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Both caffeine and beetroot juice have ergogenic effects on endurance cycling performance. We investigated whether there is an additive effect of these supplements on the performance of a cycling time trial (TT) simulating the 2012 London Olympic Games course. Twelve male and 12 female competitive cyclists each completed 4 experimental trials in a double-blind Latin square design. Trials were undertaken with a caffeinated gum (CAFF) (3 mg·kg(-1) body mass (BM), 40 min prior to the TT), concentrated beetroot juice supplementation (BJ) (8.4 mmol of nitrate (NO3(-)), 2 h prior to the TT), caffeine plus beetroot juice (CAFF+BJ), or a control (CONT). Subjects completed the TT (females: 29.35 km; males: 43.83 km) on a laboratory cycle ergometer under conditions of best practice nutrition: following a carbohydrate-rich pre-event meal, with the ingestion of a carbohydrate-electrolyte drink and regular oral carbohydrate contact during the TT. Compared with CONT, power output was significantly enhanced after CAFF+BJ and CAFF (3.0% and 3.9%, respectively, p < 0.01). There was no effect of BJ supplementation when used alone (-0.4%, p = 0.6 compared with CONT) or when combined with caffeine (-0.9%, p = 0.4 compared with CAFF). We conclude that caffeine (3 mg·kg(-1) BM) administered in the form of a caffeinated gum increased cycling TT performance lasting ∼50-60 min by ∼3%-4% in both males and females. Beetroot juice supplementation was not ergogenic under the conditions of this study.
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Single and combined effects of beetroot juice and caffeine
supplementation on cycling time trial performance
Stephen C. Lane, John A. Hawley, Ben Desbrow, Andrew M. Jones, James R. Blackwell, Megan L. Ross,
Adam J. Zemski, and Louise M. Burke
Abstract: Both caffeine and beetroot juice have ergogenic effects on endurance cycling performance. We investigated whether
there is an additive effect of these supplements on the performance of a cycling time trial (TT) simulating the 2012 London
Olympic Games course. Twelve male and 12 female competitive cyclists each completed 4 experimental trials in a double-blind
Latin square design. Trials were undertaken with a caffeinated gum (CAFF) (3 mg·kg
body mass (BM), 40 min prior to the TT),
concentrated beetroot juice supplementation (BJ) (8.4 mmol of nitrate (NO
), 2 h prior to the TT), caffeine plus beetroot juice
(CAFF+BJ), or a control (CONT). Subjects completed the TT (females: 29.35 km; males: 43.83 km) on a laboratory cycle ergometer
under conditions of best practice nutrition: following a carbohydrate-rich pre-event meal, with the ingestion of a carbohydrate–
electrolyte drink and regular oral carbohydrate contact during the TT. Compared with CONT, power output was significantly
enhanced after CAFF+BJ and CAFF (3.0% and 3.9%, respectively, p< 0.01). There was no effect of BJ supplementation when used
alone (–0.4%, p= 0.6 compared with CONT) or when combined with caffeine (–0.9%, p= 0.4 compared with CAFF). We conclude
that caffeine (3 mg·kg
BM) administered in the form of a caffeinated gum increased cycling TT performance lasting 50– 60 min
by 3%–4% in both males and females. Beetroot juice supplementation was not ergogenic under the conditions of this study.
Key words: cycling performance, nitrate, caffeine, ergogenic, time trial, carbohydrate.
Résumé : La caféine et le jus de betterave ont des effets ergogènes sur la performance d’endurance en cyclisme. Dans cette étude,
on examine l’effet additif de ces suppléments sur la performance dans une course contre-la-montre (« TT ») simulant le parcours
aux Jeux olympiques de Londres de 2012. Douze femmes et douze hommes, tous des cyclistes de compétition, participent a
essais expérimentaux, et ce, selon la méthode du carré latin et a
`double insu. Les sujets participent aux essais dans les conditions
suivantes : gomme renfermant de la caféine (« CAFF »; 3 mg·kg
masse corporelle (« BM »), 40 min avant la TT), supplément de jus
de betterave concentré (« BJ »; 8,4 mmol de NO
, 2 h pré-TT), caféine plus jus de betterave (« CAFF+BJ ») et contrôle (« CONT »). Les
sujets réalisent la TT (femmes : 29,35 km, hommes : 43,83 km) dans un laboratoire sur un cycloergomètre dans un contexte d’une
pratique nutritive standard: repas précompétitif riche en sucres, apport d’une boisson contenant des sucres et des électrolytes
et consommation régulière de sucreries durant la TT. Comparativement a
`la condition de contrôle, on observe une augmentation
de la puissance générée dans les conditions CAFF+BJ et CAFF (3,0 % et 3,9 % respectivement, p< 0,01). Consommé seul, le jus de
betterave n’a pas d’effet (–0,4 %, p= 0,6; comparativement a
`CONT) ou en combinaison avec la caféine (–0,9 %, p= 0,4;
comparativement a
`CAFF). En conclusion, la caféine (3 mg·kg
BM) administrée sous forme de gomme suscite une amélioration
de la performance (50– 60 min) de 3–4 % chez des femmes et des hommes dans un contre-la-montre. [Traduit par la Rédaction]
Mots-clés : performance cycliste, nitrate, caféine, ergogène, contre-la-montre, sucre.
Athletes continually strive to improve training capacity and
performance. Not surprisingly, widespread use of a large number
of nutritional supplements is commonplace in most sports as
athletes search for a “magic bullet” that will elevate their perfor-
mance to a higher level. Both caffeine (Desbrow et al. 2009;Irwin
et al. 2011;Lane et al. 2013a) and nitrate (NO
)(Cermak et al.
2012a;Lansley et al. 2011a;Vanhatalo et al. 2011) have been shown
to improve simulated road cycling performance in a variety of
protocols. Through mechanisms likely related to the central ner-
vous system (CNS) (Costill et al. 1978;Tarnopolsky 2008), caffeine
has been shown to improve arousal states (Backhouse et al. 2011)
and reduce perceived exertion during steady-state exercise (Backhouse
et al. 2011;Doherty and Smith 2005;Lane et al. 2013a), resulting in
enhanced performance during sustained high-intensity cycling
events (Cox et al. 2002;Lane et al. 2013a;McNaughton et al. 2008).
Contemporary protocols for caffeine use are based on evidence
that moderate intakes (3 mg·kg
) of caffeine are equally as effec-
tive as larger doses (6 mg·kg
)(Desbrow et al. 2012) for eliciting
these CNS effects, and that caffeinated gums can also provide a
Received 21 July 2013. Accepted 11 October 2013. Correction after posting 10 March 2014.
S.C. Lane. Exercise and Nutrition Research Group, School of Medical Sciences, RMIT University, Bundoora, VIC 3083, Australia.
J.A. Hawley.* Exercise and Nutrition Research Group, School of Medical Sciences, RMIT University, Bundoora, VIC 3083, Australia; Research Institute
for Sport and Exercise Sciences, Liverpool John Moores University, Liverpool, United Kingdom.
B. Desbrow. School of Public Health and Griffith Health Institute, Griffith University, Gold Coast, QLD, Australia.
A.M. Jones and J.R. Blackwell. Sport and Health Sciences, College of Life and Environmental Sciences, University of Exeter, St. Luke’s Campus, Exeter,
United Kingdom.
M.L. Ross, A.J. Zemski, and L.M. Burke. Sports Nutrition, Australian Institute of Sport, Belconnen, ACT 2626, Australia.
Corresponding author: John A. Hawley (e-mail:
*Present address: Exercise and Nutrition Research Group, Australian Catholic University, Fitzroy, VIC 3165, Australia.
This paper is a part of a Special Issue entitled Nutritional Triggers to Adaptation and Performance.
Appl. Physiol. Nutr. Metab. 39: 1050–1057 (2014) Published at on 29 October 2013.
Appl. Physiol. Nutr. Metab. Downloaded from by McMaster University on 08/27/14
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rapidly absorbed caffeine dose (Kamimori et al. 2002;Ryan et al.
2013). With regard to dietary NO
supplementation, Jones and
colleagues (Bailey et al. 2009,2010;Lansley et al. 2011a,2011b;
Vanhatalo et al. 2011) reported that ingestion of beetroot juice
increases exercise capacity through metabolic mechanisms that
improve contraction efficiency in skeletal muscle. We hypothe-
sized that the increased CNS drive and reduced perceived exertion
elicited by caffeine supplementation in combination with the pre-
viously reported improvements in metabolic efficiency resulting
from beetroot juice ingestion would result in higher sustainable
power outputs than when each supplement was taken in isola-
The specific aim of this project was to investigate the indepen-
dent and combined effects of caffeine and NO
on the performance of a cycling task simulating the physical chal-
lenges of the London 2012 Olympic Games road cycling time trial
(TT). These effects were investigated against the background of a
standardized dietary preparation, including strategies that are
typical of TT specialists; these included the intake of a small vol-
ume of fluid during the event and frequent mouth contact with
carbohydrate (CHO) in the form of a sports confectionary, a prac-
tice recently confirmed as being beneficial to performance (Carter
et al. 2004;Chambers et al. 2009;Lane et al. 2013b;Pottier et al.
2010), even when preceded by a CHO-rich pre-event meal (Lane
et al. 2013b). We hypothesized that under optimal nutritional con-
ditions (i) caffeine alone and (ii)NO
alone supplementation
would improve TT performance and (iii) the concurrent use of
caffeine and NO
supplementation would result in an additive
performance enhancement compared with when each supple-
ment was used in isolation.
