Multiple-Dose Acetaminophen Pharmacokinetics CHANDRAHAS G.SAHAJWALLA** AND JAMESw. AYRES" Received Februa 5, 1990, from the 'College of Pharmacy, Oregon State University, Cowallis, OR 97337-3507. Accepted for publication December 6, 199r *Present address: International Drug Registration, Inc., 14915 Broschart Road, Rockville, MD 20850. Abstract 0 Four different treatments of acetaminophen (Tylenol) were
administered in multiple doses to eight healthy volunteers. Each treatment (325,650, 825, and 1000 mg) was administered five times at 6-h intervals. Saliva acetaminophen concentration versus time profiles were determined. Noncompartmental pharmacokinetic parameters were calculated and compared to determine whether acetaminophen exhibited linear or dose-dependentpharmacokinetics. For doses 5 18 mg/kg, area under the curve (AUC), half-life (t,,*), mean residence time (MRT), and ratio of AUC to dose for the first dose were compared with the last dose. No statistically significant differences were observed in dose-corrected AUC for the first or last dose among subjects or treatments. Half-livesand MRT were not significantly different among treatments for the first or the last dose. Statistically significant differences in t,,* and MRT were noted (p ~0.05) among subjects for the last dose. A plot of AUC versus dose for the first and the last doses exhibited a linear relationship. Dosecorrected saliva concentration versus time curves for the treatments were superimposable. Thus, acetaminophen exhibits linear pharmacokinetics for doses of 18 mg/kg or less. Plots of AUC versus dose for one subject who received doses higher than 18 mg/kg were curved, suggesting nonlinear behavior of acetaminophen in this subject.
Acetaminophen (paracetamol,N-acetyl paraminophenol) is a widely used non-narcotic analgesic and antipyretic compound. It is rapidly absorbed and distributed after oral administration, with peak concentrations obtained within 40 to 60 min. Its pharmacokinetics after oral and iv administration can be best described by a two-compartment open model, with a rapid distribution phase and a n elimination half-life of 2 to 3 h in the usual dose range.1-B Acetaminophen is relatively uniformly distributed throughout most body tissues and fluids in appreciable concentrations, reaching a tissue-to-plasma concentration ratio of about unity, except in fat and cerebrospinal fluid.9.10 Acetaminophen is incompletely available to systemic circulation after oral administration, since it is partly metabolized during absorption, primarily to pharmacologically inactive products. Present studies suggest that -10% of acetaminophen is subject to this presystemic biotransformation at doses of 1 g or more, and the percentage may be higher (40%)at lower do~es.4~7.11~12 About 25%of the acetaminophen dose has been reported to be metabolized by a first-pass effect.5 The drug is extensively metabolized and excreted largely in urine as various conjugates: 4 6 5 5 % as glucuronide conjugates, 2&30% as sulfate, and 1 5 5 5 %as cysteine and mercapturic acid conjugates. A minor fraction of acetaminophen is converted by cytochrome P-450-dependent hepatic mixed function oxidase to a highly reactive alkylating metabolite, which is probably n-acetyl-p-benzoquinoneimine.13 This metabolite is usually rapidly inactivated by conjugation with reduced glutathione and excreted in urine as cysteine and mercapturic acid conjugates. Overdoses of acetaminophen cause acute hepatic necrosis due to saturation of conjugation pathways, and glutathione stores become depleted due to covalent binding of the excessive reactive metabolites (possibly an epoxide) to vital cell constituents.16J7 OOZZ-3549/97/0900-0855$01.00/0 0 7 99 7, American Pharmaceutical Association
Although acetaminophen is a relatively safe analgesic in high doses and over prolonged periods, in overdose its metabolism may produce a quantitatively minor metabolite of drug which can destroy the enzyme systems responsible f o r its reduction. This could paralyze the capacity of the liver to reduce other toxic substances in addition to the drug itself. Recently, Borin and Ayress conducted a study with 15 human subjects by administering five different single doses (up to 2000 mg) of acetaminophen and collected sailiva samples for 16 h post dosing. These authors concluded that statistically significant differences in elimination rates and dose-corrected area under the curve (AUC)found among doses were suggestive of dose-dependent pharmacokinetics. Multiple-dosing pharmacokinetics of acetaminophen have not been well defined in the literature. Acetaminophen has recently become available in a nonprescription sustainedrelease dosage form (Drixoral Plus, Schering Corporation, Bloomfield, NJ). If saturable presystemic biotransformcci t'ion 0ccurs,4~7JOJ1then extent of bioavailability ( F ) may be decreased for a sustained release of drug relative to immediate release of the same dose. The effect may be confounded with multiple dosing if dose-dependent variation in elimination rate occurs.8 One study of multiple rectal administration has been reported18 in which acetaminophen (300 mg) and salicylamide (200 mg) in combined repetitive (three times, every 4 h) administration of suppositories was studied in 10 male volunteers. This study concluded that acetaminophen pharmacokinetic behavior conformed to literature values for a single dose and was described by a two-compartment open model. The kinetics of acetaminophen following single- and doublestrength administration to febrile children19 have been $studied and found to be linear, but no multiple-dose study with increasing doses has been conducted in adults. The objective of this study was to determine whether acetaminophen exhibits dose-dependent pharmacokinetics when comparing single dose data with steady-state data when the maximum dose is that recommended by FDA. If dose dependency exists, there should be nonlinear accumulation of drug at steady state.
