Two-photon fluorescence probe for selectively detecting iron (III) complex in biological mercaptan

A two-photon fluorescence and biological thiol technology, which is applied in the field of two-photon fluorescent small molecule probes and two-photon fluorescent probes, can solve the problems of small types and quantities of two-photon fluorescent molecules, and achieve the effect of fluorescence recovery

Active Publication Date: 2018-07-20
NANJING UNIV OF TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the research on two-photon fluorescent probes is still in its infancy, and the types and quantities of two-photon fluorescent molecules with excellent performance are relatively small. Therefore, it is necessary to design and synthesize different types of two-photon fluorescent materials and develop two-photon fluorescent materials with excellent properties. Fluorescent probes will provide more research tools for biomedical analysis and other related fields

Method used

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  • Two-photon fluorescence probe for selectively detecting iron (III) complex in biological mercaptan
  • Two-photon fluorescence probe for selectively detecting iron (III) complex in biological mercaptan
  • Two-photon fluorescence probe for selectively detecting iron (III) complex in biological mercaptan

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Experimental program
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Effect test

Embodiment 1

[0021] A two-photon fluorescent molecule that specifically recognizes biothiol molecules. Denoted as compound TPS, the structural formula is as follows:

[0022]

[0023] The preparation steps of compound TPS are as follows:

[0024]

[0025] Compound 2 (74 mg, 0.26 mmol) was dissolved in 4 mL of anhydrous DMF at room temperature, compound 1 (70 mg, 0.26 mmol) was added to it, and EDCl (99 mg, 0.52 mmol) was added, and after stirring for 24 hours, water was added to stop the reaction . The solution was washed with saturated sodium chloride solution, then extracted three times with dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The filter residue was purified by column chromatography (PE / EA=4 / 1) to obtain pure TPS as a light brown solid with a yield of 32%.

Embodiment 2

[0027] TPFeS detects biothiol molecules in vitro, interference experiments on TPFeS probes.

[0028] First prepare the TPFeS solution in Hepes buffer with 0.1% Triton X-100, where TPS / Fe 3+ =2 / 1, add 1mL to 22 1.5mL centrifuge tubes respectively, add 2uL of 100mM amino acid aqueous solution prepared in advance to each of these centrifuge tubes, shake at room temperature for 2 hours, take 80uL and add to the enzyme label To test in the plate, the excitation light is 350nm ( figure 2 a), only three kinds of biothiol molecules have fluorescence-enhanced response to TPFeS; the statistical analysis of the experimental results shows that the probe TPFeS has significant specificity to biothiol molecules, and is not affected by other interfering substances such as amino acids. In subsequent cell experiments, the reaction of the probe with thiol molecules in the cells provided evidence.

Embodiment 3

[0030] In vitro concentration trend curves of TPFeS probe to three thiol molecules Cys(b), Hcy(c), and GSH(d). First, add TPFeS to the Hepes buffer prepared with 0.1% Triton X-100 to make the concentration reach 10uM, in which TPS / Fe 3+ = 2 / 1, divided into 1.5mL centrifuge tubes, respectively added three kinds of thiol molecules with different concentrations in the centrifuge tubes, shaken for two hours at room temperature, and took 80uL of enzyme in each centrifuge tube to 384 holes Tested in the standard board, respectively obtained figure 2 (b) (c) (d) curves, the fluorescence gradually increases with the increase of the thiol concentration, and a good linear relationship between the concentration and the fluorescence intensity is obtained.

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Abstract

The invention relates to a two-photon fluorescence probe for selectively detecting an iron (III) complex in biological mercaptan and belongs to the technical field of organic small-molecule fluorescence probes. The molecular structure of the two-photon fluorescence probe has main characteristics that molecules coordinate with Fe<3+>firstly to realize fluorescence quenching, then Fe<3+> is snatchedby mercaptan molecules due to higher binding capacity of the mercaptan molecules for Fe<3+>, and recovery of a fluorescent molecule is realized, so that recovery of fluorescence is achieved. The biological probe is low in cost, the method is simple and easy, reaction conditions are mild, treatment is easy, production cost is low, and the probe can be applied to in-vivo mercaptan detection. The structures of the fluorescent molecule TPS and the probe TPFeS are shown in the description.

Description

technical field [0001] The invention relates to a two-photon fluorescent probe for selectively detecting iron (Ⅲ) complexes of biological thiols, in particular to a two-photon fluorescent small molecule probe coordinated by iron (Ⅲ), which belongs to organic small molecule fluorescent probes. needle technology field. Background technique [0002] Fluorescence analysis has become an indispensable detection tool in the fields of biological analysis and life sciences because of its high analytical sensitivity, strong selectivity, and ease of use, and small fluorescent molecules cause little damage to cells and organisms. At present, more and more fluorescent small molecule probes have been applied to real-time detection at the molecular level. The detection objects include various ions, small molecules, free radicals, proteases, etc., and even temperature, viscosity, etc. People use the fluorescent in vivo imaging system to monitor and obtain relevant information about the det...

Claims

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Application Information

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IPC IPC(8): C07D413/12C09K11/06G01N21/64
CPCG01N21/6428G01N21/6486C09K11/06C07D413/12C09K2211/187C09K2211/1033
Inventor 李林赵燕菲张承武王刘林白磊石日日彭平平于海东刘金华黄维王延宾
Owner NANJING UNIV OF TECH
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