Materials and methods
Twelve male (mean ± SD: age 31 ± 7 years, body mass (BM) 73.4 ±
6.8 kg, height 180.8 ± 6.1 cm, maximal aerobic power (MAP) 459.4 ±
31.1 W, peak oxygen consumption (V
) 71.6 ± 4.6 mL·kg
and 12 female (age 28 ± 6 years, BM 62.1 ± 8.9 kg, height 169.1 ±
8.0 cm, MAP 327.1 ± 32.3 W, V
59.9 ± 5.1 mL·kg
competitive cyclists or triathletes volunteered to participate in
this study. Ethical clearance was obtained from the Australian
Institute of Sport Ethics Committee. Prior to participation, sub-
jects were informed of the nature and risks involved and com-
pleted a medical questionnaire before providing written informed
Study overview
On separate days following familiarization (described subse-
quently), subjects performed 4 cycling TTs under different exper-
imental conditions: caffeine and beetroot juice supplementation
(CAFF+BJ), caffeine and placebo beetroot juice (CAFF), beetroot
juice and placebo caffeine (BJ), or a control consisting of a placebo
of both caffeine and beetroot juice (CONT). All trials were sepa-
rated by 7 days, and treatments were allocated using a double-
blind Latin square design. Each ride was performed under standardized
conditions representing optimal nutritional practice: a CHO-rich
pre-event meal, ingestion of small amounts of a CHO-electrolyte
drink during the TT, and regular oral CHO contact in the form of
a sports confectionery product. All preliminary testing and exper-
imental trials were performed under standard laboratory environ-
mental conditions.
Incremental cycle test
In the 2 weeks prior to their first experimental trial, all subjects
performed a progressive maximal exercise test to exhaustion on a
cycle ergometer (Lode Excalibur Sport, Groningen, The Nether-
lands). After a 5-min warm-up, the test protocol commenced at 175
and 125 W for males and females, respectively, and increased by
25 W every 60 s until volitional fatigue. MAP was determined to be
the power output of the highest stage completed plus the fraction
of any uncompleted workload, as described previously (Ross et al.
2011,2012). Expired gases were collected into a calibrated and
customized Douglas bag gas analysis system, which incorporated
an automated piston that allowed the concentrations of O
(AEI Technologies, Pittsburg, Pa., USA) and the volume of air
displaced to be quantified. The operation and calibration of this
equipment have been described previously (Russell et al. 2002).
was calculated as the highest average O
recorded over 60 s.
Familiarization session
On the same day as the maximal test, subjects completed a
familiarization ride on the same bike and simulated course they
would complete in the subsequent experimental trials. In brief,
subjects completed the course at their own self-selected intensity
with the instruction to familiarize themselves with the course
profile, the bike set-up, and the maximal intensity they believed
they could sustain for the entire duration of the TT during subse-
quent rides. During this familiarization, dimensions for the bike
set-up were recorded for replication throughout all experimental
trials. Subjects were also familiarized with the use of the sports
confectionery product (described subsequently) to be used during
the experimental trials.
Diet and exercise control
Subjects consumed a standardized diet for the 24-h period prior
to each experimental trial using the prepackaged standardized diet
protocol described previously (Jeacocke and Burke 2010). Dietary
goals for this period were 8 g·kg
BM of CHO; 1.5 g·kg
BM of pro-
tein; 1.5 g·kg
BM of fat; and 220 kJ·kg
BM for the 24-h period.
Subjects were instructed to avoid alcohol for the 24 h prior to the
start of the TT, and to follow their habitual caffeine consumption
patterns until 12 h prior. Caffeine was not withheld for the 24-h
period because it has been shown previously that a 3-mg·kg
dose of caffeine improves cycling performance irrespective of
whether a withdrawal period is imposed on habitual caffeine us-
ers (Irwin et al. 2011). To avoid any possible effect on the experi-
mental trials, the provided pretrial standardized diets contained
no NO
-rich products.
Following an initial interview with a sports dietitian (AZ), a food
menu was prepared for each subject based on individual BM
and food preferences. During the same consultation, subjects re-
ported the ongoing or acute use of any medicine or supplement.
In any case in which the subject reported the use of a medicine or
supplement that could have influenced performance between
trials, the subject was excluded from the study. The subjects’
individual menu was prepared using Food Works Professional
Edition, version 6.0.2562 (Xyris Software, Brisbane, Australia).
Subjects were provided with all foods and drinks in portion-
controlled packages for consumption during the first 22 h of the
dietary control period and were given verbal and written instruc-
tions on how to follow the diet. Checklists were used to record
each menu item as it was consumed and to note any deviations
from the menu. Prior to undertaking each trial, each subject’s
food checklists were checked and clarified for compliance with
the standardization protocols by the sports dietitian. Using the
same software, analysis of the actual diet consumed by the sub-
jects was undertaken on completion of the study.
Experimental trials
Subjects presented to the laboratory on 4 separate occasions,
each separated by 7 days. On each occasion, subjects presented
at the same time of day, voided their bladder prior to having their
BM recorded, and then rested in a supine position for 10 min. At
this time, a Teflon cannula (Terumo, 20-22G, Tokyo, Japan) was
inserted into a vein in the antecubital fossa. A resting blood sam-
ple (8 mL) was taken, and the cannula was flushed with saline to
Lane et al. 1051
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keep the vein patent for subsequent sampling. Two hours prior to
the warm-up for each trial and immediately after the resting
blood sample, subjects consumed the remainder of the control
diet as a pre-race meal. This meal provided 2 g·kg
BM of CHO,
which was included in the total CHO quota in the 24-h standard-
ized diet. Subjects were instructed to consume their pre-race meal
within 20 min, after which time they remained in the laboratory
for the duration of that day’s experimental trial. Depending on
the trial, either the experimental or the placebo beetroot juice
concentrate was ingested in 2 separate doses (detailed below).
Forty minutes prior to commencement of the TT, subjects com-
pleted a standardized warm-up on the same bicycle on which they
would perform the TT. The caffeine gum was administered in
2 doses, the first immediately prior to commencement of the
warm-up and the second immediately after its completion. Sub-
jects then completed a TT simulating the characteristics of the
London Olympic Games cycling TT course specific to the male or
female events, under the conditions described subsequently.
Mean power output, heart rate, and rating of perceived exertion
(RPE) were recorded during each trial. During the first trial, water
was provided ad libitum for the time period leading up to com-
mencement of the TT. The volume consumed in this period was
recorded and was replicated throughout subsequent trials.
The warm-up consisted of 30 min of cycling at varying intensi-
ties (13 min at 25%, 5 min at 60%, 2 min at 70%, 3 min at 25%, 5 min
at 60%, and 2 min at 80% of MAP). Subjects then rested for 10 min
prior to commencing the TT.
Time trials
Subjects performed all experimental trials on a Velotron cycle
ergometer (Racermate, Seattle, Wash., USA) adjusted to the di-
mensions of their own bicycles. Males completed a simulated
43.83-km course, whereas females completed a 29.35-km course.
The courses were created using global positioning satellite data
collected during a prior reconnaissance of the London Olympic TT
event. Subjects were instructed to complete the TT as quickly as
possible. Financial incentives were offered to encourage maximal
Experimental interventions
Beetroot juice
During 2 of the trials, subjects received 2 separate doses of 140 mL
of concentrated NO
-rich beetroot juice delivering 8.4 mmol of
in each dose (Beet it, James White Drinks Ltd., Ipswich, UK).
Each subject ingested the first dose at a specific time 8to12h
prior to the commencement of each TT; the dose was provided in
each subject’s controlled diet, which was consumed the day prior
to each experimental trial. The second dose was ingested in the
laboratory 130 min prior to the commencement of the TT. During
the 2 placebo trials, a similar-tasting but NO
-depleted beetroot
juice product (0.006 mmol of NO
; Beet it, James White Drinks
Ltd.) (Lansley et al. 2011b) was administered at time points identi-
cal to those for the experimental trials.
During the 2 caffeine trials, a caffeinated gum (Stay Alert, Amurol
Confectioners, Yorkville, Ill., USA) was administered in 2 doses, to
deliver a total of 3 mg·kg
BM of caffeine. The gum was adminis-
tered in a nontransparent package emptied directly into the
mouth to avoid possible visual cues about the differences between
trials (experimental vs. placebo). The first dose was administered
immediately prior to the commencement of the warm-up (40 min
prior to the TT) and consisted of a caffeine dose containing
2 mg·kg
BM. Subjects were instructed to chew the gum for a total
of 10 min before it was removed and discarded. The remaining
dose containing 1 mg·kg
BM was administered with the same
instructions at the end of the warm-up (10 min prior to the TT).
During the placebo trials, noncaffeinated gum matched for taste
and texture (Jila Gum, Ferndale Confectionary Pty Ltd., Australia)
was provided under the same conditions as the caffeinated gum.
CHO ingestion
To ensure that the findings of this study would be relevant
when applied in a real-world situation in which athletes follow
current nutritional guidelines to maximize performance, a CHO
sports gel (PowerBar Gel, Powerbar Inc., Florham Park, N.J., USA)
containing 28 g CHO was ingested 15 min prior to the commence-
ment of each TT. Additionally, at the commencement of each TT,
subjects were provided with a sports confectionary product (Pow-
erBar Gel Blasts, Powerbar Inc.). Subjects were instructed to place
the confectionery item in their mouth and leave it in their cheek
cavity until it had dissolved completely, at which time another
was provided. The timing and number of confectionery pieces
used in the first trial was replicated throughout all subsequent
trials. The aim of this procedure was to provide a constant CHO
stimulus in the mouth similar to a CHO mouth rinse, which has
been shown previously to enhance cycling performance (Carter
et al. 2004;Chambers et al. 2009;Fares and Kayser 2011;Lane et al.