Experimental Section Study Design-Eight healthy volunteers rage 22 to 29 years! were administered four different treatments (325, 650, 825. or 1000 rng of acetaminophen as Tylenol). Each treatment was administered orally in fivemultipledoses with dosingintervalsof6 h each. Saliva samples were collected for 36 h after administration of the first dose. Each participant signed an informed written consent. Treatments were administered to each subject every 6 h at 0 . 6 , 12, 18, and 24 h in the following manner: ( A , 325-mg Tylenol table:s 11ot no. AFR318 12/87; McNeil, Fort Washington, PA,; [ B I 650 .ng of Tylenol (two 325-mg tablets); tC) 825 mg of Tylenol tone e ~ c hof 325-mg and 500-mgTyleno1 Extra Strength tablets; lot no. AI)GF9d6 5/86); (D)1000 mg of Tylenol (two 500-mg extra strenbqh tatdetsi. Treatments were administered on four separate occasions separated by at least a week according to a randomized block design. Su'zjects fasted at least 12 h prior and 1 h after the first dose in each Fhase. Journal of Pharmaceutical Sciences / 855 Vol. 80, No. 9, September 1991
Subjects were also required to fast 2 h prior to and 1h after each dose for subsequent doses during each phase. No alcohol was permitted during the study. Treatments were taken with 6 fluid ounces of water and, immediately after swallowing the tablets and fluid, the mouth was rinsed with 20 mL of commercial mouthwash (containing 0.045% cetylpyridium chloride and 0.005% domiphen bromide as active ingredients) followed by a water rinse to remove drug that was adsorbed to the buccal mucosa. Saliva samples were collected by chewing squares of parafilm (1 inch x 1 inch; American Can Company, Greenwich, CT) for 1 min with continuous spitting into a 12-mL glass centrifuge tube. All subjects were provided with an alarm watch to remind them of sample and dose times. Saliva was centrifuged (Beckman model TJ-6 centrifuge, Palo Alto, CA) a t 3000 rpm for 30 min to remove mucous and particulate matter. Salivary supernatant was transferred to a polypropylene container with a lock cap and frozen a t -20 "C until analyzed. All participants in the study were taking no other medication during and 1 week prior to each phase. Criterion for exclusion from the study included history of chronic disease, therapy with an enzyme-inducing agent within the previous 30 days of the study, recent myocardial infarction, allergy to acetaminophen, and subjects with bleeding gums. Standard Solutions-Stock solutions containing 20-1000 mg/mL of acetaminophen (USP reference standard; USP, Inc., Rockville, MD) were prepared in distilled, deionized water. "he internal standard was 2-acetaminodophenol (Aldrich Chemical, Milwaukee, WI) in deionized water (60 pg/mL).Standards were prepared by spiking 500 pL of blank saliva with 25 pL of stock solutions. Then, 100 pL of the standard or unknown was combined with 100 pL of internal standard in 250-pL polyethylene centrifuge tubes and vortexed for 15 s. High-Performance Liquid Chromatography Assay-Analysis of saliva samples was carried out using the HPLC method reported by Borin and Ay-res.8 Linearity of peak height ratio versus acetaminophen concentration was excellent, with correlation coefficients of >0.995 and coefficients of variation ranging from 1 to 3%. Noncompartmental Analysis-Noncompartmental pharmacokinetic parameters for first and last dose were calculated and compared to determine whether acetaminophen exhibited linear or dosedependent pharmacokinetics. Half-lives &*) were calculated using least square regression of the slopes of the terminal phase of acetaminophen concentrations versus time curves. Analyses by ANOVA were performed using SIPS (Statistical Interactive Programming System for Cyber 70173,Department of Statistics, Oregon State University) for statistical analysis for dosecorrected AUC, t1,2rterminal slopes, and mean residence times (MRT) among treatments and subjects. For regression analysis, BMDP was used (Statistical Software, Inc., Version 1988 VAXNMS, Los Angeles, CA). Daily secretion of saliva normally ranges between -800 and 1500 mL.20 Hence, 18 mg/kg (one subject only) were excluded since inclusion of said data results in a high negative intercept for
100-
5cn
80 -
0
40-
60-
3 v 3
a
20 04 0
10080 -
1
4
8 12 DOSE I WT (mg/kg)
16
20
Flgure %Area under the acetaminophen concentration-time curve
s' E
0
-I
versus dose in mg/kg body weight. Key: (0)first dose; (0)steady-state
T
60-
dose.
