DiaPharma Group, Inc.

S-2765 is a chromogenic substrate for determination of Factor Xa. It is also very sensitive to trypsin.

Reagent -- Composition, Packaging, Storage and Stability

Each vial contains the chromogenic substrate S-2765, 25 mg and mannitol 60 mg as a bulking agent.

Stability: Lyophilized substance: stable at 25°C until expiry date printed on the label. The substance is hygroscopic and should be stored in a dry place.

Solution: 2 mmol/L in H2O is stable for six months at 2 to 8°C

Suitable stock solution: 1-2 mmol/L in H2O.

** Also available in bulk sizes (500+ mg) as a special order.

Chemistry

Chemical name: N-a-Benzyloxycarbonyl-Darginyl-L-glycyl-L-arginine-pnitroaniline-dihydrochloride

Formula: N-a-Z-D-Arg-Gly-Arg-pNA·2HCl

S-2765 structure

Mol. wt.: 714.6

e316 nm: 1.27 · 104 mol-1 · L · cm-1

Solubility:

> 40 mmol/L in H2O
> 10 mmol/L in Tris buffer (pH 8.3, I 0.25)

Principle

	Enzyme
N-a-Z-D-Arg-Gly-Arg-pNA		N-a-Z-D-Arg-Gly-Arg·OH+pNA

The method for the determination of activaty is based on the difference in absorbance between the pNA formed and the original substrate. The rate of pNA formation, i.e. the increase in absorbance per second at 405 nm, is proportional to the enzymatic activity and is conveniently determined with a photometer.

Kinetic Data

Factor Xa (bovine): km=1. 10-4 mol/L, kcat=290 sec-1 in Tris buffer pH 8.3, I 0.25 at 37°C.

Factor Xa: (human plasma activated with Russel's Viper Venom): km=3 · 10-4 mol/L in Tris buffer pH 7.8, I 0.4 at 37°C.

Standardization

An activity of DA/min=0.05 (37°C) is obtained by using a substrate concentration of 2 · km and a concentration of 0.04 nkat/mL of FXa (Chromogenix).