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The Rediscovery of Methylene Blue

James Odell, OMD, ND, L.Ac.


Ever since German chemist Heinrich Caro first created methylene blue (MB) as a dye in 1876, it later became the first fully synthetic drug used in medicine. In 1891 it was applied by Paul Guttmann and Paul Ehrlich for the treatment of malaria but then ceased to be used as an anti-malarial due to its two inevitable side effects: green urine and blue sclera. MB is a dye/medication that also exhibits antioxidant, antimalarial,

antidepressant, nootropic, and cardioprotective properties. Interest in its use has recently been revived especially because of its versatility and effectiveness.1, 2, 3


Throughout many decades, a multitude of usages has been discovered among the various fields of science including clinical medicine and surgery. The famous Giemsa solution for microscopic staining and characterizing malaria parasites and blood cells contains MB, eosin A, and azure B as active components.4, 5


Numerous other microscopic discoveries including the identification of Mycobacterium tuberculosis by Robert Koch and the structural organization of nerve tissues were illuminated by the biochemical properties of MB.6, 7, 8, 9 Interestingly, staining with MB was the beginning of modern drug research. Paul Ehrlich contended that if pathogens like bacteria and parasites are preferentially stained by MB, then this staining might indicate a specific harmful effect on the pathogen which could be exploited for fighting disease. This explains why the terms “drug” and “dye” were used synonymously until World War I.


Going back to the beginning of the twentieth century, MB was used for a wide variety of medical and hygienic indications. Particularly, MB was extensively used as an antiseptic and prescribed as a treatment for malaria and gonorrhea.10


Among others, it was added to the medication of psychiatric patients to study their compliance which could be monitored by the observable color of the urine. These studies led to the discovery that MB has profound antidepressant properties and positive psychotropic effects.11, 12, 13 Consequently, MB became one of the lead compounds for other drugs including chlorpromazine and tricyclic antidepressants.


In the 1920s it proved to be an effective antidote for cyanide poisoning since its reduction potential is like that of oxygen and it can be reduced by components of the electron transport chain.14

Its versatility as an antidote has allowed MB to be used against many other poisons such as cyclophosphamide-induced encephalopathy, the exotic Jamaican ackee fruit poisoning., and to treat hypotension related to lithium toxicity.


In 1925, W. Mansfield Clark, famous for the introduction of the pH electrode and the oxygen electrode, was a co-author of an impressive 80-page review on the application of MB in engineering, industrial chemistry, biology, and medicine. A remarkable aspect of this article is the reference list of illustrious scientists including several Nobel Prize winners (Santiago Ramon y Cajal, Robert Koch, Paul Ehrlich, Alphonse Laveran, Otto Meyerhof, and Heinrich Wieland) who contributed major papers on MB. Many of the MB articles that were published 100 years ago are still relevant today.


It was soon discovered that MB miraculously reversed toxic methemoglobinemia. Methemoglobinemia is a condition when Fe2+ of hemoglobin gets oxidized to Fe3+, reducing the oxygen-carrying capacity of hemoglobin, and typically the patient presents with cyanosis of the lips and extremities, characteristic "chocolate-colored urine," fatigue, shortness of breath with hypoxia. Fe is the chemical symbol for iron. Fe2+ (ferrous) indicates iron in its +2-oxidation state as opposed to Fe3+ (ferric) or iron in its +3-oxidation state. Methemoglobinemia results from exposure to certain drugs such as dapsone, a drug indicated for the treatment of Mycoplasma leprae and Pneumocystis jirovecii prophylaxis, benzocaine (a local anesthetic), high altitude water sources, and nitrites such as nitroglycerin or amyl nitrite used for treating coronary artery disease.


MB is also used to treat the toxic effects of certain chemotherapy drugs as well as circulatory shock, but it is not approved by the FDA for the treatment of these conditions.


