Three different things, often reported as one

A comparison of what NAD+ precursor approaches and methylene blue actually change, what each body of evidence measures, and where the two overlap.

Sources: primary research read for this comparison, cited in full below.
What is being changed NAD+ precursor approaches Methylene blue
Total NAD quantity
pool size
  • The aim of the approach. Precursors feed the salvage pathway and are assembled into NAD+.
  • Animal models show tissue NAD+ repletion fairly consistently.
  • Human trials show raised measured blood NAD+ levels.
  • Not the target, and not observed as an outcome of treatment.
  • In injured tendon cells and in a rat tendinopathy model, the total NAD pool showed no significant change after methylene blue (PMID 42058931).
NAD+/NADH ratio
redox state
  • Not the primary target. Adding substrate can raise reduction, which moves the ratio toward NADH.
  • Pool measurements are reported far more often than ratio measurements in this literature.
  • The direct effect. Methylene blue accepts electrons from NADH and is reoxidised elsewhere, so NADH is emptied even when the total pool is unchanged.
  • Dose-dependent fall in NADH and in the NADH/NAD+ ratio in isolated cardiac mitochondria at 6 to 18 micromolar, with absolute NAD+ far less responsive than NADH (PMID 28303408).
  • Ratio shift is largely independent of sirtuin 3; the downstream deacetylation depends on it (PMID 28303408).
Clinical endpoints
what a trial measures
  • Human trials exist. Blood NAD+ rises. Endpoint results are mixed and generally small.
  • Animal outcomes do not always follow the ratio. Correcting the hepatic NAD+/NADH ratio in ethanol-fed rats did not reduce fatty liver (PMID 4091827).
  • Hepatic steatosis and injury markers improved in high-fat-diet mice (PMID 24486702).
  • Cardiac ejection fraction improved in diabetic rats, while complex I-supported respiration was not corrected (PMID 28303408).
  • In tendon, restoring the ratio did not prevent oxidative stress, matrix degeneration or loss of cell viability when complex I was blocked (PMID 42058931).
  • Human clinical use is acute, monitored and intravenous. No located human study measures NAD+ metabolites after methylene blue in healthy volunteers.
Direct combination evidence None located. No published study, trial, or case series found for this comparison tested methylene blue together with nicotinamide riboside, nicotinamide mononucleotide, or an NAD+ infusion. Combination claims are extrapolation from two separate bodies of evidence.

This comparison separates three quantities that are frequently reported as one. A change in the NAD+/NADH ratio is a statement about redox balance. A change in the total pool is a statement about how much NAD exists. A change in a clinical endpoint is a statement about a person, a tissue, or an animal. Evidence for one is not evidence for the others.