Determination of crystal growth rate for porcine insulin crystallization with CO2 as a volatile acidifying agent

Crystallization is controlled by two steps that determine the quality and the final size of the product, nucleation and growth, which are functions of supersaturation. Recently, Hirata et al. [1] crystallized insulin using CO2 as a volatile acid to impose supersaturation on the system. The objective of the present work was to determine the growth kinetics of insulin crystallization in 50 mM NaHCO3 solution with 0.4 mM ZnCl2 in a CO2 atmosphere at 15 °C, adjusting the parameters of the equation G = kg × Sg to the experimental data. The solubility of insulin in the NaHCO3/CO2/ZnCl2 system at 15 °C was determined as a function of pH in the range of 6.30–7.34. The crystal growth data allowed determination of the growth order “g” (g = 2.9). Although protein crystallization has some features that differ from the crystallization of less complex molecules, the apparent growth kinetics of insulin were successfully analyzed here with the same empirical methods used for small molecules, which can easily be scaled up for industrial applications to achieve specific size and purity, the goals of industrial crystallization. The method used in this work is a useful tool for describing and simplifying optimization of industrial protein crystallization processes.

The growth kinetics for insulin crystallization with CO2 were successfully analyzed here with the same empirical methods used for small molecules, although protein crystallization has some features that differ from the crystallization of less complex molecules. The G value determined (1.5 × 10−10 m/s) is in agreement with results found in the literature.Figure optionsDownload as PowerPoint slideHighlights
► Crystallization of insulin with CO2 was modeled with the equation G = kg × Sg.
► The crystal growth order g determined was equal to 2.9.
► The g = 2.9 is in agreement with g values for compounds of low solubility (>2.0).