Flash Chromatography Basics

Instructions for Sample Preparation

1. Start with TLC Screening

• Use Thin Layer Chromatography (TLC) to identify a solvent or solvent mixture that produces a good separation. We highly recommend our TLC Flash Method Development Kit TLC-FMDK1
• Aim for an Rf value between 0.13 and 0.40 with an Rf between compounds of at least 0.1.  This range strikes a balance between resolution and elution time.
• Use our Solubility Guide Based on Compound Structure
  below to choose an appropriate solvent system

2. Solubility Pre-Test

• Dissolve a small amount of your compound in the candidate solvents identified with our Sorbtech TLC Method Development Kit.
• Use visual inspection and UV light (if applicable) to check for complete dissolution.
• If solubility is poor, try warming the solution slightly or using co-solvents (e.g., DCM with a few drops of methanol).

3. Dry Loading vs. Liquid Loading

• If your compound is poorly soluble in the mobile phase, consider dry loading:
  • Dissolve in a volatile solvent (e.g., DCM), mix with silica, evaporate, and load as a dry plug.
• For well-soluble compounds, liquid loading is more straightforward and efficient.

4. Choose the Right Stationary Phase

• Most flash chromatography uses normal-phase silica gel.
• For polar or water-soluble compounds, consider reversed-phase media like C18.

5. Optimize the Mobile Phase

• Use the TLC solvent system as a starting point.
• Adjust polarity gradually if using a gradient elution.
• Ensure the mobile phase keeps your compound in solution throughout the run.

• Avoid highly viscous solvents for flash chromatography—they slow down flow rates and reduce resolution.
• Use scouting gradients with small sample amounts to refine your solvent system before full-scale purification.

Carefully record solubility observations to ensure reproducibility and assist in possible troubleshooting.

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Solubility Guide Based on Compound Structure

1. Assess Polarity of your Compound

• Polar compounds (e.g., alcohols, amines, carboxylic acids, peptides):
  • Tend to stick to polar stationary phases like silica.
  • Require more polar solvents (e.g., ethyl acetate, methanol) to move up the TLC plate or elute from a column.
• Non-polar compounds to moderately polar (e.g., hydrocarbons, aromatic rings, halogenated aromatics):
  • Travel easily with non-polar solvents (e.g., hexane, toluene). Compatible with silica-based adsorbents.

2. Functional Groups Matter

Functional Group	Polarity	Suggested Solvent System
Alcohols, Phenols	High	Ethyl acetate, MeOH
Carboxylic acids	Very high	EtOAc + AcOH or MeOH
Amines	High	DCM + MeOH or EtOAc
Ketones, Aldehydes	Moderate	DCM, EtOAc
Aromatics	Low	Hexane, Toluene
Alkanes	Very low	Pure hexane

 

3. Use LogP or Partition Coefficient (if available)

• A high LogP (lipophilic) suggests non-polar behavior → use less polar solvents.
• A low LogP (hydrophilic) suggests polar behavior → use more polar solvents.

 TLC Strategy Based on Structure

1. Start with a 1:1 mixture of hexane:ethyl acetate—a versatile baseline.
2. Adjust based on how far your compound travels:
   • If it barely moves → increase ethyl acetate (more polar).
   • If it races to the top → increase hexane (less polar).
3. For very polar compounds, try DCM:MeOH or EtOAc:MeOH blends.