A study on the simultaneous determination of nitrogen content and 15N isotope abundance in plants using peak height intensities at m/z 28 and 29.

A method for simultaneous determination of nitrogen content and 15N isotope abundance in plants was established by Elemental analysis-gas isotope ratio mass spectrometry. Taking poplar leaves and l-glutamic acid as standards, nitrogen content was determined using the standard curve established by weighted least squares regression between the mass of nitrogen element and the total peak height intensity at m/z 28 and 29. Then the 15N isotope abundance was calculated with the peak height intensity at m/z 28 and 29. Through the comparison of several sets of experiments, the impact of mass discrimination effect, tin capsule consumables, isotope memory effect, and the quality of nitrogen on the results were assessed. The results showed that with a weight of 1/x2, the standard curve has a coefficient of determination (R2) of 0.9996. Compared to the traditional Kjeldahl method, the measured nitrogen content deviated less than 0.2 %, and the standard deviation (SD) was less than 0.2 %. Compared to the sodium hypobromite method, the 15N isotopic abundances differed less than 0.2 atom%15N, and the SD was less than 0.2 atom% 15N. The established method offers the advantages of being fast, simple, accurate, and high throughput, providing a novel approach for the simultaneous determination of nitrogen content and 15N isotope abundance in plant samples.

Quantification and isotope abundance determination of 13C labeled intracellular sugar metabolites with hydrophilic interaction liquid chromatography.

Metabolic flux analysis (MFA) using stable isotope labeled tracers is a powerful tool to estimate fluxes through metabolic pathways. It finds applications in studying metabolic changes in diseases, regulation of cellular energetics, and novel strategies for metabolic engineering. Accurate and precise quantification of the concentration of metabolites and their labeling states is critical for correct MFA results. Utilizing an ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) system, an analytical method for simultaneously quantifying the concentration of sugar metabolites and their mass isotopologue distribution (MID) was developed. The method performs with good linearity and coefficient of determination (R2) > 0.99, while the detection limit ranged from 0.1 to 50 mg L-1. Seven sugar metabolites were detected in a labeled Brevibacterium flavum sample using the method. The detected quantities ranged from 6.15 to 3704.21 mg L-1, and 13C abundance was between 12.77% and 66.67% in the fermentation fluid and 16.28% and 91.93% in the bacterial body. Overall, the method is efficient, accurate, and suitable for analysis of labeled sugar metabolites in 13C MFA studies.