6-Chloro-1H-indol-3-yl beta-D-galactopyranoside
[CAS# 138182-21-5]

Identification

• Classification: Biochemical >> Carbohydrate >> Monosaccharide
• Name: 6-Chloro-1H-indol-3-yl beta-D-galactopyranoside
• Synonyms: (2S,3R,4S,5R,6R)-2-[(6-chloro-1H-indol-3-yl)oxy]-6-(hydroxymethyl)oxane-3,4,5-triol
• Molecular Formula: C₁₄H₁₆ClNO₆
• Molecular Weight: 329.73
• CAS Registry Number: 138182-21-5
• EC Number: 604-065-1
• SMILES: C1=CC2=C(C=C1Cl)NC=C2O[C@H]3[C@@H]([C@H]([C@H]([C@H](O3)CO)O)O)O

Properties

• Solubility: Slightly soluble (1.6 g/L) (25 °C), Calc.^*
• Density: 1.641±0.06 g/cm³ (20 °C 760 Torr), Calc.^*
• Boiling point: 630.2±55.0 °C 760 mmHg (Calc.)^*
• Flash point: 334.9±31.5 °C (Calc.)^*
• Index of refraction: 1.717 (Calc.)^*

^* Calculated using Advanced Chemistry Development (ACD/Labs) Software.

Safety Data

• Hazard Symbols: GHS07 Warning
• Risk Statements: H302-H315-H319
• Safety Statements: P501-P270-P264-P280-P302+P352-P337+P313-P305+P351+P338-P362+P364-P332+P313-P301+P312+P330

Discovery and Applications

6-Chloro-1H-indol-3-yl β-D-galactopyranoside is a synthetic compound belonging to the class of indolyl glycosides, specifically designed for use in enzymatic assays. The molecule comprises two key structural components: a substituted indole ring and a &betta-D-galactopyranoside moiety. The indole ring is chlorinated at the 6-position, which modifies its electronic and steric properties, while the glycosidic linkage connects this aromatic core to a &betta-D-galactopyranose sugar.

This compound has been developed as a chromogenic substrate for &betta-galactosidase, an enzyme that catalyzes the hydrolysis of &betta-galactosidic bonds. In microbiological and biochemical assays, the presence and activity of &betta-galactosidase can be monitored by incorporating such substrates into culture media or reaction mixtures. When cleaved by the enzyme, the glycosidic bond is broken, releasing the aglycone portion—here, 6-chloroindoxyl—which can undergo further oxidation to form an insoluble, colored product, typically visible as a blue or purple precipitate.

The substitution of a chlorine atom at the 6-position of the indole ring influences both the solubility and the colorimetric response of the hydrolysis product. Chlorine, being an electron-withdrawing group, alters the oxidation potential of the released indole derivative, which may enhance the stability and intensity of the resulting chromophore. This modification can be advantageous in applications requiring strong visual contrast or resistance to background interference in complex biological samples.

In practice, 6-chloro-1H-indol-3-yl &betta-D-galactopyranoside is often used in histochemical staining and microbiological media to detect the activity of &betta-galactosidase-producing organisms. For example, certain strains of Escherichia coli can be identified on plates containing this substrate, where colonies expressing the enzyme produce a distinct color change. This facilitates rapid differentiation and identification of bacterial colonies without the need for additional reagents or complex instrumentation.

The synthesis of this compound involves the glycosylation of 6-chloroindole-3-ol with an activated &betta-D-galactopyranosyl donor, such as a trichloroacetimidate or halide, under conditions that promote stereoselective formation of the &betta-anomeric linkage. Protecting groups are commonly employed to shield reactive hydroxyls on the sugar during the coupling process, and these are subsequently removed to yield the final product. Careful control of reaction conditions is necessary to ensure the desired regiochemistry and avoid decomposition of the sensitive indole nucleus.

Beyond its diagnostic applications, such compounds provide valuable tools in the study of enzyme kinetics and substrate specificity. By modifying the aglycone structure or the sugar moiety, researchers can fine-tune substrate properties to probe different aspects of enzyme function, such as catalytic efficiency, binding affinity, and response to inhibitors.

Overall, 6-chloro-1H-indol-3-yl &betta-D-galactopyranoside exemplifies the rational design of enzyme substrates for use in biological detection and analysis. Its combination of selective enzymatic reactivity and visible signal generation makes it a useful reagent in microbiology, histology, and biochemical research.

References

2012. A fast and sensitive alternative for β-galactosidase detection in mouse embryos. Development (Cambridge, England), 139(23).
DOI: 10.1242/dev.078790

2014. In Situ Hybridization Methods for Mouse Whole Mounts and Tissue Sections with and Without Additional β-Galactosidase Staining. Methods in Molecular Biology, 1092.
DOI: 10.1007/978-1-60327-292-6_1

2016. Comments on Methods to Suppress Endogenous β-Galactosidase Activity in Mouse Tissues Expressing the LacZ Reporter Gene. The Journal of Histochemistry and Cytochemistry, 64(10).
DOI: 10.1369/0022155416665337