Optimization and Validation of a Chromatographic Method for the Quantification of Isoniazid in Urine of Tuberculosis Patients According to the European Medicines Agency Guideline

Abstract

Isoniazid is a drug that is widely used against tuberculosis. However, it shows high interpatient variability in metabolism kinetics and clinical effect, which complicates the prescription of the medication and jeopardizes the success of the therapy. Therefore, in a specific patient, the pharmacokinetics of the drug must be elucidated to decide the proper dosage and intake frequency to make the drug suitable for therapeutic drug monitoring. This can be performed by the quantification of the drug in urine as this process is non-invasive and allows the effects of long-time exposure to be inferred. The paper describes the development of a micellar liquid chromatographic method to quantify isoniazid in urine samples. Extraction steps were avoided, making the procedure easy to handle and reducing the waste of toxic organic solvents. Isoniazid was eluted in less than 5 min without interference from other compounds of the urine using a mobile phase containing 0.15 SDS–12.5% 1-propanol (v/v)–Na2HPO4 0.01 M buffered at pH 7, running at 1 mL/min under isocratic mode through a C18 column with the detection wavelength at 265 nm. The method was validated by following the requirements of the Guidelines on Bioanalytical Method Validation issued by the European Medicines Agency (EMA) in terms of selectivity, calibration curve (r2 = 0.9998 in the calibration range (0.03–10.0 μg/mL), limit of detection and quantification (10 and 30 ng/mL respectively), precision (<16.0%), accuracy (−0.9 to +8.5%), carry-over, matrix effect, and robustness. The developed method was applied to quantify isoniazid in urine samples of patients of an Indian hospital with good results. The method was found to be useful for routine analysis to check the amount of isoniazid in these patients and could be used in its therapeutic monitoring.

Alternative Methods for Therapeutic Drug Monitoring and Dose Adjustment of Tuberculosis Treatment in Clinical Settings: A Systematic Review

“…Other studies qualitative 8 Reported ratios low. Poor diffusion in saliva noted [ 74 , 77 ] No data Rifapentine 6 Studies in patients with TB needed [ 28 ] 7 Urine C max , AUC not reported [ 47 ] No data No data Isoniazid 7 DBS and plasma concentrations were measured in the included study, but C max , AUC, and DBS-plasma ratio not reported [ 35 ] 7 Ratios not provided in quantitative studies [ 36 , 38 , 49 , 50 , 59 , 67 , 69 ]. Other studies qualitative 8 Wide range of reported ratios [ 50 , 74 , 77 ] 7 Wide range of reported hair concentrations at different timepoints [ 84 ] in one study.…”

Limited sampling strategy for predicting isoniazid exposure in patients with extrapulmonary tuberculosis

“…Despite the paucity of studies regarding the added value of isoniazid TDM, most of the investigators have proved a large intra‐ and interindividual variability in isoniazid pharmacokinetics in tuberculosis patients . It was shown that some factors such as age, sex, gastrointestinal disorders and drug interactions affect the pharmacokinetics of isoniazid leading to response variation .…”

Isoniazid-Gentisic acid cocrystallization: Solubility, Stability, Dissolution rate, Antioxidant and Flowability Properties Studies

“…Antioxidants herbal medicines have shown decreased oxidative stress significantly leading to the prediction of positive role of antioxidant drugs during anti-TB treatment. [3] , [4] , [5] Isoniazid [6] (INH) is a very efficient remedy against M. tuberculosis recommended by WHO.…”

“…[36,37] Crystal data: C7H6O4•C6H7N3O (M = 291.26 g/mol), triclinic, space group P1 ̅ (no. 2), a = 3.7075 (6), b = 10.9505 (18), c = 15.966(3) Å, α = 101.366 (6), β = 95.706 (6), γ = 91.327(6)°, V = 631.72(18) Å 3 , Z = 2, T = 120 K, μ(Mo-Kα) = 0.12 mm -1 , Dcalc = 1.531 g/cm 3 , 8870 reflections with 2 ≤ 50° measured, 2229 unique (Rint = 0.050), 243 refined parameters, R1 = 0.038 on 1655 reflections with I > 2σ(I), wR2 = 0.093 on all data.…”