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LogD (Micro Shake-Flask)

Lipophilicity (distribution coefficient) is an important physicochemical property of a drug as it provides an understanding of how well a drug will cross lipid membranes (and thereby penetrate tissues), how well a drug will interact with a target protein, as well as help define a drug’s pharmacokinetics and pharmacodynamics profile.

The micro shake-flask LogD7.4 assay is the gold standard assay and measures the lipophilicity of a test compound by assessing the ratio of the equilibrium concentrations of a dissolved test compound in a two-phase system consisting of an organic n-octanol phase and an aqueous buffer phase. The principal of the method is based on the Nernst partition law.

Sygnature’s micro shake-flask LogD₇.₄ assay is based on the OECD guidelines that were adopted by the council in 1995, and uses a constant temperature, a volume ratio of n‑octanol/PBS of 1:1 or 1:10 that has been pre-saturated, and centrifugation to separate the two phases after incubation (to allow distribution of the test compound between the two phases and equilibrium to be reached). The aqueous and/or Octanol phase is then sampled and quantified by LC-MS/MS. A calibration curve is generated for each test compound. Verapamil is used as a positive control.


Compound requirements 10mM DMSO, 60µL
Buffer and pH Phosphate buffered saline 7.4
Partition solvent n-octanol
Ratio of buffer : Octanol 10:1 (v:v) or 1:1 (v:v) PBS/Oct; other ratios available on request
Final DMSO % 2 %
Replicates 6
Incubation Time 600 rpm for 120 min at room temperature
Analysis Method UPLC-MS/MS
Controls Verapamil (positive control)
Data Delivery LogD₇.₄


Figure 1 Comparison of Log D7.4 values generated in Sygnature’s Log D7.4 shake flask assay with literature LogD7.4 values

Figure 2 Inter-day reproducibility of LogDpH7.4 for a single test compound. The error bars represent the standard deviation of a range of values recorded in each month.

Figure 3 Comparison of LogD7.4 values generated using Sygnature’s LogD7.4 micro shake-flask assay – automated versus manual. The error bars represent the standard deviation of six replicates.

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