相近DDA但不同分子量的四種幾丁聚醣樣品Chitosan10 (DDA 86.8%，Mv 130K)、 LMW(DDA 85.5%，Mv 800K)、MMW(DDA 87.4%，Mv 1310K)、HMW(DDA 83.0%，Mv 2290K)及分別將前述四種樣品經高溫強鹼去乙醯化至大約96% DDA，經不同醋酸酐/胺基 莫耳比例(AA/amine)的乙醯化反應後，每個樣品都可得到一系列不同DDA的幾丁聚醣產物，其AA/amine和LogDDA呈線性關係。從相同DDA但不同分子量的起始樣品經乙醯化反應所得之AA/amine和LogDDA線性關係相同，因此乙醯化反應之分子量效應不明顯。另外，我們將DDA為87.4%的MMW樣品分別進行去乙醯化和乙醯化反應，得到DDA分別為96.3%、70.6%和64.2%的樣品，前述線性關係亦成立，但其乙醯化反應之樣品DDA效應卻很明顯，亦即，幾丁聚醣初始DDA越大，則其乙醯化反應性越好。
The degree of deacetylation (DDA) of chitosan can be quickly and accurately determined in acidic buffer by capillary electrophoresis (CE). According to the electropherogram of chitosan, the DDA distribution can also be estimated from the half-width (W1 / 2, DDA ) of UV absorption peak. By using this newly developed CE-based method, we were able to investigate effects of chitosan molecular weight, DDA, concentration, reaction temperature and duration on the acetylation reaction of chitosan.
Acetylation reactions of different acetic anhydride to amine molar ratio (AA / amine) were carried out for Chitosan10 (DDA 86.8%, Mv 130K), LMW (DDA 85.5%, Mv 800K), MMW (DDA 87.4%, Mv 1310K), and HMW (DDA 83.0%, Mv 2290K), chitosan samples with different molecular weights but similar DDAs, as well as their respective derivatives obtained from deacetylation reactions to about 96% DDA in hot alkaline solution. For each sample, a series of acetylation products with different DDAs were obtained, and it was observed that AA / amine and the measured LogDDA has a linear relationship. We concluded that chitosan molecular weight has no apparent effect on acetylation reaction due to the same AA / amine – LogDDA linearity of chitosans with similar DDAs but different molecular weights. In addition, the AA / amine – LogDDA linearity also exists for the four chitosans having DDAs of 96.3%, 87.4%, 70.6% and 64.2% prepared from deacetylation and acetylation of MMW. However, the DDA effect on acetylation reaction is obvious, i.e., the higher the original DDA of chitosan, the more complete reactivity of its acetylation reaction.
On the other hand, by combining reaction quenching approach and CE analysis method, we were able to study the molecular weight effect on the kinetics of acetylation reaction. Moreover, we found that reactant concentration would affect the equilibrium DDA value of acetylation reaction. For reactions of the same AA / amine, chitosans with higher concentrations would have better reactivity and thus produce lower DDA chitosans. Nevertheless, the reaction temperature (25 ~ 60 oC) did not affect the DDA value. We also found that the acetylation reaction has reached equilibrium within 24 hours.
Through deacetylation reaction to high DDA (~ 96%) in hot alkaline solution, chitosan DDA distribution could be largely reduced. Therefore, first treating sample by deacetylation and then acetylation according to the AA / amine – LogDDA linearity, we can prepare chitosan sample with required DDA and relatively narrow distribution.