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Preparation method for 6-carboxylfluorescein

A carboxyfluorescein and reaction technology, applied in the field of preparation of 6-carboxyfluorescein, can solve the problems of low yield and large loss, and achieve the effects of reducing production cost, high application value, and simplifying purification process

Active Publication Date: 2015-01-14
WUHU HUAREN SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Even if separated, because the two isomers cross each other, the loss is large and the yield is low

Method used

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  • Preparation method for 6-carboxylfluorescein
  • Preparation method for 6-carboxylfluorescein
  • Preparation method for 6-carboxylfluorescein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020]

[0021] a. Reaction steps:

[0022] At room temperature, in a reaction vessel, the resorcinol 2 (resorcinol, 57.2 g, 0.52 mol) dissolved in methanesulfonic acid (520 ml), added trimellitic anhydride 1 (1,2,4-Benzenetricarboxylic anhydride, 50 g, 0.26 mol), after stirring at room temperature for 10 minutes, tin tetrachloride (SnCl 4 , 6.1 ml, 0.055 mol), heated, and reacted at 90 ° C. After 6 hours, HPLC showed that the reaction was complete, and the reaction was stopped. The reaction system was cooled to room temperature to obtain a reaction solution;

[0023] b. Crystallization step

[0024] The reaction solution was poured into 4 liters of ice-water solution, stirred for 10 minutes, and the product precipitated out. Filter and dry, and the dried crude product is crystallized with methanol and hexane (methanol:hexane volume ratio is 1:4). Filter and dry to obtain 6-carboxyfluorescein methylsulfonic acid adduct.

[0025] c, hydrolysis step:

[0026] Add 6-ca...

Embodiment 2

[0031]

[0032] a. Reaction steps:

[0033] At room temperature, in a reaction vessel, 4-chlororesorcinol 3 (4-Chlororesorcinol, 75.2 g, 0.52 mol) dissolved in methanesulfonic acid (620 ml), adding trimellitic anhydride 1 (1,2,4-Benzenetricarboxylic anhydride, 50 g, 0.26 mol), after stirring at room temperature for 15 minutes, tin tetrachloride (SnCl 4 , 6.7 ml, 0.057 mol), heated, and reacted at 105 °C. After 6 hours, HPLC showed that the reaction was complete, and the reaction was stopped. The reaction system was cooled to room temperature to obtain a reaction solution;

[0034] b. Crystallization step

[0035] The reaction solution was poured into 4 liters of ice-water solution, stirred for 10 minutes, and the product precipitated out. Filter and dry, and the dried crude product is crystallized with methanol and hexane (methanol:hexane volume ratio is 1:4). Filter and dry to obtain 2',7'-dichloro-6-carboxyfluorescein methanesulfonic acid adduct.

[0036] c, hydrol...

Embodiment 3

[0042]

[0043] a. Reaction steps:

[0044] At room temperature, in a reaction vessel, 4-methylresorcinol 4 (4-methylresorcinol, 64.6 g, 0.52 mol) was dissolved in methanesulfonic acid (600 ml), and trimellitic anhydride was added 1 (1,2,4-Benzenetricarboxylic anhydride, 50 g, 0.26 mol), after stirring at room temperature for 10 minutes, tin tetrachloride (SnCl 4, 5.8 ml, 0.052 mol), heated, and reacted at 95 ° C. After 6 hours, HPLC showed that the reaction was complete, and the reaction was stopped. The reaction system was cooled to room temperature to obtain a reaction solution;

[0045] b. Crystallization step

[0046] The reaction solution was poured into 4 liters of ice-water solution, stirred for 10 minutes, and the product precipitated out. Filter and dry, and the dried crude product is crystallized with methanol and hexane (methanol:hexane volume ratio is 1:4). Filter and dry to obtain 2',7'-dimethyl-6-carboxyfluorescein methanesulfonic acid adduct.

[0047]...

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Abstract

The invention discloses a preparation method for 6-carboxylfluorescein. The method comprises the following steps: a. reacting; b. crystallizing; and c. hydrolyzing. The preparation method prepares 6-carboxylfluorescein by using a metal catalyst of stannic chloride (SnCl4) to selectively catalyze the reaction, and selects an appropriate solvent mixing ratio and an appropriate purification method. Compared with the prior art, the ratio of the required product 6-carboxylfluorescein and the by-product 5-carboxylfluorescein is improved from 1:1 to 2.6:1, and the separation yield is improved from 32% to 61% (1.9 times as much as the past). The method greatly reduces the production cost, simplifies the purification process, provides a practical and efficient novel process method for the preparation of 6-carboxyfluorescein and derivatives thereof which are widely used for fluorescence labeling of biological macromolecules, and has a high application value.

Description

technical field [0001] The invention belongs to the preparation method of 6-carboxyfluorescein, in particular to the preparation method of selective metal catalysis for preparing 6-carboxyfluorescein. Background technique [0002] 6-carboxyfluorescein is a widely used fluorescent dye, and its related derivatives can be combined with biological macromolecules such as proteins, antibodies, polypeptides, nucleic acids and phospholipids, and are widely used in biological science and medical research, such as nucleic acid probes , real-time PCR and drug target labeling technology, etc. However, in the existing methods for preparing 6-carboxyfluorescein, there is a common problem, that is, when 6-carboxyfluorescein is generated, about the same amount of 5-carboxyfluorescein is generated (references: Yuichiro Ueno, Guan -Sheng Jiao & Kevin Burgess, Synthesis, 2004 (15), 2591-2593). Since 5-carboxyfluorescein and 6-carboxyfluorescein are positional isomers with similar properties,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D311/80
Inventor 石平
Owner WUHU HUAREN SCI & TECH CO LTD
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