Blood or plasma concentration and excretion of SK-827 were investigated in male rats after single intravenous injection or after constant rate intravenous infusion of ¹²⁵I-SK-827. Further, the accumulation of SK-827 was investigated in male rats after repeated intravenous injections of ¹²⁵I-SK-827.
1. After single intravenous injection of ¹²⁵I-SK-827 at doses of 1.67 ?? 15× 10^-3 PNAU (p-nitroaniline unit) /kg, the trichloroacetic acid insoluble (TCA-insoluble) radioactivity in plasma decreased rapidly. Plasma TCA-insoluble radioactivity declined with half-lives of 4 min from 2 min to 15 min, 23 min from 15 min to 60 min and 131 min from 60 min to 240 min after administration at the dose of 15×10^-3 PNAU/kg.
2. During single constant rate intravenous infusion for 30 min at doses of 1.67 ?? 15×10^-3 PNAU/kg, the TCA-insoluble radioactivity in plasma increased rapidly and reached a maximum at 29 min after starting the infusion. Following termination of the infusion, plasma TCA-insoluble radioactivity declined in similar fashion as after intravenous injection. Concentration at 29 min after starting the infusion (C_{29 min}) and AUC_{0-60 min} of plasma TCA-insoluble radioactivity increased proportionally, indicating that the plasma levels of ¹²⁵I-SK-827 fitted to the linear pharmacokinetic model within the dose range studied.
3. Within 120 hr after a single intravenous injection or constant rate intravenous infusion at a dose of 5×10^-3 PNAU/kg, essentially 100% of the administered radioactivity was excreted in urine and feces. Most of it was excreted in urine within 24 hr.
4. During repeated intravenous injections at a dose of 5×10^-3 PNAU/kg, the plasma TCA-insoluble radioactivity at 1 hr after each injection remained almost constant from the first day. There were no significant differences in the pharmacokinetic parameters of plasma TCA-insoluble radioactivity after single injection from those after the 7-day repeated injections. Excretion of radioactivity in urine and feces was nearly the same throughout the period of 7-day repeated intravenous injections. Within 120 hr after the last dose, 99% of the administered radioactivity was excreted.

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The distribution, accumulation and urinary or biliary excretion of SK-827 were investigated in male rats after single or repeated intravenous administrations of ¹²⁵I-SK-827.
1. The blood trichloroacetic acid insoluble (TCA-insoluble) radioactivity after a single intravenous injection of ¹²⁵I-SK-827 at a dose of 5×10^-3 PNAU (p-nitroaniline unit) /kg decreased rapidly within 1 hr, but thereafter in more gradual rate. At 5 min ?? 1 hr after injection, the TCA-insoluble radioactivity in the liver, adrenal, spleen, kidney and bone marrow were higher than that in the blood. At 5 min after injection, more than 90% of administered radioactivity was concentrated in the liver. The TCA-insoluble radioactivity in these tissues decreased rapidly within 1 hr, and the elimination was similar to that in the blood thereafter. Distribution profiles of radioactivity after a single constant rate intravenous infusion did not differ from those after intravenous injection.
2. TCA-insoluble radioactivity in the liver, kidney and spleen at 1, 3 and 24 hr after 4-day repeated intravenous injections of ¹²⁵I-SK-827 at the dose of 5×10^-3 PNAU/kg were 2 ?? 10 times higher than those observed after a single dose but the TCA-insoluble radioactivity in these tissues were almost constant from 4-day to 7-day repeated dosing.
3. Within 24 hr after a single intravenous injection of ¹²⁵I-SK-827 at a dose of 5×10^-3 PNAU/kg, more than 90% of administered radioactivity was excreted into urine. However, urinary excretion of TCA-insoluble radioactivity was only 1% of administered dose within 3 hr after injection.
4. Within 48 hr after a single intravenous injection of ¹²⁵I-SK-827 at a dose of 5×10^-3 PNAU/kg, about 20% of administered radioactivity was excreted into bile. However, biliary excretion of TCA-insoluble radioactivity was only 3% of administered dose within 3 hr after injection.

