Pharmacokinetics And Toxicokinetics Study Of Hydrochloric Ranolazine

Ranolazine, as a novel anti-anginal drug being invented by USA CV Therapeutics company for the first time, has been verified to have the extent anti-anginal effect in many vitro and vivo models. This paper was designed to evaluate the pharmacokinetics and toxicokinetics features of ranolazine in animals, with the dynamic process of toxicity of RNZ described, so as to seek for the toxic target organs and toxic mechanisms, which can also provide foundation for further mechanism study . 1.The establishment of analytical method of Ranolazine in biological matrixObjective To develop a rapid and sensitive liquid-chromatography -tandem mass spectrometry (HPLC-MS/MS) method for determination of the ranolazine in dog plasma and tissue homogenates using lorazepam as an inernal standard. Method Separation was obtained on a Eclipse XDB —C8 (5μm, 4.6×150mm) with multiple reaction monitoring (MRM) mode , using the transitions of m/z 428.4→279.3 and m/z 321.3→303.3 to quantify the ranolazine and lorazepam respectively. The mobile phase consisted of acetronitrile-water-formic acid (70:30:0.1,v/v), at a flow rate of 0.8 mL·min^-1. The chromatographic system used provided good separation of the compound without interfering peaks from endogenous substances. The calibration curves were linear in each sample range with correlation coefficient above 0.99. The precision of intra-day and inter-day were evaluated by analysis of variance with the result less than 15% . Conclusion The established method is suitable for ranolazine pharmacokinetics and toxicokinetics studies. 2. The pharmacokinetics study of ranolazine2.1 The pharmacokinetics study of ranolazine in beagle dogs by a single dosageObjective To investigate pharmacokinetics and toxicokinetics character -ristics of ranolazine in beagle dogs by a single dosage. Method 24 adult and healthy beagle dogs were randomly allocated to four groups , each of which were administrated with 12.5,25,50 and 120 mg·kg^-1 ranolazine respectively. Following a single oral administration of ranolazine to beagle dogs, The blood samples were collected at different time after dosing. Blood samples for determination of plasma ranolazine concentration were obtained just prior to drug administration(0 hour) and at 0.25 , 0.5, 0.75, 1.0, 1.5, 2.0 , 3.0, 4.0, 6.0, 8.0, 12.0 and 24.0 hours after administration. All collected blood samples were determined by HPLC/MS/MS method described as above. Pharmacokinetics and toxicokinetics parameters calculations were carried out using compartmental and non-compartmental analysis method. Result After a single administration of 12.5, 25, 50 and 20mg·kg^-1 ranolazine , the average peak plasma concentrations of dogs were 1.71±0.402, 4.241±1.082, 9.251 ±3.692, 28.45±12.28mg·L^-1, obtained at 1.417±0.376, 0.708±0.246, 1.083±0.204, 0.75±0.224 h respectively. The estimated elimination half-life was 4.167±0.636, 5.353±3.38, 4.341±1.427, 4.861±2.347 h, respectively. The average AUC_<o-t> of the four groups were 3.803±1.369, 8.488±1.728, 14.453±3.408, 49.684±17.11 mg·h·L^-1. Conclusion It was shown by the plasma concentration-time data that ranolazine was quickly absorbed after administration, and the peak concentration was generally obtained at about 1 hour after dosage. The average half-life is about 4～5 h, and the plasma concentrations of ranolazine were lower than detectability 24 h post dosage. The AUC_0-t and C_max of the 12.5,25,50, 120mg·kg^-1 doses was positive correlated with the doses, the correlation coefficient is 0.9847.2. 2 The phannacokinetics study of ranolazine in rats by a single dosage Objective To investigate pharmacokinetics and toxicokinetics character-ristics of ranolazine in rats by a single dosage. Method 128 adult and healthy beagle dogs were randomly allocated to four groups , each of which were administrated with 12.5,25,50 and 120mg·kg^-1 ranolazine respectively. Following a single oral administration of ranolazine to rats, The blood samples were collected at different time after dosing. Blood samples for determination of plasma ranolazine concentration were obtained just prior to drug administration (0 hour) and at 0.25 , 0.5, 0.75, 1.0, 1.5, 2.0 , 3.0, 4.0, 6.0, 8.0, 12.0 and 24.0 hours after administration. All collected blood samples were determined by HPLC/MS/MS method described as above. Pharmacokinetics and toxicokinetics parameter calculations were carried out using compartmental and non-compartmental analysis method. Result Following a single oral administration of ranolazine , the pharmacokinetics feature of ranolazine are different between male and female rats. In male rats, The peak concentrations are 0.8558, 1.94, 5.556, 7.9060mg·l^-1, obtained atl, 0.25, 0.5h respectively, T_1/2 are 0.799 , 1.545, 1.645, 2.737 h, AUC are 1.618, 2.875, 13.877, 33.64 mg·h·L^-1. In female rats, the C_maxare : 3.427, 558, 21.78, 32.566 mg·l^-1, obtained at 0.5, 0.5, 0.75, 6h respectively, T_1/2 are 2.356, 3.15, 3.81, 4.951 h; AUC_0-t are 13.899, 31.829, 113.826, 201.185 mg·h·L^-1. Conclusion It was shown by the plasma concentration-time data that ranolazine was quickly absorbed after administration, and generally, the peak concentration was obtained at about 0. 