Fractionation of transgenic corn for recovery of recombinant enzymes

Abstract

AbstractTwo germ‐separation methods, dry‐milling and density separation by flotation, were evaluated for recovering recombinant β‐glucuronidase (rGUS) that accumulated primarily in the germ of transgenic corn. The dry‐milling process consisted of (i) seed tempering, (ii) degerming with a horizontal‐drum degermer/dehuller, (iii) particle size fractionation with standard sieves, (iv) germ and endosperm separation by roller milling and sifting, and (v) removal of hulls by aspiration. Sieves nos. 5, 6, and 7 retained the majority of germ, and subfractions from these sieves were pooled as a germ‐rich fraction. Mass balances showed that the germ‐rich fraction, which constituted 17% of the total dry‐milled corn weight, contained 49% of rGUS activity and 64% of the total recoverable oil. Germ fractionation by flotation was tested as a proof‐of‐concept method aimed at separating corn fractions based on their difference in specific gravity (sp gr). The process consisted of impact‐grinding of corn kernels followed by density separation using 1.15 or 1.3 specific gravity sodium nitrate solution. The oil‐containing germ fraction floated, whereas the heavier endosperm fraction sedimented. The flotation method was simpler and resulted in higher enzyme recovery, that is, the germ‐rich fraction was 20% (w/w) of the initial corn weight, and accounted for 80% of rGUS activity and 77% of total oil. The sodium nitrate solution did not have an adverse effect on the enzyme activity.