Gassing Crystallization at Different Scales: Potential to Control Nucleation and Product Properties

Gassing crystallization is an induced nucleation process during batch cooling crystallization with the aim to control the nucleation step and thus product crystal properties. All previous studies have been made at lab scale and show that the metastable zone or the supersaturation at which gassing is started is crucial for the success of gassing crystallization. Since the metastable zone width depends on many factors, the purpose of this paper was to verify the hypothesis that, especially for parameter combinations which result in broad metastable zone widths, the success of gassing crystallization is independent of crystallizer scale and geometry. The studies were made for the substance system succinic acid/water in a 1 L lab and a 30 L pilot scale crystallizer. The effect of gassing on the metastable zone width and the median diameter was evaluated for varying process parameters (saturation concentration, gassing supersaturation, cooling rate, and stirrer speed) and compared to normal cooling crystallization. After the application of gassing, metastable zone widths were narrower, median diameters were bigger, and reproducibility was enhanced. We found that for process parameters which resulted in broad metastable zone widths the effect of gassing on the median diameter was largest, independent of crystallizer scale and geometry. Gassing crystallization induces nucleation and affects product crystal properties, which works best for process conditions resulting in broad metastable zone widths

Influence of Gassing Crystallization Parameters on Induction Time and Crystal Size Distribution

Gassing in combination with linear cooling profiles is an innovative technology to induce nucleation and control product properties. Previous work found that among gassing parameters like gassing supersaturation, gassing duration, and gas volume flow, gassing supersaturation has an effect on product properties only. This paper investigates why gassing duration and volume flow cannot be used to control product properties. Therefore, the influence of gassing parameters on induction time and final crystal size distributions were evaluated. Experiments were performed using succinic acid/water as model system in a 1 L crystallizer. Compared to normal cooling crystallization, induction time could be reduced by about 60 min by gassing. The difference in the specific bubble surface areas during gassing with the gas volume flows applied was too low to create an effect on the amount of nuclei induced and thus on induction time. Only gassing at different supersaturations resulted in different amounts of nuclei induced and thus affected induction time and crystal size distributions. Varying gassing duration did not change induction time, indicating that nuclei were induced at the beginning of the gassing period only

Design of Median Crystal Diameter Using Gassing Crystallization and Different Process Concepts

The reproducibility of product properties of normal batch cooling crystallizations is often insufficient. For a reliable design of product properties like the median diameter, it is essential to control the nucleation process. An innovative technology to induce nucleation during cooling crystallization is gassing. Therefore, quantification of the influence of gassing and process parameters is important. For this purpose, Design of Experiment approaches were used, investigating a linear cooling profile with constant cooling duration and quadratic cooling profiles with varied cooling duration. Succinic acid/water was used as the model system. The supersaturation where gassing is started was identified as most important design parameter using linear cooling profiles. Using quadratic cooling profiles, the median diameter can be mainly designed by adjusting the cooling duration. By the choice of the cooling profile and gassing supersaturation, it is possible to control the median diameter in a range between 300 and 750 μm. The results show also that independent from the cooling profile, gassing crystallization has an enlarging effect on the median diameter of product crystals. This effect can be used to reduce batch time for crystallization processes

Recombinant and endogenous ways to produce methylated phospholipids in Escherichia coli\textit {Escherichia coli}

$\textit {Escherichia coli}$ is the daily workhorse in molecular biology research labs and an important platform microorganism in white biotechnology. Its cytoplasmic membrane is primarily composed of the phospholipids phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and cardiolipin (CL). As in most other bacteria, the typical eukaryotic phosphatidylcholine (PC) is not a regular component of the $\textit {E. coli}$ membrane. PC is known to act as a substrate in various metabolic or catabolic reactions, to affect protein folding and membrane insertion, and to activate proteins that originate from eukaryotic environments. Options to manipulate the $\textit {E. coli}$ membrane to include non-native lipids such as PC might make it an even more powerful and versatile tool for biotechnology and protein biochemistry. This article outlines different strategies how $\textit {E. coli}$ can be engineered to produce PC and other methylated PE derivatives. Several of these approaches rely on the ectopic expression of genes from natural PC-producing organisms. These include PC synthases, lysolipid acyltransferases, and several phospholipid $\it N$-methyltransferases with diverse substrate and product preferences. In addition, we show that $\textit {E. coli}$ has the capacity to produce PC by its own enzyme repertoire provided that appropriate precursors are supplied. Screening of the $\textit {E. coli}$ Keio knockout collection revealed the lysophospholipid transporter LplT to be responsible for the uptake of lyso-PC, which is then further acylated to PC by the acyltransferase-acyl carrier protein synthetase Aas. Overall, our study shows that the membrane composition of the most routinely used model bacterium can readily be tailored on demand. $\textbf {Key points}$ • Escherichia coli can be engineered to produce non-native methylated PE derivatives. • These lipids can be produced by foreign and endogenous proteins. • Modification of E. coli membrane offers potential for biotechnology and research