Method

Alginic acid can be either water soluble or insoluble depending on the type of the associated salt. The salts of sodium, other alkali metals, and ammonia are soluble, whereas the salts of polyvalent cations, e.g., calcium, are water insoluble, with the exception of magnesium. The alginate polymer itself is anionic (i.e., negatively charged) overall. Polyvalent cations bind to the polymer whenever there are two neighboring guluronic acid residues. Thus, polyvalent cations are responsible for the cross-linking of both different polymer molecules and different parts of the same polymer chain. The process of gelation, simply the exchange of calcium ions for sodium ions, is carried out under relatively mild conditions. Because the method is based on the availability of guluronic acid residues, which will not vary once given a batch of the alginate, the molecular permeability does not depend on the immobilization conditions. Rather, the pore size is controlled by the choice of the starting material.

     2 Na(Alginate) + Ca++ -------> Ca(Alginate)2 + 2 Na+

Alginate is currently widely used in food, pharmaceutical, textile, and paper products. The properties of alginate utilized in these products are thickening, stabilizing, gel-forming, and film-forming. Alginate polymers isolated from different alginate sources vary in properties. Different algae, or for that matter different part of the same algae, yield alginate of different monomer composition and arrangement. There may be sections of homopolymeric blocks of only one type of monomer (-M-M-M-) (-G-G-G-), or there may be sections of alternating monomers (-M-G-M-G-M-). Different types of alginate are selected for each application on the basis of the molecular weight and the relative composition of mannuronic and guluronic acids. For example, the thickening function (viscosity property) depends mainly on the molecular weight of the polymer; whereas, gelation (affinity for cation) is closely related to the guluronic acid content. Thus, high guluronic acid content results in a stronger gel.

List of Reagents and Instruments

A. Equipment

• Beakers
• Graduated cylinder
• Balance
• Pipets
• Syringe

B. Reagents

• Alginic acid, sodium salt
• CaCl₂
• Enzyme

Procedures

1. Dissolve 30g of sodium alginate in 1 liter to make a 3% solution. See Note 1.
2. Mix approximately 0.015 g of enzyme with 10 ml of 3% (wt.) sodium alginate solution. The concentration of sodium alginate can be varied between 6-12 % depending on the desired hardness. See Note 2.
3. The beads are formed by dripping the polymer solution from a height of approximately 20 cm into an excess (100 ml) of stirred 0.2M CaCl₂ solution with a syringe and a needle at room temperature. The bead size can be controlled by pump pressure and the needle gauge. A typical hypodermic needle produces beads of 0.5-2 mm in diameter. Other shapes can be obtained by using a mold whose wall is permeable to calcium ions. Leave the beads in the calcium solution to cure for 0.5-3 hours.

Notes

1. Sodium alginate solution is best prepared by adding the powder to agitated water, rather than vice versa, to avoid the formation of clumps. Prolonged stirring may be necessary to achieve the complete dissolution of sodium alginate. After sodium alginate is completely dissolved, leave the solution undisturbed for 30 minutes to eliminate the air bubbles that can later be entrapped and cause the beads to float.
2. Although not necessary, the beads may be hardened by mixing some amines in the sodium alginate solution and cross-linking with glutaraldehyde.

Discussions

References

1. S. Ohlson, P.-O. Larsson, and K. Mosbach, Steroid transformation by living cells immobilized in calcium alginate, European J. Appl. Microbiol. Biotechnol., 7, 103, 1979.
2. J. Vaija, et al., Appl. Biochem. Biotechnol., 7, 51, 1982.
3. J. M. Lee and J. Woodward, Biotech. Bioeng., 25, 2441, 1983.