Apollo Sci. 2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride

Apollo Scientific 2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride, often abbreviated as INT (Iodonitrotetrazolium chloride), is a high-purity chemical reagent extensively employed as a sensitive redox indicator. It is commonly utilized in biochemical, microbiological, and cell biology research applications to assess metabolic activity and cellular respiration.

Chemical Information

• Chemical Name: 2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride
• Common Abbreviation: INT chloride
• CAS Number: 146-68-9
• Molecular Formula: C19H13ClIN5O2
• Molecular Weight: 505.70 g/mol
• Appearance: Pale yellow to orange crystalline powder
• Solubility: Soluble in water and aqueous buffers; soluble in ethanol and methanol.

Mechanism of Action

INT chloride acts as an electron acceptor that undergoes enzymatic reduction by active respiratory enzymes in metabolically active cells (e.g., bacterial, fungal, yeast, or mammalian cells). Upon reduction, it forms an intensely colored, water-insoluble purple to red-colored formazan precipitate. The amount of formazan produced directly correlates with the metabolic or respiratory activity present, enabling quantitative and qualitative assessment of viability and enzymatic activity.

Key Applications and Use Cases

1. Microbial Viability and Respiration Assays

Primary Use:

• Determining microbial viability and respiratory metabolism in bacterial, fungal, and yeast cultures.

Practical Example:

• Researchers evaluating antimicrobial effectiveness incubate bacterial cultures with INT chloride. Viable bacteria reduce INT chloride, producing a clearly visible formazan precipitate. Reduced INT indicates actively metabolizing organisms, whereas the absence of color development implies microbial death or inhibited metabolic function.

2. Cell Viability and Cytotoxicity Studies

Primary Use:

• Assessment of metabolic activity and cell viability in mammalian cell lines during drug screening, cytotoxicity testing, and cell culture optimization.

Practical Example:

• Pharmaceutical researchers testing novel anticancer compounds incubate cultured mammalian cells (e.g., HeLa, MCF-7, NIH/3T3) with INT chloride to rapidly measure metabolic activity post-treatment. Reduced INT to formazan correlates directly with cellular viability, facilitating rapid and reliable screening of therapeutic agents.

3. Enzyme Activity Assays and Biochemical Research

Primary Use:

• Quantitative detection and assessment of oxidoreductase enzyme activity, particularly dehydrogenases in various biological samples.

Practical Example:

• Researchers conducting metabolic pathway analyses measure mitochondrial or cytoplasmic dehydrogenase activities. In the presence of INT chloride, active enzymes produce formazan, allowing spectrophotometric quantification and facilitating comparative enzymatic studies.

4. Environmental and Soil Microbiology

Primary Use:

• Monitoring microbial respiration and activity in environmental samples, including soil, water, or wastewater samples, for ecological assessments.

Practical Example:

• Environmental microbiologists evaluate soil microbial health by adding INT chloride to soil samples. Active microbes reduce INT to formazan, visibly indicating microbial activity and enabling assessments of ecological impacts, pollution, or bioremediation efficiency.

5. Biofilm Detection and Antibiofilm Assays

Primary Use:

• Detection and quantification of biofilm formation, growth, and metabolic activity of microorganisms on various surfaces and materials.

Practical Example:

• Medical device researchers test surface materials for biofilm resistance by incubating microorganisms on samples with INT chloride. Formation of colored formazan precipitates reveals metabolically active biofilm formation, guiding materials selection for biomedical applications.

Advantages and Benefits

• Highly Sensitive Detection: Clear visual or quantitative measurement of cell viability and enzymatic activity.
• Versatility: Suitable for diverse organisms and experimental conditions.
• Rapid Results: Provides rapid, reliable metabolic activity data, ideal for high-throughput screening.
• Ease of Use: Simple protocol compatible with standard laboratory equipment (spectrophotometers, microplate readers).

Storage and Stability

• Store in a cool, dry environment (recommended storage: 2–8°C).
• Protect from light and moisture to maintain reagent stability.
• Handle with appropriate laboratory practices.

Intended Users

• Research laboratories (microbiology, cell biology, biochemistry)
• Pharmaceutical and biotechnology companies
• Environmental microbiology facilities
• Universities and academic institutions conducting biochemical research
• Biomedical device developers conducting antibiofilm assessments

Apollo Scientific’s 2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride is a versatile and indispensable reagent, providing researchers precise insights into metabolic functions and microbial activity across a wide range of research disciplines.