Agomelatine metabolite

A novel sensitive and selective LC-MS/MS method for the determination of agomelatine, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma was developed and validated. After simple protein precipitation, the analytes were separated on a Phenomenex ODS3 column (4.6×150 mm, 5μm, Phenomenex, USA) with mobile phase consisted of methanol and 5mM ammonium formate solution (containing 0.2% formic acid) at a ratio of 70:30 (v/v) with a flow rate of 0.8mL/min. The MS acquisition was performed in multiple reactions monitoring (MRM) mode with a positive electrospray ionization source. The mass transitions monitored were m/z 244.1→185.1, m/z 230.1→171.1, m/z 260.1→201.1 and m/z 180.1→110.1 for agomelatine, 7-desmethyl-agomelatine, 3-hydroxy-agomelatine and internal standard (phenacetin), respectively. The method was validated for specificity, linearity and lower limit of quantification, precision and accuracy, extraction recovery, matrix effect and stability. The calibration curves for agomelatine, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma were linear over concentration ranges of 0.0457-100ng/mL, 0.1372-300ng/mL and 0.4572-1000ng/mL, respectively. Intra- and inter-day precisions and accuracies data met the acceptance criteria of FDA guideline for bioanalytical method validation. The developed method has been successfully applied to a bioequivalence study in healthy Chinese volunteers.[2]

[1]. The novel melatonin agonist agomelatine (S20098) is an antagonist at 5-hydroxytryptamine2C receptors, blockade of which enhances the activity of frontocortical dopaminergic and adrenergic pathways. J Pharmacol Exp Ther. 2003 Sep;306(3):9.

[2]. Development and validation a LC-MS/MS method for the simultaneous determination of agomelatine and its metabolites, 7-desmethyl-agomelatine and 3-hydroxy-agomelatine in human plasma: Application to a bioequivalence study. J Chromatogr B Analy.

Agomelatin (S20098) showed pKi values of 6.4 and 6.2 on the natural (pig) and cloned human 5-HT2C receptors, respectively. It also interacted with the h5-HT2B receptor (pKi value 6.6), but exhibited low affinity for the natural (rat)/cloned human 5-HT2A (pKi values <5.0/5.3) and 5-HT1A (pKi values <5.0/5.2) receptors, and negligible affinity for other 5-HT receptors (pKi values <5.0). In antibody capture/scintillation proximity assays, agomelatin inhibited h5-HT2C receptor-mediated activation of Gq/11 and Gi3 in a concentration-dependent and competitive manner (pA2 values 6.0 and 6.1, respectively). Agomelatine eliminated the activation of phospholipase C by h5-HT2C (pKB 6.1) and h5-HT2B (pKB 6.6) receptors using the [3H]phosphatidylinositol depletion assay. In vivo experiments showed that agomelatine dose-dependently blocked penile erection induced by the 5-HT2C agonists (S)-2-(6-chloro-5-fluoroindol-1-yl)-1-methylethylamine (Ro60,0175) and 1-methyl-2-(5,8,8-trimethyl-8H-3-azacyclopenta[a]indene-3-yl)ethylamine (Ro60,0332). Furthermore, agomelatine dose-dependently enhanced dopamine dialysis levels in the frontal cortex of freely moving rats, without affecting dopamine levels in the nucleus accumbens and striatum. Although agomelatine had no effect on the electrical activity of dopaminergic neurons in the ventral tegmental area, it eliminated the inhibitory effect of Ro60,0175. Agomelatine also enhanced extracellular levels of norepinephrine in the frontal cortex in a dose-dependent manner, while simultaneously increasing the firing frequency of adrenergic neuron cell bodies in the locus coeruleus. The selective melatonin antagonist N-[2-(5-ethyl-benzo[b]thiophene-3-yl)ethyl]acetamide (S22153) had no effect on the increase in norepinephrine and dopamine levels, which may reflect its blocking effect on 5-HT2C receptors that inhibit dopaminergic and adrenergic pathways in the frontal cortex. Correspondingly, unlike agomelatine, melatonin showed negligible activity on 5-HT2C receptors and failed to alter the activity of adrenergic and dopaminergic pathways. In summary, unlike melatonin, agomelatine acts as an antagonist of 5-HT2B and 5-HT2C receptors: blocking the latter enhances adrenergic and dopaminergic transmission in the frontal cortex. [1]

C14H15NO2

229.2744

229.11

152302-45-9

7-Desmethyl-agomelatine-d3;2749427-92-5

11775984

Typically exists as White to off-white solid at room temperature

1.2±0.1 g/cm3

511.3±33.0 °C at 760 mmHg

263.0±25.4 °C

0.0±1.4 mmHg at 25°C

1.624

1.62

2

2

3

17

267

0

CC(NCCC1=CC=CC2=C1C=C(O)C=C2)=O

UNTZQBYXDYYXIY-UHFFFAOYSA-N

InChI=1S/C14H15NO2/c1-10(16)15-8-7-12-4-2-3-11-5-6-13(17)9-14(11)12/h2-6,9,17H,7-8H2,1H3,(H,15,16)

N-[2-(7-hydroxynaphthalen-1-yl)ethyl]acetamide

N-(2-(7-Hydroxynaphthalen-1-yl)ethyl)acetamide; 152302-45-9; 7-Desmethyl-agomelatine; N-[2-(7-hydroxynaphthalen-1-yl)ethyl]acetamide; 7-Desmethylagomelatine; CHEMBL109784; N-[2-(7-hydroxy-1-naphthalenyl)ethyl]acetamide; O-Demethylagomelatine;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~160 mg/mL (~697.87 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (9.07 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (9.07 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (9.07 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)