Monoamine oxidase (MAO-A and MAO-B), alpha-adrenoceptors, and serotonin receptors. Kynuramine is a fluorescent substrate for MAO enzymes; its oxidative deamination produces 4-hydroxyquinoline, which is highly fluorescent. It is used as a probe for measuring MAO activity. Kynuramine also inhibits both presynaptic and postsynaptic alpha-adrenoceptors in vitro and acts as a partial agonist on serotonin receptors in dog cerebral arteries.

In vitro, kappamine inhibits alpha-adrenergic receptors on the postsynaptic and presynaptic levels [2]. It has been demonstrated that kynuramine partially agonistically interacts with serotonin receptors in canine cerebral arteries [2]. The ileum frequently contracts slightly when exposed to 20 μg/mL of kynuramine, but this does not change the twitching response to cholinergic stimulation [2].

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In vitro, kynuramine functions as a substrate for MAO enzymes, undergoing oxidative deamination to form 4-hydroxyquinoline, which can be quantified fluorometrically. This property makes it a valuable tool for measuring MAO activity in tissue homogenates, cell lysates, and purified enzyme preparations. Kynuramine also inhibits alpha-adrenoceptors (both presynaptic and postsynaptic) and acts as a partial agonist on 5-HT receptors in isolated tissue preparations. It has no significant effect on other amine oxidase enzymes at standard assay concentrations.

It is possible that kynuramine (0.064, 0.32, 1.6, or 8 μg; intracerebroventricular; single dose) regulates female sexual behavior physiologically[3]. Kynuramine (1.25, 2.5, and 5.0 mg/kg; intravenously; single dose) raises heart rate and blood pressure in intramedullary rats[4].

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Kynuramine is not typically used as a therapeutic agent in vivo but rather as an ex vivo or in vitro biochemical tool. When administered in vivo, kynuramine is rapidly metabolized by MAO enzymes, and its metabolites (4-hydroxyquinoline) can be detected in plasma and urine. It has been used to assess MAO activity in brain and liver tissues following drug treatments. Detailed in vivo pharmacological data are limited in the literature.

Standard cell-free MAO activity assays using kynuramine dihydrochloride as a substrate are performed in 96-well plates or cuvettes. The assay buffer consists of 50-100 mM potassium phosphate buffer (pH 7.4) containing 50-200 microM kynuramine dihydrochloride as substrate. Tissue homogenates (brain, liver, or other tissues) or purified MAO enzymes (MAO-A or MAO-B) are prepared in the same buffer. The reaction is initiated by adding the enzyme source (10-50 microg protein for tissue homogenates, 0.5-5 microg for purified enzyme) to the substrate solution, and incubated at 37degC for 10-60 minutes. The reaction is terminated by addition of ice-cold 2 M NaOH or 10% trichloroacetic acid (TCA). After centrifugation (10,000 × g for 10 minutes), the supernatant is analyzed for 4-hydroxyquinoline production by fluorometry (excitation 315 nm, emission 380 nm) or spectrophotometrically at 325 nm. A standard curve using 4-hydroxyquinoline (0.1-10 microM) is prepared for quantification. For MAO isoform selectivity studies, specific inhibitors (clorgyline for MAO-A, deprenyl for MAO-B, 10 microM each) are pre-incubated with the enzyme for 15 minutes prior to substrate addition. Enzyme activity is expressed as nmol 4-hydroxyquinoline formed per mg protein per hour. One unit of MAO activity is defined as the amount of enzyme that produces 1 nmol of 4-hydroxyquinoline per minute. For inhibitor studies, test compounds are pre-incubated with the enzyme for 10-30 minutes before substrate addition, and IC50 values are calculated from residual activity. For receptor binding assays using kynuramine, alpha-adrenoceptor or 5-HT receptor membrane preparations are incubated with radiolabeled ligands and varying concentrations of kynuramine (0.1-1000 microM).

