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Pharmacokinetics of Indomethacin Octyl Ester (Prodrug) and Indomethacin Produced from the Prodrug
Pharmacokinetics of Indomethacin Octyl Ester (Prodrug) and Indomethacin Produced from the Prodrug TAROOGISO~, MASAHIROIWAKI, TAKAHIRO KINOSHITA, TADATOSHI TANINO,AND TSUYOSHIPAKU Received June 15, 1992, from the Faculty of Pharmaceutical Sciences, Kinki University, Accepted for publication April 14, 1993". 3-4- 1 Kowakae, Higashi-Osaka, Osaka 577, Japan.
concentration versus time profiles after indomethacin dosings. Indomethacin, a potent nonsteroidal anti-inflammatory drug, has been widely used in the treatment of rheumatoid arthritis, gout, osteoarthritis, and acute muscle-skeletal disorders. Unfortunately, like other nonsteroidal anti-inflammatory agents, indomethacin carries the risk of side effects, such as gastrointestinal irritation and ulceration.13 Therefore, the prodrugs acemethacin and indomethacin farnesil, were synthesized with the intention of reducing the side effects4 and increasing the tissue distribution: respectively, and have been used for clinical therapy. Although some reports on the basic pharmacokinetics of these prodrugs are published,5.6 as far as we know, there has been no report on pharmacokinetic analyses with a model for describing the blood concentration profiles of the prodrugs and parent drug after dosing. Because it is known that the estrification of acidic antiinflammatory drugs suppresses their gastrotoxicity,7Sumitomo Pharmaceutical Company and we synthesized a prodrug of indomethacin, indomethacin octyl ester (IM-OE), and the pharmacokinetics of this agent was investigated in rats after single intravenous (iv) and oral administrations. To describe the time course of the plasma indomethacin and its prodrug levels, a pharmacokinetic model consisting of central and peripheral compartments with the first-order conversion rate constant of the prodrug to indomethacin was applied. The appropriateness and significance of the models for pharmacokinetic evaluation are discussed.
Experimental Section Materials-Indomethacin (J.P. grade) and IM-OE (mp 55-56 "C; anal.-calcd for Cz~H32ClN04:C, 69.00; H, fi.86; N, 2.98;found C, 69.01; H, 6.82; N, 2.98) were a gift from Sumitomo Pharmaceutical Company (Osaka, Japan). The structure of IM-OE synthesized was confirmed on the basis of IR and lH NMR spectra. Mefenamic acid and 8-estradial dipropionate, internal standards for the HPLC assay, were obtained from Sigma Chemical Company (St. Louis, MO). All other chemicals and solvents used were of reagent grade or HPLC quality. @
Abstract published in Aduance ACS Abstracts, November 15,1993.
34 /Journal of Pharmaceutical Sciences Vot. 83, No. 1, January 1994
4
i.v.
Abstract 0 A prodrug of indomethacin, indomethacin octyl ester (IMOE), was synthesized and its pharmacokinetics was investigated in rat. To describe the time course of the plasma indomethacin and IM-OE after intravenous ( i ) and oral admlnistrations, a pharmacokinetic model with four compartmentswas developed. Indomethacin rapidly appeared in plasma after iv administration of IM-OE and declined in a monoexponential manner, with a rapid decline and low plasma levels of IM-OE. The plasma concentrations of indomethacin after oral administration of IM-OE were much lower than those after oral administration of indomethacin. The high concentrations of IM-OE compared with indomethacin were detected in liver 3 h after oral dosing of the prodrug, although IM-OE was not detected in plasma. A good fit was obtained between the observed and calculated curves based on the model, which includes a conversion rate constant of IM-OE to indomethacin for both iv and oral dosings of IM-OE. Additionally, the model could successfully describe the plasma
/
i.v.
