Cross-linked enzyme aggregates (CLEAs) have the advantages of high-enzyme activity and stability, and have an application potential in food and feed processing. In order to address the food and feed safety of CLEAs and have ideal enzymatic properties, a new design of CLEAs of lipase from Aspergillus niger is developed by using sodium tripolyphosphate (TPP) as a cross-linking agent and chitosan as an auxiliary agent. The enzyme activity recovery of the developed CLEAs reaches 126%. The half-life of the CLEAs is 11.4-fold higher than that of free enzyme at the optimum reaction temperature of CLEAs. The surface structure of the CLEAs reveals cluster and porous structure. The variations of the secondary structures of the CLEAs indicate an increase of α-helix and decrease of β-sheet, β-turn, and random coil. The adding of chitosan is beneficial to enzyme activity recovery and thermal stability of the CLEAs. TPP and chitosan are safe and nontoxic, and the CLEAs prepared with TPP and chitosan is suitable for the food and feed industries.
BACKGROUND: Chitooligosaccharides possess good physiological activity and a wide range of uses. In view of the difficulty in controlling the degree of polymerization of chitooligosaccharides prepared by chemical methods and the low efficiency of enzymatic methods, the preparation process of chitooligosaccharides prepared by H 2 O 2 oxidation pretreatment combined with ⊍-amylase catalysis was studied.RESULTS: Water-soluble chitosan was prepared by oxidation pretreatment of chitosan and then degradation by ⊍-amylase. The enzymatic hydrolysis conditions were determined as follows: pH, 6.0; temperature, 60 °C; liquid-solid ratio, 5:1. The degree of polymerization of chitooligosaccharides prepared was adjusted by the amount of enzyme and hydrolysis time. The average degree of polymerization of chitooligosaccharides obtained was 5.4 when the amount of enzyme was 20 U g −1 for a hydrolysis time of 1.5 h. The results of infrared spectroscopy showed that the oxidative pretreatment and ⊍-amylase degradation did not affect the molecular structure of the chitosan monomer. Using electrospray ionization mass spectrometry analysis, the degree of polymerization of the products was found to be mainly 2-8.CONCLUSIONS: H 2 O 2 oxidative pretreatment can significantly improve the degradation efficiency of ⊍-amylase, and the degree of polymerization of chitooligosaccharide products can be adjusted by regulating the hydrolysis conditions of the enzyme, which has ideal application prospects.
BACKGROUND: High selectivity, fewer reaction steps, and energy conservation are the most important advantages of the enzymatic synthesis of sucrose-6-acetate. To improve the enzymatic synthesis efficiency, a solvent system involving N,N-dimethylformamide (DMF) for lipase-catalyzed synthesis of sucrose-6-acetate was studied.RESULTS: Using Lipozyme TL IM as the catalyst, tert-amyl alcohol was screened out as the auxiliary solvent and complexed with DMF at 1/1 (v/v) ratio. The optimized technology conditions for the enzymatic synthesis of sucrose-6-acetate in the above solvent system were as follows: 100 ml composite solvent with sucrose (2.0%, w/v), vinyl acetate (2.90 M), and Lipozyme TL IM (20 mg/ml) was oscillated at 160 rpm in a water bath at 45 °C for 16 h. The esterification rate and the 6-position esterification rate of sucrose were 88.9 ± 1.3% and 94.2 ± 1.2%, respectively. Raising the sucrose concentration to 7.0% and reacting by mechanical stirring at 45 °C in 1000 ml composite solvent could also achieve ideal esterification effects (esterification rate: 87.2 ± 1.0%, 6-position esterification rate: 90.7 ± 1.6%). CONCLUSIONS: As a reaction medium for the enzymatic synthesis of sucrose-6-acetate, the DMF/tert-amyl alcohol solvent was significantly more efficient than the DMSO composite solvent reported in the literature; therefore, the studied solvent could be an ideal application prospect.
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