384 replies to this topic

[niner]

It is highly likely that variability in oral absorbtion is determined largely by the efficiency of initial reduction in the GI tract. One way to achieve more reliable absorbtion is therefore be to pre-reduce methylene blue with ascorbic acid. We have shown from in vitro studies that this conversion is rather slow, so that it takes 3 hours to achieve 90% reduction of methylene blue in water in the presence of 2× mg ratio of ascorbic acid. Therefore, the dosage of methylene blue which is most likely to ensure reliable absorbtion will be 3.5 mg/kg/day of methylene blue pre-reduced for at least 3 hours in the presence of 7 mg/kg/day of ascorbic acid.

This means that (according to them, anyway) if you want whatever improvement ascorbate can give you, you can't just swallow a vitamin C tablet at the same time you take the MB. You need to pre-incubate the two compounds in solution for several hours first. Considering the small doses that we're using, we could probably use a large excess of ascorbate to accelerate the reaction. To be honest, I'm kind of baffled that this should take any time at all; this sort of redox reaction is usually very fast. Also, I would think that if you see the color change, then it's done. Note that these guys are trying to get a patent on the use of leucomethylene blue as a therapeutic agent, so they are probably inclined to make it sound tricky in order to flummox the hapless patent examiner. If someone happens to have some MB and some ascorbate handy, mix a little together and let us know what happens.

There's another catch though. Leucomethylene blue is only soluble in acidic solutions (pH below 4.5ish?). So you may need to use ascorbic acid as opposed to ascorbate.
I wonder if putting a thin layer of ascorbic acid at the bottom of the diluted methylene blue solution would keep the pH sufficiently low.

http://jcs.biologist...26/113.full.pdf
page 5

Thanks for that ref. Aren't old papers great? They also pointed out that mono-, di-, and tri-methyl compounds are common impurities in MB. I don't know if that's still true today; I suppose we could try a little homebrew paper chromatography. Anyway, yeah, it looks like you want pH 4.5 or so; probably best to just have some excess ascorbic acid around in order to keep it soluble. Once it hits your stomach, you won't have to worry any more. Or maybe it's safer to just take the oxidized MB, at least that way you know it's getting in.

[rwac]

Thanks for that ref. Aren't old papers great? They also pointed out that mono-, di-, and tri-methyl compounds are common impurities in MB. I don't know if that's still true today; I suppose we could try a little homebrew paper chromatography. Anyway, yeah, it looks like you want pH 4.5 or so; probably best to just have some excess ascorbic acid around in order to keep it soluble. Once it hits your stomach, you won't have to worry any more. Or maybe it's safer to just take the oxidized MB, at least that way you know it's getting in.

Yeah, old papers are really good. Wonder why we dont see papers like this anymore. Perhaps people no longer care about making their papers accessible to the general public.

The same paper also indicates that these undermethylated compounds have a reduced affinity to nerve cells. These are all oxidation products of MB, maybe this is why Methylene Blue stops working, it partly oxidizes into Azure A,B,C and Thionine.

Please do post pictures if you do decide to do the chromatography.

The lowest dose in mice was 2.5 mg/kg. For a 70kg human that would be 175 mg without allometric scaling, or about 15 mg/day if you scale it. At this dose, the mice lived longer than controls. In fact, the mice lived longer than controls at all doses, even if there was a lymphoma trend. (Mice are cancer-prone.) The clinical trial that MarkyMark posted was 200mg/d for the active arm, and I think 16mg for the 'placebo' (lol). The placebo group probably felt better than the actives, which might be why I couldn't find a publication...

I wonder what researchers would make of a study where the placebo was more effective than the actives. Would they figure out that the dosage was wrong, or would they just put it in the circular file?

[rwac]

An overview of methylene blue.

"Lest we forget you — methylene blue …"

"MB was the very first fully synthetic drug used in med-
icine."

Also introducing selenomethylene blue, similar to, and more active than methylene blue itself.

Attached Files

•  Schirmer__Mandelkow_2011_NeurobiolAging_MethyleneBlue.pdf   648.42KB   46 downloads

Edited by rwac, 17 August 2011 - 12:14 AM.

[rwac]

I just made some Leuco-MB, and have been taking it for the last few doses instead of MB. I find that it doesn't have the same nootropic effect, I got some sort of brain fog after a few doses.

Here's 2 possibilities.

