Supervisors info:
Ευάγγελος Γκίκας, Επ.Καθηγητής, Τμ. Φαρμακευτικής, Εθνικό και Καποδιστριακό Πανεπιστήμιο Αθηνών
Ιωάννης Ντότσικας, Επ.Καθηγητής, Τμ. Φαρμακευτικής, Εθνικό και Καποδιστριακό Πανεπιστήμιο Αθηνών
Summary:
In the present study, new methods for the determination of three antihypertensive substances (hydrochlorothiazide-HCTZ, amlodipine-AML, valsartan-VAL) were achieved by RP-HPLC/UV, under isocratic conditions and using C8 and C18 columns. Mobile phase optimization was completed using three different experimental designs, Box-Behnken design (C8 column), Central Composite Design-CCD (C18 column) and D-Optimal design (C8, C18). The software model that was used was the trial version of Design Expert 10.
From each experimental design, which differ in their structural characteristics (number of experiments, levels of factors, etc.), two optimal solutions of the mobile phase were proposed depending on the elution order of the substances AML-VAL (α=1,3) or VAL-AML (α=0,77). Box-Behnken design is an exception as there was no reversal in the elution order in the pH range used in this case.
The optimum mobile phase for Box-Behnken design was characterized by ACN percentage 34,0%, an extreme pH value of 4,46 and salt concentration 56,0 mM (non-significant factor). For CCD and for selectivity factors α = 1,30 and α = 0,77, ACN percentage was higher at 38,6% and 41,2%, MeOH percentage 2,0% (lowest extreme value), pH values 3,87 and 4,18, salt concentration 57,0 mM and 26,0 mM, respectively. For D-Optimal, ACN percentage was lower 33,0%, MeOH percentage at zero, pH values 4,41 and 4,70, salt concentration 60,0 mM and 55,0 mM, for α = 1,30 and α = 0,77 respectively, with an optimal C8 column in both cases.
All three designs yielded optimal conditions, which lead to very good separation of substances, good chromatographic peaks and short time analysis. Total time analysis ranged from 6,15 min to 8,40 min. The main conclusion of their comparison is that the optimal solution for Box-Behnken design resulted with a small number of experiments and presented the shortest time analysis, so it appears as a more economical and time-consuming one. The CCD, however, showed maximum desirability and D-Optimal had the best predictive ability for the capacity factor of hydrochlorothiazide.
In total, new methods of determination of hydrochlorothiazide, amlodipine and valsartan were developed using C8 and C18 columns, for which method validation is pending.
