Orobol: Properties, Mechanisms & Lab Handling Guidelines

Orobol is a naturally occurring 3′,4′,5,7-tetrahydroxyisoflavone with documented biological activities spanning kinase inhibition, antioxidant defense, and modulation of cell signaling pathways, making it a valuable tool compound for pharmacological and biochemical research.

Chemical Class

Isoflavone (polyhydroxylated)

SKU / Catalog Number

O650798

Key Biological Activities

Kinase inhibition, antioxidant activity, modulation of apoptotic pathways

Recommended Storage

Store at -20 °C, protected from light and moisture; verify with CoA

Solubility

Soluble in DMSO; limited aqueous solubility; verify concentration with UV absorbance

Molecular Formula

C15H10O6 (MW ≈ 286.24 g/mol)

Chemical Identity and Physicochemical Properties

Orobol (SKU O650798) is a naturally occurring isoflavone belonging to the polyhydroxylated flavonoid family. Its systematic structure, 3′,4′,5,7-tetrahydroxyisoflavone, features four hydroxyl groups that confer both its biological reactivity and its moderate polarity profile.

Molecular Formula: C₁₅H₁₀O₆; Molecular Weight: approximately 286.24 g/mol. Researchers should confirm exact MW and purity from the Certificate of Analysis (CoA) supplied with each lot.

Solubility: Orobol exhibits good solubility in dimethyl sulfoxide (DMSO) and is sparingly soluble in water. For aqueous assay systems, prepare a concentrated stock in DMSO (typically 10–50 mM) and dilute into buffer or cell culture medium immediately before use, keeping the final DMSO concentration ≤0.1% (v/v) to avoid cytotoxic artefacts. Warm the DMSO stock briefly to 37 °C and vortex if particulates are observed.

Appearance: Typically a pale yellow to yellow solid or powder. Discoloration or browning may indicate oxidative degradation; discard affected material.

Biological Mechanisms and Research Applications

Orobol has been investigated for multiple biological activities that make it a versatile tool compound in cell-signaling and pharmacological research:

• Kinase Inhibition: Orobol has been reported to inhibit certain protein tyrosine kinases and other kinase targets relevant to proliferative and inflammatory signaling. Researchers exploring kinase-pathway crosstalk may use Orobol as a reference inhibitor or comparative tool compound. Specific IC₅₀ values vary by assay format and target; consult primary literature and flag for citation verification.
• Antioxidant Activity: The multiple phenolic hydroxyl groups of Orobol enable radical-scavenging activity through hydrogen-atom or electron transfer mechanisms, relevant to oxidative stress models in vitro.
• Modulation of Apoptosis: Published studies indicate Orobol can influence apoptotic pathways, including modulation of caspase activation and Bcl-2 family protein expression, though mechanistic details depend on cell type and concentration.
• Estrogenic/Anti-estrogenic Activity: As a phytoestrogenic isoflavone, Orobol may interact with estrogen receptors; researchers working in hormone-sensitive models should account for this activity.

Typical in vitro working concentrations range from low micromolar to mid-micromolar; optimize for each cell line and endpoint. Include appropriate vehicle controls (DMSO at matched concentration) in all experiments.

Storage, Stability, and Handling Guidelines

Proper storage and handling of Orobol are essential to maintain compound integrity and ensure reproducible experimental results.

Storage Conditions

• Store at −20 °C in a tightly sealed container, protected from light and moisture.
• Aliquot stock solutions into single-use volumes to minimize freeze–thaw cycles; repeated freeze–thaw degrades phenolic compounds.
• DMSO stock solutions may be stored at −20 °C for short-to-medium term (weeks to a few months); confirm stability with lot-specific CoA data.

Handling Precautions

• Handle as a general research chemical: wear appropriate PPE including lab coat, nitrile gloves, and eye protection.
• Weigh powders in a ventilated enclosure to minimize inhalation of fine particulates.
• Orobol is not classified as a controlled substance; however, as with all research reagents, follow institutional safety guidelines and SDS recommendations.