0
5
v
o 3
40-
Q
20 1
04 0
400
800
1200
DOSE (mg)
Figure 2-Mean area under the saliva acetaminophen concentrationtime curve versus dose for eight subjects. Key: (0)first dose; (0) steady-state dose.
the first dose data. First-dose data are described by y = 3.31~ - 1.025,and steady-state data are described by y = 3.17~+ 0.83, as determined by linear regression. There was no improvement in fit using polynomial regression. Slopes were compared for first-dose and steady-state data and were not found to be statistically Significantlydifferent a t the 0.05level of significance. The data euggeet that acetaminophen follows 'linear kinetics with multiple doses up to 1000mg or doses 18 mg/kg; however, this cannot be substantiated until an organized study administering doses > l 8 mgkg is conducted. A study conducted with children,lg in which multiple doses of 24 to 30 mgkg were administered every 8 h for 72 h, did not indicate nonlinear pharmacokinetics for acetaminophen. A single-dose study8 suggests that nonlinearity occurs a t doses above -20 mgkg. Fifth Dose Corrected-In calculations of the abovediscussed parameters, steady state was assumed, and AUC and AUMC were calculated without extrapolating to infinity for the fifth dose. The AUC and AUMC for the fifth dose were recalculated (corrected fifth dose; as explained in the Exper-
-
'O01
-
8o
1
0 0
Figure 44Saliva acetaminophen concentration versus time curve for eight human subjects normalized to the 325-mg dose following multiple oral administration of commercial acetaminophen tablets at 0, 6, 12, 18, and 24 h. Key: (0)325 mg; (A) 650 mg; (*) 825 mg; (0)1000 mg. 858 I Journal of Pharmaceutical Sciences Vol. 80, No. 9, September 1991
1 400
I
800
1200
DOSE (mg)
Figure &Area under the saliva acetaminophen concentration-time curve versus dose for subject No. 2. Key: (e)first dose; (0)steady-state
dose.
Table Ill-Mean Pharmacoklnetlc Parameters for the Fifth Dose' (Corrected for Residual Concentrations of Prevlous Doses) following Oral Admlnlstratlon of Acetamlnophen In Multiple Doses
Dose, mg
p, h-lb
tim
h"
MRT, h
AUC, pg . h/mLd
AUCID, mg . hImL mge
C L ,mUmin'
325
0.335 (0.072) 0.399 (0.084) 0.346 (0.042) 0.356 (0.043)
1.952
3.24 (0.99) 2.83 (0.42) 3.41 (0.51) 3.75 (0.79)
19.03 (6.54) 32.62 (8.89) 45.66 (9.15) 615 7 (18.42)
0.059 (0.020) 0.050 (0.014) 0.055 (0.011) 0.062 (0.016)
311.6 (93.2) 351.7 (85.4) 314.9 (79.6) 288.1 (68.9)
650 825 1000
1.737 2.003 1.950
a Values in parentheses are standard deviations. Slope of the terminal portion of the saliva acetaminophen concentration-lime curves. '0.693lp. Area under the curve divided by dose (0). Relative bioavailability is calculated as bioavailability for each dose relative to the 1000-mgdose; average AUC for a doselaverage AUC for 1000-mg dose. 'Apparent clearance = dose/AUC.
Table IV-Mean
Pharmacoklnetlc Parameters for the Fourth Dose' following Oral Admlnlstratlon of AcetamlnoDhen In MultlDle Doses
Dose, mg
p, h-lb
325
0.341 (0.108) 0.325 (0.1 14) 0.299 (0.079) 0.321 (0.069)
650 825 1000
4/29
hC
2.032 2.132 2.318 1.159
AUCID,
-
MRT, h
AUC, pg h/mLd
mg h/mL. mge
CL,,,, mumin'
2.33 (0.160) 2.32 (0.215) 2.46 (0.289) 2.35 (0.155)
20.09 (6.48) 35.75 (8.19) 47.87 (12.01) 60.96 (15.60)
0.062 (0.021) 0.055 (0.0126) 0.058 (0.015) 0.061 (0.016)
295.8 (95.1) 316.4 (67.7) 301.4 (66.1) 292.0 (66.1)
a Fourth doses from 16 to 24 h; values in parentheses are standard deviations. Slope of the terminal portion of the saliva acetaminophen concentration-lime curves. " 0.693Ip. Area under the curve divided by dose (0). Relative bioavailability is calculated as bioavailability for each dose relative to the 1000-mg dose; average AUC for a dose/average AUC for 1000-mgdose. 'Apparent clearance = dose/AUC.
