUCloratadine.

Statistical Analysis

Animal synchronization was verified by cosinor analysis (based on theleast-squares method) of the 24 h body temperature marker rhythm tovalidate circadian (with a trial period t ¼ 24 h) rhythm. A rhythm is vali-dated by the rejection of the hypothesis of which the amplitude is null(no rhythm [i.e., the rhythm’s amplitude ¼ 0]). Rhythm detection is con-sidered statistically significant with a p value of �0.05 (Nelson et al.,1979). The rhythm is then characterized by other parameters in additionto A: the mesor (M: the 24 h rhythm-adjusted mean) and acrophase (V: thepeak time of the cosine function, with light onset 0 HALO as V0 reference).A, M, and V are given with their 95% confidence limits when rhythmdetection is statistically significant. When the confidence limits of the circa-dian acrophase are greater than +2 h, both the V location and rhythmdetection are questionable (De Prins & Waldura, 1993), because thecurve pattern of the time series differs too greatly from a cosine function.

In this chronopharmacokinetic study, time-point data were computedas mean + standard deviation (SD) or mean and 95% confidence interval(CI 95%) from three independent determinations, and groups were com-pared by one-way ANOVA. p � 0.05 was considered to be significant.

RESULTS

Rectal Temperature

The rectal temperature rhythm was used as a marker of circadian syn-chronization of the animals. Analysis of the time series of rectal tempera-ture by the cosinor test revealed a significant (p , 0.001) circadianrhythm with a peak-time located (V) in the second half of the dark (activityspan), at 20.07 HALO + 1.20 h. The mean value (+SEM) of rectal temp-erature at the peak-time was 36.2 + 0.338C. The characteristics of the 24 hpattern in rectal temperature confirmed the physiological synchronizationof animals to the environmental 12L:12D schedule.

D. Dridi et al.538

Pharmacokinetics Irrespective of Circadian Dosing Time

The mean plasma concentration-time profiles of loratadine anddesloratadine irrespective of circadian dosing-time and the 24 h meanpharmacokinetic parameters are presented in Table 1.

After loratadine administration, the Cmax (110.66 + 8.72 ng/mL) wasquickly attained (Tmax ¼ 0.5 h). However, the main metabolite Cmax wastwo times higher (211.48 + 10.8 ng/mL) and appeared later(Tmax ¼ 2 h). The desloratadine AUC (1402.87 ng . h/mL) was almostfour times higher than that of loratadine (376.02 ng . h/mL). Conse-quently, loratadine clearance (54.35 L/h/kg) was four times higher thanthat of desloratadine (14.36 L/h/kg).

Drug Chronopharmacokinetics

Loratadine

The highest and lowest values of loratadine (see Figure 1) pharmaco-kinetic parameters (Cmax, AUC, and t1/2) occurred when the drug wasadministered at the end of light-rest (9 HALO) and the dark-activity (17HALO) or the beginning of the diurnal rest phase (1 HALO), respectively.There were significant differences between the different circadian dosingtimes in the Cmax (see Figure 1 and Table 1). Loratadine reached meanCmaxs of 104.5 + 2.61, 133.1 + 3.55, and 94.3 + 20 ng/mL at the sameTmax of 0.5 h in mice treated at 1, 9, and 17 HALO, respectively(p , 0.01 by ANOVA). Nevertheless, the post-hoc ANOVA Tukey testshowed no significant differences between 1 and 17 HALO in Cmax. Thevalues of AUC for loratadine were 345.9, 456.8, and 325.4 ng/mL.h at1, 9, and 17 HALO, respectively. The elimination half-life (t1/2) valueswere 3.26, 5.6, and 4.3 h, and the apparent clearance CL/F values were57.8, 43.8, and 61.4 L/h/kg for the 1, 9, and 17 HALO oral adminis-tration, respectively.