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The feto-placental transfer of ¹²⁵I-SK-827 or excretion with milk were investigated in rats on 12 or 18 days of gestation or lactating rats after a single intravenous bolus injection of ¹²⁵I-SK-827.
1. At 2 min after a single intravenous injection of ¹²⁵I-SK-827 at a dose of 5×10^-3 PNAU (p-nitroaniline unit) /kg, the radioactivity in the fetus was 125 ?? 840 times lower than that in maternal blood, but most of that was TCA-soluble. This result suggested that ¹²⁵I-SK-827 administered to pregnant rats was not transfered to the fetus.
2. After a single intravenous injection of ¹²⁵I-SK-827 at a dose of 5×10^-3 PNAU/kg, the TCA-insoluble radioactivity was detected in the milk at 5 hr and reached a maximum level at 24 hr. However, the properties of TCA insoluble radioactivity in the milk after injection were completely different from those of ¹²⁵I-SK-827 added to blank milk. TCA-insoluble radioactivity in the milk was considered to be endogenous high molecular weight compounds into which TCA-soluble radioactivity produced by degradation of ¹²⁵I-SK-827 was incorporated.

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We investigated the inhibition of SK-827 by human, dog and rat plasma inhibitors using gel filtration column chromatography (Sephacryl S-300 Superfine).
1. In human plasma, there was one inhibitor which was revealed to be α₁-antitrypsin (α₁-AT) based on its enzymatic and immunological propaerties and the elution volume of the complex on gel filtration column chromatography. After 2 min incubation of ¹²⁵I-SK-827 with human plasma at 37°C, 28% of ¹²⁵I-SK-827 remained free, but was not detected 30 min later.
2. In dog plasma, there were two inhibitors which were described to be α₁-AT and α₂-macrogrobulin (α₂-M) base on their enzymatic and immunological properties and the elution volume of the complex. The ratio of ¹²⁵I-SK-827-α₂-M complex and ¹²⁵I-SK-827-α₁-AT complex in the plasma was about 30 :70. After 5 min incubation of ¹²⁵I-SK-827 with dog plasma at 37°C, 36% of ¹²⁵I-SK-827 remained free, but was not detected 60 min later.
3. In rat plasma, there were two inhibitors which were revealed to be α₁-AT and α₂-M base on their enzymatic and immunological properties and the elution volume of the complex. The ratio of ¹²⁵I-SK-827-α₂-M complex and ¹²⁵I-SK-827-α₁-AT complex in the plasma was about 92 : 8. After 2 min incubation of ¹²⁵I-SK-827 with rat plasma at 37°C, only 11% of ¹²⁵I-SK-827 remained free.