1. The solution is almost transparent, so the Leuco-MB may have precipitated out.

2. Leuco-MB doesn't prefer neural cells, so it partitions differently, resulting in a lower concentration in the brain.

[niner]

I just made some Leuco-MB, and have been taking it for the last few doses instead of MB. I find that it doesn't have the same nootropic effect, I got some sort of brain fog after a few doses.

Here's 2 possibilities.

1. The solution is almost transparent, so the Leuco-MB may have precipitated out.

2. Leuco-MB doesn't prefer neural cells, so it partitions differently, resulting in a lower concentration in the brain.

Hmm. Leuco-MB is supposed to cross the BBB better than oxidized MB, at physiological pH. This makes me lean toward option 1. Was the solution acidic enough to protonate both amines? There probably wasn't enough compound to see a precipitate very easily, but did it look crystal clear, or was it slightly cloudy?

[rwac]

I just made some Leuco-MB, and have been taking it for the last few doses instead of MB. I find that it doesn't have the same nootropic effect, I got some sort of brain fog after a few doses.

Here's 2 possibilities.

1. The solution is almost transparent, so the Leuco-MB may have precipitated out.

2. Leuco-MB doesn't prefer neural cells, so it partitions differently, resulting in a lower concentration in the brain.

Hmm. Leuco-MB is supposed to cross the BBB better than oxidized MB, at physiological pH. This makes me lean toward option 1. Was the solution acidic enough to protonate both amines? There probably wasn't enough compound to see a precipitate very easily, but did it look crystal clear, or was it slightly cloudy?

It's as acidic as saturating it with vit C, with leftover powder at the bottom would make it.

I'm using a brown bottle so it's not easy to tell. But I can see little wiggles running through it if that makes sense. No obvious cloudiness though.

Ah, but Leuco-MB stains all tissue, while MB only stains neural and muscle tissue to some extent (ref: Harris and Peters). So LMB may no longer have that preference for neural cells, and distribute across the entire body ...

In pieces of excised tissue, and in whole organisms where surface damage
has produced similar conditions, oxidized methylene blue stains nerve cells
and fibres (and muscle cells, though less strongly), with increasing speed and
intensity as the pH of the staining solution is raised. In such experiments the
dye penetrates along intracellular pathways in the nervous tissue, not crossing
cell boundaries except at functional (synaptic ?) contacts, which may well re-
present regions of alteration in properties of the membrane. A simple, though
not necessarily correct, explanation of these facts could be made on the
assumption that the free dye cation of the completely ionized methylene blue
salt combined with negatively charged proteins (Singer, 1952).

Reduced methylene blue penetrates and stains such tissues, as well as un-
damaged whole organisms, much more readily, crossing cell-membrane bar-
riers in the process. It is suggested that this penetration occurs in the form
of uncharged molecules of leucomethylene blue. Below pH 5 the rate of pene-
tration falls of f very rapidly; this result agrees with a corresponding diminu-
tion in the concentration of uncharged leucomethylene blue, which ionizes at
more acid values to give successively a singly and a doubly charged cation.

[niner]

I just made some Leuco-MB, and have been taking it for the last few doses instead of MB. I find that it doesn't have the same nootropic effect, I got some sort of brain fog after a few doses.

Here's 2 possibilities.

1. The solution is almost transparent, so the Leuco-MB may have precipitated out.

2. Leuco-MB doesn't prefer neural cells, so it partitions differently, resulting in a lower concentration in the brain.

Hmm. Leuco-MB is supposed to cross the BBB better than oxidized MB, at physiological pH. This makes me lean toward option 1. Was the solution acidic enough to protonate both amines? There probably wasn't enough compound to see a precipitate very easily, but did it look crystal clear, or was it slightly cloudy?

It's as acidic as saturating it with vit C, with leftover powder at the bottom would make it.

I'm using a brown bottle so it's not easy to tell. But I can see little wiggles running through it if that makes sense. No obvious cloudiness though.

Ah, but Leuco-MB stains all tissue, while MB only stains neural and muscle tissue to some extent (ref: Harris and Peters). So LMB may no longer have that preference for neural cells, and distribute across the entire body ...

I'm pretty sure that saturated ascorbic acid should do the trick. The leftover powder and the brown glass complicate the visual issue. In the body, a significant fraction of ox-MB gets reduced, and the redox cycle is pretty facile, so some of the LMB ought to go in the other direction. I've never seen anything about the pharmacokinetics of oral LMB. Like you say, it should more happily partition into lipids of all sorts, but it's certainly possible that it just has a problem getting in through the GI tract. At this point, bad PK is my best guess... I have some MB in the mail; I'll experiment with it when it comes in.