Desloratadine

Similar circadian variations were found in the pharmacokinetic par-ameters of the major metabolite of loratadine, desloratadine (seeFigure 2). In fact, there were significant differences in desloratadine Cmax

among the three treatment-time groups (see Figure 1 and Table 1). Thehighest and lowest values of desloratadine pharmacokinetic parametersoccurred when the drug was administered at the end of the light-rest (9HALO) and dark-activity (17 HALO) or beginning of diurnal rest times (1HALO), respectively. Desloratadine reached higher mean Cmaxs of188.6+ 7.20, 258.07+ 14.45, and 187.75+ 10.79 ng/mL at the same

Chronopharmacokinetic Study of Loratadine in Mice 539

TABLE 1 Oral Pharmacokinetic Parameters of Loratadine and Desloratadine at the Three Circadian Times of Drug Administration

Parameters

1 HALO 9 HALO 17 HALO 24 h Mean

Loratadine Desloratadine Loratadine Desloratadine Loratadine Desloratadine Loratadine Desloratadine

Tmax (h) 0.5 2 0.5 2 0.5 2 0.5 2

Cmax (ng/mL)a 104.5 + 2.61 188.62 + 7.20 133.05 + 3.55 258.07+ 14.45 94.33 +20 187.75 + 10.79 110.66+ 8.72 211.48 + 10.8

AUC (ng . h/mL)a 345.92 + 20.89 1405.53 + 135.83 456.75 + 47.78 1550.57 + 280.79 325.39 + 117.98 1252.53 + 263.88 376.02+ 70.66 1402.87 + 149.04

t1/2 (h)b 3.26 [2.14–4.38] 3.27 [2.43–4.11] 5.61 [3.67–7.55] 4.08 [2.79–5.37] 4.29 [2.87–5.71] 2.60 [1.98–3.22] 4.38 [1.47–7.29] 3.31 [1.48–5.16]

CL/F (L/h/kg)a 57.81 + 3.49 14.22 +1.37 43.79 + 4.58 12.89 +2.33 61.46 + 22.29 15.97 +3.36 54.35+ 9.33 14.36 +1.54

aValues are mean+ SD of three replications.bValues are mean and 95% confidence interval (CI 95%).Abbreviation: HALO ¼ hours after light onset.

54

0

Tmax of 2 h in mice treated at 1, 9, and 17 HALO, respectively (p , 0.01 byANOVA). However, the Tukey post hoc test revealed no significant differ-ences in Cmax between 1 and 17 HALO. The AUC values for desloratadinewere three to four times higher than those for loratadine (1405.5, 1550.6,and 1252.5 ng/mL . h at 1, 9, and 17 HALO, respectively). The CL/Fvalues for desloratadine were 14.2, 12.9, and 15.9 L/h/kg (i.e., three tofour times less than loratadine), and the elimination half-life (t1/2) valueswere 3.27, 4.1, and 2.6 h for the 1, 9, and 17 HALO treatment, respectively.

FIGURE 2 Plasma concentration of desloratadine after the oral administration of loratadine (20 mg/kg) at 1, 9, and 17 HALO in mice (n ¼ 33 per circadian stage). Each value is the mean + SD (bars) oftriplicate assays.

FIGURE 1 Plasma concentration of loratadine following oral drug (20 mg/kg) administration at 1, 9, and17 HALO in mice (n ¼ 33 per circadian stage). Each value is the mean+ SD (bars) of triplicate assays.

Chronopharmacokinetic Study of Loratadine in Mice 541

DISCUSSION

To the best of our knowledge, this is the first report on circadiantime-dependent pharmacokinetics of loratadine and its major metabolite,desloratadine, in mice. The pharmacokinetics of loratadine and deslorata-dine in plasma were previously explored in non-chronobiology investi-gations, both in rats and mice. Gender-related differences in thepharmacokinetics of loratadine and desloratadine were observed in ratsbut not in mice. Higher concentrations of loratadine were observed inmale rats, while exposure to desloratadine was higher in female rats(Ramanathan et al., 2005). Other studies focused on the effect of drugassociation on loratadine pharmacokinetics, showing the concomitantadministration of loratadine and ketoconazole, cimetidine (Kosoglouet al., 2000), or erythromycin (Brannan et al., 1995) significantly increasedloratadine plasma concentrations compared with the administration of lor-atadine alone.