2013b;Pottier et al. 2010). Subjects also received a CHO-electrolyte
sports drink (Gatorade, Gatorade Co., Chicago, Ill., USA) to con-
sume at specific points during each TT. During the first trial,
males received 2 bottles, the first at 15 km and the second at 30 km
during the TT, whereas females received a single bottle at 15 km.
These points correspond to portions of the TT in which prior
reconnaissance of the course suggested it would be practical for
competitors to take a drink. During the first trial, each bottle was
preweighed, and subjects were instructed to consume as much
fluid as desired within 1 min. Each bottle was then reweighed and
the volume of fluid consumed was recorded; this was repeated
throughout all subsequent trials.
Blood collection and analysis
At each sampling time point, a total of 8 mL of whole blood was
collected in a tube containing lithium heparin. Each trial included
4 sampling time points consisting of a resting sample, a sample
taken immediately prior to commencement of the warm-up (prior
to caffeine ingestion), a third sample taken immediately after the
warm-up, and a final sample taken immediately after the TT.
Tubes were centrifuged immediately at 4 °C at 4000 r·min
(3040g) for 10 min. The resultant plasma was divided into equal
aliquots and stored at –80 °C for the subsequent analysis of caf-
feine, NO
, and nitrite (NO
) concentrations.
Plasma caffeine concentration
The quantitative analysis of plasma caffeine was performed us-
ing an automated reverse-phase, high-performance liquid chro-
matography system. Conditions were adapted from Koch, Tusscher,
Koppe, and Guchelaar (Koch et al. 1999) with subtle modifications.
The precise method has been described previously (Desbrow et al.
Plasma NO
and NO
Plasma NO
and NO
were analyzed by gas phase chemilumi-
nescence analysis. This initially required NO
and NO
to be
reduced to nitric oxide (NO) gas. For the reduction of NO
, undi-
luted plasma was injected into a glass purge vessel containing 5 mL
glacial acetic acid and 1 mL NaI solution. For the NO
plasma samples were deproteinized in an aqueous solution of zinc
sulphate (10% w/v) and 1 mol·L
sodium hydroxide, prior to reduc-
tion to NO in a solution of vanadium (III) chloride in 1 mol·L
hydrochloric acid (0.8% w/v). Quantification of NO was enabled by
the detection of light emitted during the production of nitrogen
dioxide formed upon the reaction of NO with ozone. Lumines-
cence was detected by a thermoelectrically cooled, red-sensitive
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photomultiplier tube housed in a Sievers gas-phase chemilumi-
nescence NO analyzer (Sievers NOA 280i, Analytix Ltd., Durham,
UK). The concentrations of NO
and NO
were determined by
plotting signal area (mV) against a calibration plot of 25 nmol·L
to 1 mol·L
sodium nitrite and 100 nmol·L
to 10 mol·L
dium nitrate, respectively.
Statistical analyses
Statistical analyses were performed using software package
SPSS (version 18). For all blood and physiological measures (com-
bining both male and female results), 1-way analyses of variance
(ANOVAs) for repeated measures were used to make comparisons
between time points and trials using a Bonferroni adjustment
where appropriate. Mean power outputs from the 4 trials were
analyzed for males and females separately as well as combined,
using the magnitude-based inference approach recommended for
studies in sports medicine and exercise (Hopkins et al. 2009). The
same inference-based approach was also used to compare time to
complete each trial for males and females separately. A spread-
sheet (Microsoft Excel), designed to examine post-only crossover
trials, was used to determine the clinical significance of each
treatment (available at
html), as based on guidelines outlined by Hopkins (2007). Qualita-
tive inferences are reported as the percentage chance of a positive
effect compared with the corresponding trial where a least worth-
while effect on power output of 1% was used as established previ-
ously (Paton and Hopkins 2006). Significance was set at p< 0.05.
All data are presented as means ± SD unless stated otherwise.
Body mass
Among all the trials, there was no difference in BM upon pre-
senting to the laboratory. Similarly, there was no statistical differ-
ence among trials in the change in BM before and after each trial
(Table 1).
Plasma caffeine
Figure 1A displays the plasma caffeine concentrations for all
trials. At rest, there was a small variation in plasma caffeine con-
centrations, likely because subjects were instructed to abstain
from caffeine only in the 12 h prior to the trials (Irwin et al. 2011).
Within 30 min of ingestion, plasma caffeine concentrations were
significantly increased (CAFF+BJ 9.2 ± 3.2 mol·L
and CAFF 10.0 ±
3.80 mol·L
) when compared with resting values and with the
noncaffeine trials. Peak caffeine concentrations (CAFF+BJ 16.7 ±
3.1 mol·L
and CAFF 17.2 ± 5.5 mol·L
) were recorded at the
final collection point at the end of each TT.
Plasma NO
and NO
Figure 1B shows plasma NO
concentrations for all trials. The
preloading NO
dose, administered 6–10 h prior to the resting
blood sample, increased plasma NO
concentrations in the CAFF+BJ
and BJ trials (113.1 ± 33.3 mol·L
and 123.2 ± 37.6 mol·L
, respec-
tively; p< 0.01) when compared with the non-beetroot juice trials.
Plasma NO
levels remained significantly elevated in CAFF+BJ and
BJ at all time points compared with CAFF and CONT (p< 0.05). The
second dose of beetroot juice (administered 130 min prior to the
TT) further elevated plasma NO
concentrations at 90 min
(282.7 ± 64.8 mol·L
and 295.8 ± 67.0 mol·L
) and 2 h after
ingestion (310.6 ± 58.7 mol·L
and 333.9 ± 64.7 mol·L
) when
compared with resting values (p< 0.05), with concentrations re-
maining elevated until after the TT (334.1 ± 53.3 mol·L
343.1 ± 58.4 mol·L
,p< 0.05).
Figure 1C shows plasma NO
concentrations for all trials. Con-
centrations were significantly higher in CAFF+BJ and BJ at all time
points compared with CAFF and CONT (p< 0.01). The preloading
dose consumed 6–10 h prior to the resting blood sam-
ple elevated NO
levels to 176.1 ± 90.9 nmol·L
and 174.3 ±
87.1 nmol·L
(p< 0.05) for the CAFF+BJ and BJ trials, respectively,
compared with the non-beetroot juice trials. The second dose of
-rich beetroot juice did not elevate plasma NO
tions further in the CAFF+BJ and BJ trials.
Power output
Figure 2 shows the relative mean power output combined for
males and females. On average, caffeine improved mean power
output when compared with CONT in the CAFF+BJ and CAFF trials
by 3.5% (p< 0.01). Beetroot juice supplementation had no effect on
mean power output in either CAFF+BJ vs. CAFF or BJ vs. CONT.
Using an inference-based statistical approach, caffeine was very
likely (99%) and very likely (97%) (CAFF+BJ vs. BJ and CAFF vs.
CONT, respectively) to have a positive effect on performance out-
comes during a cycling TT. NO
supplementation was most un-
likely (0%) and very unlikely (1%) (BJ vs. CONT and CAFF+BJ
vs. CAFF, respectively) to have any positive effect on performance.
When mean power output was compared for trial order rather
than intervention, no significance was detected among any trials
(Trials 1 through 4: males 298 ± 40 W, 301 ± 35 W, 306 ± 40 W, and
305 ± 37 W, respectively; females 212 ± 30 W, 210 ± 26 W, 212 ±
34 W, and 208 ± 31 W, respectively; p= 1.0), indicating that the
Table 1. Summary of cycling time trial performance and associated measures.
Power Heart rate
Body mass (kg)
Intervention Time (h:mm:ss.00) W % MAP Pre Pre-Post TT
Males (43.83 km)
CONT 1:03:30.39±0:03:16.15 303±41 66±5.8 167±11 17±0.9 73.3±6.7 −1.1±0.4
CAFF+BJ 1:02:38.04±0:03:31.00* 314±44* 68±6.3* 171±9 17±0.9 73.6±6.9 −1.4±0.3
CAFF 1:02:43.86±0:03:04.87* 313±38* 68±4.5* 172±10 17±0.8 73.5±7.0 −1.8±0.3
BJ 1:04:05.03±0:02:50.09 298±35 65±4.8 169±9 17±1.0 73.4±6.9 −1.2±0.3
Females (29.35 km)
CONT 0:51:40.10±0:02:31.71 207±29 63±6.2 171±8 17±1.2 62.2±8.9 −0.9±0.4
CAFF+BJ 0:51:11.88±0:02:22.13* 212±27* 65±5.1* 176±6 17±0.8 62.0±9.1 −0.9±0.4
CAFF 0:50:50.53±0:02:56.48* 216±34* 66±6.4* 174±9 17±1.0 62.2±9.0 −0.7±0.3
BJ 0:51:41.06±0:02:39.51 207±31 63±6.5 174±9 17±0.6 62.4±9.3 −0.8±0.5
CONT 250±57 64±5.8 ——
CAFF+BJ 258±59 66±5.6* ——
CAFF 260±58 67±5.4* ——
BJ 249±56 64±5.6 ——
Note: MAP, maximal aerobic power; RPE, rating of perceived exertion; TT, time trial; CONT, placebo of caffeine and beetroot juice;
CAFF+BJ, beetroot juice with caffeine; CAFF, caffeine; BJ, beetroot juice.
