Based on your experimental protocol, the mixture of "50% PEG300 + 50% normal saline" can achieve a suspension state at a concentration of 5 mg/mL, but sedimentation occurs after being left to stand for 10 minutes.

The "corn oil" can also form a suspension at a concentration of 5 mg/mL, yet sedimentation takes place after being left undisturbed for 10 minutes.

Both of these formulations require fresh preparation for immediate use and cannot be stored. During experiments, proper solvent controls need to be set up.

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• Biochanin A is poorly water-soluble (an isoflavone). True solutions usually require a co-solvent, surfactant, or complexation (cyclodextrin).
• Vehicle toxicity and maximum IP volume are species-dependent. Always check your institution’s IACUC / veterinary guidance before dosing. If you tell me the species later I can tailor volumes and % limits precisely; for now I will present generally accepted conservative limits (keep DMSO low, avoid high Cremophor unless necessary, prefer cyclodextrin when possible).
• Use sterile technique for IP injections: prepare under aseptic conditions, sterile filter (0.22 µm) whenever compatible, and inspect visually before dosing.

Option A — HP-β-cyclodextrin (preferred when available)

Cyclodextrins (hydroxypropyl-β-cyclodextrin, HP-β-CD) often solubilize flavonoids well and are generally well tolerated IP at moderate concentrations.

Example: 20% w/v HP-β-CD vehicle (often used in preclinical work)

• Target concentration: 5 mg/mL Biochanin A.

Recipe — 10 mL final

• HP-β-CD: 2.0 g (20% w/v)
• Biochanin A: 50 mg (5 mg/mL × 10 mL)
• Purified water (or sterile saline) q.s. to 10 mL

Prep:

Pros: often gives a clear solution, better tolerated than high DMSO or Cremophor. Cons: requires access to HP-β-CD and confirm tolerability for your species and dose.

Option B — Low-DMSO / PEG cosolvent (fast, commonly used)

Use a co-solvent blend to obtain a true solution. Keep DMSO ≤ 10% v/v in the final vehicle if possible (many labs use ≤10% for IP; lower is safer). A common blend: 10% DMSO : 40% PEG-400 : 50% saline (v/v).

Recipe — 10 mL final

• Biochanin A: 50 mg
• DMSO (100%): 1.0 mL (10% v/v)
• PEG-400: 4.0 mL (40% v/v)
• 0.9% saline: 5.0 mL to make up 10 mL

Prep:

Pros: quick, commonly effective for many hydrophobic small molecules; yields a true solution. Cons: DMSO/PEG can cause abdominal irritation/diarrhea in some species and should be kept low. Verify species tolerability and consider pilot vehicle-only controls.

Option C — Solutol/Cremophor® EL + saline or ethanol/PEG

Cremophor EL or Solutol HS15 can solubilize lipophilic drugs at low % (e.g., 5–10%). Use cautiously — Cremophor can cause hypersensitivity in some species.

Example vehicle: 5% Cremophor EL : 45% PEG-400 : 50% saline — similar prep steps to Option B but substitute Cremophor for DMSO. Sterile filter if possible.

Pros: good solubilization. Cons: possible hypersensitivity / histamine-like reactions; avoid if you lack experience or species sensitivity is unknown.

Option D — Suspension for IP (if solution impossible)

If you cannot make a clear solution, a well-prepared nanosuspension or finely micronized suspension in a viscous vehicle (0.5% CMC with a small % Tween-80) may be used, but solutions are preferred for IP because suspensions can irritate the peritoneum and give variable absorption.

If you must use a suspension:

• Micronize drug to
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