Fig. 2.

Unknown C₁₂H₁₅NO₄ is N-lac-Phe. (A) ¹H-NMR spectrum of isolated C₁₂H₁₅NO₄ recorded in D₂O at 300 MHz matches the ¹H-NMR spectrum of the synthesized reference N-lac-Phe. (B) Unknown C₁₂H₁₅NO₄ elutes at the same time as N-lac-Phe. Extracted ion chromatogram of the mass corresponding to C₁₂H₁₅NO₄ (m/z 236.09) in conditioned medium from HEK 293/ABCC5 cells before (depicted in blue) and after (depicted in red) spiking with synthesized N-lac-Phe (1 μg/mL, 4.2 μM). The high-resolution MS² fragmentation spectrum of synthesized N-lac-Phe (C) and unknown C₁₂H₁₅NO₄ in conditioned medium from HEK 293/ABCC5 (D) match. Differences in background and intensities can be attributed to the lower concentration of unknown C₁₂H₁₅NO₄. Additional NMR and MS spectra are presented in Figs. S2 and S4, respectively.