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Measurement of the 15N/14N ratio of phenylalanine in fermentation matrix by isotope ratio mass spectrometry
- Zhang, Shiwei, Zhong, Qiding, Wang, Daobing, Huang, Zhanbin, Li, Guohui
- Biotechnology letters 2017 v.39 no.12 pp. 1853-1857
- fermentation, high performance liquid chromatography, isotope fractionation, mass spectrometry, metabolic flux analysis, nitrogen, phenylalanine, stable isotopes
- OBJECTIVES: To determine the origin of ¹⁵N-labeled phenylalanine in microbial metabolic flux analysis using ¹⁵N as a tracer, a method for measuring phenylalanine δ¹⁵N using HPLC coupled with elemental analysis-isotope ratio mass spectrometry (EA-IRMS) was developed. RESULTS: The original source of the ¹⁵N-labeled phenylalanine was determined using this new method that consists of three steps: optimization of the HPLC conditions, evaluation of the isotope fractionation effects, and evaluation of the effect of pre-processing on the phenylalanine nitrogen stable isotope. In addition, the use of a ¹⁵N-labeled inorganic nitrogen source, rather than ¹⁵N-labeled amino acids, was explored using this method. CONCLUSIONS: The method described here can also be applied to the analysis of metabolic flux.