Determination of second virial coefficient of proteins using a dual-detector cell for simultaneous measurement of scattered light intensity and concentration in SEC-HPLC

被引：48

作者：

Bajaj, H ^{[1
]}

Sharma, VK ^{[1
]}

Kalonia, DS ^{[1
]}

机构：

[1] Univ Connecticut, Sch Pharm, Dept Pharmaceut Sci, Storrs, CT 06269 USA

来源：

BIOPHYSICAL JOURNAL | 2004年 / 87卷 / 06期

基金：

美国国家科学基金会;

关键词：

D O I：

10.1529/biophysj.104.048686

中图分类号：

Q6 [生物物理学];

学科分类号：

071011 ;

摘要：

A method is proposed for the measurement of the B-22 value of proteins in aqueous solutions in flow-mode that utilizes a novel fabricated dual-detector cell, which simultaneously measures protein concentration and the corresponding scattered light intensity at 90degrees, after the protein elutes from a size-exclusion column. Each data point on the chromatograms obtained from the light scattering detector and the concentration ( ultraviolet) detector is converted to Rayleigh's ratio, R-theta, and concentration, c, respectively. The B-22 value is calculated from the slope of the Debye plot (K-c/R-theta versus c) generated from a range of concentrations obtained from these chromatograms for a single protein injection. It is shown that this method provides reliable determination of the B-22 values for such proteins as lysozyme, chymotrypsinogen, and chymotrypsin in various solution conditions that agree well with those reported in literature.

引用

页码：4048 / 4055

页数：8

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