Alkaline phosphatase encapsulated in gellan-chitosan hybrid capsules.
Alkaline phosphatase (ALP) was encapsulated in gellan-chitosan polyion complex (PIC) capsules using a convenient procedure. The recovery of ALP was about 50% when the capsules were prepared by dropping a solution of ALP and gellan mixture (ALP/gellan) into a chitosan solution. When p-nitrophenyl phosphate (p-NPP) and 5-bromo-4-chloro-3-indolyl phosphate (BCIP) were incubated with ALP/gellan-chitosan capsules as substrates for ALP, the transparent colorless capsules changed to yellow and blue, respectively. The encapsulation of ALP into the PIC capsules was also confirmed by SDS-PAGE and immunoblot analyses. The ALP and polypeptides of more than 30 kDa remained without release even after incubation at 4 degrees C for 14 d. The biochemical properties of the encapsulated ALP activity were similar to those of the intact enzyme. When the solution containing p-NPP was loaded on a column packed with ALP/gellan-chitosan capsules at 27 degrees C, approximately 75% of p-NPP was hydrolyzed by passing through the column. No significant leakage of ALP was observed during the procedure, indicating that the capsules were resistant to pressure in the chromatographic operation. Furthermore, 70% of the hydrolytic activity of the packed capsules remained after storage at 4 degrees C for one month. These results suggest that the polyion complex capsules could be useful materials for protein fixation without chemical modification. [Diagram: see text] Encapsulation of ALP into PIC capsules and the morphological changes seen in the absence of the ALP substrate and in the presence of p-NPP and BICP.
Fujii
Toshihiro
T
Department of Kansei Engineering, Faculty of Textile Science and Technology, Shinshu University, Tokida 3-15-1, Ueda 386-8567, Japan. fujiit1@shinshu-u.ac.jp
Ogiwara
Daisuke
D
Ohkawa
Kousaku
K
Yamamoto
Hiroyuki
H
D000469:Alkaline Phosphatase / Q000737:chemistry*; D019149:Bioreactors; D002214:Capsules / Q000737:chemistry; D048271:Chitosan / Q000737:chemistry; D015151:Immunoblotting; D010539:Permeability; D011135:Polysaccharides, Bacterial / Q000737:chemistry
D002214:Capsules; D011135:Polysaccharides, Bacterial; C048288:gellan gum; D048271:Chitosan; D000469:Alkaline Phosphatase