SynopsisThe combination of an agarose gel (Bio-Gel A) and a dioxane-water (1:l) solvent system allowed the fractionation, on a preparative scale, of a very polydisperse, nonderivatized lignin preparation (enzymatically liberated lignin prepared from sweetgum sapwood with Lenzites trabea). Three fractions differing markedly in molecular weight were obtained. A gel of crosslinked alkylated dextran (Sephadex LH-20) with the same solvent system allowed division of the lowest molecular weight fraction into two fractions. These materials were characterized by measurements of intrinsic viscosity and number-average molecular weights in dimethylformamide and dioxane-water.It was established that the two highest molecular weight fractions were associated in an average trimeric form in dioxane-water (1 :1) as compared to the form (considered to be molecular) that occurred in dimethylformamide. Molecular size distributions and eluant volumes of the fractions were determined with a Sephadex G-1Wformamide system, the latter being one of the most powerful nonaqueous solvents for lignin. Adsorption effects were known to be absent in this case, and the lignin molecules were considered to be unassociated in formamide. The four fractions were distinguishable with the f0rmamide-G-100 system, thus indicating that the original fractionation was based on molecular size. The enzymatically liberated lignin contained molecules that comprised a continuum of molecular weights from approximately monomeric to molecules that were a t the limit of the solvating power of dioxane-water (1:l) and dimethylformamide. Limited physicochemical data were consistent with a compact, approximately spherically symmetric shape of the lignin in solution.