Determination of Degree of N-Acetylation for Chitin and Chitosan by Using 2-bromo-1-phenylethanone Derivatization and HPLC Method

• Main Authors: Cheng, Shih-Hua, 鄭詩樺
• Other Authors: Hong, Liang-Bang
• Format: Others
• Language: zh-TW
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/8kjfx6

碩士 === 國立臺灣海洋大學 === 食品科學系 === 104 === Using HPLC method to determine the degrees of N-acetylation of chitin and chitosan usually involves the step of acid hydrolysis to quantify the release of acetic acid. For samples of low degree of N-acetylation, the small acetic acid peak on the chromatogram may cause underestimation of the result. The purpose of this study is to improve the previous HPLC method proposed by our laboratory. In order to increase UV absorption and improve detection sensitivity, 2-bromo phenylethanone was used to react with acetic acid to form the ester bond. The higher sensitivity could make the determination of the degrees of N-acetylation more accurate for the samples of low degree of N-acetylation. The analysis was carried out on a C18 column. 50% acetonitrile aqueous solution was used as the mobile phase and a PDA detector setting at 243 nm was uesd. Derivatized butyric acid was applied as the internal standard. The acid hydrolysates were extracted with ethyl acetate and the extracts were then derivatized. The derivatization was held in the solution with the mixing ratio of 1:5 (the mole ratio of carboxylic acids to that of derivatization reagent). After calculation, the degrees of N-acetylation of three samples without derivatization were 5.04, 11.01, and 81.68%, respectively. The degrees of acetylation of three samples with derivatization are 5.40, 12.88, and 82.61%, respectively. Both experiment results agreed with the values labelled. However, the results with derivatization were found to be closer to the label values. Therefore, the derivatization method could increase the detection sensitivity of acetic acid, and elevate the accuracy for determining the samples of low degree of N-acetylation.