Trimukhe K D, Bachate S, Gokhale D V, Varma A J
Polymer Science and Engineering Division, National Chemical Laboratory, Pune 411008, India.
Int J Biol Macromol. 2007 Dec 1;41(5):491-6. doi: 10.1016/j.ijbiomac.2007.06.009. Epub 2007 Jun 30.
This paper investigates the behavior of crosslinked chitosans and metal-complexed crosslinked chitosans under similar hydrolytic conditions. Crosslinked chitosans with trimellitic anhydride, diisocyanatohexane, and dibromodecane as crosslinking agents under heterogenous reaction conditions were used as metal complexing agents by equilibrating them with metal salts such as ZnCl(2), MnSO(4), CuSO(4), CdSO(4), Pb(NO(3))(2), and HgCl(2). Crosslinked chitosan without metal complexation had the same hydrolytic behavior as uncrosslinked chitosan. However, when the crosslinked chitosans were complexed with metals, their rates of hydrolysis and extent of hydrolysis were significantly reduced. Thus, while for chitosan about 840microg/ml reducing sugar was produced in 4h time, and 780microg/ml was produced for diisocyanatohexane crosslinked chitosan, only 400microg/ml and 320microg/ml reducing sugars were produced for cadmium sulfate with crosslinked chitosan and diisocyanatohexane crosslinked chitosan, respectively. Similar results are obtained for other crosslinking agents. Studies on preincubation of the metal with the enzyme show that of the metals studied, Mn has no effect on preincubatioin with the enzyme, Hg, Cd, Pb, and Cu completely deactivates the enzyme, while Zn reduces the enzyme activity by about 43.3%. Preincubation of the metal salts with the chitosan shows that Hg and Cu completely deactivate the molecule from enzyme hydrolysis, Cd and Zn inactivate it to the extent of 56.8% and 43.3%, respectively, while Mn has no effect. Availability of the amino functions seems to be a key feature for the chitosanase to hydrolyze the chitosan polymer. This was also proved by the significant increase in the extent of hydrolysis for chitosan samples with 88% (final value 1120micro/ml reducing sugar) and 85% deacetylation (final value 840microg/ml reducing sugar). HPIC studies of the products show that a variety of oligomers are produced in the chitosanase enzyme hydrolytic reaction.
本文研究了交联壳聚糖和金属络合交联壳聚糖在相似水解条件下的行为。以偏苯三酸酐、二异氰酸己酯和二溴癸烷为交联剂，在非均相反应条件下制备的交联壳聚糖，通过与ZnCl₂、MnSO₄、CuSO₄、CdSO₄、Pb(NO₃)₂和HgCl₂等金属盐平衡，用作金属络合剂。未进行金属络合的交联壳聚糖具有与未交联壳聚糖相同的水解行为。然而，当交联壳聚糖与金属络合时，它们的水解速率和水解程度显著降低。因此，壳聚糖在4小时内产生约840μg/ml的还原糖，二异氰酸己酯交联壳聚糖产生780μg/ml的还原糖，而硫酸镉与交联壳聚糖和二异氰酸己酯交联壳聚糖分别仅产生400μg/ml和320μg/ml的还原糖。其他交联剂也得到了类似的结果。对金属与酶预孵育的研究表明，在所研究的金属中，Mn对与酶的预孵育没有影响，Hg、Cd、Pb和Cu完全使酶失活，而Zn使酶活性降低约43.3%。金属盐与壳聚糖的预孵育表明，Hg和Cu完全使分子不能被酶水解，Cd和Zn分别使酶失活56.8%和43.3%，而Mn没有影响。氨基官能团的可用性似乎是壳聚糖酶水解壳聚糖聚合物的关键特征。这也通过脱乙酰度为88%（最终值为1120μg/ml还原糖）和85%（最终值为840μg/ml还原糖）的壳聚糖样品水解程度的显著增加得到证明。对产物的高效离子交换色谱研究表明，壳聚糖酶水解反应中产生了多种低聚物。