1-Four-compartment pharmacokinetic models. Key: (IM) indomethacin; (IM-OE) indomethacin octyl ester; (klo and k3,,)the firstorder elimination rate constants for IM-OE and IM, respectively; (k13) the first-order conversion rate constant of IM-OE to IM; (k12,kpl,k3,, and ka) the first-order kinetic constants for the distribution between central and the first-order absorption rate peripheral compartments;(kal- and kaIM) constants for IM-OE and IM, respectively. Figure
Experimental Animals-Male W i s h rats, weighing200-280g, were used. The animals, maintained on an MF diet (a bleeding feed, Oriental Yeast Company, Tokyo, Japan) for 3-4 days prior to the experiment, were divided at random into two groups, each consisting of 3-5 rats. One group was treated with indomethacin and the second group was administered IM-OE. On the day before the experiment, the rat jugular vein was cannulated with silicon tubing.8 Intravenous and Oral Administration-Indomethacin (5 mg/kg) and IM-OE ( 5 mg/kg, indomethacin equivalent) were administered iv as a solution in a mixture of NJV-dimethy1formamide:ethanol:water( 6 22, v/v). Indomethacin (7.6 mg/kg) and the prodrug (7.6 mg/kg, indomethacin equivalent) were administered orally as an aqueous suspension in 1.5% acasia. A 0.2-mL blood sample was collected periodically after dosing through the cannula for a total time period. The plasma was separated immediately by centrifugation and stored frozen until the time of assay. Hepatic Uptake-Rats received indomethacin or IM-OE orally at the same dose as described for the above oral dosing. At 3 hafter dosing, the liver was removed from the animals immediately after sacrifice. The liver,perfused with ice-cold0.9% NaCl toremove blood, was homogenized in ice-cold 10 mM phosphate buffer (pH 7.4). The drugs in the homogenate (1.0 mL) were extracted with methanol (2.0mL) containing the internal standard. After centrifugation, the supernatant was stored frozen until assay. D e t e r m i n a t i o n of I n d o m e t h a c i n a n d IM-OE-Indomethacin and IM-OE in plasma or homogenate was determined by the HPLC method of Kwong et aL9with slight modifications. A 50-pLaliquot of plasma was mixed with 150 pL of methanol containing 8 pg/mL of mefenamic acid as an internal standard, and the mixture was centrifuged a t 15 OOO rpm. The supernatant was filtered through the membrane filter (0.45 pm, Ekikrodisc 3CR, Gelman Sciences Japan, Tokyo, Japan). The filtrate was injected into a reversed-phase Inertail ODs-2column (5 pm, 4.6 mm X 15cm; GL Sciences Inc., Tokyo, Japan) with a Shimadzu liquid chromatograph (model LC-fiA) equipped with a UV spectrophotometer (modelSPD-6AV). The mobile phase was acetonitrile:methanok Hz0:acetic acid (651025:1, v/v), pumped at a flow rate of 1.0 mL/min. Detection was at 254 nm. For the determination of IM-OE, a 100-pL aliquot of sample was mixed with 300 pL of methanol containing
0022-3549/94/ 1200-34$04.50/0
0 1994, American Chemical Society and American Pharmaceutical Association
A
,-
B
0.01
0
0.01
2
4
6
8
1
0
1
2
I i 0
2
4
6
8 1 0 1 2
Time after administration (h) Flgure 2-Plasma concentration-time curves for lndomethacin and IM-OE after (A) iv administration at a dose of 5 mg InL-methacin equivalentlkg and (B)oral administrationat a dose of 7.6 mg indomethacin equivalentlkg. Key: (R) indomethacin; (A)indomethacinafter IM-OE(0)IM-OE. Points
and vertical bars represent the mean and SD (n = 3-4), respectively. Solid lines are the simulation curves calculated with the model descrlbed In Figure 1 and the parameters listed in Table 11. Table 1-Hepatlc Plasma and Blood Concentratlons at 3 h after Oral Admlnlstratlon of Indomethacln or IM-OE.
Concentration ~~~
Drug IM
IM-OE IM after IM-OE
~~
Liver, nmol/g
Plasma, nmol/mL
nmol/mL
11.86 f 4.84 131.52 f 48.8 2.88 f 0.48
47.52 f 3.47 0 1.22 f 0.06
26.57 f 4.84 0 0.58 f 0.17
Blood,
* Dose, 7.6 mg/kg (indomethacinequivalent);values are the mean f SD (n = 3). fl-estradiol dipropionate at 20 pg/mL. After adding 1 mL of 0.01 M KH~POI,the IM-OE in samples was extractedwith 3.0 mL of chloroform; this was followed by centrifugation. The organic phase was evaporated under reduced pressure and the residue, dissolved in 0.1 mL of mobile phase, was injected into a reversed-phase Inertsil ODs-2 column. The acid (6030101, v/v). mobile phase was acetonitrile:methano~H~Oacetic The conditions used in the determination were the same as those for indomethacin. The sensitivity of the methods was 30 ng/mL for indomethacin and 50 ng/mL for IM-OE. Intra- and interday variabilities were 4 0 % . Data Analyses-Pharmacokinetic analysis was performed based on a compartmental description of the observed data, with the assumptions that all elimination and biotransformation processes are first order and that the hydrolysis of the ester bond is mainly carried out in the liver and circulation system by first-order kinetics. The compartmental representation of the pharmacokinetic model is shown in Figure 1.The model includes nine first-order rate constants. The prodrug and indomethacin are absorbed separately from the absorption site with the first-order absorption rate constants kam-oEand kam, respectively. The model includes the conversion rate constant of prodrug to indomethacin in liver and plasma (kl3). The mean plasma concentration-time course data obtained after administration were simultaneouslyfitted to multiexpo-nential equations (see Appendix) by the iterative nonlinear least-squares regression procedure PCNONLIN (SCI Software, ClinTrials, Inc., Lexington, KY), with weighting factors equal to the inverse of square of each observation. The disposition and absorption parameters (al,(12, a3, a4, kzi, k43, V I ,V3, kam-OE,and kam) obtained for each of the data of indomethacin and that of IM-OE and the biotransformation parameters (kid inputted with trial and error were utilized as initial parameters for fitting of the model to data. The mean of all data are presented with their SDs. All computations were carried out with a personal digital computer (SANYO MBC-18NVH4, Osaka, Japan).
