Citation: WU Zhichao, WU Enguo, YANG Zhongyi, TAO Yuxiang, CHEN Tan, ZHONG Yongjun. Determination of N,N'-ethylenediamine disuccinic acid in bioconversion reaction solution using high performance liquid chromatography[J]. Chinese Journal of Chromatography, ;2019, 37(10): 1118-1123. doi: 10.3724/SP.J.1123.2019.03033 shu

Determination of N,N'-ethylenediamine disuccinic acid in bioconversion reaction solution using high performance liquid chromatography

1.
Taizhou University, Taizhou 318000, China;
2.
Zhejiang Hisun Pharmaceutical Co., Ltd., Taizhou 318000, China;
3.
Taizhou Bona Chemical Co., Ltd., Taizhou 318000, China

Received Date: 26 March 2019

Fund Project: Taizhou Science and Technology Program Project (Nos. 162gy47, 1701ny06)

A method was developed for the determination of N,N'-ethylenediamine disuccinic acid (EDDS) in bioconversion samples by high performance liquid chromatography (HPLC). An InertSustain AQ-C₁₈ column (250 mm×4.6 mm, 5 μm) was used in the analysis. The mobile phase was 25% (v/v) methanol with 1.0 g/L Cu(CO₂CH₃)₂·H₂O, 2.0 g/L tetrabutylammonium hydroxide, and the pH was adjusted to 2.80 with phosphoric acid. The flow rate of the mobile phase was 1.0 mL/min, and the column temperature was set at 30℃. The detection wavelength was 254 nm. EDDS, fumaric acid, citric acid, malic acid and ethylenediaminetetraacetic acid (EDTA) were separated from one another within 8 min. EDDS showed good linearity in the range of 0.06-0.6 g/L. About 0.25 g/L EDDS was detected in the biosynthesis reaction solution catalyzed by recombinant EDDS-lyase, while 36.56 g/L malic acid was formed as the by-product from fumaric acid. The catalytic activity of the enzyme was confirmed in the hydrolysis of 10 g/L EDDS, which produced 3.05 g/L malic acid in 3 h. This analytical method is simple, rapid, sensitive, reliable, and suitable for the analysis in the research of EDDS bioconversion process.
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沈阳化工大学材料科学与工程学院沈阳 110142