A1PI Extraction from Plasma Using Salt Precipitation
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for isolating alpha-1-proteinase inhibitor (A1PI) from blood products face challenges such as protein denaturation due to the use of ethanol and salts, and require complex purification steps, including the separation of cryoprecipitate from plasma, which can lead to low yields and inefficiencies.
Innovation Solution
A method involving the addition of salts to blood products to create intermediates with specific salt concentrations for separation into supernatants and pastes, followed by affinity and ion exchange chromatography, eliminating the need for cryoprecipitate separation and reducing protein loss, allowing for high-yield extraction of A1PI without ethanol-based processes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If ethanol is used to extract A1PI from plasma proteins, then A1PI can be separated from the solution, but the desirable proteins such as A1PI are denatured
Solution Approach 1:
The patent changes the extraction parameter from ethanol to salts (sodium citrate, ammonium sulfate) with specific concentrations (11-20 wt%, 21-30 wt%). This parameter substitution maintains separation efficiency while preserving protein integrity, as salts do not denature proteins like ethanol does.
Solution Approach 2:
The patent uses simple, inexpensive salt solutions instead of expensive and harmful ethanol. The salt-based extraction system is replaceable and does not require complex recovery processes, making it a practical substitute that achieves the same separation function without the harmful effects.
2Ease of manufacture
If cryoprecipitate is separated from plasma before salt precipitation, then the purification process follows conventional methods, but the process becomes complex and yields are reduced
Solution Approach 1:
The patent merges the cryoprecipitate separation step with the salt precipitation steps by adding salts to the entire plasma mixture before freezing. This combines multiple separation functions into one integrated process, simplifying the manufacturing procedure while maintaining or improving A1PI yield by preventing protein loss during intermediate separations.
Solution Approach 2:
The salt precipitation process serves multiple functions simultaneously: it precipitates unwanted proteins, concentrates A1PI in the supernatant, and eliminates the need for separate cryoprecipitate handling. This multi-functional approach reduces process complexity and improves overall productivity.
3Quantity of substance
If high salt concentrations (above 11 wt %) are added to previously frozen blood products, then protein products can be extracted from supernatants with higher salt content, but conventional methods limit salt concentration to below 11 wt %
Solution Approach 1:
The patent changes the salt concentration parameter from the conventional limit of 11 wt% to higher concentrations (21-30 wt%). This parameter expansion allows extraction of additional protein products from the supernatants that would remain dissolved at lower salt concentrations, thereby increasing the quantity of extractable protein.
Solution Approach 2:
The patent performs salt addition and precipitation before the final extraction step, creating supernatants with predetermined high salt concentrations. This preliminary action prepares the matrix for subsequent protein extraction, allowing recovery of proteins that would otherwise be inaccessible due to solubility at lower salt concentrations.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method enables high-yield production of A1PI with minimal pre-processing, maintaining protein integrity and simplifying the purification process by using salt concentrations that minimize A1PI precipitation, thereby improving the overall efficiency and purity of the protein extraction.
Implementation Method 1
adding a salt to the blood product to produce a first intermediate, wherein the salt comprises between 11-20 wt % of the first intermediate
Implementation Method 2
separating a third intermediate from the second supernatant by affinity chromatography
Implementation Method 3
separating the third intermediate by ion exchange chromatography to produce an eluate containing the protein product
Data Source
AI summary
A method of producing protein products including alpha-1-proteinase inhibitor, gamma globulin, albumin, and other proteins from plasma includes steps of: (1) adding a salt to the blood product to produce a first intermediate, wherein the salt comprises between 11-13 wt % of the first intermediate; (2) separating the first intermediate to produce a first supernatant and a first paste; (3) adding a salt to the first intermediate to produce a second intermediate, wherein the salt comprises between 21-23 wt % of the second intermediate; (4) separating the second intermediate to produce a second supernatant and a second paste; (5) separating a third intermediate from the second supernatant by affinity chromatography; and (6) separating the third intermediate by ion exchange chromatography to produce an eluate containing the protein product. Advantageously, the inventive methods are simple and produce alpha-1-proteinase inhibitor, gamma globulin, albumin, and other proteins in high yields.