Shake-Flask equilibrium solubility assay²

This method is suitable for both, neutral an ionizable substances. The method can provide the following solubility parameters:

  • Water solubility (Sw)
  • Solubility at fixed pH (SpH)
  • Intrinsic solubility (So)
  • Solubility in a biorelevant media

Appropriate amounts of substance and volume of solvent are mixed and stirred at constant temperature for 24 hours. After 24h of rest, the pH of the aqueous solution is measured and the concentration of substance in aqueous phase is determined by an appropriated analytical method (i.e, HPLC, spectrophotometry, etc.).


  • Movil-RoD from Selecta (Abrera, Spain) rotational stirrer.
  • Ross Combination electrode Orion 8102 from Thermo Fisher Scientific (Waltham, MA,USA) in a Crison micro-pH 2002 potentiometer with a precision of ±0.1 mV (±0.002 pH units) from Crison Instruments S.A. (Alella, Spain).
  • Rotanta 460RS centrifuge (Andreas Hettich GmbH & Co. KG, Germany)
  • Instrumentation for quantification: HPLC, UHPLC, UV-vis, etc.


Required amount of substance:

It depends on the compound solubility and solvent volume to be used. Tentative values for a unique measurement involving 3 mL of solvent are given in the following Table (being S the solubility in working solution).

S (mol/L) mols/3 mL
log S < -5 0.12
-5<log S < -4 0.15 – 1.5
-4<log S < -3 1.5 – 15
-3<log S < -2 15 – 125
-2<log S < -1 125 – 1200
log S > -1 > 1500

² EPA, 1998. EPA Product Properties Test Guidelines, OPPTS 830.7840, Water Solubility: Column Elution Method; Shake Flask Method.

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