Chronobiological studies require animal synchronization. This is why,in our study, we used the L/D:12/12 alternation as a synchronizer andtemperature as a marker of circadian rhythms in mice. In fact, the temp-erature rhythm represents one of the most important end-points of tox-icity in animal experiments (Boughattas et al., 1996; Ohdo et al., 1995).In one of our recent publications, we (Dridi et al., 2005) reported that ahigh toxic dose of loratadine (4 g/kg) resulted in slight hyperthermia,which varied as a function of drug dosing-time. The safe supratherapeuticdose of loratadine (20 mg/kg) was used in this study to ensure measurablelevels of the parent drug as well as its metabolite at the considered timepoints of the pharmacokinetic curve.

Our findings related to the pharmacokinetics of loratadine reveal thatthe clearance of loratadine is significantly reduced following adminis-tration at 9 HALO, compared with 1 and 17 HALO, resulting in alonger elimination half-life and an greater AUC. These results documentcircadian time-dependent differences in the pharmacokinetics of lorata-dine. The difference in Tmax between loratadine and desloratadinemight be explained by the extensive loratadine biotransformation todesloratadine. The metabolite Tmax occurred 1.3 h later than the parentcompound, loratadine, Tmax.

After oral administration of loratadine at 9 HALO, the AUC of deslor-atadine was significantly greater (see Table 1). This could be due to a sig-nificantly greater exposure of the parent drug (AUC of loratadine) at9 HALO. This probably resulted from an increased gastrointestinalabsorption of loratadine at 9 HALO, although this circadian time corre-sponds to a relative “fasting” stage of the 24 h cycle. The rest-activity differ-ences in loratadine absorption and its major metabolite, desloratadine, canbe partly explained by the variation in food consumption and by circadian

D. Dridi et al.542

rhythms of absorption. Both factors could accentuate the variability ofmonitored drug levels. Because the effect of food per se was not investi-gated in this study, we think that a better understanding of the influenceof food could prove beneficial for explaining the mechanisms of the bioa-vailability of this medication.

Loratadine is highly (99%) bound to plasma proteins; hence, circadianchanges in the plasma protein levels (Touitou et al., 1986) might haveinfluenced the pharmacokinetics of loratadine as a function of dosing-time. Loratadine biotransformation by microsomal enzymes to hydroxy-metabolites is a major mechanism of the elimination of this drug(Yumibe et al., 1996). Circadian changes have already been shown forthe cytochrome P450 level and activity of phase I and II drug-metabolizingenzymes (Labrecque & Beauchamp, 2003; Labrecque & Belanger, 1991;Miyazaki et al., 1990) Hepatic P450 monooxygenase activities showobvious daily fluctuations in rats with high values during the dark andlow values during the light span (Hirao et al., 2006). These observationsare in accordance with the present study, in which the highest values of lor-atadine and desloratadine clearances were observed in the dark phase at17 HALO (61.46 vs. 15.97 L/h/kg) and beginning of the diurnal restphase 1 HALO (57.81 vs. 14.22 L/h/kg).

Loratadine belongs to the class of high clearance drugs due to exten-sive first-pass metabolism, large tissue uptake, and high elimination(Paton & Webster, 1985). However, dosing time-dependent variation indrug elimination cannot be explained only by circadian fluctuation ofenzyme activity; circadian rhythmicity of liver blood flow can play a roleas well. The hepatic blood flow in rats and humans shows significant24 h variation with the peak in the active phase and trough in the restphase (Bruguerolle, 1998; Labrecque & Beauchamp, 2003; Lemmer &Nold, 1991). Furthermore, circadian variation of kidney elimination isalso important for some drugs (Belanger, 1993; Bruguerolle, 1998;Bruguerolle & Lemmer, 1993). Accordingly, 24 h variation in the func-tions of metabolism and excretion, including rhythms in hepatic bloodflow and metabolic enzymes (CYP450), are likely mechanisms of the circa-dian dosing time-dependent differences in the AUC of loratadine.

We presume that circadian variations in the activity of CYP450 mayhave affected the first-pass metabolism of loratadine. This would have con-tributed to the observed changes in the Cmax and the AUC of the studydrug. The high estimated CL/F of loratadine suggests a high presystemicdrug metabolism and tissue distribution or elimination (Katchen et al.,1985). However, no significant effect of administration time (9 and 17HALO) on the loratadine metabolic ratio (AUCdesloratadine/AUCloratadine)was observed, suggesting the variability in the oral apparent clearance isprobably related to the variability in drug distribution and/or eliminationbeing circadian time-dependent.

