Synthesis
4-Nitrofluorescein was prepared as follows: 8.6 g (18.7 mmol) of ethyl 4-nitrofluorescein triacetate was dissolved in 87 mL of 10% sodium hydroxide solution and 432 mL of methanol and gently heated. The precipitate was filtered and dissolved in 500 mL of water. The solution was acidified with hydrochloric acid and allowed to stand overnight in the dark. The crystals were filtered out and washed with water. 6.9 g (18.2 mmol), 97% red crystals of 4-nitrofluorescein were obtained.C20H11NO7; molecular weight: 377.30; 1HNMR (300 MHz, DMSO-d6)δ 10.20 (brs, 2H, OH), 8.65 (s, 1H, nsi-4H), 8.57 (d, J=8.4. 1H, nsi-6H), 7.57 (d, J=8.3, 1H, nsi-7H), 6.70-6.64 (m, 4H, xa-4, 5, 2, 7H), 6.56 (d, J=8.7, 2H, xa-1, 8H). esims: 376.1 [M-H+] (100).