[unregistered_user]

I seem to recall Methylene Blue being more effective when taken with Vitamin C. Does anyone know the reason this synergy? Can I just go pick up a Vit C supplement at the store for this? Is this proven or just speculation?

[MrHappy]

I seem to recall Methylene Blue being more effective when taken with Vitamin C. Does anyone know the reason this synergy? Can I just go pick up a Vit C supplement at the store for this? Is this proven or just speculation?

It's covered in the recent bunch of posts regarding leucomethylene blue .. ascorbate or ascorbic acid transforms MB into LMB, which is absorbed more efficiently. The thing is, you need to mix the MB with ascorbic acid, wait until it changes colour (should turn clear) and then consume it.

[niner]

QUESTION: when I made up the solution, adding 1.3 ml of 2.303% MB (Kordon) to 500 ml water, it was still quite blue. Bright, dense blue. I added vitamin C, and got it to a pale blue, but still blue.

So, those people who have said that a 60 mcg/ml solution should be colorless, do you really know or are you guessing? If you're guessing, for crissakes keep your guesses to yourself. If you're not, then someone please explain to me why my 60 mcg/ml solution is so blue?

I think that the people who said it was colorless (or very pale) were talking about 60mcg in a much larger volume of water; maybe a 2-300 fold dilution over your solution.

[rwac]

I'm taking MB 250mcg x5/day.

Methylene Blue is a vasoconstrictor, so you might want to reduce the dose for a bit.

http://www.springerl...49aeb5e58jg4pp/

Pressor and vasoconstrictor effects of methylene blue in endotoxaemic rats

Xing Cheng and Catherine C. Y. Pang

Nitric oxide (NO) is a primary mediator of hypotension in sepsis. We examined the effects of methylene blue (MB), an inhibitor of the NO/cGMP pathway, on mean arterial pressure (MAP), cardiac output (CO), total peripheral resistance (TPR), mesenteric blood flow (MBF) and renal blood flow (RBF) in pentobarbitone-anaesthetised rats injected with lipopolysaccharide (LPS, 7.5 mg/kg). MB (1, 3 or 10 mg/kg.h) or vehicle was i.v. infused into four groups at 2.5 h after i.v. injection of LPS. Two other groups received MB or vehicle at 2.5 h after receiving saline. LPS reduced MAP, CO, RBF as well as MBF at 2.5 and 4 h, and increased TPR at 2.5 but not 4 h. Whereas MB alone had no effects on measured variables in control rats at 4 h, in LPS-treated rats, it elevated TPR at all doses and attenuated the fall in MAP at the two low doses. CO was unaltered by low doses of MB but reduced by the high dose. MBF was unaltered, but RBF was increased by the lowest dose but decreased by the highest dose of MB. Therefore, in endotoxaemia, a low dose of MB increases MAP and TPR but does not alter CO; a high dose of MB does not raise MAP but increases TPR and reduces CO.

Note that the "Low" dose was 1mg per kg per hour.

[niner]

There's some missing info here. How much ascorbic acid? How much water? (I assume there's water; MB is a solid in its pure state.) How long did you let it sit before you consumed it? Supposedly, it takes 3 hours for some sort of equilibration to occur. Finally, what does MB on it's own feel like for you? Can you tell if you've taken it?

Probably 800mg-1g of ascorbic asid. somewhere 50-100 ml of water. It did not sit at all - I swallowed it at once after it lost its color. So should I wait for 3 hours before taking?
After taking MB i feel like my mood increases and I become more energetic. After taking MB with ascorbic acid I did not feel any change in mood -

There is a report in the literature that you need to wait 3 hours, though they don't say why. I have a hypothesis as to what might be the reason: When the MB is first reduced by the ascorbate, it is possible that it aggregates, since it initially is converted to a neutral, hydrophobic molecule. It should, at least if it's at a pH of 4.5 or less, get protonated, and become soluble. If it aggregates at first, this step might be slow. If the pH is much above 4.5, it might not happen at all. Without doing any calculations, I think that your pH should be ok with the amount of water/vitamin C you're using, though it's possible that you need more acid. Well, anyway, that's my educated guess. You might try using more acid and waiting 3 hours.

[Brainbox]

This company is developing a drug called "rember" that is based on MB. This drug is said to target TAU aggregation to fight alzeimer. Reading the site, the drug seems to be in development, but there's a lead to more information in the resources section of the site.

Edited by Brainbox, 30 August 2011 - 09:13 PM.