*Significantly different from CONT and BJ (p< 0.05).
Lane et al. 1053
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Latin square design was successful in eliminating any possible
trial order effect.
TT completion time
Times to complete the respective distances for males and fe-
males are presented in Table 1. For males, when compared with
CONT, the time to complete the 43.83-km distance was reduced to
a similar extent of 1.3% (p< 0.05) for both the CAFF+BJ and CAFF
trials. For females, the time to complete the 29.35-km distance
was reduced by 0.9% and 1.6% (p< 0.05) for the CAFF+BJ and CAFF
trials, respectively, when compared with CONT. Beetroot juice
supplementation had no significant positive or negative effect on
time to complete the trials for either males or females in either
CAFF+BJ vs. CAFF or BJ vs. CONT. Using an inference-based statis-
tical approach, caffeine would possibly (65%) and likely (89%) for
males and possibly (42%) and likely (88%) for females (CAFF+BJ vs.
BJ and CAFF vs. CONT, respectively) produce a positive effect on
performance outcomes during a cycling TT. NO
tion was unlikely (7%) and very unlikely (1%) for males (CAFF+BJ
vs. CAFF and BJ vs. CONT, respectively) and very unlikely (0%)
under both conditions for females to have any positive effect on
performance (Table 2).
Heart rate and RPE
Table 1 shows mean heart rate and RPE for each trial for males
and females. There were no differences in mean heart rate or RPE
among any trials.
To the best of our knowledge, this is the first study to determine
the single and combined effects of caffeine and NO
tation on the performance of cycling protocols that simulated real
TT courses and was undertaken with the support of nutritional
practices considered optimal for elite TT performance. Because
each of these ergogenic aids is purported to elicit its performance-
enhancing effect via different mechanisms (i.e., central vs. periph-
eral), we hypothesized that the combination of the 2 interventions
would increase mean power output to a greater extent than when
each intervention was administered in isolation. Our results indi-
cate that caffeine supplementation provided a worthwhile en-
hancement of TT performance to both male and female cyclists,
Fig. 1. Plasma concentrations. (A) Caffeine. (B) NO
. (C) NO
. Times
are relative to ingestion. “Rest” in B and C includes a “preload”
dose 6–10 h prior to the TT. Data are presented as means ± SD.
BJ+CAFF, beetroot juice with caffeine; CAFF, caffeine; BJ, beetroot
juice; CONT, placebo of caffeine and beetroot juice. TT, time trial;
, nitrate; NO
, nitrite. *, Different from CONT and BJ (p< 0.01);
†, different from CONT and CAFF (p< 0.01); a, different from REST
and 0 min (p< 0.01); b, different from REST, 0 min, and 30 min
(p< 0.05); c, different from REST.
Fig. 2. Mean power output combined for males and females. Data
are presented as means ± SD. CAFF+BJ, beetroot juice with caffeine;
CAFF, caffeine; BJ, beetroot juice; CONT, placebo of caffeine and
beetroot juice. *, Different from CONT and BJ (p< 0.01).
1054 Appl. Physiol. Nutr. Metab. Vol. 39, 2014
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but that beetroot juice did not provide a detectable benefit under
these conditions.
In the current study, pre-event supplementation with caffeine
(3 mg·kg
BM) increased mean power output in cycling TTs last-
ing 50 min (competitive female cyclists) and 60 min (com-
petitive male cyclists) to an extent (3%–4%) similar to those
reported previously using similar caffeine doses (Cox et al. 2002;
Irwin et al. 2011;Jenkins et al. 2008;Lane et al. 2013a). In particu-
lar, these results are in agreement with the findings of Ryan et al.
(2013), who reported that caffeine administered in the form of
a gum prior to a cycling TT induced an elevation of circulating
caffeine concentrations within 30 min of intake and resulted in
significantly improved performance. We observed that the bene-
fits of ingesting caffeine 40 min prior to TTs simulating the
specific courses undertaken at the 2012 London Olympic Games
were similar for males and females, although the courses they
rode were slightly different in length and duration. Although
these results were derived specifically for the preparation of cy-
clists for the 2012 Olympic Games, they can be generalized to
other events of a similar nature.
It is worth noting that in our study, caffeine ingestion improved
performance under the standardized conditions of dietary prepa-
ration that are both recommended and typical of the practices of
cycling TT specialists. These practices included a CHO-rich pre-
event meal (Lane et al. 2013b), consumption of a small fluid intake
during the event according to the practical opportunities to drink
(Garth and Burke 2013), and frequent mouth contact with CHO
(Lane et al. 2013b). Many studies often neglect to recognize that the
real-world application of ergogenic interventions may be influ-
enced by optimal race day strategies. Indeed, a meta-analysis has
shown that the benefits of caffeine ingestion on endurance per-
formance are reduced when it is taken in combination with CHO
(Conger et al. 2011). However, under the conditions of our study,
caffeine ingestion still improved performance to the same degree
as reported previously (Cox et al. 2002;Irwin et al. 2011;Jenkins
et al. 2008;Lane et al. 2013a;Ryan et al. 2013) when guidelines for
optimal CHO ingestion for this specific type of event (Burke et al.
2011) were followed. Finally, our findings are also in agreement
with Irwin et al. (2011), who reported a similar degree of improve-
ment in cycling performance when a comparable 12-h with-
drawal from caffeine was enforced in habitual caffeine users. This
observation suggests that longer withdrawal periods (24–48 h), as
recommended previously (Burke 2008), may not be necessary.
In contrast, we found no effect of beetroot juice ingestion on a
cycling TT lasting 50–60 min despite elevated plasma NO
concentrations. Indeed, the conditions under which supple-
mentation with NO
and beetroot juice ingestion enhances exer-
cise capacity or performance remain somewhat unclear. Elements
that could be of importance include the timing and dose of the
, the intensity and duration of the exercise protocol, and the
training history or calibre of the athlete. Recently, the effect of
beetroot juice on exercise capacity has been shown to be dose
dependent, with maximal benefits being seen with the acute in-
gestion of 2 bottles of beetroot juice concentrate (acute dose of
8.4 mmol NO
)(Wylie et al. 2013). Because the cyclists in our study
ingested the same amount of the same product, both acutely
(2 h before exercise) and as an additional preload (6–10 h before
the trial), we are confident that our failure to detect benefits from
supplementation cannot be explained by a suboptimal dos-
ing protocol.
Our NO
supplementation protocol substantially elevated
plasma NO
concentrations, although the peak values in our
study were lower (225 vs. 470–687 nmol·L
) than those reported
by studies employing a similar acute dosing protocol in subjects
with a range of training histories (Cermak et al. 2012b;Lansley
et al. 2011a;Muggeridge et al. 2013;Wilkerson et al. 2012;Wylie
et al. 2013). Although it is only speculation, it is possible that the
pre-race meal, consumed shortly before the ingestion of the sec-
ond beetroot juice dose, may have affected the conversion of NO
to NO
. However, despite this observed difference, Cermak et al.
(2012b)and Wilkerson et al. (2012) reported significantly higher
peak plasma NO
concentrations (532 and 472 nmol·L
, respec-
tively) compared with the current study, but also failed to detect a
performance benefit in well-trained cyclists. Because of the range
of plasma NO
concentrations, varied training histories, and dif-
ferent performance tasks employed, it is difficult to determine
whether these observations play a role in the effectiveness of NO
The mechanism underpinning the observed benefits of NO
supplementation on exercise capacity is believed to be a reduction
in the oxygen cost of exercise, as a consequence of a reduced
energy cost of contraction or enhanced mitochondrial efficiency
(Jones et al. 2012). Whether this translates into an enhancement of
performance across a range of exercise intensities has not been
studied systematically. However, it is worth noting that the per-
formance of shorter cycling tasks (5–30 min in duration) has
been enhanced following NO
supplementation. For example, in
a study by Lansley et al. (2011a), subjects who sustained intensities
equivalent to 98% and 95% of maximal oxygen consumption
) during 4-km and 16.1-km TTs, respectively, recorded an
improvement in performance after beetroot juice supplementa-
tion. However, a 50-mile cycling TT lasting 135 min and eliciting
a sustained exercise intensity equivalent to 74% of V
not show a performance benefit following NO
despite subjects showing an improvement in power output per
oxygen volume (W/L·min
)(Wilkerson et al. 2012). Cermak et al.
(2012b)also reported no enhancement of 1-h cycling TT perfor-
mance in a cohort of well-trained cyclists using an acute NO
and timing strategy similar to that employed in the current study.
We did not measure oxygen consumption during the TT in the
current study; however, Coyle et al. (1991) reported that well-
trained cyclists completed a 1-h TT at 87% V
, suggesting
that our comparable subjects worked at a lower percentage of
their aerobic capacity than did those observed in shorter-
duration tasks as employed in the study by Lansley et al. (2011a).
Possible explanations for this observation include the effects of
exercise intensity on muscle oxygenation and motor unit re-
cruitment. Higher exercise intensities are likely to result in a
greater degree of skeletal muscle hypoxia, which would be
expected to facilitate NO production through the reduction of
(Maher et al. 2008). In addition, higher exercise intensities
would be expected to mandate a greater recruitment of type II
muscle fibres. There is evidence that the effects of NO
mentation on blood flow (Ferguson et al. 2013), muscle force,
and calcium handling (Hernandez et al. 2012) may be more
pronounced in type II fibres. These observations merit further
investigation because it appears that the effectiveness of NO
supplementation may be influenced by the intensity of the
performance task.
