Periodic wet milling as a solution to size-based separation of crystal products from biocatalyst for continuous reactive crystallization

• A 3D in situ filter is designed to separate catalyst carriers and crystals. • Wet milling decreases the size of the crystals and facilitates product discharge. • The impact of separator on slurry withdrawal is investigated. Combining the synthesis and crystallization of a compound into a single step may be beneficial for certain processes. Motivations range from process intensification to improving reaction network yield by shifting an equilibrium limited reaction or protecting an intermediate, as is the case in the biocatalytic synthesis of beta-lactam antibiotics. Adapting such a reactive crystallization process to continuous manufacturing faces a challenge: the continuous separation of crystals from biocatalyst carriers. In this study, a size-based method for this separation problem is proposed. Separation is performed using a filter placed on the reactive crystallization vessel outlet. The primary factors for filter design are carrier and crystal size dictated by the separation condition L crystal < L filter < L carrier . Enzymatic assays confirmed an increasing loss of activity and selectivity for the biocatalyst, Penicillin G Acylase, when large carrier sizes are used for immobilization, limiting the maximum L carrier that can be used in the process. Intermittent wet milling, however, is shown to be effective in reducing the crystal size and allowing the use of sufficiently small carriers while still satisfying the criterion for solid–solid separation . Slurry transfer tests were performed to check the possibility of isokinetic withdrawal when the separator filter was used. Filter size of 300 μm, a carrier size of 300–425 μm, and intermittent wet milling of 2–3 reactor volumes per residence time at 5000 RPM were found to provide an almost complete separation, acceptable biocatalyst kinetics, and an isokinetic crystal product withdrawal.