Enzyme Substrates

6-Chloro-3-indoxyl-beta-D-galactopyranoside is a chromogenic substrate for beta-galactosidase that yields a salmon or rose-coloured precipitate and hence is also known as Salmon-Gal. It can be used in conjunction with isopropyl-beta-D-thiogalactopyranoside (IPTG) as an alternative to the chromogenic substrate 5-bromo-4-chloro-3-indolyl beta-D-galactopyranoside (X-Gal) for detection of beta-galactosidase activity in bacterial colonies in colorimetric assays. Where beta-galactosidase expressed from a reporter gene in molecular biology experiments, recombinant colonies on growth plates are salmon-coloured while non-recombinants appear as white. Use of nitroblue tetrazolium salt (NBT) with Salmon-Gal may provide faster results, yielding a clear, purple precipitate, than for Salmon-Gal alone. In conjunction with 5-bromo-4-chloro-3-indolyl-beta-D-glucoside (X-Glc), Salmon-Gal is useful for simultaneous detection of GUS and Lac activities in the same medium.

AquaSpark® H2S/-SH is a chemiluminescent probe for the detection of sulfides and thiols. The chemiluminescence obtained through sulfide is 3-5 fold higher than chemiluminescence obtained through thiols. AquaSpark® H2S/-SH probe was used in a study identifying antibiotic-resistant bacterial strains by detecting the beta-lactamase-driven hydrolysis of beta-lactam antibiotics, where hydrogen sulfide is a biodegradation metabolite (Popat Gholap et al, 2021).

Glutaryl-glycyl-L-arginine 7-amido-4-methylcoumarin hydrochloride is a synthetic peptide substrate used as a fluorogenic substrate for aminopeptidase.

4-Nitrophenyl O-(N-acetyl-a-neuraminosyl)-(2-3)-b-D-galactopyranosyl-(1-4)-2-acetamido-2-deoxybDglucopyranoside is a fluorescent compound that has been used as a substrate in enzyme assays. It has been shown to be an effective ligand for the enzyme beta galactosidase, which hydrolyzes the substrate to produce 4nitrophenol and o-(Nacetyl)a-neuraminic acid. This product is CAS No. 501427-92-5 and can be used as a high purity, high quality reagent for conjugation.

4-Methylumbelliferyl-alpha-L-arabinofuranoside is a nucleic acid molecule that has an average diameter of 10 nm. It is used as a tracer in electron microscopy to detect the presence of nucleic acid molecules in cells. This compound is also used to study the interactions between DNA and RNA molecules by labeling the DNA strand with 4-methylumbelliferyl-alpha-L-arabinofuranoside, which contains an alpha-(2,4,6) arabinofuranosyl group. The recombinant expression of this compound can be carried out in corynebacterium glutamicum, and it has a bulk density that ranges from 1.7 to 2.1 g/cm3 and an average particle diameter ranging from 0.5 to 1 micron. It is not soluble in water or alcohol but is soluble in acetone or chloroform.

Applications A novel fluorigenic substrate for the determination of Biotinidase activity. References Kumugi, S., et al.: Chemistry Letters, 391 (1997)

Fluorogenic substrate for esterase yielding a blue fluorescent solution upon cleavage.

This Clips-O substrate rac-7-(3',4'-Dihydroxybutyloxy)coumarin-4'-phosphate allows the detection of single and dual enzymatic activity (phosphatase and oxidation of 4-amino-3-hydroxybutyloxy spacer either enzymatically or by exposure to sodium periodate) with a single substrate resulting in a fluorescent signal. The main virtues of Clips-O substrates are an increased stability towards non-enzymatic hydrolysis that results in a reduced background and high sensitivity.

Resorufin phosphate disodium is a fluorogenic substrate for phosphatase exhibiting a red color after cleavage that can be measured at ~ 585nm.

5-Bromo-4-chloro-3-indolyl b-D-lactopyranoside is a chromogenic substrate used to detect the specific enzymatic activity of beta-galactosidase. After cleavage, 5-bromo-4-chloro-indoxyl is released, resulting in a blue to blue-green color change in bacterial colonies or media. 5-Bromo-4-chloro-3-indolyl b-D-lactopyranoside is used in lactose intolerance screening.

2-O-(4-Nitrophenyl)-a-D-N-acetylneuraminic acid, also known as p-nitrophenyl N-acetylneuraminic acid, is a chromogenic substrate specifically designed for the detection and quantification of neuraminidase (sialidase) activity. Upon cleavage by neuraminidase, it releases a yellow-colored product, 4-nitrophenol, which can be easily monitored spectrophotometrically at 405 nm. This substrate is widely used in various applications, including enzyme kinetics studies, inhibitor screening, and the determination of neuraminidase activity in biological samples such as viruses, bacteria, and cell lysates. It is particularly useful for studying the enzymatic properties of neuraminidases from different sources and for the development of novel antiviral and antibacterial agents targeting neuraminidase activity.

2'-(4-Methylumbelliferyl)-a-D-N-glycolylneuraminic acid is a fluorogenic substrate that is used for enzymatic labeling of glycolipids. It can be used as a diagnostic or food testing agent, and also has applications in environmental testing. The CAS number for this product is 1434265-32-3.

4-Nitrophenyl 4,6-ethylidene-?-D-maltoheptaoside is a substrate used in a reference measurement method for ?-amylase activity in the diagnosis of acute pancreatitis, approved by the International Federation of Clinical Chemistry (IFCC). The substrate is hydrolysed by serum ?-amylase into shorter 4-nitrophenylated oligosaccharides and their free counterparts. These reaction products are channeled into an indicator reaction with ?-glucosidase and are degraded into glucose and the chromophore, 4-nitrophenol.

4-Methylumbelliferyl 6-O-(N-acetylneuraminyl)-b-D-glucopyranoside is a fluorescent substrate for the enzyme β-D-galactosidase. It is used in the detection of β-D-galactosidase activity in the diagnosis of various diseases, including celiac disease and AIDS. 4-Methylumbelliferyl 6-O-(N-acetylneuraminyl)-b-D-glucopyranoside can also be used as a chromogenic or fluorogenic substrate for chemiluminescent detection of β-D-galactosidase activity. The conjugate with horseradish peroxidase can be used to detect hydrogen peroxide in water samples, such as drinking water, wastewater, and surface waters. 4MUG has been shown to have high purity and is CAS No. 78761-35-4.

Chromogenic substrate targeting alpha-D-galactosidase