The failure to find a benefit of NO
supplementation in highly
trained athletes may be because of a factor that has not yet been
identified. For example, studies in which pre-event ingestion of
beetroot juice has been unable to produce a detectable improve-
Table 2. Power improvement for respective trials.
%(±90% CL) p
CAFF vs. CONT 4.0±1.7 <0.001
CAFF+BJ vs. CONT 3.1±1.9 0.01
BJ vs. CONT −0±1.3 0.6
CAFF vs. BJ 4.2±1.8 <0.001
CAFF+BJ vs. BJ 3.4±1.6 <0.001
CAFF+BJ vs. CAFF −1±1.7 0.4
Note: CL, confidence limit; CAFF, caffeine; CONT, pla-
cebo of caffeine and beetroot juice; CAFF+BJ, beetroot
juice with caffeine; BJ, beetroot juice.
Lane et al. 1055
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ment in performance have involved subelite or well-trained
cohorts (Cermak et al. 2012b;Christensen et al. 2013;Muggeridge
et al. 2013;Wilkerson et al. 2012). A recent meta-analysis of studies
of beetroot juice and NO
supplementation and endurance per-
formance published before August 2012 found that the effects
were observed more readily in inactive to recreationally active
individuals (Hoon et al. 2013). Clearly, this intriguing aspect war-
rants further study, with candidate explanations including the
optimization of arginine-mediated pathways of NO production in
highly trained individuals, or differences in muscle fibre type
(Christensen et al. 2013).
It is noteworthy that Cermak et al. (2012a)reported a significant
improvement in 10-km cycle TT performance following 6 days of
beetroot juice supplementation but no effect on 1-h TT perfor-
mance after acute beetroot juice intake (Cermak et al. 2012b).
Although this finding may be related to differences in exercise
duration and intensity, as discussed earlier, it is also possible that
longer periods of beetroot juice supplementation are necessary
for performance changes to be realized in highly trained subjects.
For example, changes in proteins related to mitochondrial effi-
ciency (Larsen et al. 2011) and muscle calcium handling (Hernandez
et al. 2012) that have been reported following NO
are likely to take several days (rather than hours) to become
Previous studies have suggested that there may be “responders”
and “nonresponders” to dietary NO
supplementation (Christensen
et al. 2013;Wilkerson et al. 2012;Wylie et al. 2013), and this obser-
vation appears to be consistent in a highly trained cohort
(Christensen et al. 2013;Wilkerson et al. 2012). In the current study,
only 2 male individuals recorded better performances in both BJ vs.
CONT and CAFF+BJ vs. CAFF, suggesting that they were possible re-
sponders. In comparison, Christensen and colleagues (2013) noted
that 2 of the 10 highly trained cyclists in their study (mean aerobic
capacity of 72.1 mL·kg
vs. 71.6 mL·kg
in the male
cyclists in the current study) appeared to benefit from a chronic
beetroot juice intake protocol, deriving a 3% improvement in per-
formance of an 18-min TT compared with a control condition. Fac-
tors to explain individual responsiveness to such supplementation
remain elusive at present.
In conclusion, we have provided evidence that a caffeine gum
containing 3 mg·kg
BM ingested in the 40 min prior to a cycling
TT lasting 45–60 min increases cycling power output in both
males and females. However, despite increasing circulating NO
and NO
concentrations, beetroot juice supplementation in-
gested 8–12 h prior to the TT as well as an acute dose ingested
2 h prior to the TT did not enhance cycling performance either
in isolation or in combination with caffeine ingestion. Based on
previous evidence that NO
supplementation can improve per-
formance in a variety of high-intensity endurance tasks, we can-
not rule out the possibility that an additive effect may still be
possible with different protocols or in specific individuals (re-
sponders). Further research is required to determine if NO
plementation, when coingested with caffeine, can further enhance
performance under shorter, more intense tasks in which the benefit
of NO
supplementation is more pronounced.
We acknowledge the hard work and commitment of the ath-
letes who gave their time to participate in this research project.
We also thank Greg Shaw for his assistance in conducting the
study and developing the placebo gum protocol, as well as all the
people who assisted in recording data during the trials.
This project was funded by a research grant from the Australian
Institute of Sport (AIS) Sports Supplement Program and AIS
Sports Nutrition.
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... Standard errors were obtained by calculating the square root of the pooled variances. Separate outcomes from the same study were considered when overall data combining all participants were not available (Algrain et al., 2015;Lane et al., 2014). For the subgroup analyses, Cochran Q-tests were performed with the "update.meta" ...
... A final sub-analysis was performed in which exercise outcomes were categorized according to the timing of Caff-gum supplementation (within15 and before15). Two studies (Lane et al., 2014;Oberlin-Brown et al., 2016) were excluded from this specific analysis because Caff-gum was provided during several time points within a Caff-gum condition. Another study (Ryan et al., 2012) had one outcome excluded because Caff-gum was provided 15 minutes after the exercise test was initiated. ...
... A total of 14 studies were included in the meta-analysis, totaling 200 participants (Table 1). Four studies included female participants, totaling 34 individuals, although only one study included values for a standalone group of women (Lane et al., 2014) ( Table 1). This study contained data from both men and women which were included separately in the meta-analysis due to different doses provided and different distances covered in the cycling time-trial. ...
Highlights: This study determined the effect of Caff-gum on exercise performance, using a systematic review and meta-analysis. Fourteen studies, totalling 200 participants performing a variety of endurance and strength/power exercise tests were included. The relative Caff-gum dose ranged from 1.27-4.26 mg/kg BM and timing ranged from 120 min prior to exercise up to intra-test application.Caff-gum was shown to be an effective ergogenic aid for trained individuals involved in both endurance and strength/power exercise.Supplement dose and timing modified the efficacy of Caff-gum. Supplementation with Caff-gum was effective when provided in doses ≥3 mg/kg BM and within 15 min prior to initiating exercise.Trained endurance or strength/power athletes seeking to benefit from caffeine in the form of chewing gum should supplement within 15 min prior to initiating an exercise task, in doses ≥3 mg/kg BM.
... For example, several used a low (≤4.9 mmol) (55,70,92,93) or high (≥15 mmol) (52, 80, 94) acute dose; others ingested <150 min before the exercise test (70,92,93,(95)(96)(97)(98)(99) or supplemented nitrate salts (80,96). Additionally, in some cases the duration of exercise was ≤60 s (37, 100) or >600 s (52,55,70,(93)(94)(95)101), or used time-trial tasks (25,52,55,94,95,99,(101)(102)(103)(104)(105). Furthermore, ovarian hormones may influence sports performance, particularly during the follicular phase of the menstrual cycle (106). ...
... For example, several used a low (≤4.9 mmol) (55,70,92,93) or high (≥15 mmol) (52, 80, 94) acute dose; others ingested <150 min before the exercise test (70,92,93,(95)(96)(97)(98)(99) or supplemented nitrate salts (80,96). Additionally, in some cases the duration of exercise was ≤60 s (37, 100) or >600 s (52,55,70,(93)(94)(95)101), or used time-trial tasks (25,52,55,94,95,99,(101)(102)(103)(104)(105). Furthermore, ovarian hormones may influence sports performance, particularly during the follicular phase of the menstrual cycle (106). ...
To identify how variables such as exercise condition, supplementation strategy, participant characteristics and demographics, and practices that control oral microbiota diversity could modify the effect of inorganic nitrate ingestion (as nitrate salt supplements, beetroot juice, and nitrate-rich vegetables) on exercise performance, we conducted a systematic review with meta-analysis. Studies were identified in PubMed, Embase, and Cochrane databases. Eligibility criteria included randomized controlled trials assessing inorganic nitrate on exercise performance in healthy adults. To assess the variation in effect size, we used meta-regression models for continuous variables and subgroup analysis for categorical variables. One hundred and twenty-three studies were included in this meta-analysis totaling 1705 participants. Nitrate was effective for improving exercise performance (Standardized Mean Difference (SMD):0.101; 95% confidence intervals (95%CI):0.051,0.151, P < 0.001, I2 = 0%), although nitrate salts supplementation was not as effective (P = 0.629) as ingestion via beetroot juice (P < 0.001) or a high nitrate diet (P = 0.005). Practices that control oral microbiota diversity influenced the nitrate effect, with practices harmful to oral bacteria decreasing the ergogenic effect of nitrate. Nitrate ingestion was most effective for exercise lasting between 2 and 10min (P < 0.001). An inverse dose-response relationship between the fraction of inspired oxygen and the effect size (coefficient: -0.045;95%CI: -0.085, -0.005, P = 0.028) suggests that nitrate was more effective in increasingly hypoxic conditions. There was a dose-response relation for acute administration (P = 0.049). The most effective acute dose was between 5–14.9mmol provided ≥150min prior to exercise (P < 0.001). An inverse dose-response for protocols ≥ 2days was observed (P = 0.025), with the optimal dose between 5–9.9mmol∙day−1 (P < 0.001). Nitrate, via beetroot juice or a high nitrate diet, improved exercise performance, particular those lasting between 2–10min. Ingestion of 5–14.9mmol⋅day−1 taken at least 150min prior to exercise appears optimal for performance gains, while athletes should be aware that practices which control oral microbiota diversity may decrease the effect of nitrate.
