Temperature stability of Poly-[hemoglobin-superoxide dismutase-catalase-carbonic anhydrase] in the form of a solution or in the lyophilized form during storage at −80 °C, 4 °C, 25 °C and 37 °C or pasteurization at 70 °C

Abstract

Polyhemoglobin-superoxide dismutase-catalase-carbonic anhydrase (Poly-[Hb-SOD-CAT-CA]) contains all three major functions of red blood cells (RBCs) at an enhanced level. It transports oxygen, removes oxygen radicals and transports carbon dioxide. Our previous studies in a 90-min 30 mm Hg Mean Arterial Pressure (MAP) sustained hemorrhagic shock rat model shows that it is more effective than blood in the lowering of elevated intracellular pCO 2 , recovery of ST-elevation and histology of the heart and intestine. This paper is to analyze the storage and temperature stability. Allowable storage time for RBC is about 1 d at room temperature and 42 d at 4C. Also, RBC cannot be pasteurized to remove infective agents like HIV and Ebola. PolyHb can be heat sterilized and can be stored for 1 year even at room temperature. However, Poly-[Hb-SOD-CAT-CA] contains both Hb and enzymes and enzymes are particularly sensitive to storage and heat. We thus carried out studies to analyze its storage stability at different temperatures and heat pasteurization stability. Results of storage stability show that lyophilization extends the storage time to 1 year at 4 C and 40 d at room temperature (compared to respectively, 42 d and 1 d for RBC). After the freeze-dry process, the enzyme activities of Poly-[SFHb-SOD-CAT-CA] was 100 ± 2% for CA, 100 ± 2% for SOD and 93 ± 3.5% for CAT. After heat pasteurization at 70 C for 2 h, lyophilized Poly-[Hb-SOD-CAT-CA] retained good enzyme activities of CA 97 ± 4%, SOD 100 ± 2.5% and CAT 63.8 ± 4%. More CAT can be added during the crosslinking process to maintain the same enzyme ratio after heat pasteurization. Heat pasteurization is possible only for the lyophilized form of Poly-[Hb-SOD-CAT-CA] and not for the solution. It can be easily reconstituted by dissolving in suitable solutions that continues to have good storage stability though less than that for the lyophilized form. According to the P50 value, Poly-[SFHb-SOD-CAT-CA] retains its oxygen carrying ability before and after long-term storage.ARTICLE HISTORY

ARTIFICIAL CELL evolves into nanomedicine, biotherapeutics, blood substitutes, drug delivery, enzyme/gene therapy, cancer therapy, cell/stem cell therapy, nanoparticles, liposomes, bioencapsulation, replicating synthetic cells, cell encapsulation/scaffold, biosorbent/immunosorbent haemoperfusion/plasmapheresis, regenerative medicine, encapsulated microbe, nanobiotechnology, nanotechnology

“…contaminated donor blood. It is only then that intense R&D on blood substitutes was belatedly carried out around the world [20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35]. It was found out too late that blood substitute requires the same long-term research as in any other medical research for cancer and other diseases.…”

“…This complex has no blood groups. The lyophilized preparation can be heat pasteurized at 68 F for 2 h [31]. This can be important if there is a need to inactivate H.I.V.…”

“…Figure 7 Enzymes inside artificial cells can act on external permeant substrates while avoiding protein sensitization, anaphylactic reaction, or antibody production with repeated injection [2][3][4]6,8,9] (Figure 13). Chang's groups has been investigating the use of artificial cells for enzyme therapy since 1964 [2][3][4][5][6][7][8][9][10]25,26,[29][30][31][32][33][51][52][53][54][55] (Figure 13).…”

Long term safety and immunological effects of a nanobiotherapeutic, bovine poly-[hemoglobin-catalase-superoxide dismutase-carbonic anhydrase], after four weekly 5% blood volume top-loading followed by a challenge of 30% exchange transfusion

“…SFHb contains Hb and all the red blood cell enzymes, including CAT, SOD and CA. To enhance the red blood cell enzymes, commercial CAT, SOD and CA are added for the preparation of poly-[Hb-CAT-SOD-CA] [20][21][22][23]. For avoiding the high cost of commercial enzymes, a cost-effective novel enzyme extraction method has been established to obtain these three enzymes from SFHb simultaneously [24,25].…”

In vivoevaluation of electron mediators for the reduction of methemoglobin encapsulated in liposomes using electron energies produced by red blood cell glycolysis

“…Moreover, its use entails various complicated, time-consuming and expensive blood tests, creating persistent conditions of blood supply shortage. Alternative blood substitutes such as haemoglobinbased oxygen carriers (HBOCs) are believed to have the capacity to ease blood supply shortages and alleviate many blood transfusion-related issues [1][2][3][4][5][6]. Actually, HBOCs can be produced in a way that eliminates viral infections such as HIV and hepatitis.…”