[MrHappy]

Incidently, people mixing MB with any of the racetam family are wasting the nitric oxide synthase inhibitor effect of MB. Racetams elevate nitric oxide levels, leading to earlier neuron death. Your bulb burns brighter, but wears out faster. Kind of the opposite of the immortality project, really..

[kassem23]

Incidently, people mixing MB with any of the racetam family are wasting the nitric oxide synthase inhibitor effect of MB. Racetams elevate nitric oxide levels, leading to earlier neuron death. Your bulb burns brighter, but wears out faster. Kind of the opposite of the immortality project, really..

Do you have any evidence of this AT ALL? It's ridiculous to state that 'racetams induce neuronal death (neuronal apoptosis) without any backing at all. There's no single study in PubMed that I've read that has said anything near what you just said. OTOH, they all seem to agree that Piracetam is protective of neuronal insult, etc.

[MrHappy]

Incidently, people mixing MB with any of the racetam family are wasting the nitric oxide synthase inhibitor effect of MB. Racetams elevate nitric oxide levels, leading to earlier neuron death. Your bulb burns brighter, but wears out faster. Kind of the opposite of the immortality project, really..

Do you have any evidence of this AT ALL? It's ridiculous to state that 'racetams induce neuronal death (neuronal apoptosis) without any backing at all. There's no single study in PubMed that I've read that has said anything near what you just said. OTOH, they all seem to agree that Piracetam is protective of neuronal insult, etc.

Sure. This is a no-brainer.. literally. <pun intended>
http://www.longecity...6-pramiracetam/

Pramiracetam increases nitric oxide synthase (NOS) activity in the cerebral cortex of the rat [1]. NOS is the enzyme that catalyzes the formation of NO from oxygen and arginine is nitric oxide synthase [2].
---
Moving right along...

Nitric oxide synthase is the primary signaller to induce cell death. It is the untimey 'killing stroke' in Parkinson's, Alzheimer's, Huntington's, ALS and practically every other neurodegenerative disease. Nitric oxide synthase inhibitors, such as MB and many others (genestein, allicin,aspirin,DHA, gingko baloba, silymarin, aminoguanidine, etc.), protect against cell death. The body normally keeps NOS in balance, but artificially elevating NOS is going to lead to the untimely death of some very useful neurons.

http://www.ncbi.nlm....pubmed/12594242
http://www.ncbi.nlm..../pubmed/9436549
http://www.google.co...thase apoptosis

<Ta-dah!>

[aaaa]

I have a question.If methylene blue really works as a nootropic.How could it not be used as a treatment for Dementia.
Although MB has been widely used by doctor for anoyher purpose like antidote or placebo for decades.
If it really work, it should be researched more ealier.Because someone of the patient wii notice the effect on mental.
Why? I really wanna know.

[manic_racetam]

(In the spirit of stopping unnecessary misinformation)

I hate to butt in here but first of all, the thread you're referencing is specific to high dose pramiracetam. You originally said "people mixing MB with any of the racetam family are wasting the nitric oxide synthase inhibitor effect". Can you produce studies on "any [other] of the racetam family" that show an increase in NOS?

Also, it's in the full abstract of the thread you quoted that "A lower dose of pramiracetam (100 mg/kg i.p.) was ineffective on NOS mRNA expression and enzyme activity". And it's uncommon for people to even dose at levels that would cause the NOS reaction (at least based on the rat study).

Edited by manic_racetam, 02 September 2011 - 10:47 PM.

[MrHappy]

Thinking about this, I can see why MB is an ideal treatment for neurodegenerative diseases:
1) NOS-I - stops/delays neuron death
2) MAO-I - elevates dopamine levels for Parkinson's.
3) Antibacterial/fungal etc and crosses the BBB easily. If it deactivates HSV1, then very, very useful for any virally caused AD and PD.
4) Untangles tau proteins.
5) Enhances neural pathways.

The only concern is the NOS-I possibly preventing apoptosis when it IS actually necessary, eg. carcinoma, neuroblastoma etc. The rat studies showed increased rates of lymphoma proportional to the dose level of MB. Since the primary absorption is the brain, lymph nodes, etc., this may explain why it was lymphoma and not other forms of cancer. It would possibly go into remission naturally when normal NOS function is restored. Perhaps this can be avoided entirely by localised or minimal doses.

[MrHappy]

(In the spirit of stopping unnecessary misinformation)

I hate to butt in here but first of all, the thread you're referencing is specific to high dose pramiracetam. You originally said "people mixing MB with any of the racetam family are wasting the nitric oxide synthase inhibitor effect". Can you produce studies on "any [other] of the racetam family" that show an increase in NOS?

