A near-infrared Nile Red fluorescence probe for discrimina-tion of biothiols by dual-channel response and its bioimaging applications in living cells and animals

Biothiols, including cysteine (Cys), homocysteine (Hcy), glutathione (GSH) and H2S, play important roles in human physiology processes. However, it is a great difficulty to distinguish biothiols from each other because of their similar chemical properties. Based on the Nile Red, we have designed and synthesized a near-infrared fluorescent probe for discriminating Cys/Hcy from GSH/H2S by a dual-channel detection method. Using an ether bond, a near-infrared Nile Red was attached to 7-nitrobenzofurazan to construct the probe. Due to the photo-induced electron transfer, the probe performed almost no fluorescence from green to red emission band. But upon addition of Cys (0-150 μM) or Hcy (0-200 μM), the probe exhibited a noteworthy fluorescence “turn-on” signal in two unique emission bands (Green and Red) with a fast response (within 30 min). In contrast, the probe displayed the increment fluorescence only in the red channel when encountering GSH (0-70 μM) or H2S (0-50 μM). And GSH/H2S could be tested respectively by different response time. The limit of detection was calculated to be 0.09 μM (Cys), 0.30 μM (Hcy), 0.24 μM (GSH), and 0.04 μM (H2S), respectively (based on S/N = 3). The desirable dual-channel detection could be achieved in the serum sample and living cells. Moreover, the probe could be applied for the bioimaging in mice, which indicated its potential application in clinic.