Agmatine targets an exceptionally wide range of molecules, earning it the description of a "molecular shotgun," and exerts functions by simultaneously modulating multiple receptor and enzyme systems. Its targets include: (1) Glutamatergic receptors: It acts as an NMDA receptor antagonist (particularly at the GluN2B subunit) and permeates ionotropic glutamate receptors (AMPA, KA, NMDA); (2) Adrenergic receptors: It serves as an α₂-adrenergic receptor agonist and endogenous imidazoline receptor ligand, with an IC₅₀ of 36.5 μM at the rat kidney imidazoline I1 receptor; (3) Nitric oxide synthase (NOS): It inhibits multiple NOS isoforms and reduces NOS-2 protein levels in astroglial cells and macrophages; (4) Ion channels: It blocks ATP-sensitive potassium channels and voltage-gated calcium channels; (5) 5-HT3 receptor: It functions as an antagonist; (6) Polyamine metabolism: It enters cells via the polyamine transport system and regulates intracellular polyamine levels.

Agmatine exhibits differential regulatory effects on various cell types in vitro. In neuroprotection studies, agmatine (1-2000 µM) significantly protects primary cerebellar granule neurons from NMDA-induced excitotoxic injury. In glomerular mesangial cells, agmatine significantly suppresses proliferation of human mesangial cells stimulated with 20% FBS or PDGF-BB while reducing intracellular polyamine levels (putrescine, spermidine, spermine), and this anti-proliferative effect is not mediated through induction of apoptosis. In macrophage culture systems, agmatine inhibits lipopolysaccharide (LPS)-induced inflammatory cytokine and NO secretion, partially through inhibition of histone deacetylase (HDAC) activity. Additionally, as an ion channel permeant probe (AGB), agmatine can be used at 5-10 mM concentrations in in vitro tissue perfusates to study permeability of glutamate receptor-gated channels.

Agmatine demonstrates broad in vivo biological activities in various animal models. In neuroprotection, agmatine shows protective effects in multiple neural injury models including cerebral ischemia, spinal cord injury, and traumatic brain injury. In an ulcerative colitis mouse model, agmatine administered at 10 mM in drinking water significantly improves colonic tissue pathology and survival rates, with mortality decreasing from 70% to 0% across 0.4, 2, and 10 mM dose groups, through mechanisms involving inhibition of M1 macrophage polarization and increase in M2 macrophage proportion. In a drug-resistant epilepsy mouse model, intraperitoneal administration of agmatine at 5 and 10 mg/kg reverses drug resistance, restores the efficacy of standard anti-epileptic drugs, and modulates neurotransmitter levels (increased GABA, 5-HT, norepinephrine; decreased glutamate) and antioxidant markers in the brain. In a retinal inflammation model, topical delivery of agmatine via extracellular vesicles attenuates LPS-induced neuroinflammation. In NMDA-evoked pain behavior models, subcutaneous or oral agmatine administration attenuates NMDA-evoked biting/scratching behavior and tail-flick hyperalgesia.

Receptor binding affinity of agmatine is assessed using radioligand competitive binding assays. For imidazoline I1 receptor binding: membrane preparations from Wistar rat kidney are incubated with fixed concentrations of radiolabeled [³H]-clonidine and increasing concentrations of agmatine (typically 1 nM-10 mM). Radioactivity is measured by liquid scintillation counting, and IC₅₀ values are calculated, with an IC₅₀ of 36.5 µM at the rat kidney I1 receptor. For monoamine oxidase (MAO) inhibition assessment, the MAO-Glo luminescence method is used: agmatine is incubated with human recombinant MAO-A or MAO-B in the presence of substrate, and enzyme activity inhibition is calculated by luminescence detection, with IC₅₀ values of 100 µM for both MAO-A and MAO-B. For organic cation transporter (OCT) inhibition assessment, OCT1-expressing 293 cells are used to measure the inhibition of 0.1 µM MPP⁺ uptake by agmatine, with Ki values of 8.6 mM for rat OCT1 and 24 mM for human OCT1.

