# NMR and LC-MS-Based Metabolomics to Study the Effect of Surfactin on the Metabolome of Flax
## 3 Results and Discussion
### Phenolic compounds:  
In the aerial parts, several phenolic compounds were identified by NMR when enough accumulated or by LC-MS when accumulated in small amounts, using standards and/or by matching the MSe spectra with the molecules cited in other studies involving flax extracts. From both NMR and LC-MS datasets (Figure 7), we found that almost all the phenolic compounds exhibited changes in concentration in the same way.  
Figure 7: Heatmap of the discriminant metabolites differently accumulated in the flax stem (**left**) and leave (**right**) extracts between the surfactin-treated and control conditions. The scale bar (from red to blue) on the right represents the ratio of metabolite content in surfactin-treated flax aerial parts to that in control flax aerial parts. For each organ, the column on the left indicates the kinetic point at 56 h, and the column on the right indicates the kinetic point at 10 D. The analytical methods used to determine discriminant metabolites are indicated on the left of the graph; the red color corresponds to the combination of NMR and LC-MS, the blue color corresponds to LC-MS, and the purple color corresponds to NMR.  
In the stems, it is noticeable that the content of isovitexin, lucenin-2, isoorientin, vitexin, orientin, carlinoside, trictuside A, and vicenin-2 strongly decreased at 56 h after surfactin treatment. All these compounds belong to the flavonoid family with one or two glycoside unit(s) linked to the flavone core via C bound. The content of chlorogenic acid isomers, DCG, shaftsoside and vicenin-1 decreased slightly or increased at 56 h. At 10 D, the content of all these phenolic compounds decreased with surfactin treatment but less strongly than at 56 h. The influence of surfactin on phenolic content has already been studied in other models, showing, for example, that surfactin treatment did not modify phenolic content in peanut in control conditions but induced phenolic increase when exposed to a pathogen. The phenolic content was evaluated in the stems 10 D after surfactin treatment, and the results agreed with our results, showing that surfactin affects phenolic content early after surfactin treatment, but this effect is attenuated late after surfactin treatment [40].  
In the leaves, some of the phenolic compounds increased with surfactin treatment at 56 h, namely caffeic acid, chlorogenic acid isomers, carlinoside, trictuside, and lucenin-2, whereas the other ones slightly decreased. At 10 D, in the leaves, all the phenolic compound content decreased. The ratios of surfactin treatment conditions vs. control conditions were lower at 10 D than at 56 h for all the phenolic compounds. In another study, the phenolic content in the sprouts of sesame seeds treated by _Bacillus clausii_ increased, which is in accordance with our results obtained at 56 h regarding the leaves treated with surfactin, produced by _Bacillus velezensis_[41].  
This suggests that the stems and leaves exhibit opposite behavior during the kinetic response of the plant after surfactin treatment. Stems react more like roots than leaves.