Purification process precipitation step

The first step of NCA polymerization is usually accomplished by the use of nucleophilic initiators. These initiators can be alkoxides, alcohols, amines, transition metals, and even water [53,54]. In order to synthesize a copolymer diblock, the polymerization of the second block and its connection to the previously formed block are performed in a single process. This is achieved by initiating the polymerization of the second NCA monomer using the first homopolypeptide as a macroinitiator. Precipitation and purification processes follow to isolate the... 

Solvent extraction—purification of wet-process phosphoric acid is based on preferential extraction of H PO by an organic solvent vs the cationic impurities present in the acid. Because selectivity of acid over anionic impurities is usually not sufficient, precipitation or evaporation steps are included in the purification process for removal. Cmde wet-process acid is typically concentrated and clarified prior to extraction to remove post-precipitated sludge and improve partition of the acid into the solvent. Concentration also partially eliminates fluoride by evaporation of HF and/or SiF. Chemical precipitation of sulfate (as Ba or Ca salts), fluorosiUcates (as Na salt), and arsenic (as sulfides) may also be used as a prepurification step preceding solvent extraction. 

To determine the efficiency of aminoacylation of [14C]Phe-tRNA, 5 fil aliquots of the aminoacylation mixture are withdrawn before and after the reaction the samples taken from the reaction mixture at the end of the incubation are spotted onto 3-MM paper discs (Schleicher Schuell) and processed by the cold TCA precipitation method, while the sample taken before the reaction is spotted on a paper disc pretreated empty by the same cold TCA procedure. Determination of the radioactivity present on these filters by liquid scintillation counting allows one to calculate the aminoacylation efficiency of the reaction (which, for phenylalanine, should be >2% of total tRNA). The specific activity of the [14C] Phe-tRNA can be determined after one-step purification of Phe-tRNA by BD cellulose chromatography (Gillam et al., 1968), followed by determination of the radioactivity and of the A260. 

For these studies a cell paste of E. coli K-12 containing the plasmid pBR322 was processed by ammonium sulfate precipitation and ion exchange chromatography to mimic typical purification process steps. The isolated proteins were then used as the immunogen for three groups of three rabbits each. 

The sequence of stages in a purification process usually starts with low resolution steps such as precipitation and liquid-liquid extraction, which... 

Precipitation is used as a separation step during the early stages of a purification process, usually followed by chromatographic separations, and also as a concentration method prior to an analysis or to a subsequent purification step. 

As an example of concentration from a large volume, a silver radioanalyte can be precipitated as AgCl in an early step. This process combines purification with concentration because only palladium follows silver under specified conditions. 

The development of a qualified down-scale model of a process module is integral to the approach of process validation using bench-scale experiments, as described earlier. We have developed down-scale models of process steps ranging from various types of process chromatography for protein purification to separation by precipitation and filtration. These down-scale models have been utilized to evaluate the effects of relevant process parameters on product-quality attributes. The normal logical sequence of process development, of course, is bench scale to pilot scale to full scale. However, for many plasma protein purification processes, a reverse order needs to be followed. As licensed full-scale processes already exist, the full-scale process steps need to be scaled down to construct small process models in order to evaluate the robustness of process parameters on the product without impacting full-scale production. These models can also be utilized to evaluate process changes, improvements, and optimizations easily and economically. 

Example 2 Robustness of Pasteurization and Precipitation Steps in a Fractionation/Purification Process... 

The production process for protein prepared by fractionation/ purification includes a heat inactivation step to ensure viral safety of the product. The heat inactivation step is followed by a precipitation step that further purifies the product. The purpose of our study was to determine whether the process steps are robust within the parameter ranges routinely used in manufacturing. 

The topic of this paper is the modeling of events occurring in the recovery of proteins and in the conditioning of the product streams for further purification using precipitation. The typical goal of downstream processing is the recovery of a desired product from a very dilute stream while minimizing the loss of the material in what is usually a multi-step separation process. Precipitation enables an early concentration of the product and can simultaneously serve to remove contaminants that would interfere with subsequent purification steps. Further, the wide variety of potential... 

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