By 2010, there were more than 11,000 entries for MB in the biomedical library PubMed, not counting the studies which had been published in the era before PubMed. Current indications for MB that are approved by the FDA are hereditary methemoglobinemia and acute acquired methemoglobinemia, prevention of urinary tract infections in elderly patients, and intraoperative visualization of nerves, nerve tissues, and endocrine glands.15 As a dye, it has also been used for the detection of intestinal, enterovesical, and bronchopleural fistulas. MB can be used to detect lung nodules during thoracoscopic procedures.16 Despite all of this, the most traditional use of MB as a dye remains the detection of sentinel nodes in different cancers 17, 18, 19, 20, 21, 22 as initially proposed by Giuliano et al. for breast cancer in 1994.23


Methylene Blue Biochemistry

Known as a heterocyclic aromatic chemical compound, methylene blue possesses a unique molecular formula that situates it well as an effective medicinal treatment. Methylene blue powder is green in color. It chemically exists as methylene blue chloride (or more technically, methylthioninium chloride). When this compound is added to water or alcohol, it breaks up into the positive methylene blue ion, which has a blue color, and the negative chloride ion, which is colorless.


Methylene Blue in Powder Form


Methylene Blue in Solution


MB is an inhibitor of nitric oxide synthase and guanylate cyclase. Nitric oxide synthases (NOS) are a family of isoforms responsible for the synthesis of the potent dilator nitric oxide (NO). Nitric oxide stimulates soluble guanylate cyclase, which converts guanosine triphosphate into cyclic guanosine monophosphate. Increases in cGMP concentration, in turn, through a cascade of protein kinases, induce smooth muscle relaxation and vasodilation. Methylene blue has direct inhibitory effects on nitric oxide synthases (NOS), both constitutive and inducible, and blocks the accumulation of cyclic guanosine monophosphate (cGMP) by inhibiting the enzyme guanylate cyclase. Also, MB blocks iron-containing enzymes such as xanthine oxidase (XO) and has antioxidant effects. One of the main mechanisms of action of MB is reducing the oxidized form of hemoglobin Fe3+ when in a state of methemoglobinemia, to Fe2+. MB protects organs from the toxic effects of free oxygen radicals by competing with molecular oxygen for the transfer of electrons by XO and is effective in attenuating ischemia-reperfusion syndrome.24, 25


Methylene blue also inhibits the enzymes endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), and guanylate cyclase, decreasing the amount of cGMP needed for nitrous oxide to be released, therefore causing vasoconstriction of blood vessels via inhibition of vascular smooth muscle relaxation.


MB is permeable through biomembranes, and it readily crosses the blood-brain barrier and easily enters brain tissue.26 Additionally, MB inhibits monoamine oxidase (MAO-A) activity, a property that may, at least in part, account for its antidepressant effects in humans. The central inhibition of MAO-A by MB has also been linked to serotonin toxicity which may arise when high-dose of MB is used in combination with serotonergic drugs.27, 28


MB is also an acetylcholine-esterase inhibitor with a preference for muscarinic acetylcholine receptors it prevents the breakdown of acetylcholine and makes this neurotransmitter more available for the central nervous system.29


MB also stimulates glucose metabolism. Taken together, increases in CMRO2 and glucose uptake mean that MB elevates oxygen consumption which helps glucose increase ATP production. Increases in ATP production provide more cellular energy for better overall brain function, including cognition, mood, and memory.

Antioxidant


MB is a powerful antioxidant and as such possesses unique biochemical properties that are neuroprotective and cardiovascular protective. MB has an affinity for tissue oxidases, so it concentrates in mitochondria (the energy-producing powerhouses of cells). There it maintains mitochondrial function by accepting electrons from blocked components of the respiratory chain thus improving mitochondrial respiration. It does this by accepting electrons from NADH and transferring them to cytochrome c. Cytochrome complex (cytochrome c) is a component of the electron transport chain in mitochondria.30

The effects of MB are reflected in increased activity in processes coupled with mitochondrial energy production such as Na/K ATPase activity and intermediary metabolism.31


For this reason, MB constitutes a metabolic cellular energy enhancer that accelerates the activity of the electron transport chain (Krebs Cycle) improving oxidation to the cells.


