There are no comments yet.

CMYK

ADME SERVICES

Our In-vitro ADME screening service offers a portfolio of assays for investigating: metabolism, distribution and toxicity. permeability, solubility & physicochemical properties, GVK BIO delivers consistent, accurate compound data with cost-efficiency.

Solubility

Solubility is one of the most important physicochemical properties. We determine equilibrium solubility by dried DMSO method. We can also determine solubility in aqueous buffer (pH 1-9), organic solvents (DMSO, Ethanol, etc), formulations and excipients by pION/Multiscreen/HPLC/UV/MS/MSMS methods.

In-vitro ADME Capabilities

Physicochemical studies · · · ·

Log D/Log P Solubility(Kinetic/Equilibrium) Chemical stability Biological matrix stability (serum/ plasma/ microsomes/ blood /hepatocytes/tissue homogenates)

Protein Binding

In-vitro binding studies with plasma have proven to be a valuable tool for predicting In-vivo protein binding. We determine the protein binding by both ultra filtration and rapid equilibrium dialysis. Using RED device we determine protein binding in microsomes. Plasma and tissue homogenate across various species (Rat/mice/human &dog).

Absorption/Distribution Assays · · · · ·

Caco2 and PAMPA permeability assay Pgp substrate / inhibitor assay Protein binding Blood/Plasma Partitioning Ratio

Caco2 Permeability Assay

Caco2 cells are the most frequently used In-vitro models to assess intestinal permeability. Permeability across Caco2 cell monolayer is used to predict human permeability of drug candidates, to perform in-depth mechanistic and absorption studies, to study the effects of transporters on permeability. We can determine apparent permeability (Papp) / efflux ratio / Unidirectional / Bidirectional by LCMS.

Metabolism/Excretion

Half life/clearance determination using microsomes/Hepatocytes /S9 fractions/Cyp across species (Human/Rat/Mouse/Dog/ monkey) CYP Inhibition (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4) Pathway determination (Phase I and Phase II)

· · · ·

CaCo-2 Cells Semi-permeable membrane

Metabolite identification using Microsomes / Hepatocytes

and Characterization of potential metabolites using microsomes and Hepatocytes across species (Human/Rat/Mouse/Dog/ monkey)

PAMPA

(Parallel artificial membrane permeability assay)

The Parallel Artificial Membrane Permeability Assay (PAMPA) assay is used as an in-vitro model of passive, transcellular permeability. As well as for the prediction of oral absorption and brain penetration. Effective permeability (log Pe) may be measured by pION/Multiscreen/LCMS.

Log D/Log P

Log D is determined by the shake-flask method, by dissolving some of the solute in a volume of octanol and water/buffer and measure the concentration of the solute in each solvent. Log D is determined by HPLC-UV with confirmation by mass.

CMYK

CMYK

ADME SERVICES

Microsomal Stability

Metabolic stability plays an important role in the success of drug candidates. First pass metabolism is one of the major causes of poor oral bioavailability and short half life and the study influences both oral bioavailability and half life. The half life / clearance / % metabolized can be determined in microsomes / S9 fractions / hepatocytes.

Microsomal Stability

CYP2D6 inhibition by Furafyline

1 2 3 4 5 6 7 8 9 10 1112 A B C D E F G H

CYP2D6 inhibition by Qunidine

96 - Well Plate

110 90 70 % Inhibition 50 30 10 0 -10 -8 -7 -6 -5 -4 EC50 3.092e-006 R2 0.9968

Chemicals

Add Hepatocytes or Microsomes

80 70 60 50 40 30 20 10 0 -10

EC50 1.015e-008 R2 0.9900

% Inhibition

-9

-8

-7

-6

-5

Incubate Extract

Log drug Con [M]

Log drug concentration [M]

CYP2D6 inhibition by Sulphaphenazole

CYP3A4 - Ketoconazole

LCMS Analysis

35 30 25 % Inhibition 15 10 5 0 -9 -8 -7 -6 -5 -4

% Inhibition

EC50 6.372e-007 2 R 0.9431

Hepatocyte metabolic stability

Per orally administered drug may undergo first-pass metabolism which influences the pharmacokinetic properties such as the clearance, the half-life or the bioavailability. Cryopreserved hepatocytes are used to calculate half life/clearance/% parent compound remaining.

20

110 100 90 80 70 60 50 40 30 20 10 0 -10 -10

EC50 1.884e-007 R2 0.9979

-9

-8

-7

-6

Log drug Concentrations [M]

Log drug concentration [M]

Human hepatocytes metabolic stability

% Parent compound remaining

100 90 80 70 60 50 40 30 20 10 0

Metabolite identification using Light Sight Software

Metabolic stability and the identification of formed metabolites has become an important tool. In-vitro metabolite identification & characterization of the test compounds is determined using Q trap with light sight software.

Bioanalysis

LC-MS/MS

Qu

Pro

Ve

Ca

Imi

pra

Ate

? 3200 QTRAP ? API3200 ? API4000

HPLC

no lol

ine

ine

mil

C 7-E

min

rap

pro

nid

Compound details

ffe

no

lol

e

CYP Inhibition

CYP inhibition occurs either as reversible inhibition, quasiirreversible inhibition or irreversible inhibition. CYP inhibition is a fluorescent based/LCMS assay. Specific isoforms of CYP (CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4) are used to determine the inhibition (IC50).

? RRLC with PDA Agilent 1200 ? Shimadzu Prominence with UV ? RRLC with HTS PAL Agilent 1200

Others

? (Molecular Devices) Spectramax ? BMG Polarstar ? Handler (Tomtec) Qudra4 Liquid

GVK Biosciences Private Limited

28A, IDA, Nacharam, Hyderabad 500 076, India. T 91 40 66281823 F 91 40 6628 1505 E-mail: bdbio@gvkbio.com Website: www.gvkbio.com

CMYK