Global Tracking of Transformation Products of Environmental Contaminants by ²H-Labeled Stable Isotope-Assisted Metabolomics

Abstract

Stable isotope-assisted metabolomics (SIAM) enables global tracking of isotopic labels in nontargeted metabolomics in living organisms. However, its application in tracking transformation products (TPs, as metabolites of contaminants) of environmental contaminants is still a challenge due to limits in methodology, unmatured algorithms, and the high cost of 13 Clabeled contaminants. Therefore, we developed a 2 H-SIAM pipeline coupled with a highly flexible algorithm 2 H-SIAM(1.0) (https://github.com/kechen1984/2H-SIAM), facilitating tracking TPs of contaminants in the environmental matrix. A detailed discussion illustrates the theory, behavior, and prospect of 2 H-SIAM. We demonstrate that the proposed 2 H-SIAM pipeline has unique advantages over 13 C-SIAM, for example, cost-effective 2 Hlabeled contaminants, easy synthesis of 2 H-labeled emerging contaminants, and providing more structural information. A pyrene soil degradation study further confirmed its high performance. It efficiently discarded 99% of noise signals and extracted 52 features from the nontargeted high resolution mass spectrometry (HRMS) data. Among them, 13 features were annotated as TPs of pyrene with identification confidence from Level 2a to Level 5, and 5 TPs were reported for the first time. In conclusion, the proposed 2 H-SIAM pipeline is powerful in tracking potential TPs of environmental contaminants with unique advantages.

H/D Exchange Coupled with 2H-labeled Stable Isotope-Assisted Metabolomics Discover Transformation Products of Contaminants of Emerging Concern

“…com/kechen1984/2H-SIAM), enabling global tracking of TPs by 2 H-labeled analytes. 16 Nevertheless, the performance of the coupling using H/D exchange and 2 H-SIAM is still unclear. In this study, they were coupled to discover TPs of 6PPD to verify the strategy's performance.…”

“…2 H-SIAM pipeline was carried out following our previous study. 16 Briefly, 6PPD and in-house synthesized 6PPD-d9 (in acetone) were added to one-quarter of the soils, and they were further mixed with the rest of the soils to obtain soils with final content of 50 mg/kg 6PPD or 6PPD-d9 (n = 6, respectively). Additionally, soils without the addition of 6PPD and 6PPD-d9 were used as method blanks (n = 6).…”

“…In the 2 H-SIAM pipeline, the Mix 1:3 and Mix 3:1 samples are analyzed by HRMS, and the feature list of the mixed samples is obtained by the algorithm that custom used, for instance, XCMS and MZmine2.53. 16 Then, the algorithm 2 H-SIAM-(1.0) filters noise features by both qualitative and quantitative settings (Figure 1). 16 The 2 H-SIAM(1.0) starts with the qualitative inspection of the feature list to find potential isotopologue pairs (M and M′, natural and isotope-labeled compounds) within an indicated duration of RT, and the mass difference between isotopologue pairs (M and M′) should be integer multiples of mass difference of the indicated labeled and unlabeled atom (for 2 H and H, Δd1 = 1.006277).…”

“…16 Then, the algorithm 2 H-SIAM-(1.0) filters noise features by both qualitative and quantitative settings (Figure 1). 16 The 2 H-SIAM(1.0) starts with the qualitative inspection of the feature list to find potential isotopologue pairs (M and M′, natural and isotope-labeled compounds) within an indicated duration of RT, and the mass difference between isotopologue pairs (M and M′) should be integer multiples of mass difference of the indicated labeled and unlabeled atom (for 2 H and H, Δd1 = 1.006277). Then, the mean intensities of the features M and M′ from samples Mix 1:3 and Mix 3:1 are used for the calculation of three actual ratios (f n ) of the features, which are used to check whether these features quantitatively follow the setting of triple filters (F n ; F 1 , F 2, and F 3 ).…”

“…An alternative solution is 2 H-SIAM, which uses 2 Hlabeled analytes and costs much less than 13 C-SIAM. 16 For instance, pyrene-13 C3 is 15,000 times more expensive than pyrene-d10 (Cambridge Isotope Laboratories, https://www. isotope.com).…”