... A study by Kamimori et al. (2002) showed that caffeine is absorbed faster from chewing gum compared with caffeine capsules, likely because absorption is facilitated through the buccal mucosa. Most studies comparing caffeinated chewing gum with placebo conditions and/or multi-ingredient caffeine supplements have shown performance improvements (Ryan et al., 2013;Lane et al., 2014;Paton et al., 2015). However, it appears that no studies have compared the effect caffeinated chewing gum with pure caffeine capsules or powder on endurance performance. ...
... However, it appears that no studies have compared the effect caffeinated chewing gum with pure caffeine capsules or powder on endurance performance. Lane et al. (2014) showed that plasma caffeine concentrations were similar between caffeinated chewing gum and beetroot juice with caffeine. Both caffeinated supplements also showed similar performance improvements compared to placebo and beetroot juice alone. ...
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Caffeine is widely accepted as an endurance-performance enhancing supplement. Most scientific research studies use doses of 3–6 mg/kg of caffeine 60 min prior to exercise based on pharmacokinetics. It is not well understood whether endurance athletes employ similar supplementation strategies in practice. The purpose of this study was to investigate caffeine supplementation protocols among endurance athletes. A survey conducted on Qualtrics returned responses regarding caffeine supplementation from 254 endurance athletes (f = 134, m =120; age = 39.4 ± 13.9 y; pro = 11, current collegiate athlete = 37, recreational = 206; running = 98, triathlon = 83, cycling = 54, other = 19; training days per week = 5.4 ± 1.3). Most participants reported habitual caffeine consumption (85.0%; 41.2% multiple times daily). However, only 24.0% used caffeine supplements. A greater proportion of men (31.7%) used caffeine supplements compared with women (17.2%; p = 0.007). Caffeine use was also more prevalent among professional (45.5%) and recreational athletes (25.1%) than in collegiate athletes (9.4%). Type of sport (p = 0.641), household income (p = 0.263), education (p = 0.570) or working with a coach (p = 0.612) did not have an impact on caffeine supplementation prevalence. Of those reporting specific timing of caffeine supplementation, 49.1% and 34.9% reported consuming caffeine within 30 min of training and races respectively; 38.6 and 36.5% used caffeine 30–60 min before training and races. Recreational athletes reported consuming smaller amounts of caffeine before training (1.6 ± 1.0 mg/kg) and races (2.0 ± 1.2 mg/kg) compared with collegiate (TRG: 2.1 ± 1.2 mg/kg; RACE: 3.6 ± 0.2 mg/kg) and professional (TRG: 2.4 ± 1.1 mg/kg; RACE: 3.5 ± 0.6 mg/kg) athletes. Overall, participants reported minor to moderate perceived effectiveness of caffeine supplementation (2.31 ± 0.9 on a four-point Likert-type scale) with greatest effectiveness during longer sessions (2.8 ± 1.1). It appears that recreational athletes use lower caffeine amounts than what has been established as ergogenic in laboratory protocols; further, they consume caffeine closer to exercise compared with typical research protocols. Thus, better education of recreational athletes and additional research into alternative supplementation strategies are warranted.
... The combined use of supplements can occur either acutely, targeting a specific competition, or chronically throughout a training program. In this regard, previous studies have investigated the combination of BJ and CAF within a laboratory environment with contrasting results [16][17][18][19]. Cognitive function is essential for athletic performance with variables such as attention, memory, and executive functions involving working memory, decision-making, and multitasking shown to be improved by CAF and BJ ingestion separately [20,21]. ...
... Finally, the lack of ergogenicity seen in these supplementation strategies may be related to external factors such as nutritional status (both acute and chronic), sleep pattern, physical activity level, training status, and small sample size [48]. Overall, this finding agrees with previous studies that reported no significant effects of combining BJ + CAF on maximal and submaximal running [19] or cycling [16][17][18]. ...
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Background Beetroot juice (BJ) and caffeine (CAF) are considered as ergogenic aids among athletes to enhance performance, however, the ergogenic effects of BJ and CAF co-ingestion are unclear during team-sport-specific performance. This study aimed to investigate the acute effects of BJ and CAF co-ingestion on team-sport-specific performance, compared with placebo (PL), BJ, and CAF alone. Method Sixteen semi-professional male soccer players (age: 19.8 ± 2.2 years, body mass: 69.2 ± 6.1 kg, height: 177.3 ± 6.0 cm) completed four experimental trials using a randomized, double-blind study design: BJ + CAF, CAF + PL, BJ + PL, and PL + PL. Countermovement jump with arm swing (CMJAS) performance and cognitive function by Stroop Word-Color test were evaluated before and after the Yo–Yo Intermittent Recovery Test level 1 (YYIR1). Also, rate of perceived exertion (RPE), heart rate, and gastrointestinal (GI) discomfort were measured during each session. Results No significant differences were shown between test conditions for total distance covered in YYIR1 (BJ + CAF: 1858 ± 455 m, CAF + PL: 1798 ± 422 m, BJ + PL: 1845 ± 408 m, PL + PL 1740 ± 362 m; p = 0.55). Moreover, CMJAS performance, cognitive function, and RPE during the YYIR1 were not significantly different among conditions (p > 0.05). However, the average heart rate during the YYIR1 was higher in CAF + PL compared to PL + PL (by 6 ± 9 beats/min; p < 0.05), and GI distress was greater in BJ + CAF compared to PL + PL (by 2.4 ± 3.6 a.u.; p < 0.05). Conclusion These results suggest, neither acute co-ingestion of BJ + CAF nor BJ or CAF supplementation alone significantly affected team-sport-specific performance compared to the PL treatment.
... A similar study in elite Olympic-level cyclists showed no effect on TT performance with two 8.4 mmol doses of nitrate both 10-12 hr and ∼2 hr prior to exercise. In those athletes also given 3 mg/kg caffeine, there were no additive benefits of nitrate compared with caffeine supplementation alone (Lane et al., 2014). The three studies which found no effect of dietary nitrate supplementation via beetroot juice all investigated cycling performance outcomes, while the two studies which found positive results investigated swimming and kayaking, both upper body heavy exercises. ...
... Similarly, 6 mmol nitrate supplemented 3 hr before exercise had no effect on repeated sprint ability in amateur female team-sport athletes despite increases in plasma nitrate concentration (Buck et al., 2015). In the previously discussed study of elite Olympic level cyclists, Lane et al. (2014) saw no effect of beetroot juice on power output, either alone or in combination with caffeine. Contrarily, the study by Pospieszna et al. (2016) found more pronounced changes in anaerobic sprint swimming performance compared with aerobic TTs results. ...
Beta-alanine, caffeine, and nitrate are dietary supplements generally recognized by the sport and exercise science community as evidence-based ergogenic performance aids. Evidence supporting the efficacy of these supplements, however, is greatly skewed due to research being conducted primarily in men. The physiological differences between men and women, most notably in sex hormones and menstrual cycle fluctuations, make generalizing male data to the female athlete inappropriate, and potentially harmful to women. This narrative review outlines the studies conducted in women regarding the efficacy of beta-alanine, caffeine, and nitrate supplementation for performance enhancement. Only nine studies on beta-alanine, 15 on caffeine, and 10 on nitrate in healthy women under the age of 40 years conducted in normoxia conditions were identified as relevant to this research question. Evidence suggests that beta-alanine may lower the rate of perceived exertion and extend training bouts in women, leading to greater functional adaptations. Studies of caffeine in women suggest the physiological responder status and caffeine habituation may contribute to caffeine’s efficacy, with a potential plateau in the dose–response relationship of performance enhancement. Nitrate appears to vary in influence based on activity type and primary muscle group examined. However, the results summarized in the limited literature for each of these three supplements provide no consensus on dosage, timing, or efficacy for women. Furthermore, the literature lacks considerations for hormonal status and its role in metabolism. This gap in sex-based knowledge necessitates further research on these ergogenic supplements in women with greater considerations for the effects of hormonal status.
... Regarding the lack of efficacy of NO 3 consumption for improving exercise performance in females, it is difficult to determine whether the lack of effect is attributable to biological differences between the sexes or the contexts of the studies. All four trials that assessed the effect of NO 3 on exercise performance in females assessed timetrial performance, and were predominantly conducted in high-level athletes, with two studies conducted in participants classified as PL4 [62] and PL5 [70], and one study conducted in national-level water polo players. Hence, the absence of effects in these trials may be attributable to the decreased efficacy demonstrated for both time-trial performance and fitter athletes within this review, although an included study by de Castro, de Assis Manoel, and Machado [59] found no effects in untrained females. ...
... Given the potential interactions of NO 3 and polyphenols with other dietary factors, it is also of interest as to whether nitric oxide-related supplements may affect the responses to other ergogenic aids such as caffeine. Four beetroot studies investigated the effects of beetroot juice both alone and in combination but indicated that beetroot juice had neither a positive effect independently, nor did it influence the ergogenic effect of caffeine [62,63,70,80]. These studies were all conducted on highlytrained athletes (all ≥PL4) and three of the four studies assessed performance via a TT, with both factors having reduced ergogenic effects within this review. ...