Also, it's in the full abstract of the thread you quoted that "A lower dose of pramiracetam (100 mg/kg i.p.) was ineffective on NOS mRNA expression and enzyme activity". And it's uncommon for people to even dose at levels that would cause the NOS reaction (at least based on the rat study).

Cool, now use allometric scaling to convert 100mg/Kg in a rat to a human dosage. It equals approx 25mg/Kg.. So for a 65Kg person, 1625mg is the dose that NOS activity increases. Isochroma would have been doing himself some damage. Also you are correct - Pramiracetam appears to be the only NOS elevator. I apologise, I over-extrapolated. The others appear to be safe or even beneficial for oxidative stress damage.

[manic_racetam]

Cool, now use allometric scaling to convert 100mg/Kg in a rat to a human dosage. It equals approx 25mg/Kg.. So for a 65Kg person, 1625mg is the dose that NOS activity increases. Isochroma would have been doing himself some damage. Also you are correct - Pramiracetam appears to be the only NOS elevator. I apologise, I over-extrapolated. The others appear to be safe or even beneficial for oxidative stress damage.

If anyone needs a simple allometric scaling calculator to do these conversions from animal to human dosages here is a nice free online version.

Thanks for reminding me of that. There are people that take prami in those dosages even though personally I consider that quite high and unnecessary. Nice seeing you around here.

[MrHappy]

If anyone needs a simple allometric scaling calculator to do these conversions from animal to human dosages here is a nice free online version.

Thanks for reminding me of that. There are people that take prami in those dosages even though personally I consider that quite high and unnecessary. Nice seeing you around here.

<chuckle> Thanks!

I'd definitely be concerned about anyone dosing that high. Mind you, supplementing at least 2g of L-Lysine with that much prami should counter the effect (more, if you eat a lot of L-Arginine rich foods - chocolate, etc). The NOS is synthesised from L-Arginine, which gets 'booted out' of cells by L-Lysine.

[longevitynow]

You could apply it topically with ascorbic acid without looking like a smurf.

Yes, I've read the web posting from Thailand saying that vitamin C makes the blue color of MB go away, but I have tried it and it doesn't. Get some MB and powdered Ascorbic acid, mix in water and observe no lessening of blue color. Not that there might not be some synergy between the two,but it doesn't make color go away in my experience.

[rwac]

Yes, I've read the web posting from Thailand saying that vitamin C makes the blue color of MB go away, but I have tried it and it doesn't. Get some MB and powdered Ascorbic acid, mix in water and observe no lessening of blue color. Not that there might not be some synergy between the two,but it doesn't make color go away in my experience.

You have to give it a few hours, and the blue color will go away.

[wolfeye]

The compound methylene blue, which has a variety of uses, markedly reduces residual symptoms of depression and mania in patients with bipolar disorder, according to results of a phase 3 study released here at the 24th Congress of the European College of Neuropsychopharmacology.

http://www.medscape....warticle/749164

[ta5]

http://www.medscape....warticle/749164

Patients were randomized to 13 weeks of treatment or either low-dose methylene blue (15 mg daily), which was a surrogate for placebo, or high-dose methylene blue (195 mg daily), and then switched to the alternate treatment for the next 13 weeks. Because methylene blue stains urine, it was not possible to use a standard placebo.

[EncyclopediaBrown]

From the article above:

Patients were randomized to 13 weeks of treatment or either low-dose methylene blue (15 mg daily), which was a surrogate for placebo, or high-dose methylene blue (195 mg daily), and then switched to the alternate treatment for the next 13 weeks

The effect of methylene blue on cognitive symptoms, like verbal memory, was weak and only marginally significant. Importantly, the compound did not worsen any of the cognitive symptoms.

Our main finding was that the active dose of methylene blue was very effective for residual problems, with patients feeling better subjectively and on objective measures, including the rating scales for anxiety and depression.


Question:

Do you believe that nootropical effect was nullified due to the high dosage?

Edited by EncyclopediaBrown, 09 September 2011 - 05:43 PM.

[EncyclopediaBrown]

*Question:

Do you believe that nootropic effect was nullified due to the high dosage?

Edited by EncyclopediaBrown, 09 September 2011 - 07:03 PM.

[MrHappy]

Definitely. Even the 'placebo' was too high

[rwac]

10 patients withdrew from the study, mostly because they disliked the blue staining of their urine

What is so bad about blue urine, that you would leave a clinical trial?