Cat. No.: HY-B1359

Methylene blue trihydrate

(Synonyms: C.I. Basic Blue 9 trihydrate)

30 Cited Publications

Customer Review

Based on 30 publication(s) in Google Scholar

Methylene blue trihydrate (C.I. Basic Blue 9 trihydrate) is a guanylyl cyclase (sGC), monoamine oxidase A (MAO-A) and NO synthase (NOS) inhibitor. Methylene blue trihydrate is a vasopressor and is often used as a dye in several medical procedures. Methylene blue trihydrate has antinociception, antimalarial, antidepressant and anxiolytic activity effects. Methylene Blue trihydrate has the potential for methemoglobinemias, neurodegenerative disorders and ifosfamide-induced encephalopathytreatment.

For research use only. We do not sell to patients.

Purity: 98.0%
CAS No.: 7220-79-3
Formula: C₁₆H₂₄ClN₃O₃S
Molecular Weight:373.90

Storage:

Powder -20°C, 3 years , 4°C, 2 years ; In solvent -80°C, 6 months , -20°C, 1 month

Methylene blue trihydrate Chemical Structure

CAS No.: 7220-79-3

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Top Publications Citing

Use of MCE Products

Fluorescent Dye

Strong Fluorescence Signal / Multi-Disciplinary Applications / High Fluorescence Specificity

Biological Activity
- Description
- IC₅₀ & Target
- Cellular Effect
- In Vitro
- In Vivo
- Clinical Trial
Chemical Information
Publications (30)
Solvent & Solubility
- In Vitro
- In Vivo
Purity & Documentation
References
Help & FAQs
Classifications
Bioz

Biological Activity

Description

IC₅₀ & Target

Plasmodium

All Monoamine Oxidase Isoforms :

MAO-A
MAO-B

All NO Synthase Isoforms :

All Parasite Isoforms :

Cellular Effect

Cell Line	Type	Value	Description	References
Caco-2	CC₅₀	7.25 μM Compound: METHYLENE BLUE	Toxicity against Caco-2 cells determined at 48 hours by intracellular ATP concentration using the CellTiter-Glo Luminescent Cell Viability Assay Toxicity against Caco-2 cells determined at 48 hours by intracellular ATP concentration using the CellTiter-Glo Luminescent Cell Viability Assay	10.21203/rs.3.rs-23951/v1
Caco-2	IC₅₀	2.03 μM Compound: METHYLENE BLUE	Determination of IC50 values for inhibition of SARS-CoV-2 induced cytotoxicity of Caco-2 cells after 48 hours by high content imaging Determination of IC50 values for inhibition of SARS-CoV-2 induced cytotoxicity of Caco-2 cells after 48 hours by high content imaging	10.21203/rs.3.rs-23951/v1

In Vitro

By acting as an alternative electron acceptor/donor Methylene blue restores mitochondrial function, improves neuronal energy production and inhibits the formation of superoxide^[1].
Methylene blue inhibits cytochrome P450 (CYP) isozymes. Methylene blue is an odorless, water soluble, dark blue-green crystalline powder, which turns blue whenever mixed in solution. Methylene blue is a vasopressor that impacts the NO synthetic pathway by inhibiting inducible NOS and inhibiting the subsequent activation of sGC. In addition, by binding to the iron heme moiety of sGC and causing enzyme inhibition, Methylene blue blocks accumulation of cyclic GMP (cGMP), competing directly with NO in its ability to activate soluble guanylyl cyclase^[3].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

In Vivo

Methylene Blue (1, 5, and 25 μg/rat) significantly decreases sevoflurane minimum alveolar anesthetic concentration (MAC) and brain cyclic guanosine monophosphate (cGMP) content in a dose-dependent manner in male Sprague-Dawley rats (7-week-old, 200-250 g)^[2].
Methylene blue is used as a dye in chromoendoscopy, and is sprayed onto the mucosa of the gastrointestinal tract in order to identify dysplasia, or pre-cancerous lesions^[2].
Methylene blue is able to restore vascular tone, normalize mean arterial pressures (MAP), and reduce vasopressor usage^[3].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

Clinical Trial

NCT Number	Sponsor	Condition	Start Date	Phase A549
NCT01329991	Plexxikon\|		2011-05	PHASE1

Chemical Information

CAS No. 7220-79-3
Appearance Solid
Molecular Weight 373.90
Formula C₁₆H₂₄ClN₃O₃S
Color Dark green to black
SMILES

CN(C1=CC(SC2=C/C(C=CC2=N3)=[N+](C)\C)=C3C=C1)C.O.[Cl-]
Synonyms

C.I. Basic Blue 9 trihydrate
Shipping

Room temperature in continental US; may vary elsewhere.