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Background Increasing nitric oxide bioavailability may induce physiological effects that enhance endurance exercise performance. This review sought to evaluate the performance effects of consuming foods containing compounds that may promote nitric oxide bioavailability. Methods Scopus, Web of Science, Ovid Medline, EMBASE and SportDiscus were searched, with included studies assessing endurance performance following consumption of foods containing nitrate, L-arginine, L-citrulline or polyphenols. Random effects meta-analysis was conducted, with subgroup analyses performed based on food sources, sex, fitness, performance test type and supplementation protocol (e.g. duration). Results One hundred and eighteen studies were included in the meta-analysis, which encompassed 59 polyphenol studies, 56 nitrate studies and three L-citrulline studies. No effect on exercise performance following consumption of foods rich in L-citrulline was identified (SMD=-0.03, p=0.24). Trivial but significant benefits were demonstrated for consumption of nitrate and polyphenol-rich foods (SMD=0.15 and 0.17, respectively, p <0.001), including performance in time-trial, time-to-exhaustion and intermittent-type tests, and following both acute and multiple-day supplementation, but no effect of nitrate or polyphenol consumption was found in females. Among nitrate-rich foods, beneficial effects were seen for beetroot, but not red spinach or Swiss chard and rhubarb. For polyphenol-rich foods, benefits were found for grape, (nitrate-depleted) beetroot, French maritime pine, Montmorency cherry and pomegranate, while no significant effects were evident for New Zealand blackcurrant, cocoa, ginseng, green tea or raisins. Considerable heterogeneity between polyphenol studies may reflect food-specific effects or differences in study designs and subject characteristics. Well-trained males (V̇O 2max ≥65 ⁻¹ ) exhibited small, significant benefits following polyphenol, but not nitrate consumption. Conclusion Foods rich in polyphenols and nitrate provide trivial benefits for endurance exercise performance, although these effects may be food dependent. Highly trained endurance athletes do not appear to benefit from consuming nitrate-rich foods but may benefit from polyphenol consumption. Further research into food sources, dosage and supplementation duration to optimise the ergogenic response to polyphenol consumption is warranted. Further studies should evaluate whether differential sex-based responses to nitrate and polyphenol consumption are attributable to physiological differences or sample size limitations. Other The review protocol was registered on the Open Science Framework ( ) and no funding was provided.
... Caffeine withdrawal appears unnecessary to benefit from caffeine supplementation. caffeine intake to determine caffeine's ergogenic effects [25][26][27][28], while a substantial part of the published literature on caffeine supplementation appears not to report habitual intake of participants [29,30] or reports wide caffeine use (30-850 mg/day) [31]. This makes it difficult to determine whether habitual caffeine intake truly is an important factor that might modify the ergogenic response to caffeine supplementation. ...
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Objective The aim was to quantify the proportion of the literature on caffeine supplementation that reports habitual caffeine consumption, and determine the influence of habitual consumption on the acute exercise response to caffeine supplementation, using a systematic review and meta-analytic approach. Methods Three databases were searched, and articles screened according to inclusion/exclusion criteria. Three-level meta-analyses and meta-regression models were used to investigate the influence of habitual caffeine consumption on caffeine’s overall ergogenic effect and within different exercise types (endurance, power, strength), in men and women, and in trained and untrained individuals. Sub-analyses were performed according to the following: acute relative dose (< 3, 3–6, > 6 mg/kg body mass [BM]); whether the acute caffeine dose provided was lower or higher than the mean daily caffeine dose; and the caffeine withdrawal period prior to the intervention (< 24, 24–48, > 48 h). Results Sixty caffeine studies included sufficient information on habitual consumption to be included in the meta-analysis. A positive overall effect of caffeine was shown in comparison to placebo (standard mean difference [SMD] = 0.25, 95% confidence interval [CI] 0.20–0.30; p < 0.001) with no influence of relative habitual caffeine consumption (p = 0.59). Subgroup analyses showed a significant ergogenic effect when the caffeine dose was < 3 mg/kg BM (SMD = 0.26, 95% CI 0.12–0.40; p = 0.003) and 3–6 mg/kg BM (SMD = 0.26, 95% CI 0.21–0.32; p < 0.0001), but not > 6 mg/kg BM (SMD = 0.11, 95% CI − 0.07 to 0.30; p = 0.23); when the dose was both higher (SMD = 0.26, 95% CI 0.20–0.31; p < 0.001) and lower (SMD = 0.21, 95% CI 0.06–0.36; p = 0.006) than the habitual caffeine dose; and when withdrawal was < 24 h, 24–48 h, and > 48 h. Caffeine was effective for endurance, power, and strength exercise, with no influence (all p ≥ 0.23) of relative habitual caffeine consumption within exercise types. Habitual caffeine consumption did not modify the ergogenic effect of caffeine in male, female, trained or untrained individuals. Conclusion Habitual caffeine consumption does not appear to influence the acute ergogenic effect of caffeine.
... Lastly, it is unknown whether NO bioavailability (or other physiological mechanisms) is influenced when dietary NO 3 − is co-ingested with other ergogenic aids. In the limited available data thus far, no synergistic effects on exercise performance have been reported when beetroot juice was co-ingested with caffeine [81][82][83] or sodium bicarbonate [84], but further research is warranted to understand whether combining multiple ergogenic aids or other nutritional strategies with dietary NO 3 − could impact the effects of dietary NO 3 − on performance in various exercise modalities and participant populations. ...
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Dietary nitrate supplementation is evidenced to induce physiological effects on skeletal muscle function in fast-twitch muscle fibers and may enhance high-intensity exercise performance. An important component of sport-specific skills is the ability to perform explosive movements; however, it is unclear if nitrate supplementation can impact explosive efforts. We examined the existing evidence to determine whether nitrate supplementation improves explosive efforts lasting ≤ 6 s. PubMed, Scopus and Directory of Open Access Journals (DOAJ) were searched for articles using the following search strategy: (nitrate OR nitrite OR beetroot) AND (supplement OR supplementation) AND (explosive OR power OR high intensity OR high-intensity OR sprint* OR “athletic performance”). Out of 810 studies, 18 were eligible according to inclusion criteria. Results showed that 4 of the 10 sprint-type studies observed improved sprint time, power output, and total work in cycling or running, whereas 4 of the 10 resistance-based exercise studies observed improvements to power and velocity of free-weight bench press as well as isokinetic knee extension and flexion at certain angular velocities. These results suggest that nitrate potentially improves explosive exercise performance, but further work is required to clarify the factors influencing the efficacy of nitrate in different exercise modalities.
... Subjects chewed two boluses of gum similar to what was described by Lane and colleagues. 37 Each bolus of gum was chewed for at least 10 minutes based on the buccal absorption of caffeine. 38 Subjects initiated a standardized warm-up 10 minutes after the second bolus of gum, followed by the tactical combat movement simulation. ...
Background: Military personnel supplement caffeine as a countermeasure during unavoidable sustained wakefulness. However, its utility in combat-relevant tasks is unknown. This study examined the effects of caffeinated gum on performance in a tactical combat movement simulation. Materials and methods: Healthy men (n = 30) and women (n = 9) (age = 25.3 ± 6.8 years; mass 75.1 ± 13.1 kg) completed a marksmanship with a cognitive workload (CWL) assessment and a fire-andmove simulation (16 6-m bounds) in experimental conditions (placebo versus caffeinated gum, 4mg/kg). Susceptibility to enemy fire was modeled on bound duration during the fireand- move simulation. Results: Across both conditions, bound duration and susceptibility to enemy fire increased by 9.3% and 7.8%, respectively (p = .001). Cognitive performance decreased after the fire-and-move simulation across both conditions (p < .05). However, bound duration, susceptibility to enemy fire, marksmanship, and cognitive performance did not differ between the caffeine and placebo conditions. Conclusion: These data do not support a benefit of using caffeinated gum to improve simulated tactical combat movements.
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This study examined if acute dietary nitrate supplementation (140 mL beetroot juice, BRJ) would reduce oxygen consumption (V̇O 2 ) during cycling at power outputs of 50 and 70% maximal oxygen consumption in 14 well-trained female Canadian University Ringette League athletes. BRJ had no effect on V̇O 2 or heart rate but significantly reduced ratings of perceived exertion (RPE) at both intensities. Individually, 4 participants responded to BRJ supplementation with a ≥3% reduction in V̇O 2 at the higher power output. Novelty: Acute BRJ supplementation did not improve exercise economy in well-trained females, but significantly reduced RPE. However, 4/14 subjects did respond with a ≥3% V̇O 2 reduction.
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Dietary supplementation with beetroot juice (BR) containing ~5-8 mmol of inorganic nitrate (NO3(-)) increases plasma nitrite concentration ([NO2(-)]), reduces blood pressure, and may positively influence the physiological responses to exercise. However, the dose-response relationship between the volume of BR ingested and the physiological effects invoked has not been investigated. In a balanced crossover design, 10 healthy males ingested 70, 140 or 280 ml of concentrated BR (containing 4.2, 8.4 and 16.8 mmol NO3-, respectively) or no supplement to establish the effects of BR on resting plasma [NO3(-)] and [NO2(-)] over 24 h. Subsequently, on six separate occasions, 10 subjects completed moderate-intensity and severe-intensity cycle exercise tests 2.5 h post-ingestion of 70, 140 and 280 ml BR, or NO3(-)-depleted BR as placebo (PL). Following acute BR ingestion, plasma [NO2(-)] increased in a dose-dependent manner, with the peak changes occurring at ~2-3 h. Compared to PL, 70 ml BR did not alter the physiological responses to exercise. However, 140 and 280 ml BR reduced the steady-state VO2 during moderate-intensity exercise by 1.7% (P=0.06) and 3.0% (P<0.05), whilst time to task failure was extended by 14% and 12% (both P<0.05), respectively, compared to PL. The results indicate that, while plasma [NO2(-)] and the O2 cost of moderate-intensity exercise are improved dose-dependently with NO3(-)-rich BR, there is no additional improvement in exercise tolerance after ingesting BR containing 16.8 compared to 8.4 mmol NO3(-). These findings have important implications for the use of BR to enhance cardiovascular health and exercise performance in young adults.
