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. 2020 Sep 21;117(40):24867–24875. doi: 10.1073/pnas.2005714117

Fig. 4.

Fig. 4.

Chromatographic detection of A1T in human and mouse retina. A1T is more abundant in mouse neural retina (NR) than in RPE. Separation by UPLC with monitoring at 450 nm. (A and C) Detection of synthetic products derived from incubation of retinaldehyde isomers with taurine: The panels in A and C present synthesized A1T isomers: 11-cis-A1T, red trace; combined 13-cis-A1T, and all-trans-A1T, blue and black traces, respectively; 9-cis-A1T, gray trace. UV/Vis absorbance maximum, λmax. (Insets) UV/visible (Vis) absorbance spectra corresponding to indicated peaks (α–δ, arrows). 11-cis-A1T (α): λmax 450 nm, retention time 11.0 min; 13-cis-A1T (β): λmax, 440 nm, retention time 11.7 min; 9-cis-A1T (γ): λmax, 440 nm, retention time 12.2 min; all-trans-A1T (δ): λmax 448 nm, retention time 13.8 min; A2T, λmax, 338, 440 nm, retention time 36.2 min. (B) Human neural retina (NR). The UPLC injectant was a methanolic extract of neural retina (age 74 y). 11-cis-A1T: λmax,450 nm, retention time 11.0 min; 13+9-cis-A1T: λmax, 440 nm, retention time 11.8∼12.2 min; all-trans-A1T: λmax 448 nm, retention time 13.8 min. (C) To enable alignment with other panels, chromatograms in A, repeated in C, are the same as in A. (D) Cyclic light-reared black C57BL/6J mice, NR (age 2 mo, 8 eyes, 30.2 mg wet weight). The UPLC injectant was a methanolic extract of neural retina. 11-cis-A1T (α): λmax 450 nm, retention time 11.0 min, 13 + 9-cis-A1T: λmax, 440 nm, retention time 11.8 ∼ 12.2 min. all-trans-A1T: λmax, 448 nm, retention time 13.8 min. I.S. (internal standard, A1-β-alanine): λmax, 450 nm, retention time 18.0 min. (E). RPE harvested from cyclic light-reared black C57BL/6J mice, (age 2 mo, 8 eyes, 19.3 mg wet weight). No A1T isomers were detected in RPE. I.S. (internal standard, A1-β-alanine): λmax, 450 nm, retention time 18.0 min. The unknown peaks (B and E) at retention time 36.2 min exhibits λmax, 440 and 290 nm. These peaks do not correspond to A2T, the latter being produced in the synthetic mixture presented in A and C. Although the retention times are similar to A2-taurine (retention time 36.2 min), they present with a single λmax (440 nm, 290 nm) in the sample of NR. Conversely, A2T has 2 absorbance maxima (λmax 338, 440 nm). This unknown peak is likely NRPE (λmax).