Reflection Interference Spectroscopy For Studying The Interactions Of Bovinehemoglobin With Small Molecules

In this paper, the immobilization process of bovine hemoglobin(BHb) to the carboxymethyl dextran(CMD) chip and the interaction of immobilized bovine hemoglobin(BHb) and glutathione(GSH), N-acetyl- L-cysteine(NAC), ascorbic acid(Vc) were studied by the reflection interference spectroscopy. The primary contents were listed as follows:(1) The research significance of the interaction between proteins and small molecules, the commonly-used methods, and the topics-choosing basis and research contents were reviewed in the dissertation.(2) The immobilization process of BHb to the CMD chip was investigated by the reflection interference spectroscopy. Through literatures and experiments, the suitable fixation conditions were explored. At 20℃, the-COOH of the CMD chip was activated by NHS/EDC. The p H = 5.70 acetate buffer solution was selected as immobilization buffer. The immobilization curve was got by BIAffinity,indicting that the –NH2 of BHb and the –COOH of CMD chip formed amide bond, namely the BHb was successfully immobilized onto the CMD chip.(3) The interaction between immobilized BHb and GSH, NAC, Vc were studied by the reflection interference spectroscopy. Through the experiments, the detection curves in different concentrations of the three small molecules were obtained. The lower limit and upper limit were determined. The relation curves of concentrations and reaction rates for the three small molecules were obtained. The method to detect small molecules using immobilized proteins was initially established.(4) The conclusions obtained from immobilization process and detection process and reaction curves were systematically summarized and the future work was prospected.