Charaterization and hydrodynamic behaviour of modified gelatin: II. Characterization by high performance size exclusion chromatography — comparison with dextrans and proteins
Laboratoire de Physicochimie des Biopolyméres, Université Paris Val de Marne, U M 27 (CNRS), 2, rue Henri Dunant, 94 320, Thiais, France
Abstract:
Gelatin samples obtained by chemical modification (succinylation) are studied by SEC on silica based chromatographic supports. The influence of the pH of eluent mixtures (potassium phosphate added to NaCl) in the range 7-3.3 shows that the void volume peak (VVP) is lowered or even vanishes at pH 3.3 with the 3000 SW (TSK) gel. A process using an ultrasound treatment before injection is reported in order to determine accurately the molecular parameters of gelatin onto TSK gel with a minimal VV P. This peak is attributed to molecular aggregation of a part of the modified gelatin. After disaggregation by ultrasound or heat treatment the results are in good accordance with those obtained by other methods. It is demonstrated that with proteins and dextrans the TSK 3000 SW gel does not agree with the universal calibration curve (logν] ·
versus Kd as reported previously. A single calibration curve is obtained when the Stokes radius is plotted versus Kd. Gelatin fractions are eluted at pH 7 close to this calibration curve. This plot shows that gelatin fractions at pH 3.3 are not eluted by a pure size exclusion mechanism on 3000 SW gel. It is concluded that hydrophobic interactions between fractions of gelatin and the gel explain the high retention of these samples.