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Development and optimization of HPLC method for determination of peroxide impurities in pharmaceutical excipients
Hrubcová, Kateřina ; Křížek, Tomáš (advisor) ; Sobotníková, Jana (referee)
The chemical stability of drugs can be affected by impurities that occur in commonly used excipients. Hydroperoxides are common trace impurities, especially in polymeric excipients, which can cause oxidative degradation of drugs. Several methods for the quantification of trace levels of hydroperoxides (hydrogen peroxide and organic hydroperoxides) are described in the literature. However, there are few studies dealing with the quantification of trace levels (ppm) of hydroperoxides in drug excipients. The goal of this thesis was the development and optimization of an HPLC method with UV detection for the determination of hydroperoxides in drug excipients, based on methods already described in the literature. The developed HPLC method is based on the determination of triphenylphosphine oxide as a product of the rapid and quantitative reaction of triphenylphosphine with hydroperoxides. Method development also included optimization of sample preparation prior to HPLC analysis. During the experimental work, the influence of individual experimental parameters and sample preparation conditions on the determination of hydroperoxides was determined. The optimized HPLC method used a Kinetex F5 column (100 × 4,6 mm, 2,6 µm) and a mixture of water and acetonitrile as the mobile phase with gradient elution. As...
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Development of HPLC separation of astaxanthin, β-carotene and lutein
Hrubcová, Kateřina ; Nováková, Eliška (advisor) ; Kozlík, Petr (referee)
This bachelor's thesis deals with the development of a method for the separation of carotenoid pigments, especially astaxanthin, β-carotene and lutein, produced by the microscopic green alga Tetracystis pulchra. The work compares published methods for the separation of carotenoid pigments and attempts to optimize a method suitable for the analysis of pigment extracts from Tetracystis pulchra cells. Optimized conditions for HPLC-DAD separation of carotenoid pigments were as follows: mobile phase A - ACN/MeOH/0,05M Tris-HCl buffer (pH = 8,0): 94/2/4, mobile phase B - MeOH/EtOAc: 68/32 under gradient elution, flow rate 0,25 ml min˗1 , column temperature 28 řC. The column used was Kinetex C18 (100 × 2,1 mm, 2,6 µm), injection volume was 20 µl and UV detection was performed at 470 nm. Total analysis time was 23 minutes. Key words: HPLC, astaxanthin, β-carotene, lutein
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