imental Section) assuming that steady state may not have been reached; these pharmacokinetic parameters are presented in Table 111. As expected, dose-corrected AUC, MRT, CLapp,and tIl2were not significantly different among treatments or compared with parameters calculated assuming steady state was reached by the fifth dose. Fourth Dose-Mean pharmacokinetic parameters calculated for the fourth dose (1Eb24 h) for individual subjects are presented in Table IV. Steady state was assumed while calculating these parameters. No significant differences were noted in dose-correctedAUC, CL, ,or tl,2 among treatments. The MRT values for 325,650, or 8% mg were not significantly different from that of steady-state MRT (Table 11). However, MRT for the fourth dose for 1000 mg was significantly different from that at steady state. Half-lives for the fourth dose were slightly higher than those at steady state, although the terminal slopes (p)were not significantly different among treatments. Parameters for the fourth dose may not be reliable as there were only six data points during the dosing interval; this could give incorrect estimates of AUC and terminal slopes.
Conclusions All subjects followed the protocol and completed the study successfully. Peak concentrations for all treatments were reached within 1h, indicating rapid absorption and distribution. The t,,, although statistically nonsignificant, tended to was increase at higher doses at steady state. The C, significantly different for treatments, as would be expected with increasing doses. Dose-corrected AUC, MRT, tIl2,and CL,, were not significantly different among treatments. Plots of saliva acetaminophen concentrations normalized to a 325-mg dose, versus time, were superimposable, indicating linearity of kinetics (for doses 18 mgkg, exhibited nonlinear kinetics based on a curved plot for AUC versus dose. Although acetaminophen m a y follow (in adults) nonlinear dose-dependent kinetics8 for doses >18 mgkg, it is possible that this one subject had impaired metabolism of the drug compared with other subjects (liver function tests were not carried out for any of the subjects participating in the study). However, nonlinearity cannot be substantiated until an organized study administering doses >18 mgkg is conducted. Bioavailability relative to the 1000-mgdose at steady state was comparable for high and low doses. However, for the first dose, the 325-mg dose was only 80% bioavailable compared with the 1000-mgfirst dose. This result and the fact that t,, increases with dose support the finding of Rawlins et al.4 and Borin and Ayress that lower doses had lower bioavailability for single doses. However, in multiple dosing when steady state is reached, bioavailability is comparable for the doses studied.
References and Notes 1. Beaver, W. T. Am. J . Med. Sci. 1966,251, 576-599. 2. Seymour, R. A.;Rawlins, M. D. Eur. J . Clin.Pharmacol. 1981,20, 215-218. 3. Albert, K. S.; Sedman, A. J.; Wagner, J. G. J . Pharmokinet. Biopharm. 1974,2,381-393. 4. Rawlins, M. D.; Henderson, D. B.; Hijab, A. R. Eur. J . Clin. Pharmacol. 1977,11,283-286. 5 . Clements, J . A.; Heading, R. C.; Nimmo, W. S.; Prescott, L. F. Clin. Pharmacol. Ther. 1978,24, 420-431.
Journal of Pharmaceutical Sciences I 859 Vol. 80, No. 9, September 7997
6. Morris, M. E.; Levy,A. J . Pharm. Sci. 1984, 73,103%1041. 7. Ameer, B.; Divoll, M.;Abernethy, D. R.; Greenblatt, D. J.; Shargel, L. J . Phurm. Sci. 1983,72,955-958. 8. Borin. M.T.: Awes. J. W. Znt. J . Pharm. 1989.54. 199-209. 9. Brodie, B.B.;&elrod, S. J. Pharmacol. Exp: Ther. 1949,97, 58-67. 10. Gwilt, P. R.;Morse, D.; Burkett, M.; Petiprin, D. J . Am. J . Pharm. 1979.43. 124-126. 11. Chiov, W.L. J. Pharm. Sci. 1975,64,1734-1735. 12. Perucca, E.; Richens, A. Br. J . Clin.Pharmacol. 1979,7,201-206. 13. Miner, D. J.; Kissenger, P. T. Biochem. Pharmacol. 1979,28, 3285-3290. 14. Mitchell, J . R.;Jollow, D. J.; Potter, W. Z.; Davis, D. C.; Gillette, J. R.; Brodie, B. B. J.Pharmacol. Exp. Ther. 1973,187,185-194. 15. Mitchell, J. R.; Thorgenrsson, S. S.; Potter, W. Z.; Jollow, D. J.; Keiser, H. Clin. Pharmacol. Ther. 1974,16.676-684. 16. Andrews, R. S.; Bond, C. C.; Burnett, J.; Saunders, A,; Watson, A. J. Znt. Med. Res. 1976,4,34. 17. Davis, M.;Laboadarricas, D.; Williams, R. S. J . Znt. Med. Res. 1976.4 40. - - - - I
- 7
18. Ebel, S.;Mibler, B.; Hasse, W.; Stein, L. Arzneim.-Forsch.lDrug
860 I Journal of Pharmaceutical Sciences Vol. 80, No. 9, September 1991
