Immobilization Of Neutrase And The Application In Cigarette Flavor

Tobacco flavor, one of the most important constituents of modern cigarette products, is mainly prepared by two ways:organic synthesis and natural plants extracting. For the existing dispute against the security of the synthesized flavor, much attention has been focused on the extractions from natural plants. Despite active substance with fragrance can be well extracted by traditional extracting technology, it is inevitable that some macromolecules like protein, cellulose, pectin and so on, will be introduced into the resulting extraction at the same time. These macromolecules will strongly affect the taste of cigarette and bring instability to the apparent character of the flavor. At this point, enzymatic degradation of those macromolecules by corresponding enzymes has been applied to improve the quality of those flavors for tobacco use. Since enzyme is a protein in chemical nature, use of free enzyme will introduce a new source of contaminate into the extraction. Herein, immobilized enzyme can solve the problem very well for its superiority over free enzyme. In this work, a series of oxidized alginate gel and a few magnetic composite silica carriers were prepared. The application of these carriers on neutrase immobilization was also assessed. The main contents of this paper are as follows:(1) A series of periodate oxidized alginate gel with different oxidation degree were prepared, and their application on neutrase immobilization was studied. The result shows that the immobilization pH, enzyme loading and the oxidation degree will affect the immobilization to a great extent.(2) A few carriers were prepared via simple copolymerization between 3-amino propyltriethoxysilane and tetraethyl orthosilicate on the surface of magnetic iron oxide, which were subsequently crosslinked by glutaraldehyde or branched by methyl acrylate. Neutrase immobilized on crosslinked carrier gave the best immobilization efficiency. The optimal conditions were: 0.5% glutaraldehyde: 30mL, crosslinking time: 2h, 15mg neutrase /0.1g carrier; the stirring time: 50 min. The activity of acquired immobilized neutrase was as high as 2500 U/g carrier.(3) Yerbadetajo herb extraction was prepared for enzymatic catalysis. The enzymatic degradation was optimized: stirring time: 8h, 3g immobilized neutrase, 6g glucose. And the subsequent Maillard reaction process was done by heating the mixture at 80℃for 4 h. In sharp contrast to the blank sample, the resulting extraction could improve the aroma quantity,taste and irritancy significantly. GC-MS analysis showed that heterocyclic volatile ingredients increase both in kinds and content.In conclusion, this immobilized neutrase showed its high efficiency in the enzymatic degradation and might have widespread application in this field.