Reduction in blood pressure following acute dietary nitrate ingestion is correlated with increased red blood cell S-nitrosothiol concentrations.

Dietary nitrate (NO3-) supplementation can enhance nitric oxide (NO) bioavailability and lower blood pressure (BP) in humans. The nitrite concentration ([NO2-]) in the plasma is the most commonly used biomarker of increased NO availability. However, it is unknown to what extent changes in other NO congeners, such as S-nitrosothiols (RSNOs), and in other blood components, such as red blood cells (RBC), also contribute to the BP lowering effects of dietary NO3-. We investigated the correlations between changes in NO biomarkers in different blood compartments and changes in BP variables following acute NO3- ingestion. Resting BP was measured and blood samples were collected at baseline, and at 1, 2, 3, 4 and 24 h following acute beetroot juice (∼12.8 mmol NO3-, ∼11 mg NO3-/kg) ingestion in 20 healthy volunteers. Spearman rank correlation coefficients were determined between the peak individual increases in NO biomarkers (NO3-, NO2-, RSNOs) in plasma, RBC and whole blood, and corresponding decreases in resting BP variables. No significant correlation was observed between increased plasma [NO2-] and reduced BP, but increased RBC [NO2-] was correlated with decreased systolic BP (rs = -0.50, P = 0.03). Notably, increased RBC [RSNOs] was significantly correlated with decreases in systolic (rs = -0.68, P = 0.001), diastolic (rs = -0.59, P = 0.008) and mean arterial pressure (rs = -0.64, P = 0.003). Fisher's z transformation indicated no difference in the strength of the correlations between increases in RBC [NO2-] or [RSNOs] and decreased systolic blood pressure. In conclusion, increased RBC [RSNOs] may be an important mediator of the reduction in resting BP observed following dietary NO3- supplementation.

15 N-labeled dietary nitrate supplementation increases human skeletal muscle nitrate concentration and improves muscle torque production.

AIM: Dietary nitrate (NO3 - ) supplementation increases nitric oxide bioavailability and can enhance exercise performance. We investigated the distribution and metabolic fate of ingested NO3 - at rest and during exercise with a focus on skeletal muscle. METHODS: In a randomized, crossover study, 10 healthy volunteers consumed 12.8 mmol 15 N-labeled potassium nitrate (K15 NO3 ; NIT) or potassium chloride placebo (PLA). Muscle biopsies were taken at baseline, at 1- and 3-h post-supplement ingestion, and immediately following the completion of 60 maximal intermittent contractions of the knee extensors. Muscle, plasma, saliva, and urine samples were analyzed using chemiluminescence to determine absolute [NO3 - ] and [NO2 - ], and by mass spectrometry to determine the proportion of NO3 - and NO2 - that was 15 N-labeled. RESULTS: Neither muscle [NO3 - ] nor [NO2 - ] were altered by PLA. Following NIT, muscle [NO3 - ] (but not [NO2 - ]) was elevated at 1-h (from ~35 to 147 nmol/g, p < 0.001) and 3-h, with almost all of the increase being 15 N-labeled. There was a significant reduction in 15 N-labeled muscle [NO3 - ] from pre- to post-exercise. Relative to PLA, mean muscle torque production was ~7% greater during the first 18 contractions following NIT. This improvement in torque was correlated with the pre-exercise 15 N-labeled muscle [NO3 - ] and the magnitude of decline in 15 N-labeled muscle [NO3 - ] during exercise (r = 0.66 and r = 0.62, respectively; p < 0.01). CONCLUSION: This study shows, for the first time, that skeletal muscle rapidly takes up dietary NO3 - , the elevated muscle [NO3 - ] following NO3 - ingestion declines during exercise, and muscle NO3 - dynamics are associated with enhanced torque production during maximal intermittent muscle contractions.

Time course of human skeletal muscle nitrate and nitrite concentration changes following dietary nitrate ingestion.

Dietary nitrate (NO3-) ingestion can be beneficial for health and exercise performance. Recently, based on animal and limited human studies, a skeletal muscle NO3- reservoir has been suggested to be important in whole body nitric oxide (NO) homeostasis. The purpose of this study was to determine the time course of changes in human skeletal muscle NO3- concentration ([NO3-]) following the ingestion of dietary NO3-. Sixteen participants were allocated to either an experimental group (NIT: n = 11) which consumed a bolus of ∼1300 mg (12.8 mmol) potassium nitrate (KNO3), or a placebo group (PLA: n = 5) which consumed a bolus of potassium chloride (KCl). Biological samples (muscle (vastus lateralis), blood, saliva and urine) were collected shortly before NIT or PLA ingestion and at intervals over the course of the subsequent 24 h. At baseline, no differences were observed for muscle [NO3-] and [NO2-] between NIT and PLA (P > 0.05). In PLA, there were no changes in muscle [NO3-] or [NO2-] over time. In NIT, muscle [NO3-] was significantly elevated above baseline (54 ± 29 nmol/g) at 0.5 h, reached a peak at 3 h (181 ± 128 nmol/g), and was not different to baseline from 9 h onwards (P > 0.05). Muscle [NO2-] did not change significantly over time. Following ingestion of a bolus of dietary NO3-, skeletal muscle [NO3-] increases rapidly, reaches a peak at ∼3 h and subsequently declines towards baseline values. Following dietary NO3- ingestion, human m. vastus lateralis [NO3-] expressed a slightly delayed pharmacokinetic profile compared to plasma [NO3-].