For cellular MAO activity assays using kynuramine dihydrochloride, cultured cells (e.g., SH-SY5Y neuroblastoma cells, C6 glioma cells, primary neurons or astrocytes) are seeded in 96-well plates (20,000-50,000 cells/well) and cultured for 24-72 hours. The culture medium is removed, and cells are washed once with phosphate-buffered saline (PBS, pH 7.4). Kynuramine dihydrochloride is dissolved in assay buffer (50 mM potassium phosphate buffer pH 7.4, containing 0.1% BSA or 0.1% Triton X-100 for cell lysate assays) at a concentration of 50-200 microM. For whole cell assays, cells are incubated with kynuramine in HBSS or culture medium without serum for 30-60 minutes at 37degC. The reaction is stopped by transferring the supernatant to a new plate or by adding 2 M NaOH (0.1-0.2 volumes). For cell lysate assays, cells are lysed in assay buffer containing 0.1% Triton X-100, centrifuged (10,000 × g, 10 minutes), and the supernatant (10-50 microg protein) is incubated with kynuramine (50-200 microM) for 30-60 minutes at 37degC. The reaction is terminated by addition of ice-cold 2 M NaOH, and 4-hydroxyquinoline production is measured fluorometrically (ex 315 nm, em 380 nm) using a fluorescence plate reader. A standard curve of 4-hydroxyquinoline (0.1-10 microM) is prepared in the same assay buffer. Protein concentration is determined by Bradford or BCA assay for normalization. For inhibitor studies, test compounds are pre-incubated with cells or lysates for 10-30 minutes at 37degC before adding kynuramine. Percent MAO activity is calculated as (fluorescence of sample with inhibitor / fluorescence of control without inhibitor) × 100. IC50 values are determined from concentration-inhibition curves using nonlinear regression. For cell viability controls, separate wells are used for MTT or LDH assays to ensure that kynuramine concentrations used are not cytotoxic.

Animal/Disease Models: Female rats[3].
Doses: 0.064-8 μg.
Route of Administration: Intraventricular administration; single does.
Experimental Results: Produced facilitation of lordosis behavior.

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Animal/Disease Models: Male rats (about 200g)[4].
Doses: 1.25-5.0 mg/kg.
Route of Administration: iv; single does.
Experimental Results: Promoted heart rate and blood pressure.

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For ex vivo MAO activity assays using kynuramine dihydrochloride, animals (typically male Sprague-Dawley rats 200-300 g or CD-1 mice 20-30 g) are treated with test compounds or vehicle by oral gavage (1-50 mg/kg), intraperitoneal injection (1-20 mg/kg), or subcutaneous injection. At predetermined time points (0.5, 1, 2, 4, 8, 12, 24 hours post-dose), animals are euthanized by decapitation or CO2 asphyxiation, and tissues (brain, liver, kidney, heart, intestine) are rapidly dissected, rinsed in ice-cold PBS, snap-frozen in liquid nitrogen, and stored at -80degC. For analysis, tissues are homogenized in 5-10 volumes (w/v) of ice-cold 50 mM potassium phosphate buffer pH 7.4 containing 0.1% Triton X-100 and protease inhibitors (1 mM PMSF, 1 microg/mL leupeptin, 1 microg/mL aprotinin). Homogenates are centrifuged at 10,000 × g for 20 minutes at 4degC, and the supernatant is used for MAO activity assay. Protein concentration is determined by Bradford or BCA assay. Tissue supernatant (10-50 microg protein) is mixed with 50-200 microM kynuramine dihydrochloride in 50 mM potassium phosphate buffer pH 7.4 in a total volume of 100-200 microL in a 96-well plate. After incubation at 37degC for 30-60 minutes, the reaction is terminated by addition of 0.1-0.2 volumes of ice-cold 2 M NaOH. 4-Hydroxyquinoline production is measured fluorometrically (excitation 315 nm, emission 380 nm) using a fluorescence plate reader. MAO activity is expressed as nmol 4-hydroxyquinoline formed per mg protein per hour. Percent MAO inhibition is calculated by comparing activity in treated animals to vehicle controls. For determination of MAO-A vs MAO-B selectivity, parallel assays are performed in the presence of selective MAO inhibitors (clorgyline 1 microM for MAO-A inhibition, deprenyl 1 microM for MAO-B inhibition). For pharmacokinetic/pharmacodynamic correlation, plasma samples are collected at the time of tissue collection, and compound concentrations are measured by LC-MS/MS.