Results and Discussion Intravenous Administration-The plasma concentrations after iv administration of indomethacin and IM-OE are shown in Figure 2A. The plasma levels of indomethacin declined in a biexponential manner, as shown in our previous paper.10 However, the plasma decay curve of indomethacin that was formed from the prodrug was found to be monoexponential,and the plasma levels were much lower than those after administration of indomethacin alone (Figure 2A). The plasma concentrations of IM-OE after iv injection also declined in a biexponentialmanner, although the levelswere much lower than those of indomethacin produced. The elimination of IM-OE in plasma was rapid and the concentration was not detected 4 h after administration, suggesting a rapid hydrolysis of the ester bond in plasma and a rapid elimination of the prodrug. The elimination profiles of indomethacin after both dosings were comparatively similar to each other, as was expected. Oral Administration-The plasma concentrationsafter oral administration of indomethacin and IM-OE are depicted in Figure 2B. The plasma levels of indomethacin after IM-OE dosing were very low compared with those obtained after administration of indomethacin alone. The peak levels of indomethacin after IM-OE dosing appeared slightly later than those after indomethacin dosing alone. IM-OEwas not detected in plasma even at the early time periods. These results suggest that the prodrug might not be absorbed in appreciable amounts through the intestine compared with indomethacin, or the large distribution into tissues, although rapid hydrolysis by the first pass effect was observed. Hepatic Uptake of Indomethacin and IM-OE after Oral Administration-Because the plasma concentration-time data after IM-OE dosing suggested high uptake of IM-OE into liver and other tissues, the hepatic uptake was determined 3 h after oral administration of both drugs. The results (Table 1) demonstrate that IM-OE was taken up by the liver much more than indomethacin. Therefore, a reason for lower plasma levels of indomethacin after administration of IM-OE is assumed to be the higher hepatic uptake. This result suggests that a considerablefraction of the prodrug would be absorbed through the intestinal mucosa in the form of ester, and the prodrug may consequentlydiminishthe risk of gastrointestinalirritation based on free indomethacin, as indicated by Whitehouse and Rainsford.7 Analysis of Plasma Drug Concentration Data with the Model-Considering the rapid hydrolysis of the ester bond of Journal of Pharmaceutical Sciences / 35 Vol. 83,No. 1, January 7994
Table 2-Pharmacoklnetlc Parameters for Indomethacln and IM-OE from Four-Compartment Model Parameter a1 82
a3 a4 klOC ki zc k13 k21
k3ac __ k34c
k43 k,lM
unit
Estimateda
hh-' h-1 hh-1 h-l hh-1 h-1 h-' h-I h-' hLI kg LI kg Ll kg LI kg -
-
IMC
LI h l kg L/h/kg Llhlkg
the plasma concentration-time course with the parameters described in the model as follows:
c;p -
SEMb
1.65 0.119 6.64
0.20 0.015 1.20
0.516 0.180 0.498 4.15 2.16 0.947 1.10 6.58 18.1 0.0933 0.179 0.355 0.050 7.24 0.0464 3.39
0.045 0.030 0.039 0.89 0.33 0.137 0.62 1.97 5.3 0.0071 0.011 0.019 0.015 2.01 0.0013 0.95
DT[ k
-a3)
e48t
- a4)e44t]
+ (" ' (a3
(Al)
- a,)
In eq Al:
1
= S[(k30
+ k34 + k43) - d ( k 3 0 + k34 + k43)'
- 4k&,I
(A31
In eqs Al-A3, k34 and 1243 are the transfer rate constants between compartments 3 and 4 and V3 is the distribution volume of the measuring site. For iv IM-OE, the time courses of IM-OE and the formed indomethacin are given as follows:
