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KMID : 1146920200500050527
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Journal of Pharmaceutical Investigation
2020 Volume.50 No. 5 p.527 ~ p.536
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A sensitive and fast HPLC method for determination of Isosorbide 5-mononitrate in human plasma using ion-pair chromatography: application to bioequivalence study
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Olyaei Elmira
Samiei Nasim
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Abstract
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Purpose: Isosorbide 5-mononitrate (5-ISMN) is a charged, hydrophilic, and low ultraviolet-absorbing molecule which is challenging to determine specifically and sensitively in biological matrices. A sensitive and fast ion-exchange chromatography method for determining 5-ISMN in plasma has been developed.
Methods: The plasma was deproteinized by one-step clean up procedure using sodium chloride and methanol. Then supernatant was injected into a Chromolith C18 column (4 μm, 3.9?×?150 mm). The mobile phase was made of acetate buffer (30 mM, pH: 4.7): methanol, 77:23 (v/v) and 1 mM tetrabutylammonium hydroxide with a flow rate of 1.2 mL/min. Ultraviolet detection was done at 225 nm. The established method was fully validated and successfully applied for pharmacokinetic and bioequivalence study of two different sustained release formulations of 5-ISMN as Elantan LA (reference product) and Mononitral (test product) in twelve healthy male volunteers. A single dose, randomized, two-period, and crossover study was performed under fasting condition.
Results: Isosorbide dinitrate was used as internal standard. 5-ISMN and internal standard were eluted at 2.6 and 3.8 min, respectively. Calibration curve was linear within the concentration range of 10?800 ng/mL for 5-ISMN in plasma. The results revealed that the precision of the method ranged from 2 to 5 and relative errors lied within 0.43?4.2% regardless of whether the analysis was performed intraday or interday. By this method, the limits of quantitation and detection of 10 ng/mL and 5 ng /mL were achieved by a sample size of 430 μL plasma. Main pharmacokinetic parameters for test and reference products were 494?±?210.2 and 500?±?202.5 for Cmax and 6359?±?1235 and 6995?±?1371 for AUC0-36, respectively. The 90% confidence intervals around ratios (test/reference) from logarithmic transformed exposure values were 99?110 for Cmax and 88?105 for AUC 0?36.
Conclusion: It can be concluded that both products are bioequivalent. In vitro dissolution test was also carried out together with bioequivalence study and in vitro/in vivo correlation was successfully established.
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KEYWORD
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Isorbide 5-mononitrate, Ion-exchange chromatography, Tetrabutylammonium hydroxide, Bioequivalence, Pharmacokinetic, In vitro/in vivo correlation
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