PMC

a, b FTIR and ¹H-NMR spectra of OPD with or without protonation treatment. c, d FTIR and ¹H-NMR spectra of CDs with or without protonation treatment (black line: spectra of samples without protonation treatment; red line: spectra of samples with protonation treatment).

a−c full-scale ultraviolet photoelectron spectroscopy (UPS), near Fermi energy (low binding energy) and secondary electron cut-off (high binding energy) regions of CDs before the protonation; d−f UPS, near Fermi energy and secondary electron cut-off regions of CDs after the protonation. g UV−Vis spectra, h one-photon fluorescence spectra; i two-photon fluorescence spectra of CDs before and after the protonation. (P-CDs: CDs after the protonation treatment).

a CDs without the protonation; b CDs treated with the edge protonation.

(A) Sequence of protonation sites of SM07 were determined by¹ H-¹⁵N HMBC experiments in 1:2 water:methanol mixture. Left: The plot of¹⁵N chemical shifts of the N-10, N-12, and N-8 resonances of SM07 vs titrated TFA-d equivalents, showing the mono-protonation of N-12 as evidenced by its large upfield chemical shifts change. Acidity of the medium increased as more equivalents of TFA-d were added. Electronic effects due to protonation of N-12 caused downfield chemical shift change of N-10 and N-8 between 0–1 equivalents of TFA-d. Right: NMR-based model of the order of dominant protonation states for SM07. The protonation event was only observed at N-12. Microstates shown in the figure are the most likely contributors to the UV-metric pK_a of 6.08 ± 0.01. (B) Sequence of protonation sites of SM14were determined by¹ H-¹⁵N HMBC experiments in acetonitrile. Left: The plot of ¹⁵N chemical shifts of N-9, N-7, and N-16 of SM14 vs titrations of TFA-d equivalents, showing two sequential protonation events. The first protonation occured at N-9; a large upfield chemical shift change of 71.6 ppm was seen between 0–1 equivalents of TFA-d. Downfield chemical shift changes observed for N-7 and N-19 in this region were due the electronic effect from the protonation of N-9. N-16 also exhibited a small upfield chemical shift change of 4.4 ppm between 2.5–10 equivalents of TFA-d, which indicated N-16 as the second site of protonation. Right: NMR based model of the order of dominant protonation states for SM14, showing two sequential protonation events. Also, two pK_a values were detected with UV-metric pK_a measurements for SM14. Assuming that the sequence of protonation events will be conserved between water and acetonitrile solvents, SM14⁰ and SM14⁺¹ microstates shown in the figure are the major contributors to the UV-metric pK_a value 5.30 ± 0.01. SM14⁺¹ and SM14⁺² microstates shown in the figure are the major pair of microstates contributing to the UV-metric pK_a value 2.58 ± 0.01. There could be minor microstates with very low populations that could not be distinguished in these NMR experiments.