* The values were obtained by the simultaneous fiing of mean data of each plasma-time course. Standard error for estimate. Secondary estimates using the computer derived parameters. IM-OE in liver and the circulation system and the high uptake of the prodrug in liver, a model was presented for describing the plasma concentration data. Figure 2 showsthe simulation curves calculated according to our model (eqs 1-10, Appendix). It is evident from these results that a good fit was obtained between the observed and calculated curves based on the model for the iv and oral dosings of indomethacin and the prodrug, although only a small disagreement was observed in the plasma IM-OE at low concentrations after iv administration and the simulation curve. The model-dependent pharmacokinetic parameters are summarized in Table 2. The high conversion clearanceof the prodrug (CLIM-OE-IM, 3.39 L/h/kg) suggesta that the ester bond of IMOE was rapidly and extensively hydrolyzed in liver and plasma. Additionally, the high values of Vl and VZ(6.58 and 18.1L/kg, respectively) indicate a large distribution of the prodrug into the central and peripheral tissues. The former may be considerably attributed to the large distribution into liver, as shown in Table 1. The kinetic parameters ( a 3 = a, 6.64 h-l; a 4 = &0.162 h-1, elimination rate constant, k30, 0.498 h-1, and distribution volume of central compartment, V3, 0.093 L/kg) for iv indomethacin calculated in this study were in rough agreement with those10 (a, 2.74 h-1; 8,0.102 h-1, elimination rate constant, 0.227 h-1; and distribution volume of central compartment, 0.121 L/kg)reported by us with Wistar rats ago. As a result, a good fit between the observed and calculated curves validated the model presented. In conclusion,the present results suggest that IM-OE is rapidly hydrolyzed in liver and plasma, and that the prodrug is taken in liver to agreater extent after oral dosing. The model presented is adequately able to describe the time course of the plasma indomethacin and IM-OE following iv and oral administrations of both drug and prodrug. The model may also be applicable to other prodrugs of indomethacin.
Appendix Intravenous Administration-For iv indomethacin (IM), the common two-exponential equations are applied to express 36 / Journal of Pharmticeutical Sciences Vol. 83,No. 1, January 1994
1
a, = s[(klo + k,,
+ kzl) - d ( k l o+ k,, + k,J2 - 4k21k,01 (A71
In eq A4, V 1 is the distribution volume of central compartment of IM-OE. O ral Administration-For oral IM, the plasma concentration of IM is expressed as follows:
In eq A8, k,'M is the absorption rate constant of IM and FIMis the fraction of dose available. The time-course equations for IM-OE and IM after oral administration Of IM-OE are described as
c
;
= D o~s e p ~ I~M ~ oIM-OE ~~k , X
~
Vl (kzl - a,) (a, - a,)(k,'M-OE
- a,)
e*It
+
(kzl - a,) IM-OE -
(a, - a,)(k,
1 (kZ1- k,'(a, - k,'M-OE)(a2 - k,'M*OE)
e*st a,)
e-k.m-O%
+
1
(A91
References and Notes 1. Prabhu, V. G.; Shah, G. F.; Parikh, S. H. Indian J . Pharm. 1967,
(A10)
In eqa A9 and A10, FIM-OE is the fraction of dose absorbed.
29, 100-101. 2. Boardman, P. L.; Hart, F. D. Ann. Rheum. Dis.1967,26,127-132. 3. Tayor, R. T.; Huskisson, E. C.; Whitehouse, G. H.; Hart, F. D.; Traphell, D. H. Br. Med. J . 1968,4, 734-737. 4. Nakamura, M.; Ywhinaka, Y.; Suzuki, H.; Wada, Y. Folia Pharmacol. Japon 1981, 78,511-519. 5. Mishiia, M.; Kobayashi, S.; Abe, S.; Yamato, C. Xenobiotiea 1990, 20,135-146. 6. Dell, V. H.-D.; Doersing, M.; Fischer, W.; Jacobi, H.; Kamp, R.; Kohler, G.; Schollnhammer, G. Arzneim.-ForschlDrug Res. 1980, 30(ZI), 1391-1398. 7. Whitehouse, M.W.;Rainsford, K. D. J . Pharm. Pharmacol. 1980, 32,795-796. 8. Upton, R. A. J. Pharm. Sci. 1975,64,112-114. 9. Kwong, E.; Pillai, G. K.; McErlane, K. M. J. Pharm. Sci. 1982,71, 828-830. 10. Ogiso, T.; Ito, Y.; Iwaki, M.; Atago, H. J . Pharm. Sci. 1989, 78, 319-323.
