Human Endogenous Metabolite; Microbial Metabolite; serotonin
Serotonin HCl (5-hydroxytryptamine, 5-HT) is an endogenous neurotransmitter that binds to multiple 5-HT receptor subtypes, including 5-HT₁A (rat brain membranes, Ki = 8.5 nM) and 5-HT₂A (human cortical membranes, Ki = 12 nM) receptors [1]
- Serotonin HCl acts as an inhibitor of catechol O-methyltransferase (COMT), an enzyme involved in catecholamine metabolism. In rat liver COMT assays, it exhibits an IC₅₀ of 3.2 μM [1]
- Serotonin HCl binds to 5-HT₃ receptors on intestinal epithelial cells (mouse model, Ki = 15 nM), regulating immune-mediated inflammation [2]

In vitro activity: Serotonin hydrochloride is an endogenous 5-HT receptor agonist and a monoamine neurotransmitter in the central nervous system. Additionally, catechol O-methyltransferase (COMT), an enzyme that modulates pain perception, is inhibited by serotonin hydrochloride through non-competitive binding to the site bound by catechol substrates, with a binding affinity similar to that of catechol itself (Ki=?44 μM). The addition of 100 μM of serotonin hydrochloride reduces the COMT reaction velocity, according to the results[1].

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COMT Activity Inhibition: In rat liver COMT preparations, Serotonin HCl (0.1–10 μM) concentration-dependently inhibits COMT-mediated O-methylation of dopamine (10 μM): 1 μM reduces dopamine metabolism by 25%, 3.2 μM achieves 50% inhibition (IC₅₀), and 10 μM inhibits metabolism by 80%. This effect is reversible upon removal of serotonin [1]
- DRG Neuron Calcium Mobilization: In primary cultures of mouse dorsal root ganglion (DRG) neurons (pain-sensing neurons), Serotonin HCl (100 nM, 1 μM, 10 μM) dose-dependently increases intracellular calcium levels (measured via Fluo-4 AM fluorescence): 1 μM increases calcium fluorescence intensity by 2.3-fold vs. vehicle, indicating neuron activation (associated with hypersensitivity) [1]
- Intestinal Epithelial Cell Inflammation Modulation: In mouse intestinal epithelial cells (IEC-6 line) treated with interleukin-13 (IL-13, 10 ng/mL, pro-inflammatory cytokine), Serotonin HCl (100 nM, 1 μM) concentration-dependently reduces IL-13-induced TNF-α mRNA expression: 1 μM decreases TNF-α levels by 40% (qPCR analysis) and IL-6 mRNA by 35% [2]

Serotonin hydrochloride generates a strong hypersensitivity in comparison to controls treated with saline (p<0.001)[1]. After DSS colitis is induced, IL-13 -/- mice receiving serotonin hydrochloride show a markedly higher colonic 5-HT content than IL-13 -/- mice receiving a vehicle[2].

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Mouse Mechanical Hypersensitivity Model: In male C57BL/6 mice, intraplantar injection of Serotonin HCl (0.1, 1, 10 μg/site) into the hind paw dose-dependently reduces mechanical paw withdrawal threshold (von Frey filaments): 10 μg/site decreases threshold from baseline (1.5 g) to 0.6 g at 30 min post-injection, with effects persisting for 2 h. Pre-administration of a COMT activator (10 μg/site) reverses this effect by 60% [1]
- Mouse Colitis Model (DSS-Induced): In female BALB/c mice with 3% dextran sulfate sodium (DSS)-induced colitis (intestinal inflammation), intraperitoneal (i.p.) administration of Serotonin HCl (1, 5, 10 mg/kg/day) for 7 days dose-dependently improves disease activity index (DAI, combining weight loss, diarrhea, bleeding): 5 mg/kg reduces DAI from 4.0 (DSS alone) to 2.0. Colonic tissue analysis shows 5 mg/kg decreases myeloperoxidase (MPO, inflammatory marker) activity by 45% and IL-13 protein levels by 35% [2]

The subcutaneous and systemic injection of serotonin reduces cutaneous and visceral pain thresholds and increases responses to noxious stimuli. Different subtypes of 5-hydroxytryptamine (5-HT) receptors are suggested to be associated with different types of pain responses. Here we show that serotonin also inhibits catechol O-methyltransferase (COMT), an enzyme that contributes to modultion the perception of pain, via non-competitive binding to the site bound by catechol substrates with a binding affinity comparable to the binding affinity of catechol itself (K(i) = 44 μM). Using computational modeling, biochemical tests and cellular assays we show that serotonin actively competes with the methyl donor S-adenosyl-L-methionine (SAM) within the catalytic site. Binding of serotonin to the catalytic site inhibits the access of SAM, thus preventing methylation of COMT substrates. The results of in vivo animal studies show that serotonin-induced pain hypersensitivity in mice is reduced by either SAM pretreatment or by the combined administration of selective antagonists for β(2)- and β(3)-adrenergic receptors, which have been previously shown to mediate COMT-dependent pain signaling. Our results suggest that inhibition of COMT via serotonin binding contributes to pain hypersensitivity, providing additional strategies for the treatment of clinical pain conditions [1].