Chronopharmacokinetic Study of Loratadine in Mice 543

In our previous study, we reported the modeling in mice of loratadinecircadian dosing time-dependent toxic effects. We showed that maximumlethality, motor incoordination, and body weight loss was observed follow-ing dosing at 9 HALO, whereas minimum lethality followed dosing at 17HALO (Dridi et al., 2005). If the results of these studies and our recentresults are greatly inter-related, it appears that the administration time-dependent toxicity caused by loratadine results from the 24 h variationin drug pharmacokinetics. Indeed, adverse effects showed high valueswhen the AUC of loratadine and its metabolite were high, and lowvalues when these AUCs were low. The circadian dosing time-dependentdifference of loratadine pharmacokinetics appear to contribute to that ofloratadine chronotoxicity. Such a study should be extended to allergicpatients so that circadian time-dependent and pharmacodynamics can befully explored. Once such data are available, along with the data pertainingto rhythms in the histamine receptor number and binding capacity, aneffective and safe chronotherapeutic schedule for the treatment ofallergy can be designed (see Smolensky et al., 2007b).

ACKNOWLEDGMENTS

We wish to express our gratitude to Prof. Michael Smolensky for hismost pertinent advice and his help in the English editing of this article.We would like to thank Mr. Adel Rdissi for proofreading this article.Special thanks are addressed to Pierre Marquet for his warm welcome tohis Laboratory of Pharmacology in the Faculty of Medicine, Limoges,France, and his valuable scientific recommendations. This work was sup-ported by Le Ministere de l’Enseignement Superieur, de la RechercheScientifique et de la Technologie de la Republique Tunisienne.

REFERENCES

Barret A. (1998 November 30). New teeth for old patents. Business Week 92–94.Belanger PM. (1993). Chronopharmacology in drug research and therapy. Adv. Drug. Res. 24:1–80.Boughattas NA, Levi F, Hecquet B, Lemaigre G, Roulon A, Fournier C, Reinberg A. (1988). Circadian

time dependence of murine tolerance for carboplatin. Toxicol. Appl. Pharmacol. 96:233–247.Boughattas NA, Levi F, Fournier C, Lemaigre G, Roulon A, Hecquet B, Mathe G, Reinberg A. (1989).

Circadian rhythm in toxicities and tissue uptake of 1,2-diamminocyclohexane (trans-1) oxalato-platinum (II) in mice. Cancer Res. 49:3362–3368.

Boughattas NA, Hecquet B, Fournier C, Bruguerolle B, Trabelsi H, Bouzouita K, Omrane B, Levi F.(1994). Comparative pharmacokinetics of oxaliplatin (L-OHP) and carboplatin (CBDCA) in micewith reference to circadian dosing time. Biopharm. Drug Dispos. 15:761–773.

Boughattas NA, Li XM, Filipski J, Lemaigre G, Filipski E, Bouzouita K, Belhadj O, Levi F. (1996).Modulation of cisplatin chronotoxicity related to reduced glutathione in mice. Hum. Exp.Toxicol. 15:563–572.

Boughattas NA, Ben Attia M, Ixart G, Lemaigre G, Mechkouri M, Reinberg A. (2002). Pharmacologicalblockage of serotonin biosynthesis and circadian changes in oxaliplatin toxicity in rats. Chronobiol.Int. 19:1121–1136.

D. Dridi et al.544

Boulamery A, Kadra G, Simon N, Besnard T, Bruguerolle B. (2007). Chronopharmacokinetics of

imipenem in the rat. Chronobiol. Int. 24:961–968.

Brannan MD, Reidenberg P, Radwanski E, Shneyer L, Lin CC, Cayen MN, Affrime MB. (1995). Lor-

atadine administered concomitantly with erythromycin: Pharmacokinetic and electrocardio-

graphic evaluations. Clin. Pharmacol. Ther. 58:269–278.

Bruguerolle B. (1998). Chronopharmacokinetics: Current status. Clin. Pharmacokinet. 35:83–94.

Bruguerolle B, Lemmer B. (1993). Recent advances in chronopharmacokinetics: methodological pro-

blems. Life Sci. 52:1809–1824.

