Methylene Blue

Methylene blue, also called methylthioninium chloride, is a synthetic phenothiazinium dye. First made in 1876, it formed the basis of early antimalarial chemotherapy and later led to the neuroleptic phenothiazines, and its ready oxidation and reduction underlies much of its activity (Wainwright and Crossley, 2002). It was the first synthetic antimalarial, used in the late 19th and early 20th centuries (Lu et al., 2018).

Chemical data and synonyms are from PubChem CIDs 6099, 104827 and 4139.

Published methylene blue research falls into six areas, and each summary below names its model. Mechanistic and neuroprotection data come from cells and rodents.

Methylene blue and mitochondria: redox cycling and electron transfer

Flavin enzymes use NAD(P)H to reduce methylene blue to leucomethylene blue, and cytochrome c reoxidizes it. In IMR90 human fibroblasts, nanomolar methylene blue delayed senescence by more than 20 population doublings and raised complex IV by 30 percent and oxygen consumption by 37 to 70 percent, which the authors tied to this cycling (Atamna et al., 2008).

In mitochondrial assays and HT-22 mouse hippocampal cells, it carried electrons from NADH to cytochrome c around blocked complexes I and III, while an N-acetylated derivative with its redox center disabled had no effect on complex activities (Wen et al., 2011). Models: cell lines and mitochondrial preparations.

Nitric oxide, guanylate cyclase and MAO-A

Methylene blue is widely used to block nitric oxide-stimulated guanylate cyclase. In blood vessels, endothelial cells and purified enzymes, it inhibited nitric oxide synthase directly, abolishing L-citrulline formation at 30 µM, while guanylate cyclase needed about 60 µM for half inhibition (Mayer et al., 1993). With purified human enzymes, it was a potent, tight-binding, reversible inhibitor of MAO-A and inhibited MAO-B only at much higher concentrations (Ramsay et al., 2007). Models: vascular tissue, endothelial cells and purified enzymes.

Neuroprotection and memory in rodents

In male Sprague-Dawley rats, methylene blue almost completely rescued rotenone-induced striatal dopamine depletion, attenuated nigral neuron loss and reduced damage after transient focal cerebral ischemia (Wen et al., 2011). Given locally with rotenone infused into the rat striatum, it shrank lesions and preserved cytochrome oxidase activity (Rojas et al., 2009).

Normal rats given 1 mg/kg after each holeboard training day made 66 percent of probe visits to baited holes versus 31 percent for saline controls, and in brain homogenates low but not high concentrations sped cytochrome c oxidation (Callaway et al., 2004). Models: rats; animal data only.

Tau aggregation and the hydromethylthionine trials

In a cell-free assay, methylene blue blocked tau-tau binding through the repeat domain, at higher concentrations than likely reachable clinically (Wischik et al., 1996). Phase 3 trials sponsored by the developer, TauRx, then tested LMTM, a stable reduced form of the methylthioninium moiety. In 891 people with mild to moderate Alzheimer's disease, two higher twice-daily amounts did not differ on either co-primary outcome from a low amount kept to mask urine discoloration (Gauthier et al., 2016).

In 800 people with mild disease, there was again no difference as randomized; revised analyses of nonrandomized monotherapy subgroups were significant, and the authors called for another randomized trial (Wilcock et al., 2018). In 598 amyloid-positive participants, hydromethylthionine mesylate (HMTM, the drug's current name) did not separate from a low methylthioninium chloride control on co-primary endpoints at 52 weeks, which the authors attributed to activity in the control arm (Wischik et al., 2026). Models: cell-free assay and human randomized trials.

Photodynamic and antimicrobial lab work

Methylene blue is a photosensitizer, and light-activated methylene blue and related dyes have been applied in antimicrobial work, especially the disinfection of blood products (Wainwright and Crossley, 2002). In Candida albicans strains that overexpress efflux pumps, ABC-type pumps cut methylene blue uptake and light-activated killing more than MFS-type pumps, and the ABC inhibitor verapamil raised both (Prates et al., 2011). Model: fungal cultures.