Effects of agmatine on cell proliferation are typically evaluated using various cell lines. Using human glomerular mesangial cells (MCs) as an example: cells are cultured in DMEM/F12 medium containing 10-20% FBS at 37°C, 5% CO₂. Logarithmically growing cells are seeded in 96-well plates (approximately 5×10³ cells per well), allowed to adhere overnight, then treated with increasing concentrations of agmatine (typically 0-1000 µM) while stimulating proliferation with 20% FBS or PDGF-BB (20 ng/ml). After 48-72 hours of treatment, cell viability is measured by MTT assay. Intracellular polyamine levels are detected by HPLC, and apoptosis is assessed by DNA fragmentation ELISA. Results show that agmatine significantly inhibits FBS or PDGF-stimulated mesangial cell proliferation, an effect associated with reduced intracellular polyamine levels and not related to apoptosis. For macrophage inflammatory response assessment, macrophages are co-incubated with LPS (100 ng/ml) and agmatine, with inflammatory cytokines detected by ELISA and NO levels measured by Griess assay.

In vivo study protocols for agmatine vary depending on experimental objectives. In a mouse ulcerative colitis model, agmatine is administered via drinking water (0.4, 2, and 10 mM) starting from day 0 of DSS induction through day 7. Body weight, stool consistency, and fecal blood are monitored daily, and animals are euthanized on day 7 for colon tissue histological scoring and inflammatory cytokine detection. In a drug-resistant epilepsy model, after establishing seizure via corneal kindling in adult albino mice, agmatine is administered intraperitoneally at 5 and 10 mg/kg (once daily for 7 days). Efficacy is evaluated by assessing seizure severity scores and resistance reversal, and following euthanasia, cortex and hippocampus are collected for neurotransmitter (GABA, glutamate, 5-HT, norepinephrine) and oxidative stress (GSH, catalase, TBARS) measurements. In NMDA-evoked pain behavior models, agmatine is administered subcutaneously or orally, with analgesic effects evaluated by assessing NMDA-evoked biting/scratching behavior duration and tail-flick latency. For pharmacokinetic studies, Sprague-Dawley rats receive IV or oral agmatine, serial blood samples are collected via indwelling catheters, and plasma drug concentrations are analyzed by HPLC-MS/MS.

The pharmacokinetic profile of agmatine has been preliminarily characterized in animal studies. Agmatine is a polar small molecule that does not efficiently diffuse passively across biological membranes, and its oral bioavailability is limited, requiring high doses (30-300 mg/kg in animal models) to achieve pharmacological effects. The compound can cross the intestinal epithelium and blood-brain barrier (BBB), but distribution is restricted. Notably, agmatine has a long half-life in the central nervous system (CNS), but its short elimination half-life in plasma restricts its efficacy as a systemic therapeutic agent. In rats, agmatine is absorbed through the intestine following oral administration and partially metabolized to urea and putrescine. Due to these pharmacokinetic limitations, agmatine-based prodrugs (Strategically Substituted Agmatines, SSAs) are being developed to improve pharmacokinetic parameters by increasing lipophilicity and prolonging plasma half-life.

Agmatine demonstrates a favorable safety profile in various animal models. In animal behavioral studies, no cardiac, motor, or neurological adverse effects have been observed with agmatine within the effective analgesic dose range. In a mouse ulcerative colitis model, agmatine (10 mM) treatment did not produce obvious systemic toxicity. Compared to arginine (ARG), agmatine (AGB) exhibits significantly lower cytotoxicity, while arginine may induce substantial cytotoxicity in certain cells, particularly those containing high levels of NO synthase. Among NMDA antagonists, agmatine's selectivity for the GluN2B subunit avoids the negative side effects observed with complete NMDA receptor inhibition. As an investigational drug, agmatine is currently being studied in clinical trials for small fiber peripheral neuropathy. Regarding physicochemical properties, agmatine free base has a melting point of 234-238°C, a LogP of approximately 1.099, and is stable under routine laboratory conditions.

