Poster: Emerging Technologies

Abs # 874: An Optimized Spectrophotometric Determination of Glutathione and Glutathione Disulfide

Presenter:	Hung, Shu Hsien , shhung@mail.dyu.edu.tw
Authors	Hung, Shu Hsien  (A)   Yu, Chih Wen  (A)   Chou, I Min  (A)   Lin, Chin Ho  (B)
Affiliations:	(A): Department of Bioindustry Tech, Inst of Agri Biotech, Da Yeh Univ, Taiwan, Republic of China (B): Life Science Department, National Chung Hsing University, Taiwan, Republic of China

A simple optimized procedure for the determination of GSH and GSSG based on enzymatic recycling of DTNB in the presence of GR was developed. The procedure was adopted from three published procedures. Three key components adopted in this combined procedures are: 1. Homogenization of the sample in metaphosphoric acid as a deproteinization agent to quickly acidify the homogenate and maintain the GSH to GSSG ratio; 2. Addition of PVPP to adsorb the phenolic compound from the extracts; 3. Use of 2-vinylpyridine as a glutathione derivatization reagent. The optimized method gives a linear reading for glutathione dosages between 10 and 1000 pmole. The reduced/oxidized glutathione ratio for mung bean leaves was between 9 to 22 with and without exposure to environmental stress. Adaptation of micro-homogenization sssystem, 30 mg mung bean leaves suffice for glutathione determination. Based on the protocol, a 96-well microplate microassay of GSH and GSSG was also established for fast and cost effective handling of samples.