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The metabolism of SK-827 was s tudied by gel filtration of plasma, urine, bile and liver homogenate after intravenous injection of ¹²⁵I-SK-827 in rats.
1. Gel filtration (Sephacryl S-300 superfine) chromatograms of radioactivity in plasma, obtained after intravenous injection of ¹²⁵I-SK-827, demonstrated that the radioactivity in plasma was mainly in the form of ¹²⁵I-SK-827 bound to an inhibitor and free ¹²⁵I-SK-827. The elimination half-lives of ¹²⁵I-SK-827-inhibitor complex and free ¹²⁵I-SK-827 in the plasma were calculated to be 5.8 min and 39 min, respectively.
2. Gel filtration chromatograms of radioactivity in the liver demonstrated that the radioactivity in the liver was mainly in the form of ¹²⁵I-SK-827 bound to an inhibitor and free ¹²⁵I-SK-827. The half-lives of ¹²⁵I-SK-827-inhibitor complex and free ¹²⁵I-SK-827 in the liver were calculated to be 6.6 min and 10.2 min, respectively.
3. Appearance of trichloroacetic acid soluble (TCA-soluble) radioactivity during incubation at 37°C for 2 hr of the liver slices obtained at 5, 15 and 30 min after intravenous injection of ¹²⁵I-SK-827 were 9%, 39% and 51% of the radioactivity present in liver slice before incubation, respectively. This result suggests that the degradation of injected ¹²⁵I-SK-827 occurred in the liver. It was found that TCA-soluble degradation product in liver slices was mainly (90%) inorganic iodine.
4. Gel filtration chromatogram of radioactivity in urine within 1 hr after injection demonstrated that the radioactivity excreted to the urine was in the form of free ¹²⁵I-SK-827 (16.8% of total radioactivity) and degraded product. No radioactivity related to the ¹²⁵I-SK-827-inhibitor complex was observed.
5. Gel filtration chromatogram of radioactivity in bile at 1 hr after injection demonstrated that the radioactivity in bile was in the form of ¹²⁵I-SK-827 bound to inhibitors (51.2%), free ¹²⁵I-SK-827 (34.3%) and degraded product (14.6%).
6. From these results, it became clear that ¹²⁵I-SK-827 administered intravenously was bound by an inhibitor, and complex was cleared from the plasma by the liver. The complex taken up by the liver was rapidly degraded into low molecular weight compounds. TCA-insoluble radioactivity detected in the liver at 24 hr after administration were considerded to be endogenous high molecular weight compounds into which TCA-soluble radioactivity produced by degradation of ¹²⁵I-SK-827 was incorporated.

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Pharmacokinetics of SK-827 was studied in dogs and rabbits after intravenous injection or after constant rate intravenous infusion of SK-827. Plasma levels of SK-827 were determined by sandwich enzymeimmunoassay.
1. After a single intravenous injectio n of SK-827 to dogs at doses of 0.649 ?? 6.25×10^-3 PNAU (p-nitroaniline unit) /kg, the plasma levels declined bi-exponentially. The plasma half-lives of the rapid phase and slower phase after intravenous injection of SK-827 (6.25×10^-3 PNAU/kg) were about 3 min (t_1/2α) and 40 min (t_1/2β), respectively.
2. During single constant rate intravenous infusion of SK-827(1.67 ?? 15×10^-3 PNAU/kg for 30 min) to dogs, the plasma levels increased rapidly. After stopping the infusion of SK-827, the plasma levels declined bi-exponentially similar to that after intravenous injection. The AUC_0-∞ values after intravenous infusion of SK-827 and C₂₉ (concentration at 29 min after starting the infusion) were proportional to the dose and there were no dose-related changes in the mean residence time (MRT_inf), plasma clearance (Clp) and steady-state distribution volume (Vdss), indicating that the plasma levels of SK-827 can be described by the linear pharmakokinetic model.
3. During repeated injec tions of SK-827 to dogs at the dose of 5×10^-3 PNAU/kg for 7 days, the plasma levels at 90 min after each injection remained almost constant from the first day. There were no differences in the plasma half-lives of β phase between single and repeated injection (for 4 or 7 days)
4. After a single intravenous injection of SK-827 to rabbits at do ses of 1.67 ?? 15×10^-3 PNAU/kg, the plasma levels declined bi-exponentially. The plasma half-lives of the rapid and slower phase were about 5 min (t_1/2α) and 120 min (t_1/2β) . There were no changes in the values of MRT, Clp and Vdss, indicating that the plasma levels of SK-827 followed the linear pharmakokinetic model.
5. During a single constant rate intravenous infusion of SK-827(5×10^-3 PNAU/kg for 30 min) to rabbits, the plasma levels increased rapidly, and declined bi-exponentially after stopping the infusion. The Vdss values and Clp values after intravenous infusion were not different from those after intravenous injection.

