Simple Hplc Validated Method for the Determination of Diltiazem Hydrochloride in Human Plasma

Objectives: To develop and validate a high-performance liquid chromatographic method for determination of diltiazem hydrochloride (DLZ) Methods: Mixture of n-hexane and 2-propanol (96:4, ratio) was added to plasma at sample preparation time followed by centrifuging the samples. The obtained upper organic layer was transferred and evaporated to dryness. The residue was reconstituted with a mobile phase and the supernatant was then injected onto the column. The mobile phase used was consisted of 0.2 M ammonium dihydrogen phosphate, acetonitrile, isopropyl alcohol and triethylamine (55:43:1.7:0.3, v/v) with pH adjusted to 4.5 using 85% phosphoric acid. The flow rate was 0.7 ml/min. UV detector set at 240 nm and samples were quantified using peak area. in human plasma. Results: A well-resolved DLZ peak and free of interference from endogenous compounds in plasma with a retention time of 6.03 min was achieved. Recovery of DLZ was satisfactory (≥ 91.3%) over the concentration range tested 0.25 20 μg/ml. LOD of this assay was 0.125 μg/ml and LOQ was 0.25 μg/ml and, at this concentration, intraand inter-day CV were 6.8 and 9.2 %, respectively. DLZ was found to be stable in plasma after storage at -80oC, over 90 days. Conclusion: The HPLC method described in this article was simple, sensitive, selective, reproducible, linear, precise, accurate, stability indicating and requires only a small sample volume, lending it suitable for the determination of DLZ concentration in routine measurements for pharmacokinetic/bioavailability studies.