Fluorescein 5-isothiocyanate → 2-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

HRID 1099115Notes

Reaction equation

HRID 1099115 reaction equation
C(=O)([O-])[O-].[CH:5]1[C:10](N=C=S)=[CH:9][C:8]2[C:14]([O:16][C:17]3([C:27]4[CH:28]=[CH:29][C:30]([OH:32])=[CH:31][C:26]=4[O:25][C:19]4[CH:20]=[C:21]([OH:24])[CH:22]=[CH:23][C:18]3=4)[C:7]=2[CH:6]=1)=[O:15]>O.CS(C)=O>[CH:5]1[CH:10]=[CH:9][C:8]([C:14]([OH:16])=[O:15])=[C:7]([C:17]2[C:18]3[CH:23]=[CH:22][C:21]([OH:24])=[CH:20][C:19]=3[O:25][C:26]3[C:27]=2[CH:28]=[CH:29][C:30]([CH:31]=3)=[O:32])[CH:6]=1

Participants

5

Reactant 2

HCID 18730Fluorescein 5-isothiocyanate2×HCID 19660Carbonate Ion2×

Product 1

HCID 33832-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

Reagents, catalysts & solvents 2

HCID 679Dimethyl SulfoxideSolventHCID 962WaterSolvent

Procedure

This HA-HZ was dissolved in distilled water (Milli Q water), and an equal amount of 100 mM carbonate buffer (Ph 9.0) was then added thereto to a final concentration of 1 mg/ml. Thereafter, FITC dissolved in DMSO that was 1/10 the volume of the HA-HZ solution was added to the mixture at an additive ratio of FITC/HA unit=3.0 mol/mol, and the obtained mixture was then reacted in the dark at room temperature for 1 hour. Thereafter, 25 ml of the reaction solution was poured in PD-10 columns (10 columns) that had previously been equilibrated with a 50 mM carbonate buffer (pH 9.0), so as to remove unreacted FITC. Succinic anhydride dissolved in DMSO (3.5 ml) was added to the thus roughly purified solution at an additive ratio of succinic anhydride/HZ=250 mol/mol, and the same above reaction was carried out. The reaction mixture was dialyzed against large excess amount of distilled water (Milli Q water) for purification, followed by freeze-drying, so as to obtain fluorescein-labeled HA-HZ-SUC, which was formed by labeling HA-HZ with FITC.

Workup

  1. customthe obtained mixture was then reacted in the dark at room temperature for 1 hour
  2. customso as to remove unreacted FITC
  3. dissolutionSuccinic anhydride dissolved in DMSO (3.5 ml)
  4. additionwas added to the thus roughly purified solution at an additive ratio of succinic anhydride/HZ=250 mol/mol
  5. customthe same above reaction
  6. distillationThe reaction mixture was dialyzed against large excess amount of distilled water (Milli Q water) for purification
  7. customby freeze-drying

Sources & evidence

ORD record
ord-a9af082ab38348538b0570fac565e6a7
Source dataset
uspto-grants-2012_03
Patent number
US08143391B2

Reaction notes

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