Journal of pharmaceutical Sciences / 37 Vol. 83, No. 1, January 1994
concentration versus time profiles after indomethacin dosings. Indomethacin, a potent nonsteroidal anti-inflammatory drug, has been widely used in the treatment of rheumatoid arthritis, gout, osteoarthritis, and acute muscle-skeletal disorders. Unfortunately, like other nonsteroidal anti-inflammatory agents, indomethacin carries the risk of side effects, such as gastrointestinal irritation and ulceration.13 Therefore, the prodrugs acemethacin and indomethacin farnesil, were synthesized with the intention of reducing the side effects4 and increasing the tissue distribution: respectively, and have been used for clinical therapy. Although some reports on the basic pharmacokinetics of these prodrugs are published,5.6 as far as we know, there has been no report on pharmacokinetic analyses with a model for describing the blood concentration profiles of the prodrugs and parent drug after dosing. Because it is known that the estrification of acidic antiinflammatory drugs suppresses their gastrotoxicity,7Sumitomo Pharmaceutical Company and we synthesized a prodrug of indomethacin, indomethacin octyl ester (IM-OE), and the pharmacokinetics of this agent was investigated in rats after single intravenous (iv) and oral administrations. To describe the time course of the plasma indomethacin and its prodrug levels, a pharmacokinetic model consisting of central and peripheral compartments with the first-order conversion rate constant of the prodrug to indomethacin was applied. The appropriateness and significance of the models for pharmacokinetic evaluation are discussed.
Experimental Section Materials-Indomethacin (J.P. grade) and IM-OE (mp 55-56 "C; anal.-calcd for Cz~H32ClN04:C, 69.00; H, fi.86; N, 2.98;found C, 69.01; H, 6.82; N, 2.98) were a gift from Sumitomo Pharmaceutical Company (Osaka, Japan). The structure of IM-OE synthesized was confirmed on the basis of IR and lH NMR spectra. Mefenamic acid and 8-estradial dipropionate, internal standards for the HPLC assay, were obtained from Sigma Chemical Company (St. Louis, MO). All other chemicals and solvents used were of reagent grade or HPLC quality. @
Abstract published in Aduance ACS Abstracts, November 15,1993.
34 /Journal of Pharmaceutical Sciences Vot. 83, No. 1, January 1994
4
i.v.
Abstract 0 A prodrug of indomethacin, indomethacin octyl ester (IMOE), was synthesized and its pharmacokinetics was investigated in rat. To describe the time course of the plasma indomethacin and IM-OE after intravenous ( i ) and oral admlnistrations, a pharmacokinetic model with four compartmentswas developed. Indomethacin rapidly appeared in plasma after iv administration of IM-OE and declined in a monoexponential manner, with a rapid decline and low plasma levels of IM-OE. The plasma concentrations of indomethacin after oral administration of IM-OE were much lower than those after oral administration of indomethacin. The high concentrations of IM-OE compared with indomethacin were detected in liver 3 h after oral dosing of the prodrug, although IM-OE was not detected in plasma. A good fit was obtained between the observed and calculated curves based on the model, which includes a conversion rate constant of IM-OE to indomethacin for both iv and oral dosings of IM-OE. Additionally, the model could successfully describe the plasma
/
i.v.