Physiological and performance effects of dietary nitrate and N-acetylcysteine supplementation during prolonged heavy-intensity cycling.

The purpose of this study was to investigate effects of concurrent and independent administration of dietary nitrate (NO3-), administered as NO3--rich beetroot juice (BR; ~12.4 mmol of NO3-), and N-acetylcysteine (NAC; 70 mg·kg-1) on physiological responses during prolonged exercise and subsequent high-intensity exercise tolerance. Sixteen recreationally active males supplemented with NO3--depleted beetroot juice (PL) or BR for 6 days and ingested an acute dose of NAC or maltodextrin (MAL) 1 h prior to performing 1 h of heavy-intensity cycling exercise immediately followed by a severe-intensity time-to-exhaustion (TTE) test in four conditions: 1) PL+MAL, 2) PL+NAC, 3) BR+MAL and 4) BR+NAC. Pre-exercise plasma [NO3-] and nitrite ([NO2-]) were elevated following BR+NAC  and BR+MAL (both P < 0.01) compared with PL+NAC and PL+MAL; plasma [cysteine] was increased in PL+NAC  and BR+NAC (both P < 0.01) compared to PL+MAL. Muscle excitability declined over time during the prolonged cycling bout in all conditions  but was better preserved in PL+NAC  compared to BR+NAC (P < 0.01) and PL+MAL (P < 0.05). There was no effect of supplementation on subsequent TTE . These findings indicate that co-ingestion of BR and NAC does not appreciably alter physiological responses during prolonged heavy-intensity cycling or enhance subsequent exercise tolerance.

Human skeletal muscle nitrate store: influence of dietary nitrate supplementation and exercise.

KEY POINTS: Nitric oxide (NO), a potent vasodilator and a regulator of many physiological processes, is produced in mammals both enzymatically and by reduction of nitrite and nitrate ions. We have previously reported that, in rodents, skeletal muscle serves as a nitrate reservoir, with nitrate levels greatly exceeding those in blood or other internal organs, and with nitrate being reduced to NO during exercise. In the current study, we show that nitrate concentration is substantially greater in skeletal muscle than in blood and is elevated further by dietary nitrate ingestion in human volunteers. We also show that high-intensity exercise results in a reduction in the skeletal muscle nitrate store following supplementation, likely as a consequence of its reduction to nitrite and NO. We also report the presence of sialin, a nitrate transporter, and xanthine oxidoreductase in human skeletal muscle, indicating that muscle has the necessary apparatus for nitrate transport, storage and metabolism. ABSTRACT: Rodent skeletal muscle contains a large store of nitrate that can be augmented by the consumption of dietary nitrate. This muscle nitrate reservoir has been found to be an important source of nitrite and nitric oxide (NO) via its reduction by tissue xanthine oxidoreductase. To explore if this pathway is also active in human skeletal muscle during exercise, and if it is sensitive to local nitrate availability, we assessed exercise-induced changes in muscle nitrate and nitrite concentrations in young healthy humans, under baseline conditions and following dietary nitrate consumption. We found that baseline nitrate and nitrite concentrations were far higher in muscle than in plasma (∼4-fold and ∼29-fold, respectively), and that the consumption of a single bolus of dietary nitrate (12.8 mmol) significantly elevated nitrate concentration in both plasma (∼19-fold) and muscle (∼5-fold). Consistent with these observations, and with previous suggestions of active muscle nitrate transport, we present western blot data to show significant expression of the active nitrate/nitrite transporter sialin in human skeletal muscle. Furthermore, we report an exercise-induced reduction in human muscle nitrate concentration (by ∼39%), but only in the presence of an increased muscle nitrate store. Our results indicate that human skeletal muscle nitrate stores are sensitive to dietary nitrate intake and may contribute to NO generation during exercise. Together, these findings suggest that skeletal muscle plays an important role in the transport, storage and metabolism of nitrate in humans.

Influence of dietary nitrate supplementation on physiological and muscle metabolic adaptations to sprint interval training.