Storage

Powder	-20°C	3 years
	4°C	2 years
In solvent	-80°C	6 months
	-20°C	1 month

Publications (30)

All Gel ElectrophoresisCell Migration/Invasion AssayWB

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Journal Impact Factor

Nat Commun

DEAD-box RNA helicase 10 is required for 18S rRNA maturation by controlling the release of U3 snoRNA from pre-rRNA in embryonic stem cells. [Abstract]

2024 Nov 27;15(1):10303. PMID: 39604362

Methylene blue trihydrate purchased from MedChemExpress. Usage Cited in: Nat Commun. 2024 Nov 27;15(1):10303. [Abstract]

Northern blot analysis of pre-rRNA intermediates using 5'ETS-1, 5'ETS-2, 5'ETS and ITS1-29 probes in DDX10-AID mESCs with IAA treatment (0 h, 6 h, 24 h, and 48 h) or treated with IAA for 48 h followed by 48 h of washing. MB represents methylene blue staining. Experiments were repeated three times independently with similar results. Membranes were stained with Methylene Blue stain solution (0.03% Methylene Blue in 0.3 M sodium acetate (pH 5.0-5.5)), and then RNA was crosslinked onto the membrane using UV light.

Methylene blue trihydrate purchased from MedChemExpress. Usage Cited in: Nat Commun. 2024 Nov 27;15(1):10303. [Abstract]

Northern blot analysis of pre-rRNA intermediate levels in DDX10-AID (+ OsTir1) mESCs overexpressing DDX10FL, DDX10ΔHBD, DDX10ΔHCD or DDX10ΔHD at 0 h and 48 h after IAA treatment. MB represents Methylene Blue staining. Experiments were repeated three times independently with similar results. Source data are provided as a Source Data file. Membranes were stained with Methylene Blue stain solution (0.03% Methylene Blue in 0.3 M sodium acetate (pH 5.0-5.5)), and then RNA was crosslinked onto the membrane using UV light.

Methylene blue trihydrate purchased from MedChemExpress. Usage Cited in: Nat Commun. 2024 Nov 27;15(1):10303. [Abstract]

Northern blot analysis of pre-rRNA intermediates in DDX10-AID (+ OsTir1) mESCs overexpressing NUP98-DDX10 or DDX10, treated with IAA at different time points (0 h, 24 h, and 48 h). MB represents Methylene Blue staining. Experiments were repeated two times independently with similar results. Source data are provided as a Source Data file. Membranes were stained with Methylene Blue stain solution (0.03% Methylene Blue in 0.3 M sodium acetate (pH 5.0-5.5)), and then RNA was crosslinked onto the membrane using UV light.
ACS Nano

Keratinocyte-Derived Apoptotic Nanovehicles Reprogram Cutaneous Dendritic Cells to Ameliorate Psoriasis. [Abstract]

2026 Jun 30;20(25):18058-18072. PMID: 42301004
Redox Biol

An effective inactivant based on singlet oxygen-mediated lipid oxidation implicates a new paradigm for broad-spectrum antivirals. [Abstract]

2020 Sep:36:101601. PMID: 32535542
Gut Microbes

Helicobacter pylori infection aggravates hepatic steatosis by lactylation-driven WTAP-mediated m6A modification. [Abstract]

2025 Dec 31;17(1):2599543. PMID: 41384837
J Nanobiotechnology

SIRT5-modified human umbilical cord mesenchymal stem cells loaded with antioxidant polydopamine nanozyme enhance parpi resistance in ovarian cancer via fatty acid metabolism reprogramming. [Abstract]

2025 Jul 4;23(1):485. PMID: 40616128
Theranostics

Establishment of tumor inflammasome clusters with distinct immunogenomic landscape aids immunotherapy. [Abstract]

2021 Oct 17;11(20):9884-9903. PMID: 34815793

Methylene blue trihydrate purchased from MedChemExpress. Usage Cited in: Theranostics. 2021 Oct 17;11(20):9884-9903. [Abstract]

Migration analysis of THP-1, co-cultured with cancer cells that were pre-treated with Methylene Blue.

