3-(N-Ethyl-3-methylanilino)-2-hydroxypropanesulfonic acid sodium salt

• CAS: 82692-93-1
• Catalog Number: ACM82692931
• Category: Main Products
• Molecular Weight: 295.33
• Molecular Formula: C12H18NNaO4S
• Synonyms: 3-(Ethyl(m-tolyl)amino)-2-hydroxypropane-1-sulfonic acid, sodium salt
• Appearance: White crystalline powder

Case Study

3-(N-Ethyl-3-Methylanilino)-2-Hydroxypropanesulfonic Acid Sodium Salt Used in Sensitive Spectrophotometric Assays

Pająk, Małgorzata. The journal of biochemistry 168.5 (2020): 509-514.

Understanding enzyme kinetics and inhibition mechanisms is fundamental in biochemical research, particularly for enzymes like L-Amino Acid Oxidase (LAAO), which has significant biological activities. TOOS (3-(N-Ethyl-3-methylanilino)-2-hydroxypropanesulfonic acid sodium salt) is used as the chromogenic coupling reagent in a peroxidase-dependent detection system. In the presence of peroxidase and 4-aminoantipyrine (4-AA), H2O2 reacts to form a quinoneimine dye that absorbs at 570 nm. TOOS therefore provides the terminal, spectrophotometrically measurable signal that directly reflects LAAO catalytic turnover.
Assay Workflow
· Buffers: McIlvaine buffer was prepared in either H2O or D2O (citric acid/Na2HPO4 mixtures) with pH/pD adjustments as needed for proton/deuteron kinetic comparisons.
· Detection mix (Solution A): 5.5 mg TOOS (≈0.019 mmol), 100 μL peroxidase solution (133 U in 1 mL buffer), 100 μL 4-AA solution (3.8 mg, 0.018 mmol in 1 mL buffer) made up to 4.8 mL with protonated or deuterated McIlvaine buffer.
· Substrate mix (Solution B): 3 mM solutions of L-Tyr, O-methyl-L-Tyr or O-methyl-[2-²H]-L-Tyr prepared in matching protonated or deuterated buffer.
· Enzyme: LAAO prepared as 1 mg enzyme protein per mL buffer.
· Reaction assembly: 133 μL Solution A, varying volumes of Solution B (25-150 μL in 25 μL increments), buffer to 743 μL final volume, and initiation with 7 μL LAAO solution.
· Monitoring: Absorbance at 570 nm recorded every minute for 30 minutes to follow quinoneimine formation. Initial rates from multiple substrate concentrations were fitted to the Michaelis-Menten equation (Enzfitter 1.05) to derive Vmax and Km, which were then used to compute KIE and SIE values.

Application of 3-(N-Ethyl-3-Methylanilino)-2-Hydroxypropanesulfonic Acid Sodium Salt in Enzymatic Assay for Inhibitor Characterization

Chang, Lei, et al. Journal of Medicinal Chemistry 57.23 (2014): 10080-10100.

TOOS (3-(N-ethyl-3-methylanilino)-2-hydroxypropanesulfonic acid sodium salt) was employed as a component of the peroxidase reagent used to quantify choline released by NPP family hydrolyze nucleotides, i.e., NPP1 (PC-1), NPP2 (autotaxin), and NPP3. In the reported workflow, TOOS is included at 2 mM in a Tris buffer together with peroxidase; after initial enzymatic conversion (e.g., LPC → choline), a coupled choline oxidase/peroxidase reaction produces a colored product that is read at 555 nm, providing a convenient spectrophotometric endpoint for inhibitor studies.
Experimental Protocol
· The inhibitory activity of compound against human NPP2 lysophospholipase D was measured in a buffer containing 5 mM MgCl2, 5 mM CaCl2, 100 mM Tris (pH 9.0), 400 μM LPC (18:1) and varying inhibitor concentrations.
· Reactions were started by adding 44 μg of human NPP2 and incubating at 37 °C for 60 minutes. Released choline was then detected spectrophotometrically at 555 nm after a 10-minute incubation at 37 °C with 50 μL each of the peroxidase reagent (50 mM Tris, pH 9.0, 2 mM TOOS, 5 U/mL peroxidase) and the choline oxidase reagent (50 mM Tris, pH 9.0, 2 mM 4-aminoantipyrine, 5 U/mL choline oxidase).
· Inhibition of human NPP3 by the same compound was assessed using the capillary electrophoresis-based NPP1 assay format with ATP as substrate, employing 43 μg of NPP3. IC50 values were obtained by fitting data from three independent experiments in Prism 5.0, and Ki values were calculated from those IC50s using the Cheng-Prusoff equation. The Michaelis-Menten constants were 138 μM for human NPP2 (with LPC (18:1)) and 43.4 μM for human NPP3.

Custom Q&A

What is the molecular formula of the keyword?

The molecular formula of the keyword is C12H18NNaO4S.

What is the molecular weight of the keyword?

The molecular weight of the keyword is 295.33 g/mol.

What are the synonyms of the keyword?

The synonyms of the keyword are 82692-93-1, TOOS, 3-(N-Ethyl-3-methylanilino)-2-hydroxypropanesulfonic acid sodium salt, TOOS sodium salt, and Sodium 3-(ethyl(m-tolyl)amino)-2-hydroxypropane-1-sulfonate.

What is the IUPAC name of the keyword?

The IUPAC name of the keyword is sodium;3-(N-ethyl-3-methylanilino)-2-hydroxypropane-1-sulfonate.

What is the InChIkey of the keyword?

The InChIkey of the keyword is IRQRBVOQGUPTLG-UHFFFAOYSA-M.

What is the Canonical SMILES of the keyword?

The Canonical SMILES of the keyword is CCN(CC(CS(=O)(=O)[O-])O)C1=CC=CC(=C1)C.[Na+].

What is the CAS number of the keyword?

The CAS number of the keyword is 82692-93-1.

What is the European Community (EC) number of the keyword?

The European Community (EC) number of the keyword is 617-377-8.

What is the hydrogen bond donor count of the keyword?

The hydrogen bond donor count of the keyword is 1.

What is the hydrogen bond acceptor count of the keyword?

The hydrogen bond acceptor count of the keyword is 5.