Construction of Hemoglobin-vesicle with long-term in vivo oxygen transporting ability

在磷脂囊泡（脂质体）中包裹血红蛋白（Hb）的血红蛋白囊泡（HbV）可作为人工氧载体。然而，当中心铁从二价氧化为三价时，Hb失去了氧结合能力（甲基形成）。在这种现象中，一个电子从中心的铁转移到结合的氧，甲基苯丙胺形成（自氧化），以及由此产生的活性氧（过氧化物）进一步形成甲基苯丙胺是连续发生的。我们发现（甲氧基血红蛋白/ l -酪氨酸）系统能迅速消除过氧化物。本研究证实，以l -酪氨酸为底物，利用甲基乙酯过氧化物酶活性的过氧化氢消除体系能有效抑制甲基乙酯的形成。此外，我们证实了在HbV的内水相中加入该体系可以显著抑制HbV的甲基乙基形成，即使过氧化氢持续加入到H…将20mL/kg的HbV分散体注入大鼠体内，与常规HbV的14小时相比，该HbV的甲基苯丙胺形成半衰期延长至44小时。此外，通过一种新的筛选方法，测量铁基Hb自由基与除Tyr外的14种候选物质的反应速率，我们发现一些含有内环的衍生物，如色氨酸或羧基，其抑制作用高于Tyr。另一方面，由自氧化引起的甲氧基血红蛋白（HbCO）不会形成甲氧基血红蛋白。如果利用血液循环中HbCO逐渐转化为HbO_2的现象，可以通过抑制含HbCO的HbV形成甲基来维持携氧能力。在本研究中，我们测量了含HbCO的HbV的甲氧基乙基生成速率的变化，并阐明了尽管HbCO的部分不同，但剩余HbO_2中甲氧基乙基乙基的生成速率是相同的。因此，建议在HbCO中去除CO的同时，构建温和的metHb生成体系。如上所述，如果这两种方法配对，预计可以有效地控制HbV的甲基乙基形成。少

The hemoglobin vesicles (HbV) encapsulating hemoglobin (Hb), which binds oxygen reversibly, in the phospholipid vesicle (liposome) function as an artificial oxygen carrier. However, Hb loses an oxygen binding capacity when central iron is oxidized from two valences to three (metHb formation). In this phenomenon, metHb formation (autoxidation) by one electron transfer from the central iron to the bound oxygen and the further metHb formation by the resulting reactive oxygen (peroxide) are taken place in a row. We have found out that a (metHb Hb/L-tyrosine) system eliminates peroxide promptly. In present study, it was confirmed that the hydrogen peroxide elimination system utilizing the peroxidase activity of metHb which uses L-tyrosine as a substrate was effective for suppression of metHb formation. Moreover, we confirmed the remarkable suppression of metHb formation of HbV which included this system in their inner aqueous phase, even if hydrogen peroxide were continuously added to the H … More bV dispersion. When the 20mL/kg of the HbV dispersion was administrated into rats, the half time of metHb formation in this HbV extended till 44 hours in comparison with 14 hours of the conventional HbV. Furthermore, from a novel screening method which measured the reaction rate of ferryl Hb radical with 14 kinds of candidate substances other than Tyr, we found that some derivatives which possesses indore rings such as tryptophane or carboxyl groups showed the suppressive effect higher than Tyr.On the other hand, metHb formation caused from autoxidation does not occur for carbonyl Hb (HbCO). And if we use the phenomenon of the gradual conversion of HbCO into HbO_2 in blood circulation, we can maintain an oxygen carrying capacity by suppressing the metHb formation of HbV containing HbCO. In present study, we measured the change of the metHb formation rate of HbV containing HbCO and clarified that the rate of metHb from the remaining HbO_2 was the same in spite of the differed portion of HbCO. Therefore, it is suggested that gentle metHb formation system can be constructed in accompanied with the elimination of CO from HbCO. As mentioned above, if these dual approaches are paired, it is expected that the metHb formation of HbV can be effectively controlled by in vivo. Less