Continuous Spherical Crystallization of Lysozyme in an Oscillatory Baffled Crystallizer Using Emulsion Solvent Diffusion in Droplets

Abstract

Continuous protein crystallization is a cost-effective alternative to traditional chromatographic purification techniques. However, proteins characteristically have slow growth rates, requiring long crystallization times to generate particles large enough for efficient isolation. In this work, spherical crystallization using an emulsion solvent diffusion (ESD) based methodology is proposed to produce large lysozyme agglomerates in an oscillatory baffled crystallizer (OBC). Process sensitivity analysis was performed to investigate the effect of the process parameters on the product's crystal size distribution at two different scales. The concentration of ethanol in the bulk phase was found to have a significant effect on the crystallization kinetics, as its diffusivity drives supersaturation at the droplet interface. Although controlling supersaturation is important in determining product quality, maintaining droplet suspension near the injection site is critical for process longevity, as fouling is most likely to occur in this high supersaturation region. Similarly, in scaling this process, the ratio of droplet diameter to tube diameter plays a significant role in the formation of encrust. Increasing the size of the droplets, relative to the diameter of the system, increases the role of wall effects in distorting the final product quality. Overall, these tunable process parameters make the OBC an ideal platform for spherical protein crystallization.

Increasing the Batch Size of a QESD Crystallization by Using a MSMPR Crystallizer

“…An abundance of publications are available in the literature that describe the use of continuous crystallizers for cooling or antisolvent crystallizations [22][23][24]; however, there are only a few that describe continuous QESD crystallizations [25,26]. Furthermore, to the best of our knowledge, only a single publication exists for QESD crystallizations using a MSMPR setup [13].…”

Optimization of an Oscillatory Flow Crystallizer Based on Heat Transfer and Supersaturation Distribution

“…However, in the traditional stirred tank crystallizer, the flow field distribution is not uniform even under the optimum conditions, typically because the turbulence intensity alters greatly as the size of the tank increases [9]. The oscillatory flow reactor combines the flow control of pulse oscillation with internal geometries to achieve effective control of the mixing and heat transfer process [10,11], which has potential applications in crystallization [8,[12][13][14][15][16].…”

Effects of Scale-Up and Impeller Types on Spherical Agglomeration of Dimethyl Fumarate

“…33 Current continuous processes of producing spherical agglomerates involve the use of (1) microfluidics for glycine by QESD, 40 (2) single-stage MSMPR with a solvent-recycling system for albuterol sulfate by QESD, 41 (3) two-stage MSMPR with decoupling SA from crystallization steps for benzoic acid, 4 (4) oscillatory flow baffled crystallizer (OFBC) for benzoic acid by SA, 42 (5) single-stage MSMPR coupling with impurity separation for fenofibrate by QESD, 43 and (6) OFBC for lysozyme, a protein, by QESD. 44 Besides, a focused beam reflectance measurement probe is the most commonly used PAT tool for online monitoring of a chord length distribution of spherical agglomerates, size determination of droplets, and further identification of agglomeration mechanisms and if a steady-state is achieved. 4,39,41,42 Although continuous operation is associated with easier scalability, higher production rates, and better waste and offspecification product management, 45 reach >50 tons/day according to standard economies of scale.…”

“…To the best of our knowledge, only two research studies have concerned scaling-up of spherical crystallization: one was from 0.5 to 10 L in a batchwise stirred tank using direct SA without crystallization taking place, 6 and the other was from 0.3 to 1.25 L in a continuous OFBC using QESD. 44 A desired size distribution of agglomerates could not be reproduced by simply fixing some of the parameters constant, such as solid loading, bridging liquid and BSR, and residence time, upon scale-up in the stirred tank. However, similar results were harvested upon scale-up in OFBC by translating some parameters, especially axial velocity based on tube diameter, into the system on a larger scale.…”

“…The first continuous SA process was introduced by Kawashima et al in a single-stage mixed suspension, mixed product removal (MSMPR) crystallizer, in which pulverized or milled sulfamethoxazole powder of less than 10 μm was dispersed in water and directly agglomerated into spherical wax matrices of sulfamethoxazole . Current continuous processes of producing spherical agglomerates involve the use of (1) microfluidics for glycine by QESD, (2) single-stage MSMPR with a solvent-recycling system for albuterol sulfate by QESD, (3) two-stage MSMPR with decoupling SA from crystallization steps for benzoic acid, (4) oscillatory flow baffled crystallizer (OFBC) for benzoic acid by SA, (5) single-stage MSMPR coupling with impurity separation for fenofibrate by QESD, and (6) OFBC for lysozyme, a protein, by QESD . Besides, a focused beam reflectance measurement probe is the most commonly used PAT tool for online monitoring of a chord length distribution of spherical agglomerates, size determination of droplets, and further identification of agglomeration mechanisms and if a steady-state is achieved. ,,, …”