Methylene blue trihydrate purchased from MedChemExpress. Usage Cited in: Theranostics. 2021 Oct 17;11(20):9884-9903. [Abstract]

Western blotting analysis of PD-L1 expression in THP-1 which was co-cultured with cancer cells pre-treated with Methylene Blue.
Biomaterials

Manganese-enriched photonic/catalytic nanomedicine augments synergistic anti-TNBC photothermal/nanocatalytic/immuno-therapy via activating cGAS-STING pathway. [Abstract]

2023 Feb:293:121988. PMID: 36580716
Chem Eng J

2025 Nov 24;526:171230.
Phytomedicine

Arnicolide C induces ROS-mediated modulation of PI3K/Akt and MAPK pathways to suppress MYC in hepatocellular carcinoma. [Abstract]

2025 Nov 25:148:157423. PMID: 41138574
Nano Res

2026 Feb 6.
Phytother Res

Carnosol inhibits KGN cells oxidative stress and apoptosis and attenuates polycystic ovary syndrome phenotypes in mice through Keap1-mediated Nrf2/HO-1 activation. [Abstract]

2023 Apr;37(4):1405-1421. PMID: 36786429
Cell Rep

WDFY2 promotes MRN complex formation required for homologous recombination-mediated DNA repair. [Abstract]

2025 Nov 25;44(11):116520. PMID: 41196680
Cell Rep

The ARID1A-METTL3-m6A axis ensures effective RNase H1-mediated resolution of R-loops and genome stability. [Abstract]

2024 Feb 13;43(2):113779. PMID: 38358891
Nanophotonics

Disentangling the absorption lineshape of methylene blue for nanocavity strong coupling. [Abstract]

2025 Nov 25;14(27):4993-5001. PMID: 41426083
J Environ Sci

2025 Dec 25.
Cell Signal

YTHDF2 drives oral squamous cell carcinoma progression via m6A-dependent degradation of MTUS1/ATIP1 mRNA and mitochondrial dysregulation. [Abstract]

2025 Dec:136:112145. PMID: 40975502
Bioengineering (Basel)

LIPUS Enhances Gallbladder Motility via ANO1 in Acute Cholecystitis Guinea Pigs. [Abstract]

2025 Oct 27;12(11):1164. PMID: 41301120
BMC Cancer

FTO-mediated demethylation of MTUS1/ATIP1 promotes tumor progression in head and neck squamous cell carcinoma. [Abstract]

2024 Dec 3;24(1):1489. PMID: 39627705
Chemistry

2026 May;8(5):65.
Biomed Res Int

Gentiopicroside Produces Endothelium-Independent Vasodilation by Deactivating the PI3K/Akt/Rho-Kinase Pathway in Isolated Rat Thoracic Aorta. [Abstract]

2021 May 14:2021:5565748. PMID: 34095301
Exp Ther Med

MicroRNA-139-5p regulates chronic inflammation by suppressing nuclear factor-κB activity to inhibit cell proliferation and invasion in colorectal cancer. [Abstract]

2019 Nov;18(5):4049-4057. PMID: 31616518
World J Gastrointest Surg

Impact of interstitial cells of Cajal on slow wave and gallbladder contractility in a guinea pig model of acute cholecystitis. [Abstract]

2023 Jun 27;15(6):1068-1079. PMID: 37405098
Hematology

Downregulation of Smad4 expression confers chemoresistance against imatinib mesylate to chronic myeloid leukemia K562 cells. [Abstract]

2022 Dec;27(1):43-52. PMID: 34957936
Res Sq

2025 Nov 12.
Patent

US20240325538A1.
Patent

US20240238271A1.
Patent

US20230226111A1.
Research Square Print

2023 Feb 21.
Curr Res Virol Sci

Identification of broad anti-coronavirus chemical agents for repurposing against SARS-CoV-2 and variants of concern. [Abstract]

2022;3:100019. PMID: 35072124
Evid Based Complement Alternat Med

Acupuncture at Gastric Back-Shu and Front-Mu Acupoints Enhances Gastric Motility via the Inhibition of the Glutamatergic System in the Hippocampus. [Abstract]

2020 Mar 10:2020:3524641. PMID: 32215036

Solvent & Solubility

In Vitro:

DMSO : 100 mg/mL (267.45 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)

View the Complete Stock Solution Preparation Table

Preparing Stock Solutions

View the Complete Stock Solution Preparation Table

Concentration Solvent Mass

1 mg

5 mg

10 mg

Concentration

Solvent

Mass

1 mM

2.6745 mL

13.3726 mL

26.7451 mL

5 mM

0.5349 mL

2.6745 mL

5.3490 mL

10 mM

0.2675 mL

1.3373 mL

2.6745 mL

Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.

Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

Molarity Calculator

Mass (g) = Concentration (mol/L) × Volume (L) × Molecular Weight (g/mol)

Mass Mass

Concentration Concentration

Volume Volume

Molecular Weight Molecular Weight

Dilution Calculator

Concentration _(start) × Volume _(start) = Concentration _(final) × Volume _(final)

Concentration _(start) Concentration _(start)

Volume _(start) Volume _(start)

Concentration _(final) Concentration _(final)

Volume _(final) Volume _(final)

In Vivo:

Select the appropriate dissolution method based on your experimental animal and administration route.

