Determination of hydroxyproline using ion-exchange liquid chromatography with vis detector and high performance liquid chromatography with fluorescence detector

Abstract

The first aim of the experiment was an optimization of the method for determination of hydroxyproline in a pig skin using high-performance liquid chromatography with fluorescence detector (HPLC-FLD) and ion-exchange liquid chromatography with VIS detector (IEC-VIS). On the basis of the experiments performed, it was found that HPLC-FLD method is three times more sensitive than IEC-VIS method. For IEC analysis of hydroxyproline, the limit of detection (LOD) was 4.10 g/ml whereas the limit of quantification (LOQ) was 13.50 g/ml. For HPLC analysis of hydroxyproline, the limit of detection (LOD) was 1 ng/ml whereas the limit of quantification (LOQ) was 3 ng/ml. The second aim was the testing of influence of different volume of 6M HCl on extraction of sample (50 mg) for analysis using HPLC-FLD. Here it was found that the best volume was 250 µl 6M HCl.The first aim of the experiment was an optimization of the method for determination of hydroxyproline in a pig skin using high-performance liquid chromatography with fluorescence detector (HPLC-FLD) and ion-exchange liquid chromatography with VIS detector (IEC-VIS). On the basis of the experiments performed, it was found that HPLC-FLD method is three times more sensitive than IEC-VIS method. For IEC analysis of hydroxyproline, the limit of detection (LOD) was 4.10 g/ml whereas the limit of quantification (LOQ) was 13.50 g/ml. For HPLC analysis of hydroxyproline, the limit of detection (LOD) was 1 ng/ml whereas the limit of quantification (LOQ) was 3 ng/ml. The second aim was the testing of influence of different volume of 6M HCl on extraction of sample (50 mg) for analysis using HPLC-FLD. Here it was found that the best volume was 250 µl 6M HCl.