Intramolecular Cross-linked Hemoglobin for Stable Oxygen Delivery
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Solution Overview
Problem
Existing methods for stabilizing hemoglobin outside red blood cells result in instability, leading to renal injury, vasoconstriction, and the formation of met-hemoglobin, which cannot oxygenate tissues, due to the breakdown of tetramers into dimers and the presence of protein impurities.
Innovation Solution
A method involving controlled hypotonic lysis, ultrafiltration, and cross-linking with bis-3,5-dibromosalicy fumarate to create high-temperature stable, nonpolymeric cross-linked tetrameric hemoglobin, blocking sulfhydryl groups to prevent met-hemoglobin formation, and packaging in oxygen-impermeable materials to maintain stability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If cross-linking techniques are used to create polymeric hemoglobin to increase circulatory half-life, then the circulatory half-life is improved, but met-hemoglobin formation increases and oxygenation capability deteriorates
Solution Approach 1:
The patent segments the cross-linking approach by using intramolecular cross-linking within tetrameric hemoglobin units rather than intermolecular cross-linking that forms polymers. This is achieved through controlled cross-linking conditions that target intra-tetramer bonds while preventing inter-tetramer polymerization, thus maintaining circulatory half-life extension without compromising oxygenation capability through met-hemoglobin formation
Solution Approach 2:
The patent changes the parameters of cross-linking by using specific cross-linking agents and controlled reaction conditions that favor intramolecular bonding over intermolecular bonding. This parameter control allows tetrameric stabilization without polymeric formation, resolving the contradiction between extended circulation and maintained oxygenation function
2Stability of the object's composition
If tetrameric hemoglobin is stabilized to prevent breakdown, then circulatory stability is improved, but dimer formation increases causing renal injury
Solution Approach 1:
The patent applies preliminary cross-linking action to stabilize tetrameric hemoglobin before it can break down into dimers during circulation. By pre-establishing intramolecular cross-links within the tetramer structure, the hemoglobin is protected from dissociation into harmful dimeric units that would otherwise cause renal injury upon filtration
Solution Approach 2:
The patent converts the potential harm of cross-linking (which could lead to polymerization and loss of function) into a benefit by carefully controlling cross-linking parameters to achieve intramolecular stabilization. This controlled approach transforms what could be a harmful polymeric formation into a beneficial tetramer stabilization that prevents renal injury
3Adaptability or versatility
If sulfhydryl groups are left unblocked, then natural hemoglobin function is maintained, but vasoconstriction occurs due to endothelium-derived relaxing factor binding
Solution Approach 1:
The patent introduces sulfhydryl-blocking agents as intermediaries that selectively bind to reactive sulfhydryl groups on hemoglobin. This intermediary action prevents direct binding between endothelium-derived relaxing factor and hemoglobin sulfhydryl groups, thereby eliminating vasoconstriction while preserving oxygen transport function through the blocked cysteine sites
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
The method produces a stable, purified tetrameric hemoglobin that prevents renal injury, vasoconstriction, and met-hemoglobin formation, effectively oxygenating tissues and enhancing cancer treatment and organ preservation.
Implementation Method 1
cross-linking with bis-3,5-dibromosalicy fumarate to create high-temperature stable, nonpolymeric cross-linked tetrameric hemoglobin
Implementation Method 2
blocking sulfhydryl groups to prevent met-hemoglobin formation
Implementation Method 3
A first ultrafiltration process is performed using an ultrafiltration filter configured to remove impurities
Implementation Method 4
The filtered red blood cell fraction is washed to remove plasma protein impurities. The washed red blood cells are disrupted by a controlled hypotonic lysis
Implementation Method 5
packaging in oxygen-impermeable materials to maintain stability
Data Source
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AI summary
A high temperature-stable and highly purified cross-linked (optionally > 70% ß-ß linked) tetrameric hemoglobin with high efficiency of oxygen delivery suitable for use in mammals without causing renal injury and vasoconstriction is provided. The dimeric form of hemoglobin is degenerated and purification processes are performed on red blood cells from whole blood. Controlled hypotonic lysis in an instant cytolysis apparatus prevents lysis of white blood cells. Nucleic acids from white blood cells and phospholipids impurities are not detected. Blocking of reactive sulfhydryl groups by a sulfhydryl reagent is performed in an oxygenated environment. Flowthrough column chromatography removes different plasma protein impurities. N-acetyl cysteine is added to the cross-linked tetrameric hemoglobin to maintain a low level of met-hemoglobin. The stabilized hemoglobin is preserved in an infusion bag with aluminum overwrap to prevent formation of inactive met-hemoglobin from oxygen intrusion. The product finds use in tissue oxygenation and cancer treatment.