Selective Removal of DNA from Bioproducts by Polycation‐Immobilized Cellulose Beads

Abstract

This paper describes a method for the selective removal of DNA from various cellular products using columns packed with polycation-immobilized cellulose beads. Polyethyleneimine (PEI), poly-N,N-dimethylaminopropylacrylamide (poly(DAPA)) and poly(1-lysine) (P1L), all of which have cationic properties, were used as the ligands on the beads. Cellufine-GC15 w and -CPC w were used as cellulose matrices. Adsorption of DNA by the beads was determined using a batchwise method or a column method. Each bead type showed high DNA adsorbing activity at pH 7.0 and ionic strengths of m ¼ 0.05 -0.8. The larger the pore size of the beads, the larger the DNA-adsorbing activity. The DNA adsorbing capacities per wet mL of PEI-, poly(DAPA)-and P1L immobilized Cellufine-CPC with large pore sizes, were 3.7, 3.2, and 1.8 mg, respectively. When a protein, such as bovine serum albumin (BSA) or g-globulin, was present in solution with the DNA under physiological conditions (pH 7.0, m ¼ 0.2), the DNA selectivity of the PEI immobilized Cellufine-CPC columns was unsatisfactory, because both the DNA and the protein were adsorbed into the column. In contrast, the poly(DAPA) immobilized Cellufine-CPC column selectively removed DNA from each protein solution contaminated with DNA under similar conditions: the DNA concentration in each treated protein solution was below 10 ng mL 21 , and high recovery of each protein (.92%) was obtained.

EFFECT OF ADSORBENT pKa ON SELECTIVE REMOVAL OF GLUCOAMYLASE FROM UNPASTEURIZED SAKE MATERIALS

“…We already reported [12,13] that polycation-immobilized Cellufine was able to separate DNA and proteins by a column method. The Cellufine was then easily regenerated by washing with 0.2 M sodium hydroxide, followed by 2.0 M sodium chloride.…”

“…To reduce adsorption of the organic acid, it is important to control the adsorbent's electrostatic activities, such as its anion-exchange capacity (AEC) and apparent pK a (pK a,app ). We previously reported [12,13] that the pK a,app of a polymer bead's surface was easily adjusted by changing the type of functional groups and their content.…”

Chromatographic Removal of DNA from Protein Solutions by Cationic Polymer Beads

“…DNA charge is anionic at pH values greater than its pKa (<2) [13]. PEI (pK a : 8.6) [48], poly-N,N-dimethylamino-propylacrylamide (poly(DAPA)) (pK a : 8.9) [49] and poly( -lysine) (P L) (pK a : 7.6) [50][51][52], all have cationic properties, and were used as ligands on the matrices. PEI (pK a : 8.6) [48], poly-N,N-dimethylamino-propylacrylamide (poly(DAPA)) (pK a : 8.9) [49] and poly( -lysine) (P L) (pK a : 7.6) [50][51][52], all have cationic properties, and were used as ligands on the matrices.…”

“…The effect of ionic strength (μ) on the selective adsorption of DNA from a BSA-containing solution by various cationic beads was examined using a batchwise method [13,40,41,49]. The effect of ionic strength (μ) on the selective adsorption of DNA from a BSA-containing solution by various cationic beads was examined using a batchwise method [13,40,41,49].…”

“…It is also necessary to adjust the pore size (M lim ) of the beads for selective adsorption of DNA without adsorption of protein [13,49]. The effect of the pore size of beads on the adsorption capacity of DNA was examined using a column method.…”