吲哚基二聚体菁类探针同步荧光测定蛋白质的研究

doi:10.3964/j.issn.1000-0593(2008)11-2615-04

Abstract
References
Related Citation (15)

Download: PDF (680 KB)
Export: BibTeX | EndNote (RIS)

Abstract Using a new homodimeric hydrophilic indole dye (Dye-I) as fluorescence probe, a sensitive synchronous spectrofluorometric determination for protein was developed. Characteristics of the fluorescence reaction between DYE-1 and BSA protein were investigated. Effects of the concentration of the hydrophilic dye, pH value of the buffer solution, and ion-intensity of NaCl were also studied and the optimum condition was gained. At pH of 2.50, electrostatic interactions of positive charges of the BSA chain and negative charges on the sulfonic groups of DYE-1 were carried out. The interactions of the indole group of DYE-1 and some active groups of BSA (viz. amido, carboxyl or sulfhydryl) were also achieved, and resulted in the combination of indole group of dye at the chain of BSA, which caused a notable increase in synchronous fluorescence with an observable shift to the longer emission wavelength. Effects of the concentration of indole dye on the determination of BSA were also investigated. With the augmentation of BSA, the α-helix structure of BSA molecular would change from the unwrapped state to the enfolded state, which was in favor of restraining free-oscillation of fluorescence probe in the solution and maintaining a high energy transfer efficiency. Such a fact would fuel a high fluorescence enhancement. and the change in fluorescence intensity (ΔF) gained the peak at 3.00 μmol·L^-1. The influences of ion-intensity of NaCl on the fluorescence of BSA-DYE-1 system was visible. Effects of coexistent substances such as amino acid and metal ions such as Cu²⁺, K⁺, Ca²⁺, Mg²⁺, Al³⁺, and Zn²⁺ were also investigated. Most substances showed no notable influences on the determination of BSA except Zn²⁺ and Cu²⁺ ions. Under the optimum conditions, good calibration curves of the protein were also obtained in the range of 5.00×10^-7-2.50×10^-5 g·mL^-1 (BSA) with a detection limit of 3×10^-8 g·mL^-1. Applied to simulant protein samples at the level of 1.00, 2.00, and 5.00 μg·mL^-1 of BSA, the recoveries were in the range of 98.6%-103.0% with the RSD of 1.1%-1.9%。Compared with the UV standard method, the relative deviation was obtained in the range of 0.4%-3.9%.

Key words：Synchronous fluorescence;Homodimeric indole cyanine probe;Determination of protein

Received: 2007-08-08 Accepted: 2007-11-16

ZTFLH:

O657.3

Corresponding Authors: XIE Zeng-hong E-mail: zhxie@fzu.edu.cn

Cite this article:

LIN Xu-cong,YAN Jin,GUO Liang-qia, et al. Determination of Protein by Synchronous Fluorometric Method with a New Indole Homodimeric Cyanine as Fluorescence Probe[J]. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2008, 28(11): 2615-2618.

URL:

https://www.gpxygpfx.com/EN/10.3964/j.issn.1000-0593(2008)11-2615-04 OR https://www.gpxygpfx.com/EN/Y2008/V28/I11/2615

[1] ZHAO Dan-hua, LI Yong-xin, ZHU Chang-qing(赵丹华，李永新，朱昌青). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2006，26(7)：1318.
[2] LIN Xu-cong, GUO Liang-qia, LIN Yan-xia, et al(林旭聪, 郭良洽, 林艳霞，等). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2007，27(9)：1775.
[3] CAO Shu-xia, ZHAO Yu-fen(曹书霞，赵玉芬). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2004, 24(10)：1197.
[4] HUANG Jian-hua, MA Hong-min, SUN Shu-ting, et al(黄建华, 马洪敏, 孙舒婷，等). Spectroscopy and Spectral Analysis(光谱学与光谱分析)，2006, 26(10)：1899.
[5] Glunde K, Foss C A, Takagi T, et al. Bioconjugate Chem.，2005，16(4)：843.
[6] SHI Yan, ZHENG Wei-wan, WU Xiao-xing, et al(石燕, 郑为完, 吴晓星，等). Spectroscopy and Spectral Analysis(光谱学与光谱分析)，2006, 26(9)：1653.
[7] LIN Xu-cong, WU Meng-hui, GUO Liang-qia, et al(林旭聪, 吴孟辉，郭良洽，等). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2007，27(8)：1587.
[8] LI Yong-xin，ZHAO Dan-hua，ZHUO Shu-juan, et al(李永新, 赵丹华, 卓淑娟，等). Chinese Journal of Analytical Chemistry(分析化学)，2003，31(5)：638.
[9] SUN Wei，HAN Jun-ying，JIAO Kui(孙伟, 韩军英, 焦奎). Journal of Analytical Science(分析科学学报)，2006，22(2)：145.
[10] CHEN Ying，YAO Min-na，TANG Yao-ji, et al(陈莹, 姚闽娜, 唐尧基, 等). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2005，25(12)：2048.
[11] ZHANG Yong, ZHU Ya-xian, et al(张勇，朱亚先, 等). China Environmental Science(中国环境科学)，2002，22(4)：289.
[12] JIAN Ju，DU Jiang-yan， FENG Yu-ying， et al(剑菊，杜江燕，冯玉英，等). Chinese Journal of Analytieal Chemistry(分析化学)，2001, 29(2)：219.
[13] SHI Yan，YAN Yuan，HUANG Jian-feng(石燕，鄢远，黄坚锋). Journal of Nanchang University (Natural Science)(南昌大学学报·理科版)，2000，24(3)：282.
[14] Funovics M， Weissleder R，Tung C H. Anal. Bioanal. Chem.，2003，377(6)：956.
[15] Laurence P G， Wakelin X Y， Bu Alexand-ra Eleftheriou，et al. J. Med. Chem.，2003，46：5790.
[16] Zheng Hong, Chen Xiao-lan, Hua Ming-hui, et al. Analytica Chimica Acta，2002，461：235.
[17] Schaberlea F A, KuZmin V A, Borissevitch I E. Biochimica and Biophysica Acta，2003，1621：183.
[18] Ogul’chansky T Yu，Losytskyy M Yu， Kovalska V B，et al. Spectrochimica Acta Part A，2001，57：2705.
[19] Pham W, Lai W F, Weissleder R, et al. Bioconjugate Chem.，2003，14：1048.
[20] Rouzina L， Bloomfied V A. J. Phys. Chem.，1996，100：4301.