Succinic anhydride → 2-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

HRID 1055279Notes

Reaction equation

HRID 1055279 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].C1(=O)OC(=O)CC1>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

6

Reactant 3

HCID 7922Succinic anhydrideHCID 18730Fluorescein 5-isothiocyanateHCID 19660Carbonate Ion

Product 1

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

Reagents, catalysts & solvents 2

HCID 679Dimethyl SulfoxideSolvent4×HCID 962WaterSolvent

Procedure

Each of the HA-DETA, HA-DAHP, and HA-DAP obtained in Example 4-2 and Example 4-3-1 was dissolved in distilled water (Milli Q water), so as to prepare a solution of a concentration of 20.0 mg/ml. Thereafter, a 0.2 M carbonate buffer (pH 9.0) was added to the prepared solution, resulting in a concentration of 10.0 mg/ml. Thereafter, fluorescein isothiocyanate that was 0.07 mole times to an HA unit was dissolved in DMSO that was 1/10 the volume of a modified HA solution. The DMSO solution was then added to the above solution, and the obtained mixture was then stirred in the dark at room temperature for 1 hour. Thereafter, succinic anhydride that was 40 mole times to the HA unit was dissolved in DMSO that was 1/10 the volume of the HA-AM solution. The DMSO solution was then added to the above solution, and the obtained mixture was then stirred in the dark at room temperature for 30 minutes. Thereafter, each sample was dialyzed against large excess amount of 0.5 M NaCl aqueous solution for purification, and the external dialysis solution was then exchanged with distilled water (Milli Q water), followed by further dialysis and purification (Spectrapore 4; molecular weight cut off (MWCO): 12 k-14 kDa). The obtained aqueous solution was freeze-dried, so as to obtain a fluorescein-labeled modified HA.

Workup

  1. customso as to prepare a solution of a concentration of 20.0 mg/ml
  2. customresulting in a concentration of 10.0 mg/ml
  3. stirringthe obtained mixture was then stirred in the dark at room temperature for 30 minutes
  4. customThereafter, each sample was dialyzed against large excess amount of 0.5 M NaCl aqueous solution for purification
  5. customfollowed by further dialysis and purification (Spectrapore 4; molecular weight cut off (MWCO): 12 k-14 kDa)
  6. customThe obtained aqueous solution was freeze-dried

Sources & evidence

ORD record
ord-5959ae29aecd43fd991c12aeab8ccbbb
Source dataset
uspto-grants-2016_07
Patent number
US09394379B2

Reaction notes

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