Physicochemical screening
Is a critical early-stage process that evaluates the fundamental physical and chemical properties of potential drug candidates. These properties influence a compound’s pharmacokinetics, pharmacodynamics, and overall drug-likeness, helping researchers identify molecules with the best chance of success in later development stages.
Solubility
| Protocol | |
|---|---|
| Buffer and pH | As per client request |
| Incubation concentration | 100 µM |
| Incubation time | 2 h |
| Replicates | n=3 |
| Analysis | LC-MS/MS or HPLC |
| Assay controls | Carvedilol, Diethylstilbestrol |
| Final DMSO concentration | 1-2% v/v |
| Compound requirement | 10 mM DMSO stock 30 µL |
Buffers and pH’s
- Hanks Balanced salt solution (HBSS; pH 6.5 or 7.4)
- Phosphate buffer saline (pH 7.4)
- Phosphate buffer (pH 7.4)
- Fasted State Simulated gastric fluid- FaSSGF (pH 1.6)
- Fed state Simulated intestinal fluid - FeSSIF (pH 5.0)
- Fasted State Simulated intestinal fluid - FaSSIF (pH 6.5)
| Protocol | |
|---|---|
| Incubation time | 24 hours |
| Buffer pH | As per client request |
| Incubation concentration | 1 mg/mL or 500 µM |
| Replicates | n=3 or 2 |
| Analysis | UPLC |
| QC compounds | Carbamazepine and Diclofenac sodium |
| Compound requirement | 50 mM DMSO stock 20 µL or 5 mg (n=3) or 4 mg (n=2) per buffer condition |
Buffers and pH’s
- Phosphate buffer (pH 7.4)
- Fasted State Simulated gastric fluid- FaSSGF (pH 1.6)
- Fed state Simulated intestinal fluid - FeSSIF (pH 5.0)
- Fasted State Simulated intestinal fluid - FaSSIF (pH 6.5)
Partition coefficient
| Protocol | |
|---|---|
| Assay System | Octanol saturated Phosphate Buffer pH 7.4 and phosphate buffer pH 7.4 saturated Octanol |
| Buffer | Phosphate buffer, pH 7.4 |
| Partitioning phases | Buffer saturated with octanol/octanol saturated with buffer (24 h) prior to study |
| Compound concentration | 100 µM |
| Equilibration period | 2 hr |
| Incubation Temperature | Room temperature |
| Control compounds | Caffeine, Warfarin, Chlorpromazine |
| Final DMSO | 1 % |
| Analysis | LC-MS/MS |
| Compound requirement | 10 mM DMSO stock 40 µL |
Octanol: Water (O/W) partition coefficient
- Log D at 7.4 pH
- Log P
| Protocol - UPLC method | |
|---|---|
| Mobile Phase-A | 50 mM Ammonium Acetate |
| Mobile phase pH | pH 6.8 or 7.4 |
| Compound concentration | 0.125 mg/mL |
| Reference compounds | Theophylline, Benzimidazole, Colchicine, 1-3 Dimethyl-8-Phenyxanthine (8-phenyltheophylline), Indole, Acetophenone, Butyrophenone and Valerophenone |
| Analysis | UPLC |
| Compound requirement | 10 mM DMSO stock 40 µL |
Chemical stability
| Protocol | |
|---|---|
| Assay Controls | Chlorambucil, Erythromycin |
| Incubation time | up to 24hr at room temperature or 37±1 °C |
| Buffer pH | As per client request |
| Time points | 0, 1, 2, 4, 6 and 24 hr at room temperature or 37±1 °C |
| Test compound Incubation concentration | 10 µM or any client recommended concentration |
| Replicates | n=2 |
| Analysis | LC-MS/MS |
| Final DMSO concentration | ≤1% |
| Test compound requirement | ~1 mg of powder compound or 10 µL of 10 mM DMSO stock per pH |
Buffers and pH’s
- Phosphate buffer – (1.2, 2.0, 3.0, 4.0, 5.0, 6.0, 6.8, 7.0, 7.4, 8.0)
- Fasted State Simulated gastric fluid- FaSSGF (pH 1.6)
- Fed state Simulated intestinal fluid - FeSSIF (pH 5.0)
- Fasted State Simulated intestinal fluid - FaSSIF (pH 6.5)
- Phosphate buffer saline (pH 7.4)
Chemical reactivity
| Protocol | |
|---|---|
| Buffer pH | 100 mM potassium phosphate buffer, pH 7.4 |
| GSH Concentration | 5.5 mM |
| Preincubation time | 15 min (To ensure all reagents reaches 37±1 °C, before assay initiation) |
| Incubation time | 60 min @ 37±1 °C |
| Time points | 0, 15, 30, 60 min (+GSH / NAC) and 60 min (-GSH / NAC) |
| Test compound Conc. | 10 µM |
| Replicates | n=2 |
| Analysis | LC-MS/MS: Parent degradation or Adduct formation |
| Final DMSO concentration | ≤1% |
| Assay Controls | Afatinib and Ibrutinib |
| Test compound requirement | ~1 mg of powder compound or 10 µL of 10 mM DMSO stock |
Glutathione (GSH)
- Adduct formation
- Parent depletion
N-Acetyl Cysteine (NAC)
- Parent depletion
