3,5-Bis(acetylamino)benzoic Acid: Handling & Use Protocol

3,5-bis(acetylamino)benzoic acid is a bifunctionally substituted aromatic compound bearing two acetylamino groups and a carboxylic acid on a benzene ring, widely employed as a synthetic intermediate and building block in pharmaceutical and materials research.

Chemical Name

3,5-bis(acetylamino)benzoic acid

CAS Number

Please verify with Aladdin product datasheet (B300853)

Molecular Formula

C₁₁H₁₂N₂O₄

Molecular Weight

252.23 g/mol

Appearance

White to off-white crystalline solid (verify with CoA)

Recommended Storage

Store at room temperature in a cool, dry place, away from light

Solubility

Sparingly soluble in water; soluble in DMSO and dilute alkaline solutions

SKU

B300853

Overview and Chemical Properties

3,5-Bis(acetylamino)benzoic acid (SKU: B300853) is a symmetric aromatic compound derived from 3,5-diaminobenzoic acid by N-acetylation of both amine groups. The resulting molecule presents a central benzoic acid scaffold flanked by two acetylamino (acetamido) substituents at the 3- and 5-positions, conferring a high degree of molecular symmetry.

The carboxylic acid group (–COOH) provides a versatile handle for further functionalization, including esterification, amide bond formation, and coupling reactions, making this compound a valuable building block in medicinal chemistry, polymer science, and supramolecular chemistry. The acetylamino groups can participate in hydrogen bonding, influencing crystal packing, solubility, and intermolecular interactions in designed materials.

Key physicochemical parameters (verify all values against the current Aladdin Certificate of Analysis for lot-specific data):

• Molecular formula: C₁₁H₁₂N₂O₄
• Molecular weight: 252.23 g/mol
• Melting point: confirm with CoA
• pKa (COOH): estimated ~3–4 (typical aryl carboxylic acid; verify experimentally)

Safety, Handling, and Storage

Always consult the Safety Data Sheet (SDS) for 3,5-bis(acetylamino)benzoic acid (B300853) before use. The following precautions represent general best practice for fine aromatic amide reagents and must be confirmed against the current SDS provided by Aladdin Scientific.

Personal Protective Equipment (PPE)

• Wear appropriate chemical-resistant gloves (e.g., nitrile).
• Use safety glasses or chemical splash goggles.
• Wear a laboratory coat and closed-toe shoes.
• Handle in a well-ventilated area or fume hood when weighing powders to avoid inhalation of dust.

Storage Conditions

• Store in a tightly sealed container at room temperature (15–25 °C), in a cool, dry location away from direct light and moisture.
• Keep away from strong oxidizers, strong acids, and strong bases.
• Container should be clearly labeled with compound name, CAS number, and date received.

Disposal

• Dispose of waste material in accordance with local, regional, and national regulations for chemical waste.
• Do not dispose of down the drain without checking applicable regulations.

Solution Preparation Protocol

The following protocol describes the preparation of stock and working solutions of 3,5-bis(acetylamino)benzoic acid. All concentrations and volumes should be adapted to the specific experimental requirements.

Materials Required

• 3,5-Bis(acetylamino)benzoic acid (Aladdin B300853)
• Analytical balance (±0.1 mg precision)
• Appropriate solvent: DMSO (preferred for stock), dilute NaOH or Na₂CO₃ solution for aqueous applications
• Volumetric flask or calibrated vessel
• 0.22 µm syringe filter (if sterile-filtered solution is required)
• Amber glass vials for storage of light-sensitive solutions

Procedure

1. Calculate required mass: Determine the target concentration and volume. Use the formula: mass (mg) = concentration (mM) × volume (mL) × molecular weight (g/mol) ÷ 1000.
2. Weigh compound: Tare a clean, dry weighing vessel on the analytical balance. Carefully weigh the required amount of 3,5-bis(acetylamino)benzoic acid. Perform weighing in a fume hood to minimize dust exposure.
3. Dissolve in DMSO (for organic stock): Transfer the weighed compound to an appropriate vessel. Add approximately 80% of the target DMSO volume and vortex or sonicate (30–60 seconds) until fully dissolved. Adjust to final volume. Note: warming to 37–40 °C in a water bath may aid dissolution if needed.
4. Dissolve in aqueous base (for aqueous applications): Dissolve the compound in a minimal volume of 0.1 M NaOH or 0.1 M Na₂CO₃, then dilute with the appropriate aqueous buffer to achieve the desired pH and concentration. Adjust pH as required with dilute HCl or NaOH.
5. Filter if required: Pass the solution through a 0.22 µm syringe filter for sterile applications.
6. Aliquot and store: Distribute into single-use aliquots in amber vials. Store at –20 °C for long-term stability of DMSO stock solutions; use fresh aqueous solutions or store at 4 °C short-term. Avoid repeated freeze-thaw cycles.

Note: Solubility data should be verified experimentally for each specific solvent system and concentration used in your application.

Applications in Research

3,5-Bis(acetylamino)benzoic acid finds application across multiple research domains due to its unique combination of a reactive carboxylic acid and two hydrogen-bond-capable acetylamino groups on a symmetric benzene ring.

