Applied Biochemistry and Biotechnology

, Volume 55, Issue 2, pp 95–121 | Cite as

High-Alkaline protease fromBacillus PB92 entrapped in calcium alginate gel

Physicochemical and microscopic studies
  • M. G. Roig
  • D. H. Rashid
  • J. F. Kennedy
Original Articles


High-alkaline protease (HAP) has been entrapped in Manugel DMB (an alginate gel) and assayed with two sizes and types of substrates: neutral protein casein and synthetic chromogenic tripeptide substrate, Z-Gly-Pro-Cit-PNA. Increasing the concentration of calcium chloride used for capsule formation decreased the measured enzyme activity with both substrates. Capsules were found to be stable in water for long periods of time, but they dissolved in both phosphate and carbonate-bicarbonate buffers. The pH vs activity profiles of encapsulated enzyme showed pH optima between 10 and 11 with both substrates. The calcium alginate matrix surrounding the enzyme was quite effective in stabilizing the enzyme at 20–25 °C and even more so at 4°C. Enzyme stability at 50 °C was quite impressive, some enzyme activity being evident even after remaining for 1 wk at this temperature in water. Increasing concentrations of sodium dodecyl sulfate (SDS) were also found to inhibit the protease progressively, whereas a polyhexamethylene biguanidium chloride (PHMBH+Cl-) and SDS:PHMBH+Cl- combination showed the opposite effect.

Optical microscopy, especially polarized light microscopy, provided a sensitive physical means of ascertaining some of the structural properties (sphericity, disorganization or organization, distinct layer enveloping the capsules, intensity of the maltese cross) of the capsules with and without enzyme before and after different chemical treatments and the presence or absense of the substrate.

Index Entries

Protease high-alkaline protease immobilized alginate pH stability microscopy SDS polyhexamethylene biguanidium 


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Copyright information

© Humana Press Inc 1995

Authors and Affiliations

  • M. G. Roig
    • 1
  • D. H. Rashid
    • 2
  • J. F. Kennedy
    • 2
  1. 1.Departamento de Quimica Fisica, Facultad de FarmaciaUniversidad de SalamancaSpain
  2. 2.Birmingham Carbohydrate and Protein Technology Group, Research Laboratory for the Chemistry of Bioactive Carbohydrates and Proteins, School of ChemistryBirmingham UniversityUK

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