OPTIMIZED HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY-FLUORESCENCE DETECTION METHOD FOR THE MEASUREMENT OF GLYCINE, PROLINE, AND HYDROXYPROLINE CONCENTRATIONS IN PORCINE GELATIN

  • Aryaty Ekasary Department of Pharmacy, Faculty of Pharmacy, Universitas Indonesia, Depok 16424, Indonesia
  • Harmita . Department of Pharmacy, Faculty of Pharmacy, Universitas Indonesia, Depok 16424, Indonesia
  • Baitha Palanggatan Maggadani Department of Pharmacy, Faculty of Pharmacy, Universitas Indonesia, Depok 16424, Indonesia

Abstract

Objective: The aim of this study is to develop an optimized method for glycine, proline, and hydroxyproline content quantitation in porcine skin
gelatin.
Methods: Gelatin was isolated from porcine skin by hydrolysis for 24 h in 0.5 M acetic acid, heating in distilled water at 55°C for 3 h, and drying at
60°C. The extract was evaluated by organoleptic tests, Fourier-transform infrared spectroscopy, moisture assay, ash assay, and viscosity test. Gelatin
amino acids were derivatized using 9-fluorenylmethylchloroformate-chloride and measured by high-performance liquid chromatography (HPLC)
with fluorescence detection using a C18 column after the optimization of the mobile phase composition, flow rate, and detection wavelengths.
Results: The optimized parameters for the quantitation of glycine, proline, and hydroxyproline by HPLC with fluorescence detection were as
follows: Excitation wavelength, 265 nm; emission wavelength, 320 nm; mobile phase composition acetic buffer: acetonitrile, 55:45; and flow
rate, 0.8 mL/min. The average proportional amino acid contents were 28.57±0.74%, 19.24±0.48%, and 2.89±0.33% for glycine, proline, and
hydroxyproline, respectively.
Conclusion: This method allows for sensitive and accurate quantitation of glycine, proline, and hydroxyproline in porcine skin gelatin samples for
quality control and source determination.

Keywords: Derivatization, Fluorenylmethoxycarbonyl chloride, Glycine, High-performance liquid chromatography, Hydroxyproline, Optimization, Porcine gelatin, Proline.

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Ekasary, A., ., H., & Maggadani, B. P. (2018). OPTIMIZED HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY-FLUORESCENCE DETECTION METHOD FOR THE MEASUREMENT OF GLYCINE, PROLINE, AND HYDROXYPROLINE CONCENTRATIONS IN PORCINE GELATIN. International Journal of Applied Pharmaceutics, 10(1), 352-330. https://doi.org/10.22159/ijap.2018.v10s1.72
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