氧合、碳氧及高铁血红蛋白拉曼光谱的测定

目的测定比较氧合、碳氧和脱氧血红蛋白的拉曼光谱。方法分别向氧合血红蛋白(HbO2)中通入一氧化碳(CO)、加入高铁氰化钾(K3Fe(CN)6)制备得到碳氧血红蛋白(COHb)及高铁血红蛋白(MetHb);在785 nm波长激发光下,测定HbO2、COHb及MetHb的拉曼光谱。结果 HbO2、COHb及MetHb分别在1 210～1 230、1 375～1 379和1 550～1 650 cm-1处的差异可在拉曼光谱中分辨。COHb相较于HbO2在1 550～1 650 cm-1处特征峰(与自旋态及卟啉环中心孔径大小相关)发生变化,即1 559 cm-1特征峰峰强增高,1 589和1 645 cm-1特征峰强度降低;MetHb相较于HbO2在1 210～1 230 cm-1(与自旋态及次甲基键变形振动相关)、1 375～1 379 cm-1(与血红蛋白氧化态相关)和1 550～1 650 cm-1处特征峰变化,即1 228 cm-1特征峰移至1 215 cm-1处,1 375～1 379 cm-1处特征峰峰强增高,1 550 cm-1特征峰峰强增高,1 589和1 645 cm-1特征峰强度降低。结论用785 nm激发的拉曼光谱可以鉴别HbO2、COHb及MetHb,可作为1种针对血红蛋白样品快速、准确、无损的在线监测方法。

Raman spectral measurement of oxyhemoglobin, carboxyhemoglobin and methemoglobin

Objective To study comparatively the Raman spectra of oxyhemoglobin,carboxyhemoglobin and methemoglobin.Methods The solution of carboxyhemoglobin was prepared by flowing dry carbon monoxide in the purpose high oxygenated hemoglobin solution.The solution of methemoglobin was prepared by making the high oxygenated hemoglobin solution reacting with the potassium ferricyanide(K3Fe(CN) 6).Raman spectra of oxyhemoglobin,carboxyhemoglobin and methemoglobin recorded using 785 nm laser excitation were presented.Results The spectral differences between the three hemoglobin derivatives were exhibited in several relevant peaks in the 1 210 ～ 1 230 cm-1,1 375 ～1 379 cm-1 and 1 550 ～ 1 650 cm-1.Compared to oxyhemoglobin,a number of changes were apparent in the range of 1 550 ～ 1 650 cm-1(related to spin state and the heme core size) in the Raman specrtra of carboxyhemoglobin.Furthermore,compared to oxyhemoglobin,in the spectra of carboxyhemoglobin,the band at 1 559 cm-1 appears was more intense,while the 1 589 cm-1 and 1 645 cm-1 band was less intense.In the spectra of methemoglobin,a number of changes were exhibited in the range of 1 210 ～ 1 230 cm-1(assigned to spin state and the C-H methine bending vibration),1 375 ～1 379 cm-1(assigned to redox state) and 1 550 ～ 1 650 cm-1,by comparing with oxyhemoglobin.Particularly,by comparing the spectra of methemoglobin to oxyhemoglobin,the 1 228 cm-1 band shifted to 1 215 cm-1,and the band intensity at 1 375 ～1 379 cm-1 was enhanced.Meanwhile,compared to oxyhemoglobin,in the spectra of carboxyhemoglobin,the band at 1 550 cm-1 appears was more intense,while the 1 589 cm-1 and 1 645 cm-1 band was less intense.Conclusion Raman spectroscopy can provide a simple way to distinguish oxyhemoglobin,carboxyhemoglobin and methemoglobin,which may potentially become a quick,convenient and nondestructive process method applied to at-spot detection in the blood substitute field by on-line control.