ABSTRACT
【Objective】 To study the red blood cell substitute preparation method of glutaraldehyde(GDA) and Bis(3, 5-dibromosalicyl) fumarate(DBBF) double polymerized, and also observe its curative effect. 【Methods】 The affecting factors of the crosslink of DBBF and human placental hemoglobin(Hb) were selected, including solution pH, inositol hexaphosphoric acid concentration, and molar ratios of DBBF/Hb. The changes of P50, Hill Co and methemoglobin(MetHb) content during the preparation process were studied and the molar ratio of GDA to DBBF-Hb were determined. The final product was obtained through the steps of ultrafiltration, filtration, and deoxygenation, etc., and the final product was subjected to a study on the curative effect of anti-hemorrhagic shock in rats. 【Results】 Under the condition of pH 8.0, the concentration of inositol hexaphosphoric acid was 10mM/L, the molar ratio of DBBF/hemoglobin was 6∶1, the content of dimer was(22.67±3.28)%, the content of tetramer was (74.64±7.05)%, and its P50 was (25.25±2.31) mmHg. As the molar ratio of DBBF/Hb increased, P50 and MetHb content would gradually increase, while the Hill Co will gradually decrease. The molar ratio of GDA and DBBF-Hb was 7∶1, the content of dimer was (8.23±0.79)%, the content of tetramer was (27.87±3.63)%, the content of super-macromolecule was (1.05±0.31)%. As the molar ratio of GDA and DBBF/Hb increased, MetHb content would increase, while the P50 and Hill Co would gradually decrease. The 24h survival rate of the product in rats with hemorrhagic shock was 83.33%(10/12), which was significantly higher than that in the negative control group[41.67%(5/12)], while similar with the positive control group [91.67%(11/12)]. 【Conclusion】 The red blood cell substitutes with GDA and DBBF double polymerized can effectively reduce the dimer content, which is more conducive to industrial production, and has a positive effect on the treatment of hemorrhagic shock.
ABSTRACT
【Objective】 To explore the protective effects of hemoglobin base on oxygen carries (HBOCs) with different oxygen affinity on isolated rat hearts. 【Methods】 Using Langendorff isolated heart perfusion model, 45 adult male SD rats (SPF grade), perfused with 30 min KH solution baseline, were randomly divided into sham operation group and control group: St. Thomas (STS) solution perfusion volume was 3mL/100g body weight; high P50 HBOCs group: [STS + high P50 HBOCs (P50=35.0 mmHg, 2.5 mg/100 g) product] perfusion volume was 3mL/100g body weight; medium P50 HBOCs group: [STS + medium P50HBOCs (P50=26.5.0 mmHg, 2.5 mg/100 g) product] perfusion volume was 3 mL/100 g body weight; low P50 HBOCs group: [STS + low P50 HBOCs (P50=11.0 mmHg, 2.5 mg/100 g) product] perfusion volume was 3mL/100g body weight, and the heart was arrested and placed in a 37℃ water bath to make the heart ischemic for 35 minutes, and then reperfused for 2 hours. The left ventricular development pressure (LVDevP), left ventricular end diastolic pressure (LVEDP), the rate of change of left ventricular pressure (LVPCR) and heart rate (HR) in the rat heart during reperfusion were observed and recorded. 1 min perfusion fluid from each rat in the basic and reperfusion phase was taken, and blood gas analyzer was used to measure the blood gas indexes of rats, and the myocardial injury marker enzymes [cardiac enzyme creatine kinase (CK-MB), lactate dehydrogenase (LDH) and the release of α-hydroxybutyrate dehydrogenase (α-HBDH)] were measured by ELISA kit. 【Results】 The cardiac function and the release of myocardial enzymes in the 5 groups of rats in the basal cardiac perfusion stage were similar (P>0.05). However, in the reperfusion stage, except for the insignificant changes in HR (P>0.05), the heart LVDevP (mmHg) of the three P50 HBOCs groups and the control group were 10.69±3.65 vs 8.50±2.88, 23.26 ±5.62 vs 8.50±2.88, 35.60±3.82 vs 8.50±2.88, LVEDP (mmHg) were 43.34±8.08 vs 54.64±7.42, 39.43±8.30 vs 54.64±7.42, 31.46±4.11 vs 54.64±7.42, dp/dt were 12.09±9.96 vs 6.09±0.98, 25.65±8.87 vs 6.08±0.98, 35.32±9.33 vs 6.09±0.98, -dp/dt were 17.53±11.28 vs 11.39±2.16, 28.80±13.70 vs 11.39±2.16, 43.36±3.83 vs 11.39±2.16, respectively (all P<0.05); the rebound situation and the release of CK-MB, LDH, and α-HBDH in the three P50 HBOCs groups were better than those in the control group (P<0.05). Among the three P50HBOCs products, the low P50HBOCs group had the best cardiac function indexes. The myocardial enzyme indexes of the high, medium and low HBOCs groups were CK-MB (ng/mL): 110.47±4.04, 90.2±2.46, 77.1±3.51; LDH (U/L): 162.23±7.71, 135.13±23.69, 92.20±4.21; a-HBDH (U/L): 228.00±8.03, 172.30±8.99, 131.00±2.02. 【Conclusion】 STS solution containing HBOCs products can improve the function of the reperfused heart at normal temperature ischemia for 35 min and 2 h reperfusion, and reduce heart damage. The STS solution containing low P50 HBOCs has the most obvious protective effect in rat isolated heart perfusion.
ABSTRACT
Objective To study the optimization process of matching conditions using glutaraldehyde (GDA) as crosslinking agent of hemoglobin based oxygen carriers (HBOCs),to further reduce the average molecular weight and the content of super-weight molecular,and improve the conversion ratio of polymerization.Methods The orthogonal designs were done on the basis of the previous single influencing factor research of human placenta hemoglobin crosslinking GDA.Three factors were selected including molar ratio of GDA to hemoglobin,mass concentration of hemoglobin and the rate of the feeding GDA.Results The molar ratio of GDA to hemoglobin is the most important influencing factor on the molecular weight distribution of polymerized hemoglobin,followed by the mass concentration of hemoglobin and the rate of feeding GDA.When analyzing the impact on the mean molecular weight,there were significant differences between mean molecular weight corresponding to different molar ratios of GDA to hemoglobin (P＜0.05),while there was no statistical significance between mean molecular weight corresponding to different mass concentrations of hemoglobin and the rates of feeding GDA (P＞0.05).When analyzing the impact on the effective conversion ratio,there were significant differences between effective conversion ratios corresponding to different molar ratios of GDA to hemoglobin and different mass concentrations of hemoglobin (P＜0.05),while there were no statistical significances between effective conversion ratios corresponding to different rates of feeding GDA (P＞0.05).When analyzing the impact on the content of super-weight molecular,there were significant differences between content of super-weight molecular corresponding to different molar ratios of GDA to hemoglobin,while there were no statistical significances between content of super-weight molecular corresponding to different mass concentrations of hemoglobin and different rates of feeding GDA.Conclusions The optimal matching conditions of hemoglobin polymerization process were determined by orthogonal designs.