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Stroma-free hemoglobin solutions purified by chloroform and pasteurization.
T I Pristoupil1, T Marík, J Suttnar
1Institute of Hematology and Blood Transfusion, Prague, Czechoslovakia.
The International Journal of Artificial Organs
|June 1, 1990
Summary
Chloroform treatment offers a stable method for processing stroma-free hemoglobin (SFH) solutions, balancing purification with oxidative stability. Freeze-drying with fructose or sucrose further protects SFH, presenting a viable production compromise.
Area of Science:
- Biochemistry
- Biotechnology
- Materials Science
Background:
- Stroma-free hemoglobin (SFH) solutions are potential blood substitutes, but their production requires efficient purification methods.
- Current methods for processing outdated red blood cells (RBCs) into SFH need evaluation for efficacy, speed, and quality control.
- Ensuring the stability and minimizing oxidation of SFH during storage and processing is crucial for its application.
Purpose of the Study:
- To re-evaluate processing methods for native stroma-free hemoglobin (SFH) solutions based on current production and quality standards.
- To compare the efficacy of chloroform treatment and pasteurization in removing impurities from RBC hemolysates.
- To assess the impact of these purification methods on the stability and spontaneous methemoglobin formation of SFH during storage, particularly after freeze-drying.
Main Methods:
- Evaluated chloroform treatment and pasteurization for processing outdated red blood cells (RBCs) into stroma-free hemoglobin (SFH).
- Assessed purification efficacy in removing stromata, phospholipids, and non-heme proteins using techniques like SDS-PAGE and isoelectric focusing.
- Compared the influence of both treatments on spontaneous methemoglobin formation in freeze-dried SFH, with and without cryoprotectants (fructose, sucrose).
Main Results:
- Pasteurization yielded superior purification and homogeneity but increased susceptibility to oxidation, likely due to heat-inactivated radical scavengers.
- Chloroform treatment resulted in less purified SFH but demonstrated enhanced stability against oxidation.
- Both fructose and sucrose effectively protected SFH from oxidation during freeze-drying, showing equal efficacy.
Conclusions:
- Chloroform treatment presents a technically feasible and rapid method for SFH purification, offering better oxidative stability compared to pasteurization.
- Pasteurization provides higher purity but compromises SFH stability due to potential inactivation of protective enzymes.
- Combining chloroform treatment with freeze-drying using fructose or sucrose represents a promising technological compromise for SFH production, warranting further investigation.