Res. 1980,30,1295-1298. 19. Nahata, M.C.; Powell, D. A. Clin. Res. 1982,634A. 20. Guyton, A.C. Textbook of Medical Physicology, 6th 4.W. ;B. Saunders; Philadelphia, PA, 1981;pp 803-805. 21. Yamaoka, K.;Nakagawa, T.; Uno, T. J. Phurmacokinet. Biopharm. 1978,6,547-558. 22. Riegelman, S.; Collier, P. J . Phurmacokinet. Bwphurm. 1980,8, 509-534. 23. Glynn, J. P.; Bastain, W. J . Pharm. Pharmacol. 1973,25,420421. 24. Ahmed, M.;Enever, R. P. J . Clin. Hosp. Phurm. 1981,6,27-28. 25. Adithan, C.; Thangam, J. Br. J . Clin. Phurmucol. 1982, 14, 107-109. 26. Gibaldi, M. J . Clin. Pharmacol. 1986,26,330-331. 27. Hayton, W. L. J . Pharm. Sci. 1985,74 (lo), 1134. 28. Muckow, J. C.; Bending, M. R.; Kahn, A. G.; Dallery, C . T.Clin. Pharmacol. Ther. 1978,563-570. 29. Mucklow, J. C. Therapeutic Drug Monitoring, Raven: New York, 1982;Volume 4,pp 229-247. 30. Prescott, L. F.Br. J . Clin. Phurmncol. 1980,2915-2985.
administered in multiple doses to eight healthy volunteers. Each treatment (325,650, 825, and 1000 mg) was administered five times at 6-h intervals. Saliva acetaminophen concentration versus time profiles were determined. Noncompartmental pharmacokinetic parameters were calculated and compared to determine whether acetaminophen exhibited linear or dose-dependentpharmacokinetics. For doses 5 18 mg/kg, area under the curve (AUC), half-life (t,,*), mean residence time (MRT), and ratio of AUC to dose for the first dose were compared with the last dose. No statistically significant differences were observed in dose-corrected AUC for the first or last dose among subjects or treatments. Half-livesand MRT were not significantly different among treatments for the first or the last dose. Statistically significant differences in t,,* and MRT were noted (p ~0.05) among subjects for the last dose. A plot of AUC versus dose for the first and the last doses exhibited a linear relationship. Dosecorrected saliva concentration versus time curves for the treatments were superimposable. Thus, acetaminophen exhibits linear pharmacokinetics for doses of 18 mg/kg or less. Plots of AUC versus dose for one subject who received doses higher than 18 mg/kg were curved, suggesting nonlinear behavior of acetaminophen in this subject.
Acetaminophen (paracetamol,N-acetyl paraminophenol) is a widely used non-narcotic analgesic and antipyretic compound. It is rapidly absorbed and distributed after oral administration, with peak concentrations obtained within 40 to 60 min. Its pharmacokinetics after oral and iv administration can be best described by a two-compartment open model, with a rapid distribution phase and a n elimination half-life of 2 to 3 h in the usual dose range.1-B Acetaminophen is relatively uniformly distributed throughout most body tissues and fluids in appreciable concentrations, reaching a tissue-to-plasma concentration ratio of about unity, except in fat and cerebrospinal fluid.9.10 Acetaminophen is incompletely available to systemic circulation after oral administration, since it is partly metabolized during absorption, primarily to pharmacologically inactive products. Present studies suggest that -10% of acetaminophen is subject to this presystemic biotransformation at doses of 1 g or more, and the percentage may be higher (40%)at lower do~es.4~7.11~12 About 25%of the acetaminophen dose has been reported to be metabolized by a first-pass effect.5 The drug is extensively metabolized and excreted largely in urine as various conjugates: 4 6 5 5 % as glucuronide conjugates, 2&30% as sulfate, and 1 5 5 5 %as cysteine and mercapturic acid conjugates. A minor fraction of acetaminophen is converted by cytochrome P-450-dependent hepatic mixed function oxidase to a highly reactive alkylating metabolite, which is probably n-acetyl-p-benzoquinoneimine.13 This metabolite is usually rapidly inactivated by conjugation with reduced glutathione and excreted in urine as cysteine and mercapturic acid conjugates. Overdoses of acetaminophen cause acute hepatic necrosis due to saturation of conjugation pathways, and glutathione stores become depleted due to covalent binding of the excessive reactive metabolites (possibly an epoxide) to vital cell constituents.16J7 OOZZ-3549/97/0900-0855$01.00/0 0 7 99 7, American Pharmaceutical Association
