Question

explain precipitation process based on the working principle, type of equipment used and operating condition in downstream process of protein

Answer 1

Precipitation in a strictly chemical sense is the transition of a substance from the dissolved state to the non-dissolved state upon the addition of other (dissolved) reagents that lead to the formation of precipitates.

The precipitation process is described by use of the equilibrium or the end-point of the reaction for specified boundary conditions. If precipitation is defined as the transformation of two or more dissolved components to a non-dissolved substance, the so-called precipitate, then dissolution processes and precipitation processes are similar reactions but of opposite directions. The solubility of a product or vice versa the degree to which precipitation will control the dissolved species is determined by the capacity of the solute to accommodate specific ions. The mass law describes the solubility and the corresponding precipitation reaction in terms of a solubility product. The reaction rate with which the precipitation occurs or with which disturbed solubility/precipitation equilibrium is balanced again is finite. There are situations where a change in the stoichiometric parameters in terms of an overdosing of the precipitation-causing reagent leads to improved reaction rates and to increased efficiency.

The application of precipitation as wastewater-treatment process involves a combination of three unit operations: (1) the addition of chemicals to obtain a precipitation,

(2) mixing and flocculation of the chemicals to produce flocs, which settle or float readily, and

(3) the separation process, whereby the precipitated components are removed from the water.

Downstream processing operations, i.e. the processes used to turn a product from its natural state into a pure protein, can be divided into four steps.

The first step involves the isolation, concentration and stabilisation of the target protein (capturing), followed by the removal of viruses. The next step involves the removal of contaminations, such as nucleic acids, unwanted proteins and endotoxins (intermediate purification).

The final processing step involves the removal of trace contaminants that might compromise the safety of the product (polishing). The typical operations are filtration and precipitation, as well as (column) chromatrophic methods, wherein a broad range of methods and materials are available.

The purification of the target protein or molecule depends on its form, size, solubility, surface charge or its biospecific affinity to binding partners. This also increases the number of processes that can be used to purify the proteins. Considering the enormous cost and time pressure, as well as the production-plant-specific constraints, manufacturers are having a tough time developing optimal purification steps for the target molecule or protein. Usually, these processes are developed according to the trial and error principle and are established as soon as the process "functions".

It is difficult to omit steps in the purification process and not compromise thereby the safety of the protein (such as the removal of viruses or DNA). However, representatives of the pharmaceutical industry believe that it is possible to optimise the procedure and potentially also combine individual or partial steps of the process. Column chromatography is usually regarded as the major cost driver. Besides for the fixed costs, the column membranes that are used for the purification of the proteins swallow a large proportion of the costs.

This sheet contains the recommended operating conditions and the specifications for TSKgel Protein A-5PW columns

OPERATING CONDITIONS

1. Shipping Solvent: Aqueous solution contains 20% ethanol

2. Max. flow rate: 4.0 mL/min (standard operating flow rate: 2.0 mL/min)

3. Max. pressure: 2.0 MPa