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Red blood cells (RBCs) possess intact cyto-architectures while haemoglobin (Hb) is a cell-free, homogeneous solution. Both RBCs and Hb are generalized oxygen carriers. In this paper, kinetic studies on oxygen-releasing of high concentration of Hb and RBCs under various conditions were carried out regarding Hb and RBCs as fluids. Among them, Hb under specific conditions was seen as the simplest Hb-based oxygen carrier (HBOC), Also, factors affecting the oxygen releasing of Hb and RBCs, including osmotic pressure, viscosity and allosteric agent, have been well studied. Analysis of the results from the measurement above showed that kinetics of oxygen releasing of either pure Hb or the simplest HBOCs was obviously different from that of RBCs. The oxygen-releasing time of Hb was shorter and the oxygen-releasing rates of Hb were quicker than those of RBCs under various conditions. Therefore, as fluids, only by changing the milieus it exists in, Hb could not achieve the expected oxygen-releasing effect on the microcirculation so well as RBCs do in the same system, irrespective of the interaction between the fluids and blood vessels. Furthermore, kinetic properties of HBOCs must be considered and matched with those of RBCs in the study of HBOCs.

A rapid and sensitive high-performance liquid chromatography (HPLC) method for vitamin C (Vc) determination was developed in hemoglobin-based oxygen carriers (HBOCs) solution. After polymerized hemoglobin precipitation, Vc could be tested out within 5 min. The standard curve was linear in Vc quantity of 0.00-3.00 μg. Vc recovery was 103.56-108.68%, and relative standard deviation values of the repeatability and precision were below 1.00%. The limits of detection and limits of quantification were 0.002 ppm and 0.01 ppm, respectively. Furthermore, the application of established HPLC methods in methemoglobin reduction by Vc under different pH indicated that higher pH values could accelerate the reduction. This study suggested the potential application of established HPLC method in HBOCs research and development.

This study was designed to investigate the effect of polymerized human placenta haemoglobin (PolyPHb) on cardiac dysfunction after severe burns. A total of 60 male Sprague-Dawley rats were randomly divided into 3 groups: Sham, Burn and Burn + PolyPHb groups. Rats were subjected to third-degree burns to 30% of total body surface area and the haemodynamics, cardiac enzyme release and aortic endothelium ultrastructure/function were measured. PolyPHb (0.5 gHb/kg) greatly improved mean arterial pressure, left ventricular developed pressure (LVDP), maximum LVDP increase and decrease rate and reduced left ventricular end-diastolic pressure as compared to the Burn group. The plasma levels of cardiac enzyme including CK-MB and troponin I were also significantly down-regulated in the Burn + PolyPHb group. In addition, PolyPHb treatment markedly restored the endothelium-dependent relaxation impaired by severe burns and pathological changes of endothelium in aorta. Therefore, our data suggest that PolyPHb can limit severe burn-induced myocardial injury, which is associated with protection of aortic endothelium.

The new research tried to improve the distribution of molecular weight of Hb-based oxygen carriers (HBOC), a bottleneck of glutaraldehyde (GDA)-polymerization process. The orthogonal experiments were done on the basis of the early study of human placenta Hemoglobin (Hb)-crosslinked-GDA and three factors were selected including the molar ratio of GDA and Hb, Hb concentration, and the rate of the feeding GDA. The optimal match condition of polymerization process prepared for the purpose of lower mean molecular weight, content of super-weight molecule, and the content of dimer. The results showed that the molar ratio of GDA and Hb was the greatest influencing factor on the molecular weight distribution of polymerized-Hb, followed by the Hb concentration, and the last is the rate of feeding GDA. The optimum matching conditions had reached the objective that the mean molecular weight with 155.54 ± 5.79, the content of dimer with 17.23 ± 3.71, and content of super-weight molecule with 0.17 ± 0.09, and the results can be repeated in the 30 times expansion experiments.

The molybdenum blue method was used to determine the content of reduced vitamin C (Vc) in a solution of polymerized hemoglobin-based oxygen carriers (HBOCs) of the human placenta. The conditions of absorption wavelength, HCl addition, and reaction time, were investigated. The results of validation experiments showed that under the optimized conditions, a standard curve was confirmed with good linearity of 0.9985, for the Vc amount ranging from 0–200 μg. The values for relative standard deviation (RSD) of the precision and repeatability were both below 5%. Vc recovery was in the range of 97–102%. The conclusion could be made that a reduction in Vc content could be tested effectively by the molybdenum blue method.

Oxidative reactions of hemoglobin (Hb) are still a serious problem for Hb-based blood substitute development. Although varieties of antioxidant strategies have been suggested, this in vitro study examined the ability of the ascorbate, N-Acetyl-L-Cysteine (NAC), 4-hydroxy-2, 2, 6, 6-tetramethylpiperidine-1-oxygen free radicals (TEMPO) and their reductant system in preventing Hb oxidation. The content of ferric Hb is monitored in the process of vitamin C (Vc), NAC, TEMPO and their reductant system. The results suggest that ascorbate is effective in reducing ferryl Hb, and TEMPO with Vc/NAC could obviously shorten the reaction time, but it does not play the role of Met-Hb reductases. It demonstrates that TEMPO did little to recover Hb under oxidative stress.

