Hydrophilic 1,8-Naphthalimide Fluorescent Ph Probes And Their Application To Living Cells Imaging

Four fluorescent pH probes were synthesized from 1,8-naphthalic anhydride and aliphatic amines: 4-(2'-N,N-dimethylaminoethylamino)-N-n-butyl-1,8-naphthalimide (MBN), 4-(2'-N,N-diethylaminoethylamino)-N-n-butyl-1,8-naphthalimide (EBN), 4-(2'-N,N-dimethylaminoethylamino)-N-(2'-dimethylaminoethyl)-1,8-naphthalimide (MMN) and 4-(2'-N,N-diethylaminoethylamino)-N-(2'-diethylaminoethyl)-1,8- naphthalimide (EEN). Their structures were characterized by FTIR, NMR, MS and elementary analysis.MBN, EBN, MMN, EEN have good hydrophilicity and thermal stability, their pKa values are 8.50, 9.02, 8.50 and 8.96. The maximum absorption wavelengthλmax (molar extinction coefficientε) of 5.0*10-5 mol/L MBN and EBN ethanol water (V:V=1:19) solution are 438 nm (1.29 M-1·cm^-1) and 436 nm (1.32 M-1·cm^-1), the corresponding maximum fluorescence wavelengthλFL (fluorescence quantum yieldsφ) are 529 nm (0.599) and 529 nm (0.759). Theλmax (ε) of 5.0*10-5 mol/L MMN and EEN water solution are 444 nm (1.60 M-1·cm^-1) and 444 nm (1.36 M-1·cm^-1), the correspondingλFL (φ) are 534 nm (0.558) and 536 nm (0.647).MBN, EBN, MMN, EEN have good selectivity to H+ in Britton-Robinson buffer solution with coexisting ions Na+, K+ (200 times), Mg²⁺, Ca²⁺ (100 times), Cr³⁺, Mn²⁺, Fe³⁺, Fe²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺, Cd²⁺, Hg²⁺, Pb²⁺ (10 times), Cl- (560 times), PO43- (800 times), SO42- (10 times), NO3- (60 times), and the coexisting ions have no obvious influence on the H+ detection. The fluorescence enhancement of MBN, EBN, MMN and EEN are 12.9, 11.8, 13.6 and 25.6 times respectively after combining protons. EEN can be used for living cells imaging. When the extracellular pH value increased from 5.51 to 7.55, the fluorescence intensity of the cells increased gradually because more EEN molecules entered into the cells and bound more Intracellular H+.