1-Four-compartment pharmacokinetic models. Key: (IM) indomethacin; (IM-OE) indomethacin octyl ester; (klo and k3,,)the firstorder elimination rate constants for IM-OE and IM, respectively; (k13) the first-order conversion rate constant of IM-OE to IM; (k12,kpl,k3,, and ka) the first-order kinetic constants for the distribution between central and the first-order absorption rate peripheral compartments;(kal- and kaIM) constants for IM-OE and IM, respectively. Figure
Experimental Animals-Male W i s h rats, weighing200-280g, were used. The animals, maintained on an MF diet (a bleeding feed, Oriental Yeast Company, Tokyo, Japan) for 3-4 days prior to the experiment, were divided at random into two groups, each consisting of 3-5 rats. One group was treated with indomethacin and the second group was administered IM-OE. On the day before the experiment, the rat jugular vein was cannulated with silicon tubing.8 Intravenous and Oral Administration-Indomethacin (5 mg/kg) and IM-OE ( 5 mg/kg, indomethacin equivalent) were administered iv as a solution in a mixture of NJV-dimethy1formamide:ethanol:water( 6 22, v/v). Indomethacin (7.6 mg/kg) and the prodrug (7.6 mg/kg, indomethacin equivalent) were administered orally as an aqueous suspension in 1.5% acasia. A 0.2-mL blood sample was collected periodically after dosing through the cannula for a total time period. The plasma was separated immediately by centrifugation and stored frozen until the time of assay. Hepatic Uptake-Rats received indomethacin or IM-OE orally at the same dose as described for the above oral dosing. At 3 hafter dosing, the liver was removed from the animals immediately after sacrifice. The liver,perfused with ice-cold0.9% NaCl toremove blood, was homogenized in ice-cold 10 mM phosphate buffer (pH 7.4). The drugs in the homogenate (1.0 mL) were extracted with methanol (2.0mL) containing the internal standard. After centrifugation, the supernatant was stored frozen until assay. D e t e r m i n a t i o n of I n d o m e t h a c i n a n d IM-OE-Indomethacin and IM-OE in plasma or homogenate was determined by the HPLC method of Kwong et aL9with slight modifications. A 50-pLaliquot of plasma was mixed with 150 pL of methanol containing 8 pg/mL of mefenamic acid as an internal standard, and the mixture was centrifuged a t 15 OOO rpm. The supernatant was filtered through the membrane filter (0.45 pm, Ekikrodisc 3CR, Gelman Sciences Japan, Tokyo, Japan). The filtrate was injected into a reversed-phase Inertail ODs-2column (5 pm, 4.6 mm X 15cm; GL Sciences Inc., Tokyo, Japan) with a Shimadzu liquid chromatograph (model LC-fiA) equipped with a UV spectrophotometer (modelSPD-6AV). The mobile phase was acetonitrile:methanok Hz0:acetic acid (651025:1, v/v), pumped at a flow rate of 1.0 mL/min. Detection was at 254 nm. For the determination of IM-OE, a 100-pL aliquot of sample was mixed with 300 pL of methanol containing
0022-3549/94/ 1200-34$04.50/0
0 1994, American Chemical Society and American Pharmaceutical Association
A
,-
B
0.01
0
0.01
2
4
6
8
1
0
1
2
I i 0
2
4
6
8 1 0 1 2
Time after administration (h) Flgure 2-Plasma concentration-time curves for lndomethacin and IM-OE after (A) iv administration at a dose of 5 mg InL-methacin equivalentlkg and (B)oral administrationat a dose of 7.6 mg indomethacin equivalentlkg. Key: (R) indomethacin; (A)indomethacinafter IM-OE(0)IM-OE. Points
and vertical bars represent the mean and SD (n = 3-4), respectively. Solid lines are the simulation curves calculated with the model descrlbed In Figure 1 and the parameters listed in Table 11. Table 1-Hepatlc Plasma and Blood Concentratlons at 3 h after Oral Admlnlstratlon of Indomethacln or IM-OE.
Concentration ~~~
Drug IM
IM-OE IM after IM-OE
~~
Liver, nmol/g
Plasma, nmol/mL
nmol/mL
11.86 f 4.84 131.52 f 48.8 2.88 f 0.48
47.52 f 3.47 0 1.22 f 0.06
26.57 f 4.84 0 0.58 f 0.17
Blood,
* Dose, 7.6 mg/kg (indomethacinequivalent);values are the mean f SD (n = 3). fl-estradiol dipropionate at 20 pg/mL. After adding 1 mL of 0.01 M KH~POI,the IM-OE in samples was extractedwith 3.0 mL of chloroform; this was followed by centrifugation. The organic phase was evaporated under reduced pressure and the residue, dissolved in 0.1 mL of mobile phase, was injected into a reversed-phase Inertsil ODs-2 column. The acid (6030101, v/v). mobile phase was acetonitrile:methano~H~Oacetic The conditions used in the determination were the same as those for indomethacin. The sensitivity of the methods was 30 ng/mL for indomethacin and 50 ng/mL for IM-OE. Intra- and interday variabilities were 4 0 % . Data Analyses-Pharmacokinetic analysis was performed based on a compartmental description of the observed data, with the assumptions that all elimination and biotransformation processes are first order and that the hydrolysis of the ester bond is mainly carried out in the liver and circulation system by first-order kinetics. The compartmental representation of the pharmacokinetic model is shown in Figure 1.The model includes nine first-order rate constants. The prodrug and indomethacin are absorbed separately from the absorption site with the first-order absorption rate constants kam-oEand kam, respectively. The model includes the conversion rate constant of prodrug to indomethacin in liver and plasma (kl3). The mean plasma concentration-time course data obtained after administration were simultaneouslyfitted to multiexpo-nential equations (see Appendix) by the iterative nonlinear least-squares regression procedure PCNONLIN (SCI Software, ClinTrials, Inc., Lexington, KY), with weighting factors equal to the inverse of square of each observation. The disposition and absorption parameters (al,(12, a3, a4, kzi, k43, V I ,V3, kam-OE,and kam) obtained for each of the data of indomethacin and that of IM-OE and the biotransformation parameters (kid inputted with trial and error were utilized as initial parameters for fitting of the model to data. The mean of all data are presented with their SDs. All computations were carried out with a personal digital computer (SANYO MBC-18NVH4, Osaka, Japan).