Although acetaminophen is a relatively safe analgesic in high doses and over prolonged periods, in overdose its metabolism may produce a quantitatively minor metabolite of drug which can destroy the enzyme systems responsible f o r its reduction. This could paralyze the capacity of the liver to reduce other toxic substances in addition to the drug itself. Recently, Borin and Ayress conducted a study with 15 human subjects by administering five different single doses (up to 2000 mg) of acetaminophen and collected sailiva samples for 16 h post dosing. These authors concluded that statistically significant differences in elimination rates and dose-corrected area under the curve (AUC)found among doses were suggestive of dose-dependent pharmacokinetics. Multiple-dosing pharmacokinetics of acetaminophen have not been well defined in the literature. Acetaminophen has recently become available in a nonprescription sustainedrelease dosage form (Drixoral Plus, Schering Corporation, Bloomfield, NJ). If saturable presystemic biotransformcci t'ion 0ccurs,4~7JOJ1then extent of bioavailability ( F ) may be decreased for a sustained release of drug relative to immediate release of the same dose. The effect may be confounded with multiple dosing if dose-dependent variation in elimination rate occurs.8 One study of multiple rectal administration has been reported18 in which acetaminophen (300 mg) and salicylamide (200 mg) in combined repetitive (three times, every 4 h) administration of suppositories was studied in 10 male volunteers. This study concluded that acetaminophen pharmacokinetic behavior conformed to literature values for a single dose and was described by a two-compartment open model. The kinetics of acetaminophen following single- and doublestrength administration to febrile children19 have been $studied and found to be linear, but no multiple-dose study with increasing doses has been conducted in adults. The objective of this study was to determine whether acetaminophen exhibits dose-dependent pharmacokinetics when comparing single dose data with steady-state data when the maximum dose is that recommended by FDA. If dose dependency exists, there should be nonlinear accumulation of drug at steady state.
Experimental Section Study Design-Eight healthy volunteers rage 22 to 29 years! were administered four different treatments (325, 650, 825. or 1000 rng of acetaminophen as Tylenol). Each treatment was administered orally in fivemultipledoses with dosingintervalsof6 h each. Saliva samples were collected for 36 h after administration of the first dose. Each participant signed an informed written consent. Treatments were administered to each subject every 6 h at 0 . 6 , 12, 18, and 24 h in the following manner: ( A , 325-mg Tylenol table:s 11ot no. AFR318 12/87; McNeil, Fort Washington, PA,; [ B I 650 .ng of Tylenol (two 325-mg tablets); tC) 825 mg of Tylenol tone e ~ c hof 325-mg and 500-mgTyleno1 Extra Strength tablets; lot no. AI)GF9d6 5/86); (D)1000 mg of Tylenol (two 500-mg extra strenbqh tatdetsi. Treatments were administered on four separate occasions separated by at least a week according to a randomized block design. Su'zjects fasted at least 12 h prior and 1 h after the first dose in each Fhase. Journal of Pharmaceutical Sciences / 855 Vol. 80, No. 9, September 1991
Subjects were also required to fast 2 h prior to and 1h after each dose for subsequent doses during each phase. No alcohol was permitted during the study. Treatments were taken with 6 fluid ounces of water and, immediately after swallowing the tablets and fluid, the mouth was rinsed with 20 mL of commercial mouthwash (containing 0.045% cetylpyridium chloride and 0.005% domiphen bromide as active ingredients) followed by a water rinse to remove drug that was adsorbed to the buccal mucosa. Saliva samples were collected by chewing squares of parafilm (1 inch x 1 inch; American Can Company, Greenwich, CT) for 1 min with continuous spitting into a 12-mL glass centrifuge tube. All subjects were provided with an alarm watch to remind them of sample and dose times. Saliva was centrifuged (Beckman model TJ-6 centrifuge, Palo Alto, CA) a t 3000 rpm for 30 min to remove mucous and particulate matter. Salivary supernatant was transferred to a polypropylene container with a lock cap and frozen a t -20 "C until analyzed. All participants in the study were taking no other medication during and 1 week prior to each phase. Criterion for exclusion from the study included history of chronic disease, therapy with an enzyme-inducing agent within the previous 30 days of the study, recent myocardial infarction, allergy to acetaminophen, and subjects with bleeding gums. Standard Solutions-Stock solutions containing 20-1000 mg/mL of acetaminophen (USP reference standard; USP, Inc., Rockville, MD) were prepared in distilled, deionized water. "he internal standard was 2-acetaminodophenol (Aldrich Chemical, Milwaukee, WI) in