We hypothesized that 4 wk of dietary nitrate supplementation would enhance exercise performance and muscle metabolic adaptations to sprint interval training (SIT). Thirty-six recreationally active subjects, matched on key variables at baseline, completed a series of exercise tests before and following a 4-wk period in which they were allocated to one of the following groups: 1) SIT and [Formula: see text]-depleted beetroot juice as a placebo (SIT+PL); 2) SIT and [Formula: see text]-rich beetroot juice (~13 mmol [Formula: see text]/day; SIT+BR); or 3) no training and [Formula: see text]-rich beetroot juice (NT+BR). During moderate-intensity exercise, pulmonary oxygen uptake was reduced by 4% following 4 wk of SIT+BR and NT+BR (P < 0.05) but not SIT+PL. The peak work rate attained during incremental exercise increased more in SIT+BR than in SIT+PL (P < 0.05) or NT+BR (P < 0.001). The reduction in muscle and blood [lactate] and the increase in muscle pH from preintervention to postintervention were greater at 3 min of severe-intensity exercise in SIT+BR compared with SIT+PL and NT+BR (P < 0.05). However, the change in severe-intensity exercise performance was not different between SIT+BR and SIT+PL (P > 0.05). The relative proportion of type IIx muscle fibers in the vastus lateralis muscle was reduced in SIT+BR only (P < 0.05). These findings suggest that BR supplementation may enhance some aspects of the physiological adaptations to SIT.NEW & NOTEWORTHY We investigated the influence of nitrate-rich and nitrate-depleted beetroot juice on the muscle metabolic and physiological adaptations to 4 wk of sprint interval training. Compared with placebo, dietary nitrate supplementation reduced the O2 cost of submaximal exercise, resulted in greater improvement in incremental (but not severe-intensity) exercise performance, and augmented some muscle metabolic adaptations to training. Nitrate supplementation may facilitate some of the physiological responses to sprint interval training.

Dietary nitrate improves sprint performance and cognitive function during prolonged intermittent exercise.

UNLABELLED: It is possible that dietary nitrate (NO3 (-)) supplementation may improve both physical and cognitive performance via its influence on blood flow and cellular energetics. PURPOSE: To investigate the effects of dietary NO3 (-) supplementation on exercise performance and cognitive function during a prolonged intermittent sprint test (IST) protocol, which was designed to reflect typical work patterns during team sports. METHODS: In a double-blind randomised crossover study, 16 male team-sport players received NO3 (-)-rich (BR; 140 mL day(-1); 12.8 mmol of NO3 (-)), and NO3 (-)-depleted (PL; 140 mL day(-1); 0.08 mmol NO3 (-)) beetroot juice for 7 days. On day 7 of supplementation, subjects completed the IST (two 40-min "halves" of repeated 2-min blocks consisting of a 6-s "all-out" sprint, 100-s active recovery and 20 s of rest), on a cycle ergometer during which cognitive tasks were simultaneously performed. RESULTS: Total work done during the sprints of the IST was greater in BR (123 ± 19 kJ) compared to PL (119 ± 17 kJ; P < 0.05). Reaction time of response to the cognitive tasks in the second half of the IST was improved in BR compared to PL (BR first half: 820 ± 96 vs. second half: 817 ± 86 ms; PL first half: 824 ± 114 vs. second half: 847 ± 118 ms; P < 0.05). There was no difference in response accuracy. CONCLUSIONS: These findings suggest that dietary NO3 (-) enhances repeated sprint performance and may attenuate the decline in cognitive function (and specifically reaction time) that may occur during prolonged intermittent exercise.

Dietary nitrate supplementation improves team sport-specific intense intermittent exercise performance.

Recent studies have suggested that dietary inorganic nitrate (NO3(-)) supplementation may improve muscle efficiency and endurance exercise tolerance but possible effects during team sport-specific intense intermittent exercise have not been examined. We hypothesized that NO3(-) supplementation would enhance high-intensity intermittent exercise performance. Fourteen male recreational team-sport players were assigned in a double-blind, randomized, crossover design to consume 490 mL of concentrated, nitrate-rich beetroot juice (BR) and nitrate-depleted placebo juice (PL) over ~30 h preceding the completion of a Yo-Yo intermittent recovery level 1 test (Yo-Yo IR1). Resting plasma nitrite concentration ([NO2(-)]) was ~400% greater in BR compared to PL. Plasma [NO2(-)] declined by 20% in PL (P < 0.05) and by 54 % in BR (P < 0.05) from pre-exercise to end-exercise. Performance in the Yo-Yo IR1 was 4.2% greater (P < 0.05) with BR (1,704 ± 304 m) compared to PL (1,636 ± 288 m). Blood [lactate] was not different between BR and PL, but the mean blood [glucose] was lower (3.8 ± 0.8 vs. 4.2 ± 1.1 mM, P < 0.05) and the rise in plasma [K(+)] tended to be reduced in BR compared to PL (P = 0.08). These findings suggest that NO3(-) supplementation may promote NO production via the nitrate-nitrite-NO pathway and enhance Yo-Yo IR1 test performance, perhaps by facilitating greater muscle glucose uptake or by better maintaining muscle excitability. Dietary NO3(-) supplementation improves performance during intense intermittent exercise and may be a useful ergogenic aid for team sports players.