Stock Solution Working Solution

For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.

Protocol 1

Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline

Solubility: ≥ 2.5 mg/mL (6.69 mM); Clear solution

This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).

Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.

Protocol 2

Add each solvent one by one: 10% DMSO 90% (20% SBE-β-CD in Saline)

Solubility: ≥ 2.5 mg/mL (6.69 mM); Clear solution

This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).

Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.

In Vivo Dissolution Calculator

Please enter the basic information of animal experiments:

Dosage Dosage
mg/kg
Animal Weight (Per Animal) Animal Weight (Per Animal)
g
Dosing Volume (Per Animal) Dosing Volume (Per Animal)
μL
Number of Animals Number of Animals

Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.

Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.

Please enter your animal formula composition:

%
DMSO +

Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
%
+
+
%
Tween-80 + +
%
Saline +

The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).

Working solution concentration: 0.22 mg/mL

Purity & Documentation

Select Batch:

Purity: 99.98%

Assay: 99.98%

ee.: 99.98%

Data Sheet (275 KB)
SDS (558 KB)
- English - EN (558 KB)
- Français - FR (558 KB)
- Deutsch - DE (558 KB)
- Norwegian - NO (558 KB)
- Español - ES (558 KB)
- Swedish - SV (558 KB)
- Italian - IT (558 KB)
- Korean - KR (558 KB)
- Portuguese - PT (558 KB)
COA (2659 KB)
Elemental Analysis Report (2659 KB)
HNMR (2659 KB)
CNMR (2659 KB)
RP-HPLC (2659 KB)
NP-HPLC (2659 KB)
MS (2659 KB)
LCMS (2659 KB)
IR (2659 KB)
GC (2659 KB)
OR (2659 KB)
SDS-PAGE (2659 KB)
SEC-HPLC (2659 KB)
Other Report (2659 KB)
Handling Instructions (2659 KB)

References

[1]. Delport A, et al. Methylene blue and its analogues as antidepressant compounds. Metab Brain Dis. 2017 Oct;32(5):1357-1382. [Content Brief]

[2]. Masaki E, et al. Methylene blue, a soluble guanylyl cyclase inhibitor, reduces the sevoflurane minimum alveolar anesthetic concentration and decreases the brain cyclic guanosine monophosphate content in rats. Anesth Analg. 1999 Aug;89(2):484-9. [Content Brief]

[3]. McCartney SL, et al. Intraoperative vasoplegia: methylene blue to the rescue! Curr Opin Anaesthesiol. 2018 Feb;31(1):43-49. [Content Brief]

Help & FAQs

How should lyophilized recombinant proteins be reconstituted and stored?

1. Before opening the cap, centrifuge the vial at 13000 rpm for 20-30 seconds. This step will ensure that any lyophilized powder that may have adhered to the cap or walls is collected at the bottom of the vial, minimizing the risk of product loss.

Complete Stock Solution Preparation Table

Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

Optional Solvent	Concentration Solvent Mass	1 mg	5 mg	10 mg	25 mg
DMSO	1 mM	2.6745 mL	13.3726 mL	26.7451 mL	66.8628 mL
	5 mM	0.5349 mL	2.6745 mL	5.3490 mL	13.3726 mL
	10 mM	0.2675 mL	1.3373 mL	2.6745 mL	6.6863 mL
	15 mM	0.1783 mL	0.8915 mL	1.7830 mL	4.4575 mL
	20 mM	0.1337 mL	0.6686 mL	1.3373 mL	3.3431 mL
	25 mM	0.1070 mL	0.5349 mL	1.0698 mL	2.6745 mL
	30 mM	0.0892 mL	0.4458 mL	0.8915 mL	2.2288 mL
	40 mM	0.0669 mL	0.3343 mL	0.6686 mL	1.6716 mL
	50 mM	0.0535 mL	0.2675 mL	0.5349 mL	1.3373 mL
	60 mM	0.0446 mL	0.2229 mL	0.4458 mL	1.1144 mL
	80 mM	0.0334 mL	0.1672 mL	0.3343 mL	0.8358 mL
	100 mM	0.0267 mL	0.1337 mL	0.2675 mL	0.6686 mL

Methylene blue trihydrate

30 Cited Publications

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Methylene blue trihydrate

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Publications Citing Use of MedChemExpress (MCE) Methylene blue trihydrate

All Monoamine Oxidase Isoforms

All NO Synthase Isoforms

All Parasite Isoforms

Biological Activity

Chemical Information

Publications (30)

Solvent & Solubility

Purity & Documentation

References

Help & FAQs

Complete Stock Solution Preparation Table

Methylene blue trihydrate Related Classifications

HY-B1359 Related Classifications