Inhibits Platelet Aggregation


MB prevents abnormal platelet formation that can lead to heart attacks and strokes. It does this in part by inhibiting the arachidonic acid metabolism in human blood platelets.32 When human platelets are incubated with MB that oxidizes cellular NADPH, a marked inhibition of platelet aggregation and of the metabolism of the arachidonic acid in both the cyclooxygenase and lipoxygenase pathway is found. MB possesses inhibitory effects on platelet activation, adhesion, and aggregation synergistically with inhibition of platelet thromboxane A2 and endothelial prostacyclin I2 (PGI-2) production.33, 34, 35


Thus, MB works on a cellular level within the mitochondria where it affects energy production and may also act as an antioxidant to protect cells against oxidative stress. Since oxidative stress is thought to hasten the aging process in humans, this means that MB’s antioxidant effects may have some anti-aging activity.


Memory Enhancement and Neuroprotection


Methylene blue crosses the blood-brain barrier, and once in the brain, it can reversibly inhibit MAO-A. This means that methylene blue can prevent dopamine and serotonin from being broken down, leading to increased levels of these neurotransmitters.36


Dopamine and serotonin are essential for cognitive function since they play primary roles in memory, focus, learning, mood, and many other aspects of brain health. By increasing levels of these neurotransmitters, methylene blue can provide significant benefits to cognitive performance and overall well-being. Methylene blue has been found to improve memory in many studies.37, 38, 39, 40 41


Recent research has shown that MB is neuroprotective against several neurodegenerative diseases such as Alzheimer’s disease (AD). 42, 43, 44, Parkinson’s disease,45, 46, and even ischemic stroke.47, 48 The relationship between and AD has recently attracted increasing scientific attention since it has been suggested that MB may slow down the progression of this disease. One paper showed that MB can reverse forgetfulness and enhance cognitive function in people with early-stage AD.49


In another study, MB was shown to be effective at preventing age-related memory decline. It was even able to restore some lost memories in aging rats.50 Another study showed that MB was even able to reduce amyloid plaque build-up in the brains of mice with Alzheimer’s disease. Amyloid plaque build-up is a hallmark sign of Alzheimer’s.51 Other studies have shown MB to attenuate the formations of amyloid plaques and neurofibrillary tangles and to partially repair impairments in mitochondrial function and cellular metabolism.52 Multiple studies demonstrate MB’s protective effects on the brain by blocking oxidative free radicals.53, 54


Glutamate is an essential (and the main excitatory) neurotransmitter in the brain and central nervous system (CNS). However, glutamate can become toxic in a process called glutamate excitotoxicity. In certain circumstances, if there is excess glutamate in the brain or the glutamate receptors are overstimulated this can greatly impair important CNS enzymes and proteins. Thus, glutamate excitotoxicity can play a key role in neurodegeneration. It has been demonstrated that MB can protect cells from glutamate excitotoxicity that leads to neurotoxicity.55, 56 This can help prevent or slow down neurodegenerative diseases like AD, Parkinson’s disease, and age-related dementia.


In addition to diseases and conditions like those listed above, MB can prevent and manage brain damage in relation to certain tumors. Alkylating agents such as ifosfamide are used in the treatment of some types of solid tumors but can cause brain damage that may be mitigated by MB.57 It is not entirely clear how MB prevents brain damage in these cases, but it is likely due to its role in the oxidation of NADH and the restoration of mitochondrial enzymes.


Mood Enhancement


As previously discussed, MB was used in early psychiatry as an anti-depressive. Recently, one study found that methylene blue could even effectively improve symptoms of bipolar disorder.58 Methylene blue continues to show promise in treating anxiety and depression primarily by increasing the neurotransmitters dopamine and serotonin. Even more promising, is that one animal study suggested that MB may also be helpful as an alternative treatment for schizophrenia.59


Antimicrobial


Methylene blue is also a photosensitizer, which means that it has increased activity when exposed to certain wavelengths of light and as such can act as an antimicrobial agent. 60, 61 Light-activated MB is used to treat donated blood products as it is effective in killing certain viruses including HIV and herpes.62, 63 Due to its antiviral activity, methylene blue has also been explored as a potential treatment for coronaviruses,64, 65, influenza viruses,66, and antibiotic-resistant biofilm.67, 68, 69


Regarding malaria treatment, MB has been shown to improve the chloroquine response to malaria by decreasing its resistance and inhibiting Plasmodium falciparum glutathione reductase, an enzyme that prevents the formation of byproducts formed by the Plasmodium species that triggers the body's immune response.70, 71, 72, 73


Methylene Blue Administration and Dosage


Methylene blue dosage depends on the condition that is being treated and as with all drug dosages, it should be individually tailored. MB exhibits very different effects at low doses than it does at high doses. They call this the “hermetic dose-response”, where the effects of low dose are opposite than at high dose. The preponderance of scientific evidence shows that a low dose is the best dose for effectiveness while avoiding side effect issues. When doses above 2 mg/kg are taken, MB begins to function as a monoamine oxidase inhibitor (MAOI), which increases the action of serotonin and can lead to unwanted side effects.