Bruguerolle B, Prat M. (1989). Temporal variations in the erythrocyte permeability to bupivacaine, eti-

docaine and mepivacaine in mice. Life Sci. 45:2587–2590.

Bruguerolle B, Boulamery A, Simon N. (2008). Biological rhythms: A neglected factor of variability in

pharmacokinetic studies. J. Pharm. Sci. 97:1099–1108.

Burioka N, Sasaki T. (1996). Chronopharmacology and chronotherapy for asthma by using PEF.

Nippon Rinsho 54:2956–2961.

Burioka N, Miyata M, Endo M, Fukuoka Y, Suyama H, Nakazaki H, Igawa K, Shimizu E. (2005a).

Alteration of the circadian rhythm in peak expiratory flow of nocturnal asthma following night-

time transdermal beta2-adrenoceptor agonist tulobuterol chronotherapy. Chronobiol. Int. 22:

383–390.

Burioka N, Takata M, Okano Y, Ohdo S, Fukuoka Y, Miyata M, Takane H, Endo M, Suyama H,

Shimizu E. (2005b). Dexamethasone influences human clock gene expression in bronchial epi-

thelium and peripheral blood mononuclear cells in vitro. Chronobiol. Int. 22:585–590.

Burioka N, Takata M, Endo M, Miyata M, Takeda K, Chikumi H, Tomita K, Fukuoka Y, Nakazaki H,

Sano H, Shimizu E. (2007). Treatment with beta2-adrenoceptor agonist in vivo induces human

clock gene, Per1, mRNA expression in peripheral blood. Chronobiol. Int. 24:183–189.

Cloutier M, Labrecque G, Rivard GB. (1993). Chronopharmacology of albuterol in hospitalized asth-

matic children. Chronobiol. Int. 10:290–297.

Cui Y, Sugimoto K, Araki N, Sudoh T, Fujimura A. (2005). Chronopharmacology of morphine in mice.

Chronobiol. Int. 22:515–522.

De Prins J, Waldura J. (1993). Sightseeing around the single cosinor. Chronobiol. Int. 10:395–400.

Dridi D, Boughattas NA, Aouam K, Reinberg A, Ben Attia M. (2005). Circadian time-dependent differ-

ences in murine tolerance to the antihistaminic agent loratadine. Chronobiol. Int. 22:499–514.

Gabrielsson J, Weiner D. (2000). Pharmacokinetic and Pharmacodynamic Data Analysis: Concepts and Appli-cations, 3rd edn. Stockholm, Sweden: Swedish Pharmaceutical Press, pp. 141–153.

Han KS, Lee MG. (1998). Circadian changes in the pharmacokinetics and pharmacodynamics of azo-

semide in rats. J. Pharm. Pharmacol. 50:767–774.

Haus E. (2007a). Chronobiology in the endocrine system. Adv. Drug Deliv. Rev. 59:985–1014.

Haus E. (2007b). Chronobiology of hemostasis and inferences for the chronotherapy of coagulation

disorders and thrombosis prevention. Adv. Drug Deliv. Rev. 59:966–984.

Hermida RC. (2007). Ambulatory blood pressure monitoring in the prediction of cardiovascular events

and effects of chronotherapy: Rationale and design of the MAPEC study. Chronobiol. Int. 24:

749–775.

Hermida RC Ayala DE, Calvo C, Portaluppi F, Smolensky MH. (2007). Chronotherapy of hyperten-

sion: Administation-time-dependent effects of treatment on the circadian pattern of blood

pressure. Adv. Drug Deliv. Res. 59:923–939.

Hilbert J, Radwanski E, Weglein R, Luc V, Perentesis G, Symchowicz S, Zampaglione N. (1987). Phar-

macokinetics and dose proportionality of loratadine. J. Clin. Pharmacol. 27:694–698.

Hirao J, Arakawa S, Watanabe K, Ito K, Furukawa T. (2006). Effects of restricted feeding on daily fluc-

tuations of hepatic functions including P450 monooxygenase activities in rats. J. Biol. Chem. 281:

3165–3171.

Holzberg D, Albrecht U. (2003). The circadian clock: a manager of biochemical processes within the

organism. J. Neuroendocrinol. 15:339–343.

