Water → Agarose low melting and low gelling temperature

HRID 1436989Notes

Reaction equation

HRID 1436989 reaction equation
[Cl-].[Na+].[Cl-].[Ca+2].[Cl-].[OH2:6].C1O[C@H]2[C@H](O)[C@H]([O:15][C@@H:16]3[C@@H:21]([OH:22])[C@H:20]([O:23][C@H:24]4[C@@H:28]5[C@H:29]([OH:44])[C@H:30]([O:32][C@@H]6[C@@H](O)[C@H](O)O[C@H](CO)[C@@H]6O)[O:31][C@H:25]4[CH2:26][O:27]5)[O:19][C@H:18]([CH2:45][OH:46])[C@@H:17]3[OH:47])O[C@@H]1[C@H]2O>C(O)C(N)(CO)CO.Cl>[CH2:26]1[O:6][C@H:28]2[C@H:29]([OH:44])[C@H:30]([OH:32])[O:31][C@@H:25]1[C@H:24]2[O:23][C@@H:20]1[O:19][C@H:18]([CH2:45][OH:46])[C@H:17]([OH:47])[C@H:16]([O:32][C@@H:30]2[O:31][C@H:25]3[CH2:26][O:27][C@H:28]([C@@H:24]3[O:23][C@@H:20]3[O:19][C@H:18]([CH2:45][OH:46])[C@H:17]([OH:47])[C@H:16]([OH:15])[C@H:21]3[OH:22])[C@@H:29]2[OH:44])[C@H:21]1[OH:22]

Participants

6

Reactant 4

HCID 962WaterHCID 5234Sodium Chloride2×HCID 5284359Calcium ChlorideHCID 54758609HCID 54758609, compound

Product 1

HCID 11966311Agarose low melting and low gelling temperature

Reagents, catalysts & solvents 1

HCID 93573Tromethamine hydrochlorideSolvent2×

Procedure

A solution containing agarose at a concentration of 0.2% (w/v) or 0.5% (w/v) in 20 mM Tris-HCl buffer (pH 7.2) containing 10 mM sodium chloride (in case of Agarase I) or 20 mM Tris-HCl buffer (pH 7.2) containing 10 mM calcium chloride and 10 mM sodium chloride (in case of Agarase II) was prepared. 180 μl of this solution as a substrate was mixed with 20 μl of an enzyme solution. The mixture was reacted at 50° C. for 5 to 30 minutes, preferably 10 minutes, and then heated in boiling water or at 75° C. for 1 minute to stop the reaction. 30 μl of the reaction mixture was subjected to a TSKgel α-2500 column (inner diameter: 7.8 mm; length: 300 mm; Tosoh). Elution was carried out using a 70% acetonitrile solution as an eluent at a flow rate of 0.8 ml/minute. Agarotetraose eluted at a retention time of about 25 minutes or neoagarotetraose eluted at a retention time of about 24 minutes, which was generated by the enzymatic reaction, was quantified. One unit (1 U) is defined as the amount of the enzyme that produces 1 μmol of agarotetraose or neoagarotetraose in 10 minutes.

Workup

  1. customwas prepared
  2. customThe mixture was reacted at 50° C. for 5 to 30 minutes, preferably 10 minutes
  3. temperatureheated
  4. customthe reaction
  5. washElution
  6. washAgarotetraose eluted at a retention time of about 25 minutes or neoagarotetraose
  7. washeluted at a retention time of about 24 minutes, which
  8. customwas generated by the enzymatic reaction

Sources & evidence

ORD record
ord-64fd8a23ad04468aa51de6507cc09c9f
Source dataset
uspto-grants-2009_11
Patent number
US07622291B2

Reaction notes

Loading…