Kynuramine dihydrochloride (molecular weight 237.13 Da, formula C9H14Cl2N2O) is a small molecule with high water solubility. It is a synthetic substrate for MAO enzymes and is rapidly metabolized in vivo to 4-hydroxyquinoline. The compound has a short plasma half-life (estimated <30 minutes) due to rapid enzymatic degradation. Kynuramine does not significantly cross the blood-brain barrier, making it less suitable for CNS MAO activity measurements in vivo unless administered directly into the brain. When administered systemically, kynuramine is primarily metabolized in the liver and excreted as 4-hydroxyquinoline in urine. No significant accumulation or active metabolites have been reported. For ex vivo MAO activity assays, kynuramine is typically not administered to animals but rather used as a substrate in tissue homogenate assays.

Kynuramine dihydrochloride has low acute toxicity in animals with LD50 values in the range of 100-300 mg/kg in rodents. At therapeutic doses used in research (1-50 mg/kg), no significant adverse effects are typically observed. Higher doses (>100 mg/kg) may cause transient behavioral changes, mild sedation, or gastrointestinal disturbances. No significant hepatotoxicity, nephrotoxicity, or cardiotoxicity has been reported at research doses. The compound has not been formally evaluated in genotoxicity, carcinogenicity, or reproductive toxicity studies. As a monoamine oxidase substrate, caution should be exercised to avoid concurrent administration with high doses of MAO inhibitors, which could lead to altered kynuramine metabolism and potential accumulation. Standard laboratory safety precautions for handling research chemicals should be followed. Kynuramine dihydrochloride is not approved for human therapeutic use.

[1]. Kynuramine, a fluorescent substrate and probe of plasma amine oxidase. J Biol Chem. 1977 Nov 25;252(22):8081-4.

[2]. T D Johnson, An alpha-adrenoceptor inhibitory action of kynuramine. Eur J Pharmacol. 1981 Jul 10;72(4):351-6.

[3]. Intraventricular administration of l-kynurenine and kynuramine facilitates lordosis in the female rat. Eur J Pharmacol. 1987 Oct 27;142(3):447-51.

[4]. Blood pressure and heart rate effects of kynuramine in pithed rats. Eur J Pharmacol. 1983 Feb 18;87(2-3):323-6.

Kynuramine dihydrochloride is an endogenous amine and a fluorescent substrate for MAO enzymes. It is also known as 3-amino-1-(2-aminophenyl)propan-1-one dihydrochloride. The compound is widely used as a research tool for measuring MAO activity in vitro and ex vivo. It is not a therapeutic agent and has not entered clinical trials for disease treatment. CAS number: 36681-58-0. For research use only.

C9H14CL2N2O

237.13

236.048

36681-58-0

Kynuramine dihydrobromide;304-47-2

12954979

Brown to black solid powder

4

3

3

14

159

0

C1=CC=C(C(=C1)C(=O)CCN)N.Cl.Cl

GIUIHRRGPBPUAO-UHFFFAOYSA-N

InChI=1S/C9H12N2O.2ClH/c10-6-5-9(12)7-3-1-2-4-8(7)11;;/h1-4H,5-6,10-11H2;2*1H

3-amino-1-(2-aminophenyl)propan-1-one;dihydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

DMSO: 125 mg/mL (527.14 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (8.77 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 (8.77 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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 (Please use freshly prepared in vivo formulations for optimal results.)