Katchen B, Cramer J, Chung M, Gural R, Hilbert J, Luc V, Mortizen V, D’Sousa R, Symchowicz S,

Zampaglione N. (1985). Disposition of 14C-SSCH29851 in humans. Ann. Allergy 55:393–395.

Kay GG, Harris AG. (1999). Loratadine: A non-sedating antihistamine. Review of its effects on cogni-

tion, psychomotor performance, mood and sedation. Clin. Exp. Allergy 29(Suppl. 3):147–150.

Chronopharmacokinetic Study of Loratadine in Mice 545

Kosoglou T, Salfi M, Lim JM, Batra VK, Cayen MN, Affrime MB. (2000). Evaluation of the

pharmacokinetics and electrocardiographic pharmacodynamics of loratadine with concomitant

administration of ketoconazole or cimetidine. Br. J. Clin. Pharmacol. 50:581–589.

Labrecque G, Beauchamp D. (2003). Rhythms and pharmacokinetics. In Redfern PH (ed.) Chronother-apeutics. London, England: Pharmaceutical Press, pp. 75–110.

Labrecque G, Belanger PM. (1991). Biological rhythms in the absorption, distribution, metabolism and

excretion of drugs. Pharmacol. Ther. 52:95–107.

Lemmer B. (1991). Principles and concepts of chronopharmacology: Anti-asthma drugs—cardiovascu-

lar agents—H2 blockers. Internist. (Berl.) 32:380–388.

Lemmer B. (2005). Chronopharmacology and controlled drug release. Expert Opin. Drug Deliv. 2:

667–681.

Lemmer B. (2006a). Clinical chronopharmacology of the cardiovascular system: Hypertension and

coronary heart disease. Clin. Ter. 157:41–52.

Lemmer B. (2006b). The importance of circadian rhythms on drug response in hypertension and cor-

onary heart disease—from mice and man. Pharmacol. Ther. 111:629–651.

Lemmer B, Nold G. (1991). Circadian changes in estimated hepatic blood flow in healthy subjects. Br.

J. Clin. Pharmacol. 32:627–629.

Martin RJ. (1993). Nocturnal asthma: Circadian rhythms and therapeutic interventions. Am. Rev.Respir. Dis. 147(6Pt2):S25–S28.

Milano G, Chamorey AL. (2002). Clinical pharmacokinetics of 5-fluorouracil with consideration of

chronopharmacokinetics. Chronobiol. Int. 19:177–189.

Min DI, Chen HY, Fabrega A, Ukah FO, Wu YM, Corwin C, Ashton Ml, Martin M. (1996). Circadian

variation of tacrolimus disposition in liver allograft recipients. Transplantation 62:1190–1192.

Miyazaki Y, Yatagai M, Imaoka S, Funae Y, Motohashi Y, Kobayashi Y. (1990). Temporal variations in

hepatic cytochrome P450 isoenzymes in rats. Annu. Rev. Chronopharmacol. 7:149–153.

Nelson W, Tong YL, Lee JK, Halberg F. (1979). Methods for cosinor rhythmometry. Chronobiologia 6:

305–323.

Noehr-Jensen L, Damkier P, Bidstrup TB, Pedersen RS, Nielsen F, Brosen K. (2006). The relative

bioavailability of loratadine administered as a chewing gum formulation in healthy volunteers.

Eur. J. Clin. Pharmacol. 62:437–445.

Ohdo S. (2007). Chronopharmacology focused on biological clock. Drug Metab. Pharmacokinet. 22:3–14.

Ohdo S, Ogawa N, Song JG. (1995). Chronopharmacological study of acetylsalicylic acid in mice. Eur. J.Pharmacol. 293:151–157.

Paton DM, Webster DR. (1985). Clinical pharmacokinetics of H1-receptor antagonists (the antihista-

mines). Clin. Pharmacokinet. 10:477–497.

Portaluppi F, Lemmer B. (2007). Circadian rhythms in cardiac arrhythmias and opportunities for their

chronotherapy. Adv. Drug Deliv. Rev. 59:940–951.

