Novel fluorescence detection reagent and technology for quantitatively detecting in-vivo and in-vitro CO

A fluorescence detection and reagent technology, applied in the field of analysis and detection, can solve the problems of limited large-scale use, weak tissue penetration ability, short emission wavelength, etc., and achieve strong practical application value, low cytotoxicity, and high specificity

Pending Publication Date: 2022-03-01
HUNAN TARGETING DETECTION TECH CO LTD
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AI-Extracted Technical Summary

Problems solved by technology

However, the maximum emission wavelength of this probe is relatively short, resulting in its weak tissue penetration ab...
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Method used

Preparation 5mL concentration is that the PBS aqueous solution of various conventional ions and amino acid of 50mM and concentration is that the fluorescent detection reagent that 1mM embodiment 1 prepares is as standby. Dilute with PBS buffer solution (10mM, pH=7.4), adjust to the concentration of the probe compound in the solution is 10μM, the concentration of CO is 30μM, and O2-, OH, H2O2, HOCl, ONOO-, CN-, H2S , The concentration of GSH was 100 μM. After incubation at room temperature for 20 min, the fluorescence spectra of different systems were tested in 10 mm cuvettes (Figure 3). The results showed that other analytes had almost no effect on the fluorescence of the fluorescent molecular probes, but the addition of CO significantly enhanced the fluorescence of the fluorescent molecular probes.
Preparation 5mL concentration is that the aqueous solution of 1mM CO and concentration are the fluorescent detection re...
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Abstract

The invention discloses a novel fluorescence detection reagent and a technology for quantitatively detecting in-vivo and in-vitro CO, and relates to a near-infrared fluorescence probe, the chemical structural formula of the near-infrared fluorescence probe is shown in the specification, and the fluorescence detection reagent can realize high-sensitivity and high-specificity detection of CO in environment, cells or organisms. Meanwhile, the probe used in the invention is a near-infrared probe, has good biological tissue permeability, low cytotoxicity and excellent water solubility, can avoid interference from biomacromolecule background fluorescence, can also be applied to detection of endogenous CO in living cells so as to realize in-vivo and in-vitro quantitative analysis and detection of CO, and has wide application.

Application Domain

Organic chemistryFluorescence/phosphorescence +1

Technology Topic

Biological tissueBackground fluorescence +14

Image

  • Novel fluorescence detection reagent and technology for quantitatively detecting in-vivo and in-vitro CO
  • Novel fluorescence detection reagent and technology for quantitatively detecting in-vivo and in-vitro CO
  • Novel fluorescence detection reagent and technology for quantitatively detecting in-vivo and in-vitro CO

Examples

  • Experimental program(5)