Results and Discussion Intravenous Administration-The plasma concentrations after iv administration of indomethacin and IM-OE are shown in Figure 2A. The plasma levels of indomethacin declined in a biexponential manner, as shown in our previous paper.10 However, the plasma decay curve of indomethacin that was formed from the prodrug was found to be monoexponential,and the plasma levels were much lower than those after administration of indomethacin alone (Figure 2A). The plasma concentrations of IM-OE after iv injection also declined in a biexponentialmanner, although the levelswere much lower than those of indomethacin produced. The elimination of IM-OE in plasma was rapid and the concentration was not detected 4 h after administration, suggesting a rapid hydrolysis of the ester bond in plasma and a rapid elimination of the prodrug. The elimination profiles of indomethacin after both dosings were comparatively similar to each other, as was expected. Oral Administration-The plasma concentrationsafter oral administration of indomethacin and IM-OE are depicted in Figure 2B. The plasma levels of indomethacin after IM-OE dosing were very low compared with those obtained after administration of indomethacin alone. The peak levels of indomethacin after IM-OE dosing appeared slightly later than those after indomethacin dosing alone. IM-OEwas not detected in plasma even at the early time periods. These results suggest that the prodrug might not be absorbed in appreciable amounts through the intestine compared with indomethacin, or the large distribution into tissues, although rapid hydrolysis by the first pass effect was observed. Hepatic Uptake of Indomethacin and IM-OE after Oral Administration-Because the plasma concentration-time data after IM-OE dosing suggested high uptake of IM-OE into liver and other tissues, the hepatic uptake was determined 3 h after oral administration of both drugs. The results (Table 1) demonstrate that IM-OE was taken up by the liver much more than indomethacin. Therefore, a reason for lower plasma levels of indomethacin after administration of IM-OE is assumed to be the higher hepatic uptake. This result suggests that a considerablefraction of the prodrug would be absorbed through the intestinal mucosa in the form of ester, and the prodrug may consequentlydiminishthe risk of gastrointestinalirritation based on free indomethacin, as indicated by Whitehouse and Rainsford.7 Analysis of Plasma Drug Concentration Data with the Model-Considering the rapid hydrolysis of the ester bond of Journal of Pharmaceutical Sciences / 35 Vol. 83,No. 1, January 7994
Table 2-Pharmacoklnetlc Parameters for Indomethacln and IM-OE from Four-Compartment Model Parameter a1 82
a3 a4 klOC ki zc k13 k21
k3ac __ k34c
k43 k,lM
unit
Estimateda
hh-' h-1 hh-1 h-l hh-1 h-1 h-' h-I h-' hLI kg LI kg Ll kg LI kg -
-
IMC
LI h l kg L/h/kg Llhlkg
the plasma concentration-time course with the parameters described in the model as follows:
c;p -
SEMb
1.65 0.119 6.64
0.20 0.015 1.20
0.516 0.180 0.498 4.15 2.16 0.947 1.10 6.58 18.1 0.0933 0.179 0.355 0.050 7.24 0.0464 3.39
0.045 0.030 0.039 0.89 0.33 0.137 0.62 1.97 5.3 0.0071 0.011 0.019 0.015 2.01 0.0013 0.95
DT[ k
-a3)
e48t
- a4)e44t]
+ (" ' (a3
(Al)
- a,)
In eq Al:
1
= S[(k30
+ k34 + k43) - d ( k 3 0 + k34 + k43)'
- 4k&,I
(A31
In eqs Al-A3, k34 and 1243 are the transfer rate constants between compartments 3 and 4 and V3 is the distribution volume of the measuring site. For iv IM-OE, the time courses of IM-OE and the formed indomethacin are given as follows:
