High-Purity ACTH Analogue Synthesis for Large-Scale Production
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing methods for synthesizing adrenocorticotropic hormone (ACTH) of human sequence face challenges such as low yield, low purity, and the presence of many impurities, particularly due to the instability of the N-25 Asn residue under alkaline conditions and the use of corrosive and toxic chemicals in traditional synthesis techniques, making large-scale production difficult.
Innovation Solution
The use of Fmoc solid-phase synthesis technology coupled with optimized conditions for peptide assembly and purification, including specific reagents and chromatographic methods, results in a high-purity ACTH or ACTH analogue with ≥99% purity and ≤0.5% maximum single impurity, achieved through a process involving Fmoc solid-phase synthesis, cleavage with a cocktail, precipitation, and reversed-phase high-pressure liquid chromatography.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If traditional chemical synthesis methods are used to produce ACTH, then production capacity is achieved, but purity is low and impurities are high
Solution Approach 1:
The synthesis process is divided into modular stages: Fmoc-protected amino acid coupling, intermediate purification, and final deprotection. This segmentation allows each stage to be optimized independently, achieving both high purity through controlled synthesis steps and maintained productivity through efficient process flow.
Solution Approach 2:
Amino acids are pre-protected with Fmoc groups before synthesis begins. This preliminary protection prevents unwanted side reactions and ensures high purity of the final product, while the protected intermediates can be synthesized and stored, enabling scalable production.
2Manufacturing precision
If Fmoc solid-phase synthesis is used, then purity is improved to ≥99%, but production scale is limited
Solution Approach 1:
The Fmoc protecting group is selectively removed (taken out) from the synthesized peptide using mild base treatment. This extraction approach allows the protected high-purity intermediate to be synthesized on any scale, then deprotected to yield the final pure product, decoupling purity achievement from production scale limitations.
Solution Approach 2:
The Fmoc-protected ACTH intermediate serves as a mediator that can be synthesized with high purity through solid-phase methods, then converted to the final product. This intermediary approach allows the benefits of solid-phase synthesis (high purity) to be applied to large-scale production through efficient deprotection and purification steps.
3Productivity
If animal extraction is used for ACTH production, then large-scale production is achieved, but immunogenicity risk increases
Solution Approach 1:
Instead of extracting ACTH from animal pituitary glands, the invention synthesizes a human-sequence ACTH copy using Fmoc-protected amino acids. This synthetic copy has identical structure and function to natural ACTH but eliminates immunogenicity risks associated with animal-derived products, while maintaining large-scale production capability through efficient synthesis and purification.
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 significantly enhances the purity and yield of ACTH or its analogue, reducing impurities to ≤1% total, thus addressing the limitations of traditional synthesis methods and enabling large-scale production with improved safety and efficiency.
Implementation Method 1
amino acids are coupled from the C-terminal to the N-terminal according to the amino acid sequence by Fmoc solid-phase synthesis method
Implementation Method 2
the peptidyl-resin is treated with a cleavage cocktail, a solution containing ACTH or ACTH analogue is obtained after cleaving ACTH or ACTH analogue from the resin
Implementation Method 3
The solution containing ACTH or ACTH analogue is treated with precipitation reagent to obtain crude product of ACTH or ACTH analogue
Implementation Method 4
purified by liquid chromatography to obtain a composition containing the ACTH or ACTH analogue
Implementation Method 5
reversed-phase high-pressure liquid chromatography
Data Source
AI summary
Polypeptide preparation methods of preparing high-purity ACTH (human sequence) or analogue and large-scale preparation method thereof are described. The main steps include: amino acids are coupled from the C-terminal to the N-terminal by Fmoc solid-phase synthesis method to obtain the crude ACTH (human sequence) or analogue peptidyl-resin with protective groups, wherein the reaction temperature of C-15 peptide synthesis is 40-60° C. After cleavage and precipitation, the crude product of ACTH (human sequence) or analogue is obtained, and then the high-purity product is obtained by liquid chromatography. The chromatographic purity of ACTH (human sequence) or analogue prepared by the invention is more than 99%, the stability is good, and the yield of the target peptide is ≥63%.