Ramanathan R, Alvarez N, Su AD, Chowdhury S, Alton K, Stauber K, Patrick J. (2005). Metabolism and

excretion of loratadine in male and female mice, rats and monkeys. Xenobiotica 35:155–189.

Reinberg A. (1992). Concepts in chronopharmacology. Annu. Rev. Pharmacol. Toxicol. 32:51–66.

Reinberg A, Smolensky M. (1982). Circadian changes of drug disposition in man. Clin. Pharmacokinet. 7:

401–420.

Reinberg A, Smolensky M. (1983). Chronobiology and thermoregulation. Pharmacol. Ther. 22:

425–464.

Reinberg A, Levi F, Smolensky M. (1984). Clinical chronopharmacokinetics. J. Pharmacol. 15(Suppl. 1):

95–124.

Satoh S, Tada H, Murakami M, Tsuchiya N, Li Z, Numakura K, Saito M, Inoue T, Miura M, Hayase Y,

Suzuki T, Habuchi T. (2006). Circadian pharmacokinetics of mycophenolic acid and implication

of genetic polymorphisms for early clinical events in renal transplant recipients. Transplantation82:486–493.

Sauvage FL, Gaulier JM, Lachatre G, Marquet P. (2006). A fully automated turbulent-flow liquid

chromatography-tandem mass spectrometry technique for monitoring antidepressants in

human serum. Ther. Drug Monit. 28:123–130.

Smolensky MH, Peppas NA. (2007). Chronobiology, drug delivery and chronotherapy. Adv. DrugDeliv. Rev. 59:828–851.

D. Dridi et al.546

Smolensky MH, Portaluppi F. (1999). Chronopharmacology and chronotherapy of cardiovascularmedications: Relevance to prevention and treatment of coronary heart disease. Am. Heart. J.137(4Pt2):S14–S24.

Smolensky MH, Hermida RC, Portaluppi F. (2007a). Comparison of the efficacy of morning versusevening administration of olmesartan in uncomplicated essential hypertension. Chronobiol. Int.24:171–181.

Smolensky MH, Lemmer B, Reinberg A. (2007b). Chronobiology and chronotherapy of allergic rhini-tis and bronchial asthma. Adv. Drug Deliv. Rev. 59:852–882.

Takata M, Burioka N, Ohdo S, Fukuoka Y, Miyata M, Endo M, Suyama H, Shimizu E. (2005). Beta2-adrenoceptor agonists induce the mammalian clock gene, hPer1, mRNA in cultured human bron-chial epithelium cells in vitro. Chronobiol. Int. 22:777–783.

Touitou Y, Touitou C, Bogdan A, Reinberg A, Auzeby A, Beck H, Guillet P. (1986). Differencesbetween young and elderly subjects in seasonal and circadian variations of total plasma proteinsand blood volume as reflected by hemoglobin, hematocrit, and erythrocyte count. Clin. Chem. 32:801–804.

Touitou Y, Smolensky MH, Portaluppi F. (2006). Ethics, standards, and procedures of animal andhuman chronobiology research. Chronobiol. Int. 23:1083–1096.

White WB, LaRocca GM. (2002). Chronopharmacology of cardiovascular therapy. Blood Press. Monit. 7:199–207.

Widerhon N, Dıaz D, Picco E, Rebuelto M, Encinas T, Carlos Boggio J. (2005). Chronopharmacoki-netic study of gentamicin in dogs. Chronobiol. Int. 22:731–739.

Yao Z, DuBois DC, Almon RR, Jusko WJ. (2006). Modeling circadian rhythms of glucocorticoid recep-tor and glutamine synthetase expression in rat skeletal muscle. Pharm. Res. 23:670–679.

Yegnanarayan R, Mahesh SD, Sangle S. (2006). Chronotherapeutic dose schedule of phenytoin andcarbamazepine in epileptic patients. Chronobiol. Int. 23:1035–1046.

Yumibe N, Huie K, Chen KJ, Snow M, Clement RP, Cayen MN. (1996). Identification of human livercytochrome P450 enzymes that metabolize the nonsedating antihistamine loratadine. Formationof descarboethoxyloratadine by CYP3A4 and CYP2D6. Biochem. Pharmacol. 51:165–172.

Chronopharmacokinetic Study of Loratadine in Mice 547