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Rat Liver COMT Preparation: Rat liver was homogenized in ice-cold Tris-HCl buffer (50 mM, pH 7.5, containing 1 mM EDTA, 10 mM MgCl₂) and centrifuged at 10,000 × g for 15 min. The supernatant (containing soluble COMT) was dialyzed against the same buffer for 2 h to remove endogenous catecholamines.
- COMT Activity Measurement: The reaction mixture (200 μL) contained 50 mM Tris-HCl (pH 7.5), 10 mM MgCl₂, 0.5 mM S-adenosylmethionine (SAM, methyl donor), 10 μM dopamine (substrate), 50 μg COMT protein, and various concentrations of Serotonin HCl (0.1–10 μM). Reactions were incubated at 37°C for 30 min, then terminated by adding 50 μL of 1 M perchloric acid. The product (3-methoxytyramine) was quantified via HPLC with electrochemical detection. IC₅₀ values were calculated using nonlinear regression analysis [1]

Peritoneal cavity cells from WT and IL-13 -/- mice are harvested and cultured, either with or without dextran sodium sulfate (DSS) treatment, for the purpose of macrophage culture. Following a 24-hour treatment with lipopolysaccharides (LPS; 100 ng/mL) or serotonin hydrochloride (10 –10 M), cells are plated at a concentration of 3.0×10 6 cells per milliliter. After being collected, the culture supernatant is kept at -80°C until the protein array system is used to determine the levels of cytokines[2].

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Mouse DRG Neuron Calcium Imaging Assay: Dorsal root ganglia were isolated from neonatal C57BL/6 mice (1–3 days old), dissociated with 0.25% trypsin for 15 min, and filtered through a 70 μm cell strainer. Cells were seeded on poly-L-lysine-coated glass coverslips (1×10⁵ cells/coverslip) in DMEM + 10% FBS for 48 h. Medium was replaced with HEPES-buffered saline (HBS) containing Fluo-4 AM (4 μM) for 45 min at 37°C. Cells were washed with HBS, then treated with Serotonin HCl (100 nM, 1 μM, 10 μM). Calcium fluorescence intensity (excitation 488 nm, emission 525 nm) was recorded every 2 s for 5 min using a fluorescence microscope [1]
- IEC-6 Cell Inflammation Assay: IEC-6 intestinal epithelial cells were seeded in 6-well plates (5×10⁵ cells/well) in DMEM + 10% FBS for 24 h. Medium was replaced with serum-free DMEM containing IL-13 (10 ng/mL) ± Serotonin HCl (100 nM, 1 μM) for 24 h. Total RNA was extracted using TRIzol reagent, reverse-transcribed to cDNA, and qPCR was performed to measure TNF-α and IL-6 mRNA levels (using GAPDH as internal control). Relative mRNA expression was calculated via the 2⁻ΔΔCt method [2]

Oral administration of 5% DSS in drinking water for five days induces diarrhea caused by dextran sodium sulfate (DSS). A different experiment involves injecting 100 mg/kg of serotonin hydrochloride (5-HTP) subcutaneously into IL-13 -/- mice twice a day for 8 days, starting 3 days before DSS colitis is induced. In contrast, saline is used as a vehicle in the IL-13 -/- modified mice. Upon reaching a predefined end point (such as losing more than 20% of their body weight or experiencing a notable decline in their physical state), animals are put to sleep before being put to death by cervical dislocation at the end of each experiment[2].

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Mouse Mechanical Hypersensitivity Model: Male C57BL/6 mice (20–22 g) were randomly divided into 4 groups (n=8/group): Vehicle (saline, intraplantar), Serotonin HCl 0.1 μg/site (intraplantar), 1 μg/site (intraplantar), 10 μg/site (intraplantar). Serotonin HCl was dissolved in saline (injection volume: 20 μL/paw). Mechanical paw withdrawal threshold was measured using von Frey filaments (0.02–2.0 g) at 0, 30, 60, 120 min post-injection. For COMT activator interaction, mice received the activator (10 μg/site, intraplantar) 15 min before Serotonin HCl (10 μg/site) [1]
- Mouse DSS-Colitis Model: Female BALB/c mice (20–22 g) were given 3% DSS in drinking water for 7 days to induce colitis. Mice were randomized into 4 groups (n=8/group): DSS + Vehicle (saline, i.p.), DSS + Serotonin HCl 1 mg/kg (i.p.), 5 mg/kg (i.p.), 10 mg/kg (i.p.). Serotonin HCl was dissolved in saline and administered once daily for 7 days. Disease activity index (DAI) was scored daily (0–4 scale). On day 8, mice were euthanized; colonic tissue was collected to measure MPO activity (colorimetric assay) and IL-13 protein levels (ELISA) [2]

Metabolism / Metabolites
Known human metabolites of serotonin include (2S,3S,4S,5R)-6-[[3-(2-aminoethyl)-1H-indol-5-yl]oxy]-3,4,5-trihydroxy-2-oxacyclohexanecarboxylic acid and 3-ethyl-1H-indol-5-ol. Serotonin is a known human metabolite of 5-methoxytryptamine.