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The aim of the current study was to determine the effects of dietary nitrate ingestion on parameters of submaximal and supramaximal exercise and time trial (TT) performance in trained kayakers. Eight male kayakers completed four exercise trials consisting of an initial discontinuous graded exercise test to exhaustion and three performance trials using a kayak ergometer. The performance trials were composed of 15 min paddling at 60% of maximum work rate, five x ten-second all-out sprints and a 1 km TT. The second and third trials were preceded by ingestion of either 70 ml nitrate-rich concentrated beetroot juice (BR) or tomato juice (placebo [PLA]) 3 h prior to exercise using a randomised cross-over design. Plasma nitrate (PLA: 33.8 ± 1.9 μM, BR: 152 ± 3.5 μM) and nitrite (PLA: 519.8 ± 25.8, BR: 687.9 ± 20 nM) were higher following ingestion of BR compared to PLA (both P < 0.001). VO2 during steady-state exercise was lower in the BR trial than in the PLA trial (P = 0.010). There was no difference in either peak power in the sprints (P = 0.590) or TT performance between conditions (PLA: 277 ± 5 s, BR: 276 ± 5 s, P = 0.539). Despite a reduction in VO2, BR ingestion appears to have no effect on repeated supramaximal sprint or 1 km TT kayaking performance. A smaller elevation in plasma nitrite following a single dose of nitrate and the individual variability in this response may partly account for these findings.
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The purpose of this review was to examine the effect of nitrate supplementation on exercise performance by systematic review and meta-analysis of controlled human studies. A search of four electronic databases and cross-referencing found 17 studies investigating the effect of inorganic nitrate supplementation on exercise performance that met the inclusion criteria. Beetroot juice and sodium nitrate were the most common supplements, with doses ranging from 300 - 600 mg nitrate and prescribed in a manner ranging from a single bolus to 15 days of regular ingestion. Pooled analysis showed a significant moderate benefit (ES = 0.79, 95% CI: 0.23 to 1.35) of nitrate supplementation on performance for time to exhaustion tests (p = 0.006). There was a small but insignificant beneficial effect on performance for time trials (ES = 0.11, 95% CI: -0.16 to 0.37) and graded exercise tests (ES = 0.26, 95% CI: -0.10 to 0.62). Qualitative analysis suggested that performance benefits are more often observed in inactive to recreationally active individuals and when a chronic loading of nitrate over several days is undertaken. Overall these results suggest that nitrate supplementation is associated with a moderate improvement in constant load time to exhaustion tasks. Despite not reaching statistical significance, the small positive effect on time trial or graded exercise performance may be meaningful in an elite sport context. More data are required to clarify the effect of nitrate supplementation on exercise performance and to elucidate the optimal way to implement supplementation.
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Although expert groups have developed guidelines for fluid intake during sports, there is debate about their real-world application. We reviewed the literature on self-selected hydration strategies during sporting competitions to determine what is apparently practical and valued by athletes. We found few studies of drinking practices involving elite or highly competitive athletes, even in popular sports. The available literature revealed wide variability in fluid intake and sweat losses across and within different events with varied strategies to allow fluid intake. Typical drinking practices appear to limit body mass (BM) losses to ~2 % in non-elite competitors. There are events, however, in which mean losses are greater, particularly among elite competitors and in hot weather, and evidence that individual participants fail to meet current guidelines by gaining BM or losing >2 % BM over the competition activity. Substantial (>5 %) BM loss is noted in the few studies of elite competitors in endurance and ultra-endurance events; while this may be consistent with winning outcomes, such observations cannot judge whether performance was optimal for that individual. A complex array of factors influence opportunities to drink during continuous competitive activities, many of which are outside the athlete's control: these include event rules and tactics, regulated availability of fluid, need to maintain optimal technique or speed, and gastrointestinal comfort. Therefore, it is questionable, particularly for top competitors, whether drinking can be truly ad libitum (defined as "whenever and in whatever volumes chosen by the athlete"). While there are variable relationships between fluid intake, fluid balance across races, and finishing times, in many situations it appears that top athletes take calculated risks in emphasizing the costs of drinking against the benefits. However, some non-elite competitors may need to be mindful of the disadvantages of drinking beyond requirements during long events. Across the sparse literature on competition hydration practices in other sports, there are examples of planned and/or ad hoc opportunities to consume fluid, where enhanced access to drinks may allow situations at least close to ad libitum drinking. However, this situation is not universal and, again, the complex array of factors that influence the opportunity to drink during an event is also often beyond the athletes' control. Additionally, some competition formats result in athletes commencing the event with a body fluid deficit because of their failure to rehydrate from a previous bout of training/competition or weight-making strategies. Finally, since fluids consumed during exercise may also be a source of other ingredients (e.g., carbohydrate, electrolytes, or caffeine) or characteristics (e.g., temperature) that can increase palatability or performance, there may be both desirable volumes and patterns of intake that are independent of hydration concerns or thirst, as well as benefits from undertaking a "paced" fluid plan. Further studies of real-life hydration practices in sports including information on motives for drinking or not, along with intervention studies that simulate the actual nature of real-life sport, are needed before conclusions can be made about ideal drinking strategies for sports. Different interpretations may be needed for elite competitors and recreational participants.
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It is presently unclear whether the reported ergogenic effect of a carbohydrate (CHO) mouth rinse on cycling time-trial performance is affected by the acute nutritional status of an individual. Hence, the aim of this study was to investigate the effect of a CHO mouth rinse on a 60-min simulated cycling time-trial performance commenced in a fed or fasted state. Twelve competitive male cyclists each completed 4 experimental trials using a double-blinded Latin square design. Two trials were commenced 2 h after a meal that contained 2.5 g·kg(-1) body mass of CHO (FED) and 2 after an overnight fast (FST). Prior to and after every 12.5% of total time during a performance ride, either a 10% maltodextrin (CHO) or a taste-matched placebo (PLB) solution was mouth rinsed for 10 s then immediately expectorated. There were significant main effects for both pre-ride nutritional status (FED vs. FST; p < 0.01) and CHO mouth rinse (CHO vs. PLB; p < 0.01) on power output with an interaction evident between the interventions (p < 0.05). The CHO mouth rinse improved mean power to a greater extent after an overnight fast (282 vs. 273 W, 3.4%; p < 0.01) compared with a fed state (286 vs. 281 W, 1.8%; p < 0.05). We concluded that a CHO mouth rinse improved performance to a greater extent in a fasted compared with a fed state; however, optimal performance was achieved in a fed state with the addition of a CHO mouth rinse.
Dietary nitrate supplementation, usually in the form of beetroot juice, has been heralded as a possible new ergogenic aid for sport and exercise performance. Early studies in recreationally active participants indicated that nitrate ingestion significantly reduces the O2 cost of submaximal exercise and improves performance during high-intensity endurance exercise. Subsequent studies have begun to address the physiological mechanisms underpinning these observations and to investigate the human populations in whom, and the exercise conditions (high- vs. low-intensity, long- vs. short-duration, continuous vs. intermittent, normoxic vs. hypoxic) under which, nitrate supplementation may be beneficial. Moreover, the optimal nitrate loading regimen in terms of nitrate dose and duration of supplementation has been explored. Depending on these factors, nitrate supplementation has been shown to exert physiological effects that could be conducive to exercise performance enhancement, at least in recreationally active or sub-élite athletes. This article provides a "state-of-the-art" review of the literature pertinent to the evaluation of the efficacy of nitrate supplementation in altering the physiological determinants of sport and exercise performance.
Purpose: Commencing selected workouts with low muscle glycogen availability augments several markers of training adaptation compared with undertaking the same sessions with normal glycogen content. However, low glycogen availability reduces the capacity to perform high-intensity (>85% of peak aerobic power (VO2 peak)) endurance exercise. We determined whether a low dose of caffeine could partially rescue the reduction in maximal self-selected power output observed when individuals commenced high-intensity interval training with low (LOW) compared with normal (NORM) glycogen availability. Methods: Twelve endurance-trained cyclists/triathletes performed four experimental trials using a double-blind Latin square design. Muscle glycogen content was manipulated via exercise-diet interventions so that two experimental trials were commenced with LOW and two with NORM muscle glycogen availability. Sixty minutes before an experimental trial, subjects ingested a capsule containing anhydrous caffeine (CAFF, 3 mg · kg(-1) body mass) or placebo (PLBO). Instantaneous power output was measured throughout high-intensity interval training (8 × 5-min bouts at maximum self-selected intensity with 1-min recovery). Results: There were significant main effects for both preexercise glycogen content and caffeine ingestion on power output. LOW reduced power output by approximately 8% compared with NORM (P < 0.01), whereas caffeine increased power output by 2.8% and 3.5% for NORM and LOW, respectively, (P < 0.01). Conclusion: We conclude that caffeine enhanced power output independently of muscle glycogen concentration but could not fully restore power output to levels commensurate with that when subjects commenced exercise with normal glycogen availability. However, the reported increase in power output does provide a likely performance benefit and may provide a means to further enhance the already augmented training response observed when selected sessions are commenced with reduced muscle glycogen availability.