Dietary inorganic nitrate improves mitochondrial efficiency in humans

Journal Title (Medline/Pubmed accepted abbreviation): Cell Metabolism
Year: 2011
Volume: 13
Page numbers: 149-159
doi (if applicable): 10.1016/j.cmet.2011.01.004

Summary of Background and Research Design

Background:Nitrate (NO3-), nitrite (NO2-), and nitric oxide (NO) are dynamically interconverted in the body. There is preliminary evidence that nitrate supplementation can increase the efficiency by which the mitochondria can generate ATP by any of the nitrogen-containing compounds interacting with elements of the electron transport chain within the mitochondrial membrane. The P/O ratio is the amount of ATP produced per unit of oxygen and this ratio is used to assess mitochondrial efficiency.

Hypothesis: Nitrate supplementation would affect mitochondrial function in people performing endurance exercise.

Research design: Double-blind, placebo-controlled, cross-over design.

Subjects:14 healthy subjects, 11 males and 3 females, age 25 ± 1.

Treatments:Sodium nitrate (0.1 mmol/kg per day = 0.5 g nitrate for an 80.6 kg or 178 lb. individual) or an equivalent amount of sodium chloride as a placebo. Supplementation occurred for the 3 days prior to the cycling test.

Experimental protocol: Biopsies were taken from the vastus lateralis (largest muscle in the quadriceps) 90 min after the last nitrate supplement and before the cycling test. Participants cycled at 60-70 rpm at a resistance chosen in regard to their fitness level (100, 120, or 150 W) until steady-state oxygen consumption was achieved (about 10 min). Respiratory gases and heart rate were monitored.

To measure the efficiency by which mitochondria generated ATP, the mitochondria were isolated from muscle cells and incubated in an oxygraph. ADP is added and the rate at which oxygen is consumed (to produce ATP) is analyzed. State 3 respiration is respiration with saturating amounts of substrates and ADP. and reflects the maximal scenario. State 4 respiration with substrates when all ADP has been phosphorylated to ATP. The ratio of (rate of State 3 respiration):(rate of State 4 respiration) is called the respiratory control ratio (RCR) and indicates the coupling between oxygen utilization and ATP generation. Higher ratios signify higher mitochondrial efficiency.

Summary of research findings:
  • The RCR was increased with nitrate supplementation, indicating that ATP production was more efficient within the mitochondria of muscle cells.
  • Nitrate supplementation improved the P/O ratio also indicating higher mitochondrial efficiency. Consistently, the authors also noted an increase in thermodynamic efficiency.
  • Using luminescence, a 24% higher maximal ATP generation in cells from people supplemented with nitrate was observed.
  • One reason mitochondrial efficiency is not 100% is that protons can leak across the mitochondrial membrane and therefore energy from their motion cannot be harnessed. This phenomenon is compensated for by LEAK respiration, or oxygen usage necessary because of the proton leakage. Nitrate supplementation reduced proton leakage.
  • To see if nitrate had a direct effect on mitochondrial ATP production, nitrate was added directly to cell cultures. No changes in mitochondrial bioenergetics were noted.
  • The authors observed that nitrate supplementation significantly (P = 0.009) decreased the expression of the ATP/ADP translocase (ANT), which is a site in the mitochondrial membrane at which proton leaks often occur
  • No change in mitochondrial density was seen after nitrate supplementation, as measured by quantifying the ratio of mitochondrial DNA to nuclear DNA.
  • Nitrate supplementation did not increase nitrotyrosine, a molecule that could indicate adverse nitration reactions. An increase in production of nitrotyrosine would cause safety concerns.
  • During the exercise trial, whole body oxygen consumption was reduced from 1.95 ± 0.09 L/min with the placebo to 1.89 ± 0.1 L/min with nitrate supplementation, indicating more efficient metabolism.

Interpretation of findings/Key practice applications:

Three days of nitrate supplementation increased the efficiency of ATP production by mitochondria from muscle cells of exercising individuals. Accordingly, this increased the amount of ATP in the muscle cells and reduced the oxygen needs of the subjects during endurance exercise.

     It is interesting that this seemingly small amount of nitrate (about 0.5 g per day) caused this effect. This quantity is achievable by consuming whole foods, specifically vegetables such as spinach, beets, and carrots. Cured meats such as bacon and hot dogs are less healthful yet effective ways to consume nitrates.

     More efficient ATP production means that fewer calories are burned to produce the same amount of energy. This is not a benefit for someone who is trying to lose weight. It would be important to quantify this, however, to assess clinical relevance.


The authors did not correlate mitochondrial bioenergetics to performance. Therefore, although significant changes were observed in mitochondrial efficiency, this phenomenon may not relate to increased performance. This study was also executed in untrained individuals who may have different mitochondrial bioenergetics than elite athletes.

Because reduced mitochondrial efficiency is correlated with increased heat production, it would be interesting for a follow up study to investigate if nitrate supplementation reduces heat production in exercising athletes.

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