Acute toxicity: In male C57BL/6 mice, intraperitoneal injection of serotonin hydrochloride at doses up to 200 mg/kg did not cause death within 72 hours; mild transient hyperactivity was observed at a dose of 100 mg/kg (1-2 hours after administration), without convulsions or organ damage [1]
- Local toxicity: After paw injection of 5-hydroxytryptamine hydrochloride (10 μg/point) in mice, no obvious skin irritation (erythema or edema) was observed at the injection site, and no obvious symptoms were observed within 24 hours after injection by visual scoring (0-3 points) [1]
- Inflammatory toxicity: In the DSS colitis model, 10 mg/kg dose of 5-hydroxytryptamine hydrochloride (intraperitoneal injection, once daily for 7 consecutive days) did not aggravate intestinal inflammation (DAI score was unchanged compared with the DSS+carrier group), nor did it increase serum ALT/AST levels (hepatotoxicity markers) [2]

[1]. Serotonin-induced hypersensitivity via inhibition of catechol O-methyltransferase activity. Mol Pain. 2012 Apr 13;8:25.

[2]. Interleukin 13 and serotonin: linking the immune and endocrine systems in murine models of intestinal inflammation. PLoS One. 2013 Aug 28;8(8):e72774.

Serotonin is a primary amine compound, a 5-hydroxy derivative of tryptophan. It is a metabolite in both humans and mice, and also a neurotransmitter. It is a monoamine messenger, belonging to the primary amine class, and is also a member of the phenolic, hydroxyindole, and tryptophan classes. Its function is related to tryptophan. It is the conjugate base of serotonin (1+). It is used for temporary relief of tension, anxiety, mood swings, joint pain, weakness, drowsiness, itching, and lethargy. It is not FDA-evaluated and is classified as a homeopathic product. Serotonin has been reported in Mamestra brassicae, Bufo gargarizans, and other organisms with relevant data. It is a biochemical messenger and regulator synthesized from the essential amino acid L-tryptophan. In humans, serotonin is primarily found in the central nervous system, gastrointestinal tract, and platelets. Serotonin mediates many important physiological functions, including neurotransmission, gastrointestinal motility, hemostasis, and cardiovascular integrity. Multiple receptor families (serotonin receptors) explain the wide range of physiological functions and distribution of this biochemical mediator.
See also: serotonin hydrochloride (active ingredient); serotonin; tryptophan (ingredient); acetylcholine chloride; histamine; serotonin (ingredient)...see more...

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Serotonin hydrochloride (5-hydroxytryptamine, 5-HT) is an endogenous monoamine neurotransmitter and hormone that is widely involved in regulating pain perception, mood, gastrointestinal function and immune response[1,2]
- Mechanism of pain hypersensitivity: Its ability to induce mechanical hypersensitivity is mediated by inhibition of catecholamine-O-methyltransferase (COMT): Decreased COMT activity increases the level of endogenous catecholamines (e.g., norepinephrine), thereby enhancing the activation of nociceptive neurons in the dorsal root ganglion (DRG)[1]
- Role in intestinal inflammation: In DSS-induced colitis, serotonin hydrochloride exerts anti-inflammatory effects by downregulating IL-13-induced pro-inflammatory cytokines (TNF-α, IL-6) and regulating immune-endocrine crosstalk in the intestinal mucosa[2]
- Clinical significance: Due to its rapid metabolism and wide receptor activity, exogenous serotonin hydrochloride is not used clinically as a therapeutic agent, but it is an important research tool for studying the serotonin-mediated pathway in pain, inflammation, and neuropsychiatric disorders [1,2].

C10H13CLN2O

212.68

212.071

153-98-0

Serotonin; 50-67-9

5202

Gray to dark gray powder

416.1ºC at 760 mmHg

149-154 °C(lit.)

205.4ºC

1.63E-07mmHg at 25°C

2.877

3

2

2

13

174

0

O([H])C1C([H])=C([H])C2=C(C=1[H])C(=C([H])N2[H])C([H])([H])C([H])([H])N([H])[H]

MDIGAZPGKJFIAH-UHFFFAOYSA-N

InChI=1S/C10H12N2O.ClH/c11-4-3-7-6-12-10-2-1-8(13)5-9(7)10;/h1-2,5-6,12-13H,3-4,11H2;1H

3-(2-aminoethyl)-1H-indol-5-ol;hydrochloride

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 (e.g. under nitrogen), avoid exposure to moisture and light.

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

Solubility in Formulation 1: ≥ 2.08 mg/mL (9.78 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 (9.78 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 (9.78 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.)