This study was to investigate whether glutaraldehyde-polymerized human placenta hemoglobin (PolyPHb) could attenuate ischemia/reperfusion (I/R)-induced liver injury. Isovolemic hemodilution of SD rats was performed by exchanging 15% total blood volume with PolyPHb. I/R was induced by left liver lobes pedicle cross-clamping for 60 min and reperfusion for 2 h. Blood pressure moderately elevated after PolyPHb infusion and returned to basal level within 10 min. The hepatic histopathological damage and the activities of liver injury markers were reduced by PolyPHb. The TUNEL staining and caspase assay indicated hepatic apoptosis was also inhibited. Therefore, our findings suggest PolyPHb can reduce liver I/R injury.

This study was designed to investigate whether polymerized human placenta hemoglobin (PolyPHb) attenuated hemorrhagic shock-induced lung injury. A mean arterial pressure (MAP) of 30mmHg was maintained for 60 min. Then, all the rats were randomly resuscitated with hetastarch, whole blood, or PolyPHb. The result indicated that PolyPHb greatly improved the MAP and pulmonary function, and significantly reduced the release of inflammatory cytokines, histopathological changes, and pulmonary edema. Therefore, our findings suggest that PolyPHb could reduce pulmonary injury after hemorrhagic shock, and this effect was probably associated with the depressed inflammatory response.

This study was designed to investigate whether resuscitation with polymerized human placenta hemoglobin (PolyPHb) extended survival and the efficacy of hydroxyethyl starch (HES) in a rat model of hemorrhagic shock. After withdrawal of 39 ml of blood/kg, rats were randomly resuscitated with PolyPHb/HES or HES alone. Systemic, hemodynamic, and blood gas parameters, tissue oxygenation, and plasma erythropoietin (EPO) were evaluated. The results showed that resuscitation with PolyPHb/HES doubled the survival rate and maintained relatively high blood pressure and tissue oxygenation when compared with the results obtained using HES 130 alone. It is concluded that infusion of PolyPHb improves the resuscitative efficacy of HES in the present experimental conditions.

The exact mechanism of hemoglobin-based oxygen carrier (HBOC)-related vasoactivity is still unclear. This study measured the isometric tension of dog arteries and large conductance Ca(2+)-activated K(+) (BKCa) channel currents in vascular smooth muscle cells after exposure to HBOC with increasing concentrations. Data indicated that the net tensions of arteries were dramatically elevated and this elevation was more prominent in coronary artery. Moreover, HBOC exhibited inhibitory effect on BKCa channel, which is strongly correlated with changes in vascular tension. Collectively, HBOC-induced vasoconstriction in a dose-dependent manner and inhibition of BKCa channel is at least partially contributing to this effect.

In order to investigate whether hemoglobin-based oxygen carrier (HBOC) attenuates cardiopulmonary bypass (CPB)-induced cerebral damage. Male adult Beagle dogs were randomly divided into sham, control, and HBOC groups. After establishment of CPB model, hearts were arrested for 2 h and reperfused for 2 h. HBOC improved intracerebral O2 tensions and reduced the releases of biomarkers for cerebral damage, including neuron-specific enolase and S100β in both cerebrospinal fluid and serum. Moreover, HBOC attenuated the releases of tumor necrosis factor-α and interleukin-1β after CPB. Therefore, our findings suggest that HBOC could reduce cerebral damage after CPB, which was probably associated with improvement of tissue O2 preload.

In this study, lyophilization was applied to polymerized human placenta hemoglobin (PolyPHb) solution. The lyophilization process was carried out with freezing at − 70°C for 5 h and two phases of drying: the first phase of drying was carried out at −35°C–35°C for 16 h and the second phase at 35°C for 8 h. Hemoglobin (Hb) concentration, methemoglobin (MetHb) content, oxygen-carrying capacity, Fe3+ content, pH, UV spectrum, average molecular weight and its distribution, circular dichroism, SDS–PAGE, P50, crystal osmotic pressure, colloid osmotic pressure, zeta potential, average particle size, and other indicators were measured before and after lyophilization. Residual water content and rehydration time of the lyophilized products were also evaluated. All the indicators of lyophilized samples showed that the physical and biochemical properties of PolyPHb are not markedly changed before and after lyophilization. In this light, lyophilization may be a promising method for the preservation of PolyPHb s...

To reduce the mean molecular weight [Formula: see text] and to increase the effective polymerization ratio (REff) of polymerized human placenta hemoglobin (PolyPHb).Three factors of GDA-PolyPHb process such as the approach of feeding GDA (FGDA), hemoglobin concentration ([Hb]) and the molar ratio of GDA, and hemoglobin(RGDA:Hb) were investigated. Finally, the expansion experiments were conducted with optimal conditions.The data showed that the HBOCs with the REff of 67.35% and lower [Formula: see text] of 162.70 kDa were prepared by optimal conditions.Compared to original process, the optimal process greatly decreased the [Formula: see text] and increased the REff.