Synthetic Chemistry and Medicinal Chemistry

The compound serves as a versatile building block for the synthesis of more complex molecules. The carboxylic acid can be activated and coupled with amines to form amide bonds, or converted to esters, acyl halides, or anhydrides. The acetylamino groups provide sites for hydrogen bonding in designed drug-like molecules and can be hydrolyzed to free amines under acidic or basic conditions if further functionalization is required.

Polymer and Materials Science

Symmetric bifunctional aromatic monomers bearing both carboxylic acid and amide groups are used in the design of polyamides, metal-organic frameworks (MOFs), and supramolecular gels. The hydrogen-bonding capacity of the acetylamido groups enables self-assembly into ordered nanostructures.

Analytical Reference Standards

High-purity lots of 3,5-bis(acetylamino)benzoic acid may be used as reference standards or internal standards in HPLC, LC-MS, or other analytical methods where a chemically stable, UV-absorbing aromatic compound is needed.

Researchers are encouraged to consult the primary literature for specific application protocols and to verify suitability for their experimental systems.

Quality Control and Troubleshooting

To ensure experimental reproducibility and data integrity, implement the following quality control steps when working with 3,5-bis(acetylamino)benzoic acid (B300853).

Verification of Compound Identity and Purity

• Review the Certificate of Analysis (CoA) supplied with each lot for purity (typically assessed by HPLC), melting point, and spectroscopic data (NMR, MS).
• For critical applications, independently verify identity by ¹H NMR and LC-MS prior to use.
• Check for lot-to-lot consistency when reordering.

Dissolution Troubleshooting

• Poor dissolution in DMSO: Warm gently to 37–40 °C and sonicate. If still insoluble, consider reducing the target concentration.
• Precipitation in aqueous buffer: Ensure adequate pH (alkaline conditions improve solubility of the carboxylate form). Pre-dissolve in DMSO at high concentration, then dilute into buffer; keep final DMSO content ≤1% v/v for most biological assays.
• Turbid solutions: Filter through 0.22 µm membrane and re-check concentration by UV absorbance if possible.

Stability Considerations

• The acetylamino groups are susceptible to hydrolysis under prolonged exposure to strongly acidic or strongly basic aqueous conditions; use freshly prepared solutions where possible.
• Protect from prolonged UV exposure; store stock solutions in amber vials.
• For long-term storage, keep the neat solid desiccated and sealed.

Frequently asked questions

What solvents are suitable for dissolving 3,5-bis(acetylamino)benzoic acid?

3,5-Bis(acetylamino)benzoic acid is sparingly soluble in water but dissolves in DMSO and in dilute alkaline aqueous solutions (e.g., dilute NaOH or Na₂CO₃). For biological assays requiring aqueous conditions, prepare a concentrated DMSO stock and dilute into the appropriate buffer, keeping the final DMSO content at or below 1% v/v. Exact solubility should be verified experimentally.

How should 3,5-bis(acetylamino)benzoic acid be stored to ensure long-term stability?

The neat solid should be stored in a tightly sealed, desiccated container at room temperature (15–25 °C), away from light, moisture, and incompatible materials such as strong oxidizers and strong acids or bases. DMSO stock solutions are best stored at –20 °C in amber vials and protected from repeated freeze-thaw cycles. Aqueous solutions should be prepared fresh or stored at 4 °C for short-term use only.

What are the main research applications of 3,5-bis(acetylamino)benzoic acid?

This compound is primarily used as a synthetic building block in medicinal chemistry and materials science, taking advantage of its reactive carboxylic acid for amide or ester bond formation and its acetylamino groups for hydrogen-bond-mediated self-assembly. It is also applied in the design of polyamides and supramolecular materials, and high-purity lots may serve as analytical reference standards in HPLC and LC-MS methods.

How do I calculate the mass needed to prepare a specific molar concentration solution?

Use the formula: mass (mg) = concentration (mM) × volume (mL) × molecular weight (252.23 g/mol) ÷ 1000. For example, to prepare 10 mL of a 10 mM solution, you need 10 × 10 × 252.23 ÷ 1000 = 25.22 mg. Always verify the molecular weight against the lot-specific Certificate of Analysis.

Can 3,5-bis(acetylamino)benzoic acid be hydrolyzed to give the free diamine?

Yes, the acetylamino groups in 3,5-bis(acetylamino)benzoic acid can be hydrolyzed to the corresponding free amino groups (giving 3,5-diaminobenzoic acid) under prolonged treatment with strongly acidic (e.g., concentrated HCl) or strongly basic (e.g., concentrated NaOH) aqueous conditions at elevated temperatures. This reactivity should be considered when selecting reaction or storage conditions.

References

1. Aladdin Scientific. Certificate of Analysis and Safety Data Sheet for 3,5-bis(acetylamino)benzoic acid, SKU B300853. Aladdin Scientific, Shanghai.
2. March, J. Advanced Organic Chemistry: Reactions, Mechanisms, and Structure. 4th ed. Wiley-Interscience, 1992. (General reference for amide bond formation and carboxylic acid chemistry.)
3. Clayden, J.; Greeves, N.; Warren, S. Organic Chemistry. 2nd ed. Oxford University Press, 2012. (General reference for acetamide hydrolysis and aromatic chemistry.)

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.