deionized water (60 pg/mL).Standards were prepared by spiking 500 pL of blank saliva with 25 pL of stock solutions. Then, 100 pL of the standard or unknown was combined with 100 pL of internal standard in 250-pL polyethylene centrifuge tubes and vortexed for 15 s. High-Performance Liquid Chromatography Assay-Analysis of saliva samples was carried out using the HPLC method reported by Borin and Ay-res.8 Linearity of peak height ratio versus acetaminophen concentration was excellent, with correlation coefficients of >0.995 and coefficients of variation ranging from 1 to 3%. Noncompartmental Analysis-Noncompartmental pharmacokinetic parameters for first and last dose were calculated and compared to determine whether acetaminophen exhibited linear or dosedependent pharmacokinetics. Half-lives &*) were calculated using least square regression of the slopes of the terminal phase of acetaminophen concentrations versus time curves. Analyses by ANOVA were performed using SIPS (Statistical Interactive Programming System for Cyber 70173,Department of Statistics, Oregon State University) for statistical analysis for dosecorrected AUC, t1,2rterminal slopes, and mean residence times (MRT) among treatments and subjects. For regression analysis, BMDP was used (Statistical Software, Inc., Version 1988 VAXNMS, Los Angeles, CA). Daily secretion of saliva normally ranges between -800 and 1500 mL.20 Hence, 18 mg/kg (one subject only) were excluded since inclusion of said data results in a high negative intercept for
100-
5cn
80 -
0
40-
60-
3 v 3
a
20 04 0
10080 -
1
4
8 12 DOSE I WT (mg/kg)
16
20
Flgure %Area under the acetaminophen concentration-time curve
s' E
0
-I
versus dose in mg/kg body weight. Key: (0)first dose; (0)steady-state
T
60-
dose.
0
5
v
o 3
40-
Q
20 1
04 0
400
800
1200
DOSE (mg)
Figure 2-Mean area under the saliva acetaminophen concentrationtime curve versus dose for eight subjects. Key: (0)first dose; (0) steady-state dose.
the first dose data. First-dose data are described by y = 3.31~ - 1.025,and steady-state data are described by y = 3.17~+ 0.83, as determined by linear regression. There was no improvement in fit using polynomial regression. Slopes were compared for first-dose and steady-state data and were not found to be statistically Significantlydifferent a t the 0.05level of significance. The data euggeet that acetaminophen follows 'linear kinetics with multiple doses up to 1000mg or doses 18 mg/kg; however, this cannot be substantiated until an organized study administering doses > l 8 mgkg is conducted. A study conducted with children,lg in which multiple doses of 24 to 30 mgkg were administered every 8 h for 72 h, did not indicate nonlinear pharmacokinetics for acetaminophen. A single-dose study8 suggests that nonlinearity occurs a t doses above -20 mgkg. Fifth Dose Corrected-In calculations of the abovediscussed parameters, steady state was assumed, and AUC and AUMC were calculated without extrapolating to infinity for the fifth dose. The AUC and AUMC for the fifth dose were recalculated (corrected fifth dose; as explained in the Exper-
-
'O01
-
8o
1
0 0
Figure 44Saliva acetaminophen concentration versus time curve for eight human subjects normalized to the 325-mg dose following multiple oral administration of commercial acetaminophen tablets at 0, 6, 12, 18, and 24 h. Key: (0)325 mg; (A) 650 mg; (*) 825 mg; (0)1000 mg. 858 I Journal of Pharmaceutical Sciences Vol. 80, No. 9, September 1991
1 400
I
800
1200
DOSE (mg)
Figure &Area under the saliva acetaminophen concentration-time curve versus dose for subject No. 2. Key: (e)first dose; (0)steady-state
dose.
Table Ill-Mean Pharmacoklnetlc Parameters for the Fifth Dose' (Corrected for Residual Concentrations of Prevlous Doses) following Oral Admlnlstratlon of Acetamlnophen In Multiple Doses
Dose, mg
p, h-lb
tim
h"
MRT, h
AUC, pg . h/mLd
AUCID, mg . hImL mge
C L ,mUmin'
325
0.335 (0.072) 0.399 (0.084) 0.346 (0.042) 0.356 (0.043)
1.952
3.24 (0.99) 2.83 (0.42) 3.41 (0.51) 3.75 (0.79)
19.03 (6.54) 32.62 (8.89) 45.66 (9.15) 615 7 (18.42)
0.059 (0.020) 0.050 (0.014) 0.055 (0.011) 0.062 (0.016)
311.6 (93.2) 351.7 (85.4) 314.9 (79.6) 288.1 (68.9)
650 825 1000
1.737 2.003 1.950
a Values in parentheses are standard deviations. Slope of the terminal portion of the saliva acetaminophen concentration-lime curves. '0.693lp. Area under the curve divided by dose (0). Relative bioavailability is calculated as bioavailability for each dose relative to the 1000-mgdose; average AUC for a doselaverage AUC for 1000-mg dose. 'Apparent clearance = dose/AUC.
