ULTRASOUND-ASSISTED EXTRACTION AND CHARACTERIZATION OF SOME EXTRACELLULAR MATRIX COMPONENTS (COLLAGEN AND GAG) FROM LAMB FEET
Project no. 1402 · Status: Kapanmış · 2017
Summary
In this thesis, ultrasound-assisted extraction and purification of collagen and glucosaminoglycans (GAG), one of the extracellular matrix components, was aimed. Within the scope of the study, lamb legs were used as raw material source and suitable extraction conditions were determined for obtaining the highest yield The effects of ultrasound treatment on collagen and GAG structure were investigated at different temperatures and durations. In the first stage of the study, suitable extraction parameters for the production of collagen were determined. Enzymatic hydrolysis method was selected for extraction and pepsin enzyme was used. The lambs which were pretreated were left for 6 hours in a 1:20 (w / v) 5% lactic acid solution containing 0.1% (w / v) pepsin for extraction. Then, ultrasound-assisted extraction was performed at 3 different temperatures (20± 2°C, 25± 2°C and 30± 2°C) and time (20, 40, 60 min). After extraction, the samples were dialyzed for purification and freeze-dried at -56 ° C for 32 hours. The highest collagen parameter was reached at 25°C-60 min (5061,75±171,41 µg/µl). The results of SDS-PAGE electrophoresis and FTIR analysis showed that ultrasound applied at different temperatures and times did not damage the collagen structure and retain the structural integrity of all collagen. As a result of SEM analysis, it was observed that all collagen samples have fibrous and porous structure. The difference between b * and L * values of collagen samples was statistically significant(p<0.05). The difference between the a * values of the samples was significant only for the four groups. In the second stage of the study, suitable extraction parameters for GAG were determined. Papain enzyme and sodium acetate solution were used in the extraction process. The solid to solvent ratio was 1:20 (w / v), the amount of enzyme (5 mg papain / 1 g lamb's foot), temperature (60°C) and extraction time was determined as 20 hours. After 20 hours of extraction, ultrasound assisted extraction was performed in 3 different temperatures (60± 2°C, 65± 2°C and 70±2°C) and time (20, 40, 60 min) to examine the effects of ultrasound application. After extraction, the samples were dialyzed for purification and freeze-dried at -56°C for 24 hours. The highest chondroitin sulfate concentration was reached at 60°C-20 min (471,38±49,96 mg/L). As a result of FTIR analysis, it was seen that GAG samples obtained under different extraction conditions were similar to the spectra of the standard. The results of SEM analysis showed that the clustered structure of GAG samples changed depending on the ultrasound time and temperature.