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The absorption, distribution and excretion of 4-methoxyphenyl 4-(3, 4, 5-trimethoxybenzyl)-1-piperzineacetate monofumarate monohydrate (KB-5492) were studied in rats after a single oral administration of ¹⁴C-KB-5492.
The plasma radioactivity reached a maximum level at 0.21 h and then declined with terminal half-life of 9.0 h at a dose of 2.0 mg/kg. Maximum plasma concentration (Cmax) and the area under the plasma concentration-time curve (AUC) increased proportionally to administered dose, up to 10.0 mg/kg.
The extent of absorption after oral administration of KB-5492 was considered to be approximately 30% of the dose. The in situ experiment using the ligated loops of various parts of the gastrointestinal tract revealed that KB-5492 was absorbed from the stomach in 10%, and from the small intestine in about 40% within 1 h.
The cumulative excretion of radioactivity in urine and feces within 72 h were 25% and 73% of the dose, respectively. Most of the radioactivity was excreted in urine and feces within 24 h. Nearly all of the radioactive compounds in the urine and bile might be a 4-(3, 4, 5-trimethoxybenzyl)-1-piperazineacetic acid.
High levels of the radioactivity were observed in the liver, kidney and gastrointestinal tracts, while the radioactivity in the other tissue was lower than that in plasma. In the stomach, radioactivity might be located in the surface of gastric wall.

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The absorption, distribution and excretion of 4-methoxyphenyl 4-(3, 4, 5-trimethoxybenzyl)-1-piperazineacetate monofumarate monohydrate (KB-5492) and its effects on liver microsomal drug-metabolizing enzyme activities were studied in rats after multiple oral administration of ¹⁴C-KB-5492, respectively.
After multiple administration of ¹⁴C-KB-5492, the plasma and tissue radioactivity reached the steady state after the 7th to 14th administration, however no remarkable accumulation was observed in any tissues.
Within 96 h after the 14th administration, the cumulative excretion of radioactivity into urine and feces were 34.6% and 61.8% of cumulative dose, respectively and the ratios of cumulative excretion into urire and feces were constant during the multiple administration.
Gastrointestinal absorption of KB-5492 seemed to be influenced by food intake.
After multiple oral administration of KB-5492 (50, 250 mg/kg/day) to rats, there was no influence on liver microsomal drug-metabolizing enzyme activities.

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Plasma concentration, distribution and excretion of radioactivity were investigated after a single and repeated intravenous administration of 5 mg/kg of ¹⁴C-milrinone to male rats.
1. After a single dosing, the plasma concentrations declined bi-phasically with the elimination half life (t_1/2) of 3.5 hr; the area under the plasma concentration-time curve (AUC), volume of distribution (V_dss) and total clearance (CL_total) were 7.19 μg equiv. of milrinone·hr/ml, 873 ml/kg and 11.6 ml/min/kg, respectively. Plasma concentration-time curve of unchaged drug was similar to that of radioactivity. The AUC, t_1/2, V_dss and CL_total for unchaged drug were 3.2 hr, 7.35 μg·hr/ml, 727 ml/kg and 11.3 ml/min/kg, respectively.
2. Radioactivity was distributed rapidly to various tissues, showing peak levels 15 min after a single administration. At 15 min, levels of radioactivity in the kidney was approximately 4 times higer than that in the plasma, while concentrations in other tissues were similar to or lower than plasma concentrations. Radioactivity disappeared rapidly from most tissues, except the skin. After repeated administration of ¹⁴C-milrinone for 7 days, no radioactivity was accumulated in any tissues measured however, disappearance from the skin was relative ly slow. On the whole body autoradiogram 24 hr after repeated dosing, revealed that radioactivity was not observed in skin but was seen in the hair. At 72 hr, radioactivity was detected in the hair only, but the level was low.
3. Within 168 hr after a single dosing, 79.2% and 21.9% of dosed radioactivity were excreted in urine and feces, respectively, and 101.1% of the dose was recovered by both routes. After repeated dosing with ¹⁴C-milrinone for 7 days, cumulative excretion rates of radioactivity in urine and feces were 74.9% and 21.4%, being similar to those after a single dosing. Within 48 hr after administration of ¹⁴C-milrinone to bile duct-cannulated rats, 65.1%, 10.4% and 18.9% of dosed radioactivity were excreted in the urine, feces and bile, respectively, indicating that renal excretion was the main excretion route of milrinone.

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