Table IV-Mean
Pharmacoklnetlc Parameters for the Fourth Dose' following Oral Admlnlstratlon of AcetamlnoDhen In MultlDle Doses
Dose, mg
p, h-lb
325
0.341 (0.108) 0.325 (0.1 14) 0.299 (0.079) 0.321 (0.069)
650 825 1000
4/29
hC
2.032 2.132 2.318 1.159
AUCID,
-
MRT, h
AUC, pg h/mLd
mg h/mL. mge
CL,,,, mumin'
2.33 (0.160) 2.32 (0.215) 2.46 (0.289) 2.35 (0.155)
20.09 (6.48) 35.75 (8.19) 47.87 (12.01) 60.96 (15.60)
0.062 (0.021) 0.055 (0.0126) 0.058 (0.015) 0.061 (0.016)
295.8 (95.1) 316.4 (67.7) 301.4 (66.1) 292.0 (66.1)
a Fourth doses from 16 to 24 h; values in parentheses are standard deviations. Slope of the terminal portion of the saliva acetaminophen concentration-lime curves. " 0.693Ip. Area under the curve divided by dose (0). Relative bioavailability is calculated as bioavailability for each dose relative to the 1000-mg dose; average AUC for a dose/average AUC for 1000-mgdose. 'Apparent clearance = dose/AUC.
imental Section) assuming that steady state may not have been reached; these pharmacokinetic parameters are presented in Table 111. As expected, dose-corrected AUC, MRT, CLapp,and tIl2were not significantly different among treatments or compared with parameters calculated assuming steady state was reached by the fifth dose. Fourth Dose-Mean pharmacokinetic parameters calculated for the fourth dose (1Eb24 h) for individual subjects are presented in Table IV. Steady state was assumed while calculating these parameters. No significant differences were noted in dose-correctedAUC, CL, ,or tl,2 among treatments. The MRT values for 325,650, or 8% mg were not significantly different from that of steady-state MRT (Table 11). However, MRT for the fourth dose for 1000 mg was significantly different from that at steady state. Half-lives for the fourth dose were slightly higher than those at steady state, although the terminal slopes (p)were not significantly different among treatments. Parameters for the fourth dose may not be reliable as there were only six data points during the dosing interval; this could give incorrect estimates of AUC and terminal slopes.
Conclusions All subjects followed the protocol and completed the study successfully. Peak concentrations for all treatments were reached within 1h, indicating rapid absorption and distribution. The t,,, although statistically nonsignificant, tended to was increase at higher doses at steady state. The C, significantly different for treatments, as would be expected with increasing doses. Dose-corrected AUC, MRT, tIl2,and CL,, were not significantly different among treatments. Plots of saliva acetaminophen concentrations normalized to a 325-mg dose, versus time, were superimposable, indicating linearity of kinetics (for doses 18 mgkg, exhibited nonlinear kinetics based on a curved plot for AUC versus dose. Although acetaminophen m a y follow (in adults) nonlinear dose-dependent kinetics8 for doses >18 mgkg, it is possible that this one subject had impaired metabolism of the drug compared with other subjects (liver function tests were not carried out for any of the subjects participating in the study). However, nonlinearity cannot be substantiated until an organized study administering doses >18 mgkg is conducted. Bioavailability relative to the 1000-mgdose at steady state was comparable for high and low doses. However, for the first dose, the 325-mg dose was only 80% bioavailable compared with the 1000-mgfirst dose. This result and the fact that t,, increases with dose support the finding of Rawlins et al.4 and Borin and Ayress that lower doses had lower bioavailability for single doses. However, in multiple dosing when steady state is reached, bioavailability is comparable for the doses studied.
References and Notes 1. Beaver, W. T. Am. J . Med. Sci. 1966,251, 576-599. 2. Seymour, R. A.;Rawlins, M. D. Eur. J . Clin.Pharmacol. 1981,20, 215-218. 3. Albert, K. S.; Sedman, A. J.; Wagner, J. G. J . Pharmokinet. Biopharm. 1974,2,381-393. 4. Rawlins, M. D.; Henderson, D. B.; Hijab, A. R. Eur. J . Clin. Pharmacol. 1977,11,283-286. 5 . Clements, J . A.; Heading, R. C.; Nimmo, W. S.; Prescott, L. F. Clin. Pharmacol. Ther. 1978,24, 420-431.
Journal of Pharmaceutical Sciences I 859 Vol. 80, No. 9, September 7997
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