Quality Verification

• Verify purity by HPLC and identity by NMR or HRMS as documented in the supplied CoA before commencing experiments.
• If solubility appears compromised, perform a solubility test at the intended working concentration before use in biological assays.

Experimental Protocol Recommendations

The following protocol guidance is intended to assist researchers in designing robust in vitro experiments with Orobol (O650798).

Stock Solution Preparation

1. Allow the vial to equilibrate to room temperature before opening to prevent condensation on the powder.
2. Dissolve Orobol in anhydrous DMSO to prepare a 10–50 mM stock solution. Vortex and, if necessary, sonicate briefly in a water bath sonicator.
3. Confirm solubilization by visual inspection; a clear, homogeneous solution should result. Verify concentration spectrophotometrically if the molar extinction coefficient is known for your solvent system.
4. Aliquot into low-binding microtubes and store at −20 °C under inert atmosphere (argon or nitrogen) if extended storage is required.

Working Dilution

1. Prepare serial dilutions in complete cell culture medium or assay buffer immediately before use.
2. Ensure the final DMSO concentration does not exceed 0.1% (v/v) to avoid confounding cytotoxic effects.
3. Always include: (a) a vehicle control (DMSO at the maximum concentration used), (b) a positive control (validated inhibitor or antioxidant), and (c) untreated cells.

Assay Considerations

• Orobol's UV absorbance may interfere with colorimetric assays (e.g., MTT, MTS) at high concentrations; subtract compound-only blanks or use alternative viability assays (e.g., CellTiter-Fluor).
• In antioxidant assays (DPPH, ABTS), ensure compound dilutions remain within the linear range of the assay.
• For kinase inhibition assays, pre-incubate Orobol with the enzyme for 15–30 minutes at room temperature before substrate addition, per standard kinase assay practice.

Frequently asked questions

What is the recommended solvent for preparing Orobol stock solutions?

DMSO is the recommended solvent for Orobol stock solutions, typically yielding clear solutions at 10–50 mM. Aqueous solubility is limited; dilute DMSO stocks into aqueous buffers or media immediately before use, keeping final DMSO ≤0.1% (v/v).

How should Orobol be stored to maintain stability?

Store Orobol as a dry powder or aliquoted DMSO solution at −20 °C in a sealed, light-protected container. Minimize freeze–thaw cycles by working with single-use aliquots. Always confirm lot-specific stability data on the Certificate of Analysis.

Can Orobol interfere with common colorimetric cell viability assays?

Yes, Orobol's phenolic chromophore can absorb light in wavelength ranges overlapping with MTT or MTS formazan read-out windows at higher compound concentrations. Subtract compound-only (no-cell) blanks at each concentration tested, or use fluorescence-based viability assays to minimize this artefact.

What biological targets have been studied for Orobol?

Orobol has been investigated as an inhibitor of protein tyrosine kinases and has demonstrated antioxidant, pro-apoptotic, and potential estrogenic/anti-estrogenic activities in vitro. Specific targets and IC50 values should be verified against current peer-reviewed literature, as data can vary by assay format.

Is Orobol suitable for in vivo studies?

Orobol (O650798) is supplied as a research-grade reagent intended primarily for in vitro laboratory use. For in vivo applications, researchers must independently validate appropriate formulation strategies, dosing, pharmacokinetics, and institutional ethical approvals. Consult current literature for reported in vivo data.

References

1. Sigma-Aldrich / MilliporeSigma. Orobol product information and safety data sheet. General physicochemical data for isoflavones.
2. Cushman M, Nagarathnam D, Burg DL, Geahlen RL. Synthesis and protein-tyrosine kinase inhibitory activities of flavonoid analogues. J Med Chem. 1991. [Reported involvement of polyhydroxyisoflavones in kinase inhibition — specific data need verification.]
3. Harborne JB, Williams CA. Advances in flavonoid research since 1992. Phytochemistry. 2000;55(6):481-504. [General isoflavone chemistry reference — Orobol-specific data to be verified.]
4. MedChemExpress (MCE). Orobol, HY-N3127 product datasheet. [Referenced in product description; specific biological activity data require primary literature verification.]

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