Continuous Protein Crystallization in Mixed-Suspension Mixed-Product-Removal Crystallizers

Abstract

The metastable needlelike form of lysozyme can be obtained from continuous crystallization in mixed-suspension mixed-product-removal (MSMPR) crystallizers for conditions at which the stable tetragonal form is obtained from batch crystallization. A 3D-printed airlift crystallizer and stirred tank crystallizer are used, which both approximate an MSMPR crystallizer well. The influence of residence time, precipitant concentration, and agitation mechanism on the attainable solid-state form is presented. The needlelike crystals appear after nucleation of the tetragonal form. An earlier transition from the tetragonal form to the needlelike form is observed in an airlift crystallizer compared to a conventional stirred crystallizer, which is possibly caused by reduced attrition and the presence of a gas–liquid interface in the airlift crystallizer. Furthermore, a long residence time and high precipitant concentration favor the formation of the needlelike crystals. This work demonstrates how the mode of operation of well-mixed crystallizers affects the attainable solid-state form of a model protein. The results generally show the importance of obtaining a quantitative understanding of the dynamic interplay between stochastic primary nucleation processes and deterministic growth and secondary nucleation processes so that future continuous MSMPR crystallizers can be designed and operated robustly to produce protein crystals of a desired solid-state form.

Improving the reproducibility of size distribution of protein crystals produced in continuous slug flow crystallizer operated at short residence time

“…Another recent continuous crystallization using MSMPRC was reported by Mathew Thomas et al [48] By conducting crystallization of lysozyme in both batch and MSMPRC under the same condition (30 g/L lysozyme and 35 or 45 g/L NaCl in pH 4.6, 0.2 M buffer at 18 °C), the authors found that the metastable needlelike form of lysozyme can be obtained from the MSMPRC while only tetragonal from can be produced in batch. The same transition of lysozyme crystals from tetragonal to the needlelike form was seen in a 3D-printed airlift crystallizer that closely mimicked the MSMPRC.…”

“…Conversely, another principally different strategy is moving the bulk crystallization into a continuous platform [10]. By segmentation of nucleation and growth events with precisely controlled supersaturation levels in each segment, improved CSD could be realized [46][47][48]. Continuous platforms can be mainly classified into mixed-suspension mixed-product removal crystallizer (MSMPRC)…”

Electrochemical Oscillatory Baffled Reactors Fabricated with Additive Manufacturing for Efficient Continuous-Flow Oxidations

“…In this regard, additive manufacturing (AM), commonly known as 3D printing, can offer new opportunities for the development of continuous-flow reactors, − advanced reactor architectures, and other continuous-flow applications like crystallizers, , calorimeters, or magnetic resonance probes . The digitalization of the fabrication process enables the coupling of advanced and lean design techniques, like computational fluid dynamics (CFD), to allow the generation of optimized reactor structures and the production of reactor geometries that are easily adaptable to commercial setups .…”

Electrochemical oscillatory baffled reactors fabricated with additive manufacturing for efficient continuous-flow oxidations

“…The higher reactor volume would in turn enable the integration of other enabling technologies for novel applications. 8 In this regard, additive manufacturing (AM), commonly known as 3D printing, can offer new opportunities for the development of continuous-flow reactors, [9][10][11][12] advanced reactor architectures 13 and other continuous-flow applications like crystallizers, 14,15 calorimeters 16 or magnetic resonance probes. 17 The digitalisation of the fabrication process enables the coupling of advanced and lean design techniques, like computational fluid dynamics (CFD), to allow the generation of optimised reactor structures 18 and the production of reactor geometries which are easily adaptable to commercial set-ups.…”