| BACKGROUNDIn recent years,new oral anticoagulants are gradually replacing traditional anticoagulants such as vitamin K antagonists or heparin in some areas,due to their effectivity,safety and convenience.Rivaroxaban,as a novel oral anticoagulant drug with high selectivity and direct inhibition of factor Xa,has favorable pharmacokinetic and pharmacodynamics properties and is widely used in clinical practice.There is not standardized laboratory monitoring for rivaroxaban.Although clinical trials shown that rivaroxaban has equal or superior efficacy and an improved safety profile compared with warfarin or enoxaparin[1,2],fixed-dose therapy also has some problems in clinical application.It cannot be effectively monitored when the drug is overdose and there is no specific reversal agent once patients bleed.There are many situations when and populations for whom clinicians would like to assess the precise anticoagulant effect for the refinement of treatment decisions.The potential indications for coagulation testing include emergency situations,urgent surgery,urgent invasive procedures,major bleeding,overdose,acute thrombosis,renal failure,adherence verification,and potential drug-drug interactions[3,4].The study has suggested that unmonitored DOAC therapy should be only for relatively young and healthy patients,whereas patients 75 years of age and older should have coagulation laboratory monitoring 5～10 times per year[5].The clinical studies of rivaroxaban were conducted with fixed doses and did not evaluate clinical outcomes based on drug concentrations or coagulation assays.No evidence-based recommendations for drug concentration measurements,coagulation tests,assay standardization/calibration,or target therapeutic ranges have been established for rivaroxaban[6].The most studies of the pharmacokinetics are the phase I and phase II clinical studies in abroad,and the pharmacokinetics is mainly in healthy people in China.No study is found for the pharmacokinetics in patients with deep venous thrombosis.The anticoagulant clinics about warfarin are relatively mature and the quality of anticoagulant therapy has been significantly improved in abroad.They also explore actively the model of anticoagulant pharmacy services for NOACs.In China,the pharmacy service for warfarin is still under development and pharmaceutical care for NOACs has not yet been carried out.OBJECTIVETo develop an UPLC-MS/MS method for the determination of rivaroxaban in human plasma and preliminarily study the pharmacokinetics of rivaroxaban in patients with deep venous thrombosis.Then to determined the steady-state tough concentration and peak concentration of rivaroxaban in patients with deep venous thrombosis and aim to find an assay can be used to clinically evaluate rivaroxaban.And we managed to optimize the anticoagulant pharmaceutical service model.METHODSAn UPLC-MS/MS method for the determination of rivaroxaban in human plasma was developed and validated.The patients meeting the inclusion criteria,from the department of vascular surgery in our hospital,were gave rivaroxaban with the meal every time.The clinicians made the dosage regimen based on the conditions of the patients.Blood samples were collected at 0,1,2,3,8,24h after patients with deep venous thrombosis received rivaroxaban as the first oral dose,and at Oh and 2h after patients received rivaroxaban for 7 days or more.The plasma concentration was determined by UPLC-MS/MS method and the PT,APTT,anti-factor X a activity was determined in the laboratory medicine.The pharmacokinetic parameters were calculated according to non-compartmental model by using Winnonlin(version 6.3,Pharsight).The correlation between plasma concentrations and PT,APTT,and anti-Xa activity were analyzed by SPSS 22.0.At the same time,we manage the medications and educate the patients received anticoagulants in the vascular surgery wards and outpatient clinics.RESULTSAn UPLC-MS/MS method using d4-rivaroxaban as internal standard compound and electronic spray ion with positive ionization ions(ESI")monitored in the multiple reaction monitoring(MRM)mode was developed and validated for rivaroxaban in human plasma.The linear range of rivaroxaban was 0.5～400 ng·mL-1(r=0.998).The lowest limit of quantitation was 0.5 ng·mL-1 and the intra-day and inter-day relative standard deviations were<15%.The recovery rates were 75.18%～87.53%and the RSD of six-batches matrix effects were less than 3%.Twelve patients with deep venous thrombosis received rivaroxaban 20mg(n=6),15mg(n=3)or 10mg(n=3)as the first oral dose.The maximum plasma concentration of rivaroxaban 20mg,15mg and 10mg in patients with deep venous thrombosis was 317.4ng·mL-1,272.2ng·mL-1,153.3ng·mL-1,and the time to Cmax was 2-3h,and the elimination half-lifein plasma was 6～7h.The Cmax and AUC profiles were dose-dependent.Forty-four patients with deep venous thrombosis received rivaroxaban 15mg bid(n=3),10mg bid(n=9),20mg qd(n=8),15mg qd(n=7),10mg qd(n=12).The average steady-state tough concentration in patients with deep venous thrombosis was 168.5ng·mL-1,74.2ng·mL-1,25.7ng·mL-1,24.5ng·mL-1 and 15.4ng·mL-1,and the average steady-state peak concentration was 280.1ng·mL-1,271.4ng·mL-1,306.8ng·mL-1,306.4 ng·ml-1 and 229.2ng·mL-1.There was a statistically significant difference(p=0.008)in the trough concentration between the two dose group of 10mg bid and 20mg qd;there was no statistically significant difference(p=0.521)in the peak concentration.And the trough and peak concentration of the three dose group of 20mg qd,15mg qd and 10mg qd didn’t show statistically significant difference(p=0.218 and p=0.122).The Spearman correlation coefficient between plasma concentration of rivaroxaban and PT,APTT,anti-factor Xa activity were r=0.815(p<0.001),r=0.833(p<0.001),r=0.985(p<0.001).There was a linear relationship between plasma concentration and anti-factor X a activity.Compared with the retrospective data of the patients without pharmacists’intervention in our hospital,the average TTR of patients received warfarin with pharmacists’ intervention was increased by 12.6%,and the difference was statistically significant(p=0.03);moreover,more than half of the patients in the intervention group(51.22%)got the best therapy.CONCLUSIONThis UPLC-MS/MS method is rapid,specific,reliable and suitable for the determination of rivaroxaban in human plasma and pharmacokinetic study.The pharmacokinetics in patients with deep venous thrombosis is consistent with that in healthy Chinese.The plasma concentration profile is dose-dependent.The plasma concentration in patients with deep venous thrombosis received rivaroxaban 10mg bid is more stable than that in patients received rivaroxaban 20mg qd.The plasma concentration of rivaroxaban in patients of China is higher and the clearance is faster than foreigner.There was a statistical correlation between plasma concentration of rivaroxaban and PT,APTT,anti-factor Xa activity,and the linear relationship between plasma concentration and anti-factor X a activity is better than that between concentration and PT or APTT.The anti-factor X a activity and plasma concentration can be used to evaluate the anticoagulant effect clinically.However,the range of plasma concentration obtained in this study is only for reference and does not fully represent the pharmacokinetics of rivaroxaban in Chinese patients with deep vein thrombosis,due to the small sample size.Clinical pharmacist can provide series of pharmaceutical services for patients,such as educating patients,monitoring adverse drug reactions,evaluating and managing drug-drug/food interactions,assisting doctors in adjusting warfarin dose,regular follow-up and supervision of treatment adherence,and so on,to improve the quality of treatment with anticoagulation. |
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