Orally: 0.5 to 4 mg/kg is the common amount used in both human and animal studies.

IV: 0.1 to 0.2 mL per kilogram of body weight is a common amount used in both human and animal studies.


Methylene blue injection 1% is a sterile solution of Phenothiazin-5-ium, 3, 7-bis (dimethylamino)-, chloride, trihydrate, and is indicated for drug-induced methemoglobinemia. It must be administered intravenously very slowly over a period of several minutes. Administration of methylene blue for both children and adults experiencing methemoglobinemia is performed intravenously at a dose of 1 mg/kg of a 1% solution over 5 to 30 minutes.


Only a very small dose is required for nitric oxidation inhibition and to scavenge free-floating nitric oxide in the blood. Most clinicians use 1-2 mg of MB per day for conditions listed in this article. However, doses can range up to 10 mg per day for certain conditions and individuals. Generally, the dosage theme is “less is more”.


With a 1% solution, each drop contains 0.5 mg of MB. which means to achieve 2 mg it would take 4 drops; to achieve 10 mg you would need 20 drops. Higher doses should be divided into 2 or 3X daily. Some clinicians recommend higher doses, but this should always be monitored by a licensed professional familiar with MB.


Purchasing Methylene blue


It is important to purchase pharmaceutical-grade MB to avoid ingesting any impurities such as toxic metals.


Safety Considerations


Methylene blue is generally safe and well-tolerated in the recommended dosage of <2 mg/kg; however, when levels >7 mg/kg are used, many of the adverse effects it exhibits may occur.74


Potential side effects may include nausea, vomiting, dizziness, headache, anxiety, and confusion. Most of these side effects are mild and go away after a few days or weeks. Adverse symptoms from oral usage may also include green-colored urine.


However, it is essential to consult with a healthcare practitioner versed in MB therapy before starting a course of treatment. Particularly, if there are any preexisting conditions that could worsen these side effects. Methylene blue is toxic in high doses. So, it is essential not to take more than the recommended dose – doing so could lead to serious health problems.


MB is not to be used during pregnancy. There is epidemiologic evidence that methylene blue is a teratogen, and the drug can potentially cause fetal harm if administered during pregnancy.75

Patients can experience slight pain after intravenous injections, which usually resolves after flushing the access site with saline. If side effects occur, the administration of MB should stop immediately and promptly followed by supportive care, although severe anaphylactic shock with MB given IV is quite rare.


Serotonin syndrome has been found to occur when combining serotonergic agents with methylene blue at a dose of 5 mg/kg. MB use also requires caution in patients with renal failure due to its ability to reduce renal blood flow. Also, as noted in adverse effects, patients taking any drug with serotonergic activity such as SSRIs should avoid the administration of methylene blue due to the risk of serotonin syndrome.


Additionally, avoid MB in combination with the following drugs and supplements:

  • 5-HTP

  • Vyvanse (lisdexamfetamine)

  • Wellbutrin XL (bupropion)

  • Norco (acetaminophen / hydrocodone)

Conclusion

Methylene blue has been used in the field of medicine for decades as a medical imaging dye and a therapeutic drug. The compound is considered a mild antiseptic, so it can be used to kill pathogenic bacteria, viruses, and parasites in the body, particularly in conjunction with other treatments. Additionally, the compound can be used to treat memory problems, Alzheimer’s disease, Parkinson’s disease, and other neurodegenerative conditions. It has also been shown to be useful for mood and memory enhancement. It is an effective antidote to several poisons such as cyanide, and carbon monoxide poisoning, and helps mitigate the toxic side effects of certain cancer chemotherapy agents.

While several applications for MB treatment have already been identified, continued research will likely reveal even more usages. Due to its wide safety profile and versatility, MB shows tremendous promise for the treatment of numerous conditions.


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