In Vitro ADME Services
Characterization of a chemical series or drug candidate's ADME properties and its potential for drug interactions is best when determined early. This helps to de-risk candidate molecules and improve project productivity through more targeted chemical synthesis and progression of the right compounds
2020 FDA In Vitro Drug Interaction Study Guidance

The guidance focuses on in vitro approaches to evaluate the interaction potential between investigational drugs with cytochrome P450 enzymes (CYPs) and transporters and how the results can inform future clinical DDI studies.
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In Vitro ADME Assays
Many of our in vitro ADME assays have multiple formats available to suit the different stages of the drug discovery and development pipeline, providing a flexible approach to ADME screening. Choosing from a range of formats, we can select the assays that best meet your throughput, data resolution, and quality needs and achieve your project specific goals.
Below is a list of our in vitro ADME screening assays.
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Physicochemical screening
- Kinetic solubility
- Thermodynamic solubility
- Log D7.4
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CYP450 Inhibition
- Human liver enzymes or recombinant enzymes
- Major CYP450 isozymes: 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 3A4 and 2E1
- Reversible, metabolic and time-dependent inhibition (IC50 shift)
- Initial screening assessments (% inhibition) or definitive determinations (IC50, Ki)
- Kinact/KI determinations
- Determine formation of MIC and/or covalent binding
- Activity of CYP450 enzymes measured via LC-MS/MS analysis
- Drug-drug interaction experiments with known co-medications
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CYP450 Reaction Phenotyping
- Human liver microsomes or recombinant enzymes
- CYP450-selective chemical inhibitors to determine which enzyme(s) are involved in metabolism
- Metabolite-monitoring option
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CYP450 Induction
- Multiple human CYP450 enzymes: 1A2, 2B6, 2C9, 2C19 and 3A4
- Enzyme catalytic activity or mRNA levels using human cryopreserved hepatocytes
- Catalytic activity of CYP450 isoforms measured via LC-MS/MS analysis
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Protein Binding – Rapid Equilibrium Dialysis (RED), Ultrafiltration or Ultracentrifugation (UC)
- Plasma, serum, whole blood, microsomes, tissue homogenates or purified proteins (albumin, AGP)
- Human, rodent and nonrodent species
- Initial screening assessments or definitive binding determinations
- Ex vivo samples (plasma, tissue homogenates)
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Melanin Binding
- Separate free compound from melanin-bound by centrifugation
- Synthetic melanin
- Also available from sepia (cuttlefish)
- Initial screening assessments (% bound) or definitive binding determinations (kd, Bmax)
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Metabolic Stability – Microsomes, S9
- Liver microsomes, S9
- Non-hepatic microsomes available
- Human, rodent and nonrodent species
- Range of formats: % remaining or intrinsic clearance (T1/2, CLint)
- Metabolite-monitoring option
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Metabolic Stability – Hepatocytes
- Pooled cryopreserved hepatocytes
- Fresh primary hepatocytes available
- Human, rodent and nonrodent species
- Range of formats: % remaining or intrinsic clearance (T1/2, CLint)
- Metabolic monitoring option
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Metabolite ID/Profiling for in vitro ADME screening
- Microsomes, S9, hepatocytes, ex vivo samples, plant or sediment material
- Human, rodent and nonrodent species
- Initial screening assessments (profiling) or more definitive determinations (ID) using accurate mass
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Matrix Stability (Matrix, Buffer)
- Ex vivo matrices (plasma, serum, blood, tissue, homogenates, CSF), aqueous buffers, formulations
- Range of formats: % remaining pr T1/2 determinations
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Red Blood Cell (RBC) Partitioning (Blood/Plasma Ratio)
- Fresh whole blood
- Human, rodent and nonrodent species
- Blood/plasma ratio, Kp
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Permeability and Efflux (Caco-2, MDCK, MDCK-MDRI, MDCK-BCRP)
- Caco-2 (human epithelial colorectal adenocarcinoma cell line), MDCK (wild type, canine), MDCK-MDRI (transfected with human Pgp) and MDCK-BCRP (transfected with human breast cancer resistance protein)
- Cell monolayers available every week
- Uni-directional (A-B flux) or bi-directional (efflux)
- Measurement of analyte levels on apical and basolateral sides of cell monolayer using LC-MS/MS bioanalysis
- Caco-2 (human epithelial colorectal adenocarcinoma cell line), MDCK (wild type, canine), MDCK-MDRI (transfected with human Pgp) and MDCK-BCRP (transfected with human breast cancer resistance protein)
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Influx Transporters
- TransportoCells (HEK 293) from Corning over expressing: OATP1B1, OATP1B3, OAT1, OAT3, OCT1 or OCT2
- Substrate and inhibition assay formats
- Initial screening assessments or definitive determinations
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hERG Inhibition
- hERG Inhibition using the Charles River ChanTest® hERG-HEK stably transfected cell line on the Sophion Qube automated electrophysiology platform
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Cell Proliferation and Cytotoxicity
- Cell Proliferation and Cytotoxicity in HepG2 cells
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In Vitro Genotoxicity Assays Related to In Vitro ADME Assays
- Ames MPF, Ames II, mini Ames
- Bacterial Mutation Test (Ames, OECD 471)
- Mammalian Cell Gene Mutation Test (HPRT Gene, OECD 476)
- Mammalian Chromosome Aberration Test (OECD 473)
- Mammalian Cell Gene Mutation Test (TK Gene, OECD 490)
- Mammalian Cell Micronucleus Assay (OECD 487)
- Fluorescence In Situ Hybridization (FISH)
Delivering Data In Vitro ADME Assays “Even Faster”

Learn how data from in vitro ADME assays are used to triage and prioritize NCEs to enable in vivo PK, PK/PD, efficacy and TK studies.
Frequently Asked Questions (FAQs) for In Vitro ADME Assays
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Why are in vitro ADME assays important?
As compound potency improves during hit-to-lead and lead optimization, in vitro ADME and DMPK assays provide necessary data to establish insight into the key physicochemical properties and structural motifs, ensuring the efficacy and safety of compounds.
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Does Charles River offer services for custom in vitro ADME assays?
Yes, while standard protocols help to maximize throughput and reduce lead times, Charles River can assist in customizing a comprehensive strategy that provides the targeted candidate ADME profile.
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How does Charles River determine physicochemical properties?
Kinetic solubility interprets the performance of compounds within in vitro ADME assays and biology screens while also assessing the effect the structural modifications have upon solubility within chemical series. Experimental logP/logD assessments are also available using miniaturized shake-flask methodology to reinforce in silico predictions.
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What methodologies does Charles River use to determine metabolic stability for in vitro ADME assays?
Charles River uses validated metabolic stability screens to assess a compound’s vulnerability to metabolic instability whether through CYP450-mediated or other types of oxidations, conjugations or other biotransformations. High resolution mass spectrometry is used to identify putative metabolites, elucidate their structures, and characterize the metabolic fate of compounds.