[1]. https://pubchem.ncbi.nlm.nih.gov/compound/199

Agmatine is a primary amino compound belonging to the Agmatine family. It is a metabolite of E. coli and mice and is the conjugate base of the Agmatine ion (2+). Agmatine is a natural metabolite of the amino acid arginine. It is produced by the decarboxylation of arginine catalyzed by arginine decarboxylase and is naturally found in ragweed pollen, ergot, octopus muscle, herring sperm, sponges, and mammalian brain tissue. Agmatine is currently in the experimental and research stage. As an investigational drug, a non-blinded prospective case study in the United States is evaluating it in patients aged 18 to 75 years diagnosed with small fiber peripheral neuropathy. As of July 2013, the results of this study have not been published. As an experimental drug, arginine is being investigated for the treatment of various diseases, such as cardioprotection, diabetes, decreased renal function, neuroprotection (stroke, severe central nervous system injury, epilepsy, glaucoma, and neuropathic pain), and mental illnesses (depression, anxiety, schizophrenia, and cognitive impairment). The exact mechanism of action of arginine is still under investigation to evaluate all its potential indications. Arginine is a metabolite found in or produced by Escherichia coli (K12, MG1655 strains). It has also been reported to be present in soybeans, scallops, and other organisms with relevant data. Arginine is decarboxylated arginine and can be isolated from various plant and animal sources, such as pollen, ergot, herring sperm, and octopus muscle. Drug Indications Experimental studies are currently underway to evaluate the application of arginine in various indications, such as cardioprotection, diabetes, renal impairment, neuroprotection (stroke, severe central nervous system injury, epilepsy, glaucoma, and neuropathic pain), and mental illnesses (depression, anxiety, schizophrenia, and cognitive impairment). As an investigational drug, Agmatine is undergoing a non-blinded prospective case study in the United States in patients diagnosed with small fiber peripheral neuropathy. Mechanism of Action The exact mechanism of action for all potential indications of Agmatine is still under investigation. Several biochemical mechanisms have been identified that are related to the indications of Agmatine in diabetes, neuroprotection, and mental illness. In diabetes, Agmatine increases cellular glucose uptake by increasing insulin release from pancreatic islet cells and by increasing adrenal endorphin release, thus producing a hypoglycemic effect. In neuroprotection, the effects of Agmatine are thought to involve the regulation of receptors (NMDA, α2, and imidazoline receptors) and ion channels (ATP-sensitive potassium channels and voltage-gated calcium channels), as well as blocking nitric oxide synthesis. Arginine blocks nitric oxide synthesis by reducing the protein levels of nitric oxide synthase-2 (NOS-2) in astrocytes and macrophages. Regarding the therapeutic effects of arginine in mental illness, studies suggest that its mechanism involves the regulation of neurotransmitter receptors, including NMDA receptors, α2 receptors, serotonin receptors, opioid receptors, and imidazoline receptors. Specifically, when arginine binds to imidazoline and α2 receptors, it acts as a neurotransmitter, prompting the adrenal glands to release catecholamines.

C5H14N4

130.19

130.122

C, 46.13; H, 10.84; N, 43.03

306-60-5

306-60-5 ( free base);2482-00-0 (sulfate);

199

Typically exists as solid at room temperature

1.2 g/cm3

281.4ºC at 760 mmHg

234-238 degress Celcius

124ºC

1.099

3

2

4

9

85

0

NCCCCNC(N)=N

QYPPJABKJHAVHS-UHFFFAOYSA-N

InChI=1S/C5H14N4/c6-3-1-2-4-9-5(7)8/h1-4,6H2,(H4,7,8,9)

2-(4-aminobutyl)guanidine

NSC56332; agmatine; 1-(4-Aminobutyl)guanidine; 306-60-5; (4-Aminobutyl)guanidine; N-(4-aminobutyl)guanidine; NSC 56332; Agmatine; NSC-56332

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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