CNP-GlcNAc

CNP-GlcNAc (2-Chloro-4-nitrophenyl β-D-N-acetylglucosaminide, CAS 103614-82-0) is a chromogenic substrate specifically designed for quantitative spectrophotometric detection of β-N-acetylglucosaminidase (NAGase, EC 3.2.1.52; also known as β-hexosaminidase) activity through enzymatic hydrolysis of the β-glycosidic bond, releasing 2-chloro-4-nitrophenolate anion with strong yellow absorbance at 405 nm (ε ≈ 14,300 M⁻¹cm⁻¹) for sensitive kinetic assays down to 0.001 U/L enzyme concentrations. With molecular formula C14H17ClN2O8 (MW 376.75 g/mol), this white to off-white crystalline solid features β-D-N-acetylglucosamine (GlcNAc) O-linked via C1 to the electron-deficient 2-chloro-4-nitrophenyl aglycone, where ortho-chloro substitution enhances leaving group efficiency (pKa ~7.1) and hydrolysis kinetics (kcat/Km ~10^4 M⁻¹s⁻¹ for human HexA/HexB isoforms) while maintaining strict β-anomeric specificity over α-GlcNAc linkages. Routinely employed in clinical diagnostics for lysosomal storage disorders (Tay-Sachs/Sandhoff disease via HexA deficiency), urinary NAGase screening for renal tubular damage (diabetes/nephrotoxicity), and glycobiology research profiling GH3/GH20 family enzymes in chitin degradation/microbiome studies, it supports 96-well formats (0.2-1 mM substrate, pH 4.3-4.8, 37°C) with excellent linearity (0.001-2 U/L) and minimal interference from endogenous chromophores. Superior to p-nitrophenyl-β-GlcNAc by 15-25% faster turnover due to chlorine enhancement, the yellow product’s pH stability (6-10) enables reliable endpoint reads, positioning CNP-GlcNAc as the preferred reagent for therapeutic monitoring, enzyme replacement therapy evaluation, and high-throughput screening of chitinase/NAGase inhibitors targeting fungal infections, cancer metastasis, and inflammatory joint diseases.

Appearance

• White to off-white crystalline powder.
• Hydrolysis yields bright yellow solution (λmax 405 nm).​

Source

• Koenigs-Knorr synthesis: protected β-D-GlcNAc donor + 2-chloro-4-nitrophenol.
• Commercial analytical grade (Hepattack/GeneSeqTools >98%).

Molecular Weight and Structure

• Formula: C14H17ClN2O8; MW 376.75 g/mol.
• (2R,3R,4R,5S,6R)-2-(2-acetamido-2-(2-chloro-4-nitrophenoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol.​

Sugar Specificity

• Selective β-D-N-acetylglucosaminide for β-N-acetylhexosaminidases (HexA/B).
• 500x preference over α-GlcNAc linkages.​

Biological Activity

• Chromogenic NAGase substrate; 1 U = 1 μmol CNP/min, pH 4.5, 37°C.
• Km 0.2-0.8 mM (human HexA).​

Purity and Microbial Contamination

• ≥98% (HPLC); research/diagnostic grade.
• Synthetic; non-sterile.​

Identity and Quality Control

• ¹H-NMR (β-anomeric H 4.9 ppm d, J=8.0 Hz); HRMS m/z 377 [M+H]+.
• Quantitative hydrolysis yield at A₄₀₅.​

Shelf Life and Storage

• 3 years -20°C desiccated; aqueous 10 mM stable 3 months -80°C.
• Moisture/light protected.​

Applications

• Lysosomal storage disorder diagnostics (Tay-Sachs); renal NAGase screening.
• Chitinase assays, HTS inhibitors.​

Key Characteristics

• 15-25% faster than pNP-β-GlcNAc; high sensitivity.
• pH-stable yellow chromophore.​

Citation Links

• https://www.hepattack.com/en/product03_cid_217_pid_592.html​
• http://www.geneseqtools.cn/cn/product03_cid_217_pid_592.html​
• http://www.hepattack.com/en/product03_cid_217_pid_592.html​
• https://pmc.ncbi.nlm.nih.gov/articles/PMC5095700/​
• https://pmc.ncbi.nlm.nih.gov/articles/PMC2953398/​
• https://go.drugbank.com/drugs/DB00141​
• https://en.wikipedia.org/wiki/N-Acetylglucosamine​
• http://exposome-explorer.iarc.fr/compounds/2163​
• http://www.level.com.tw/_i/assets/upload/files/Agency/CBR/Catalog/Enzyme%20Substrate%20and%20Detergent%20Catalogue.pdf​
• https://pubchem.ncbi.nlm.nih.gov/compound/N-Acetyl-D-Glucosamine