* The values were obtained by the simultaneous fiing of mean data of each plasma-time course. Standard error for estimate. Secondary estimates using the computer derived parameters. IM-OE in liver and the circulation system and the high uptake of the prodrug in liver, a model was presented for describing the plasma concentration data. Figure 2 showsthe simulation curves calculated according to our model (eqs 1-10, Appendix). It is evident from these results that a good fit was obtained between the observed and calculated curves based on the model for the iv and oral dosings of indomethacin and the prodrug, although only a small disagreement was observed in the plasma IM-OE at low concentrations after iv administration and the simulation curve. The model-dependent pharmacokinetic parameters are summarized in Table 2. The high conversion clearanceof the prodrug (CLIM-OE-IM, 3.39 L/h/kg) suggesta that the ester bond of IMOE was rapidly and extensively hydrolyzed in liver and plasma. Additionally, the high values of Vl and VZ(6.58 and 18.1L/kg, respectively) indicate a large distribution of the prodrug into the central and peripheral tissues. The former may be considerably attributed to the large distribution into liver, as shown in Table 1. The kinetic parameters ( a 3 = a, 6.64 h-l; a 4 = &0.162 h-1, elimination rate constant, k30, 0.498 h-1, and distribution volume of central compartment, V3, 0.093 L/kg) for iv indomethacin calculated in this study were in rough agreement with those10 (a, 2.74 h-1; 8,0.102 h-1, elimination rate constant, 0.227 h-1; and distribution volume of central compartment, 0.121 L/kg)reported by us with Wistar rats ago. As a result, a good fit between the observed and calculated curves validated the model presented. In conclusion,the present results suggest that IM-OE is rapidly hydrolyzed in liver and plasma, and that the prodrug is taken in liver to agreater extent after oral dosing. The model presented is adequately able to describe the time course of the plasma indomethacin and IM-OE following iv and oral administrations of both drug and prodrug. The model may also be applicable to other prodrugs of indomethacin.
Appendix Intravenous Administration-For iv indomethacin (IM), the common two-exponential equations are applied to express 36 / Journal of Pharmticeutical Sciences Vol. 83,No. 1, January 1994
1
a, = s[(klo + k,,
+ kzl) - d ( k l o+ k,, + k,J2 - 4k21k,01 (A71
In eq A4, V 1 is the distribution volume of central compartment of IM-OE. O ral Administration-For oral IM, the plasma concentration of IM is expressed as follows:
In eq A8, k,'M is the absorption rate constant of IM and FIMis the fraction of dose available. The time-course equations for IM-OE and IM after oral administration Of IM-OE are described as
c
;
= D o~s e p ~ I~M ~ oIM-OE ~~k , X
~
Vl (kzl - a,) (a, - a,)(k,'M-OE
- a,)
e*It
+
(kzl - a,) IM-OE -
(a, - a,)(k,
1 (kZ1- k,'(a, - k,'M-OE)(a2 - k,'M*OE)
e*st a,)
e-k.m-O%
+
1
(A91
References and Notes 1. Prabhu, V. G.; Shah, G. F.; Parikh, S. H. Indian J . Pharm. 1967,
(A10)
In eqa A9 and A10, FIM-OE is the fraction of dose absorbed.
29, 100-101. 2. Boardman, P. L.; Hart, F. D. Ann. Rheum. Dis.1967,26,127-132. 3. Tayor, R. T.; Huskisson, E. C.; Whitehouse, G. H.; Hart, F. D.; Traphell, D. H. Br. Med. J . 1968,4, 734-737. 4. Nakamura, M.; Ywhinaka, Y.; Suzuki, H.; Wada, Y. Folia Pharmacol. Japon 1981, 78,511-519. 5. Mishiia, M.; Kobayashi, S.; Abe, S.; Yamato, C. Xenobiotiea 1990, 20,135-146. 6. Dell, V. H.-D.; Doersing, M.; Fischer, W.; Jacobi, H.; Kamp, R.; Kohler, G.; Schollnhammer, G. Arzneim.-ForschlDrug Res. 1980, 30(ZI), 1391-1398. 7. Whitehouse, M.W.;Rainsford, K. D. J . Pharm. Pharmacol. 1980, 32,795-796. 8. Upton, R. A. J. Pharm. Sci. 1975,64,112-114. 9. Kwong, E.; Pillai, G. K.; McErlane, K. M. J. Pharm. Sci. 1982,71, 828-830. 10. Ogiso, T.; Ito, Y.; Iwaki, M.; Atago, H. J . Pharm. Sci. 1989, 78, 319-323.
Journal of pharmaceutical Sciences / 37 Vol. 83, No. 1, January 1994