25¹ hour after administration. Rnoalzine is more exposured in female rats, whose AUC is 5. 98～11.07 times bigger than male's; and the t_1/2 in female is also 2³ times longer than the male one. The AUCs of different doses in both male and female rats are was positive correlated with the doses, the correlation coefficients are 0.9903 and 0.98 respectively.2. 3. The tissue distribution study of ranolazine in rats Objective To investigate the distribution of ranolazine in rats after oral administration. Method Rats were killed at 0.25, 0.5, 1, 3, 6, 12 and 24 h after oral administration of ranolazine 50mg·kg^-1. Tissues were taken out and made into homogenates and the concentrations of ranolazine in the tissue were determined by HPLC/MS/MS. Result The concentrations of ranolazine in plasma were obviously higher than in tissues. The concentration of ranolazine in all the rat tissues were detectable 0.25 h after oral administration. In male rats, concentrations descending in a order of stomach, liver, kidney, lung, heart, spleen, muscle , small intestine, testis, fat, brain, adrenal gland 0. 5 h after oral adminis -tration, concentrations descending in a order of stomach, small intestine, kidney, liver, lung, Heart, muscle, adrenal gland, testis, brain, fat 1 h post dosing, then degraded gradually with time, except kidney, descending in order of kidney, heart, stomach, liver, lung, spleen, small intestine, adrenal gland, fat, muscle, testis, brain 6 h after administration. In female rats, concentrations descending in a order of stomach, liver, kidney, lung, small intestine, heart, spleen, muscle, ovary, adrenal gland , fat, brain 0.5 h after oral administration, concentrations descending in a order of stomach, kidney, small intestine, spleen, liver, Heart, ovary, adrenal gland, Lung, muscle, fat, brain 1 h after administration, then degraded gradually with time , except kidney and lung, descending in order of kidney, liver, small intestine, spleen, lung, adrenal gland, stomach, heart, fat, brain, muscle, ovary 6 h after administration Conclusion Ranolazine was quickly distributed to the tested twelve rat organs after oral administration, especially in stomach, small intestine, kidney . It mostly took a half or an hour for tissues to obtaine the peak concentrations. Besides these , it also showed that ranolazine had obvious distribution deferences between female and male rats. After oral administration, ranolazine generally has higher exposure and slower elimination in female tissues than that in male rats, particularly in liver, lung, spleen and adrenal gland. 3 The toxicokinetics study of ranolazine in beagle dogs following multiple dosage for three monthes.Objective To detect the accumulation of ranolazine in beagle dogs after multiple dosage. Method Beagle dogs were randomly allocated to three groups: low dose (L group), mi d group (M group), and high group (H group), each group was orally administrated with 20, 40 and 80 mg·kg·d^-1 ranolazine for three monthes, from monday to Saturday, once a day. The blood samples were collected at different time after dosing. Blood samples for determination of plasma ranolazine concentration were obtained just prior to drug administration on 1^st day and at 0.25 , 0.5, 0.75, 1.0, 1.5, 2.0 , 3.0, 4.0, 6.0, 8.0, 12.0 and 24.0 hours on 1^st day and 91^st day after administration. All collected blood samples were determined by HPLC/MS/MS method described as above. Toxicokinetics parameter calculations were carried out using compartmental and non-compartmental analysis method. Result The average peak plasma concentration of raolazine on 1^st day was 1.997±0.249, 3.49±0.401, 16.105±1.74mg·L^-1, obtained at 0.958± 0.102, 1.333±0.258, 1.083±0.204 h respectively for 20, 40 and 80mg·kg^-1 doses. The AUC_0-t of the three doses were 3.428±0.733, 9.481±1.339, 33.944±5.4mg·h·L^-1. The average peak plasma concentration of raolazine on 91^st day was 2.27±0.209, 4.973±0.491, 22.46±4.972mg·L^-1, obtained at 0.958±0.102, 0.958±0.102, 0.958±0.102 h respectively for 20, 40 and 80mg·kg^-1 doses. The AUC_0-t of the three doses were 3.909±0.824, 11.06 ±0.676, 50.611±12.658 mg·h·L^-1. Conclusion The safe dose of ranolazine in dog is 20 mg·kg^-1, and the adrenal gland is the toxicity target organ of ranolazine. After dosage, ranolazine was quickly absorbed and generally obtained peak concentration in about one hour. The peak concentration and AUC of the safe dose 20 mg·kg^-1 provided the safty plasma-concentration ans AUC of ranolazine in dogs wer 0².27±0.209 mg·L^-1 and 0～3.909 ±0.824mg·h·L^-1 respectively, which were obtained in this study. The peak concentraton of 40 and 80 mg·kg^-1 obtained on 91^st day were bigger than the datas obtained on 1^st day, especially the 80 mg·kg^-1 group, of which the datas had no statistics meanings.