Disopyramide pharmacokinetics in non-smoking and smoking volunteers : analytical development and effect of phenobarbital on elimination and protein-binding
Creators: name: Kapil, Ram Prakash
Publisher: name: University of British Columbia
Year: 2010

description: Disopyramide is a class IA antiarrhythmic agent which shares a number of pharmacological properties with quinidine and procainamide. Clinically, it is effective in the treatment of supraventricular tachycardia, atrial fibrillation, ventricular tachycardia and premature ventricular contractions. Disopyramide, a basic drug, has been shown in the past to be bound to acute phase reactant, alpha₁-acid glycoprotein (AAG) in a concentration-dependent manner. Disopyramide has been known to demonstrate a steep serum concentration-pharmacological effect relationship. The primary metabolic pathway of disopyramide in humans is N-dealkylation which is susceptible to enzyme induction,as indicated by a few preliminary studies. The pharmacological role of the mono-N-dealkylated metabolite (MND) of disopyramide in humans is not fully established. This thesis describes the development of an improved gas-liquid chromatographic (GLC) assay for the simultaneous quantitation of trace levels of disopyramide and MND in human biological fluids and the application of the method in in vitro plasma protein-binding of disopyramide. This thesis also describes and discusses the effect of phenobarbital treatment on the serum protein-binding and single dose pharmacokinetics of disopyramide in non-smokers and smokers. A GLC-nitrogen/phosphorus specific detection (NPD) assay has been developed which provides improved selectivity and sensitivity for disopyramide and MND, employing a fused-silica capillary column for chromatographic separation from biological specimen components. The quantitation of disopyramide and MND in human serum, saliva and urine was accomplished by injecting trifluoro-acetic anhydride (TFAA)-treated samples containing the internal standard, p-chlorodisopyramide (PC-Dis), into a gas chromatograph equipped with a nitrogen/phosphorus detector (NPD) and an automatic liquid sampler (ALS). A 25 m (i.d. = 0.31 mm) fused-silica column, coated with crosslinked 5 % phenyl methyl silicone fluid was utilized and samples were injected using the splitless sample injection mode. Linearity was observed in the serum concentration range from 0.05 to 5.0 mcg/ml for disopyramide and from 0.02 to 3.0 mcg/ml for MND. The average coefficient of variation was 5 and 8 % for disopyramide and MND, respectively, over the concentration range studied. The capillary GLC-NPD assay was utilized to measure disopyramide concentration, unbound to plasma protein, following equilibrium dialysis of plasma samples collected from healthy volunteers. Various concentrations of disopyramide ranging from 0.5 to 10 mcg/ml in 0.4 ml of isotonic phosphate buffer (pH 7.4) were dialysed for 6 hours at 37°C, against 0.4 ml blank plasma from five healthy volunteers. The concentration-dependent binding of disopyramide was verified. The average unbound fractions for disopyramide, at concentrations 0.5, 1, 2, 3, 4, 5 and 10 mcg/ml, were 0.14, 0.15, 0.20, 0.28, 0.30, 0.35 and 0.55, respectively. The effect of phenobarbital treatment on the pharmacokinetics and binding of disopyramide was studied in non-smoking volunteers: (1) Because of the potential pharmacological and toxicological significance of the major metabolite of disopyramide, (2) due to the need to develop a better understanding of the effect of phenobarbital treatment on the serum concentration of alpha₁-acid glycoprotein (AAG), the principal binding protein of disopyramide and other basic drugs and (3) to provide insight into the selectivity of phenobarbital induction as compared to literature data on other inducers (rifampin, phenytoin, carbamazepine and spironolactone). Eight healthy male subjects received a 200 mg oral dose of disopyramide on 2 occasions separated by a 3 week period during which phenobarbital was taken (100 mg p.o.) at bedtime yielding a mean steady-state concentration of 14 ± 2 mcg/ml. Serum, saliva and urine collections were obtained following each disopyramide dose. The serum concentration versus time data were analysed by AU TOAN and NONLIN computer programs and were found to fit a one-compartment open pharmacokinetic model. The apparent terminal phase elimination half-life (t½) of disopyramide was shorter (p &lt; 0.05) in subjects during phenobarbital treatment (4.6 ± 0.7 hr) (mean ± S.D.) than during the control experiment (6.5 ± 1.5). Furthermore, an index of the amount of drug in the body, the area under the disopyramide serum concentration versus time curve (AUC), was markedly reduced from control (27 ± 6 hr.mcg/ml) after phenobarbital (17 ± 6 hr.mcg/ml, p &lt; 0.05). The AUC₀₋₂₄ʰʳ for the principal metabolite of disopyramide (mono-N-dealkyl disopyramide or MND) increased (3.8 ± 1.6 hr.mcg/ml for control vs 4.1 ± 2.3 hr.mcg/ml in pheno-barbital-treated subjects) but this change was not statistically significant. Phenobarbital treatment caused no observable trend towards a change in disopyramide unbound fraction (0.23 ± 0.02 vs 0.21 ± 0.02) or in serum AAG concentration (50 ± 15 vs 54 ± 18 mg/100 ml), as has been observed in dogs in the past. The ratio of MND to disopyramide in saliva was consistently higher in phenobarbital-treated subjects. The average percentage of dose recovered in urine as unchanged disopyramide in 48 hrs was 43 ± 6 % and 25 ± 5 % (p &lt; 0.05) in control and phenobarbital-treated subjects, respectively. The average percentage of dose recovered as MND in 48 hours was 25 ± 6 % in controls and 33 ± 7 % in phenobarbital-treated subjects. It can be concluded from our study that the enzyme inducer phenobarbital enhances disopyramide metabolism but does not alter the serum AAG levels or the binding of disopyramide to this protein in normal subjects. The effect of chronic cigarette smoking and the combined effect of smoking and phenobarbital treatment on the disopyramide pharmacokinetics and serum AAG levels were also investigated. The general procedures such as dosing and physiological monitoring of smokers, sampling protocols of smokers before and during phenobarbital-treatment, sample collection and analytical methods were identical to the non-smoking study. There was no difference in the pharmacokinetic parameters or serum AAG concentrations between the smoking and the non-smoking volunteers when studied and compared in the absence and presence of phenobarbital treatment. Therefore, it appears that the metabolic bio-transformation of disopyramide remains unaffected by chronic cigarette smoking. These findings support the general view that the induction of hepatic microsomal oxidation of a particular drug is highly selective in humans and is a function of substrate or drug specificity for a particular form of cytochrome P-450.