ISOLATION AND CHARACTERISTICS OF BOVINE SKIN GELATIN AND ANALYSIS OF GLYCINE, PROLINE, AND HYDROXYPROLINE BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHYFLUORESCENCE
Keywords:Bovine gelatin, Glycine, Proline, Hydroxyproline, Derivatization, 9-fluorenyl-methoxycarbonyl chloride, High-performance liquid chromatography, Optimization
Objective: This study aimed to isolate gelatin, determine its characteristics, and obtain analytical methods optimum for the determination of glycine,
proline, and hydroxyproline levels in bovine gelatin.
Methods: Bovine hide was hydrolyzed using 2% sodium hydroxide at an extraction temperature of 70Â°C for 3 h and a drying temperature of 60Â°C.
Then, the gelatin extracts were evaluated using an organoleptic test; Fourier transform infrared analysis; pH measurement; and ash content, moisture
content, and viscosity tests. Optimum analysis conditions for the determination of glycine, proline, and hydroxyproline levels in bovine gelatin using
high-performance liquid chromatography with a fluorescence detector were as follows: Excitation wavelength, 265 nm; emission wavelength, 320 nm;
mobile phase composition of acetic buffer: acetonitrile, 55:45; flow rate, 0.8 mL/min; C18 column with length 250 mm and inner diameter 4.6 mm;
and particle size, 5 Î¼m. Derivatization of amino acids was performed using the reagent 9-fluorenylmethoxycarbonyl chloride.
Results: The results showed average levels of glycine, proline, and hydroxyproline in bovine gelatin, i.e., 25.10Â±0.09%, 14.28Â±0.11%, and 13.0Â±0.05%,
Conclusion: The optimum conditions for bovine gelatin hydrolysis included HCl with heating for 22 h at 110Â°C. The hydroxyproline, glycine, and
proline levels obtained for analysis in bovine gelatin samples were 13.50Â±0.05%, 25.11Â±0.09%, and 14.28Â±0.11%, respectively.
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