Sodium Hydroxide → L-Tyrosine

Participants

Reaction Steps

HA (1 g, 2.5 mmol) was dissolved in 100 ml of distilled water, forming an initial solution. Tyramine hydrochloride (162 mg, 0.93 mmol) and 5-aminofluorescein (81 mg, 0.23 mmol in 1.62 ml DMSO) were added to this solution. EDC.HCl (479 mg, 2.5 mmol) and NHS (290 mg, 2.5 mmol) were then added and the pH of the mixture was maintained at 4.7 with 0.1 M NaOH. The solution was stirred overnight at room temperature and then brought to pH 7.0. The solution was next filtered with grade 1 Whatman™ cellulose filter paper to remove unconjugated aminofluorescein that had precipitated. The filtrate was collected into dialysis tubes of molecular cut-off 3500 Da. Then the dialysis and lyophilization procedures described in Example I-A were carried out. The degree of substitution of tyramine was calculated from 1H NMR and the degree of aminofluorescein conjugated was estimated by comparing the absorbance value at 490 nm of 1 mg/ml fluorescence-conjugated HA-Tyr solution to a set of aminofluorescein standards. The degrees of substitution of tyramine and aminofluorescein were 4 and 0.4, respectively.

Provenance & Evidence