Example Embodiment

[0028] Example 1
[0029] Compound 3 synthesis
[0030] Compound 1 (1.56 g, 5 mmol) and compound 2 (1.30 g, 5 mmol) were dissolved in 5 ml of concentrated sulfuric acid, and at 90 o CHer heat reflow, TLC was monitored to the end of the reaction, remove the reaction, cooled to room temperature, poured into ice water, and added 1 ml of hydrochloric acid, see a large amount of precipitation, filtrate, collected, washing the filter, recrystallization with ethanol Deep green solid 1.50 g, the yield is 55.7%.
[0031] Compound 4 synthesis
[0032] At room temperature, Compound 3 (1.34 g, 2.5 mmol), hydrazine (1.64 mL, 25 mmol) and BOP (1.13 g, 2.6 mmol) were added to 50 ml of round bottom flask, and 10 ml of dichloromethane dissolved. The reaction system was stirred at room temperature, and TLC was monitored to the end of the reaction. The reaction was removed, the solvent was removed under reduced pressure. After purification of the column chromatography, the yellow solid was obtained from 1.12 g, and the yield was 78.3%. 1 H NMR (400 MHz, DMSO-D 6 ): δ = 7.93 (D, J = 7.6 Hz, 1H), 7.78 (T, J = 7.2, 1H), 7.58 (T, J = 7.2, 1H), 7.53 (S, 1H), 7.49 (D, J = 7.6 Hz, 1H), 7.41 (T, J = 7.8 Hz, 1H), 6.70-6.67 (m, 2H), 6.52 (S, 1H), 6.22 (D, J = 6.8 Hz, 1H), 6.15 (S, 1H), 5.47 (S, 1H), 3.38 (Q, J = 6.8 hz, 4h), 3.31 (q, J = 6.8 hz, 4h), 1.17-1.13 (m, 12h).
[0033] Synthesis of target molecular probe
[0034] Compound 4 (550.65 mg, 1 mmol) and 2-pyridine formaldehyde (240.05 mg, 1 mmol) were dissolved in 10 ml of anhydrous methanol, and the mixture was completed at room temperature to end. After the end of the reaction, the solvent was removed under reduced pressure. After purification of the column chromatography, the red solid was 638.63 mg, and the yield was 81.7%. 1 H NMR (400 MHz, CDCL 3 : δ = 9.13 (D, J = 7.2, 1H), 8.48 (T, J = 8.2, 1H), 8.01 (T, J = 8.2, 1H), 7.91 (D, J = 7.6 Hz, 1H), 7.82 (T, J = 7.2, 1H), 7.60-7.56 (M, 2H), 7.50 (S, 1H), 7.43-7.40 (m, 2H), 7.18 (D, J = 7.6Hz, 1H), 6.88 (D, J = 8.2 Hz, 1H), 6.69 (D, J = 8.0 Hz, 1H), 6.54 (S, 1H), 6.20 (D, J = 6.8 hz, 1H), 6.13 (S, 1H), 5.47 (S, 1H), 4.39 (S, 3H), 3.41 (Q, J = 7.2Hz, 4H), 3.31 (q, J= 7.1 Hz, 4H), 1.16 (T, J = 7.0 hz, 6h), 1.21 (t, J = 7.1 Hz, 6H). HRMS-ESI M / Z: Calc. For [m-i] + : 654.3074; Found, 654.3719.

Example Embodiment

[0035] Example 2
[0036] Response of fluorescence detection for different concentrations CO
[0037] The fluorescent detection test agents prepared by the aqueous solution of 5 ml of a concentration of 1 mM CO and a concentration of 1 mm were prepared were used as a spare. Dilution was diluted with PBS buffer solution (10 mM, pH = 7.4), the concentration of 10 μm contained in the solution was adjusted to the solution, and the concentration of CO was 0 μm, 1 μm, 2 μm, 4 μm, 7 μm, 10 μm, and. After 20 min at room temperature, the fluorescence spectrum of different systems was tested in 10 mm cuvette. figure 1. Calculate the fluorescence intensity in each system and establish a standard curve of fluorescence intensity and CO concentration. Such as figure 2 As shown, when the concentration of CO is in the range of 0-4 μm, the fluorescence intensity is a good linear relationship with the concentration of CO, and the regression linear equation is Y = 18.56897X + 6.60021, the linear correlation coefficient is: 0.99153, and calculate the test The limit (LOD) is 1.02 μm (S / N = 3), indicating that the fluorescent probe has good sensitivity.

Example Embodiment

[0038] Example 3
[0039] Psychic detection reagent to CO selectivity
[0040] The PBS aqueous solution of various conventional ions and amino acids prepared for 5 ml of concentration of 50 mm and a concentration of a fluorescent detection tester prepared as 1 mM Example 1 were used as spare. Dilution was diluted with PBS buffer solution (10 mM, pH = 7.4), adjusted to a solution of a probe compound having a concentration of 10 μm, CO concentration of 30 μm, and O 2 - , OH, H 2 O 2 , HOCL, OONOO - , CN - H 2 S, GSH concentration is 100 μm. After 20 min at room temperature, the fluorescence spectrum of different systems was tested in 10 mm cuvette. image 3. The results showed that other objects to be tested had little effect on the fluorescence of the fluorescent molecular probe, while CO was added to increase the fluorescence of the fluorescent molecule probe.

PUM

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