Ci a ion: Vi ibay, A.;
Fe nández-Landa, J.;
Cas añeda-Baba o, A.; Collado, P.S.;
Fe nández-Láza o, D.; Mielgo-Ayuso,
J. E ec s o Ci ulline
Supplemen a ion on Di e en
Ae obic Exe cise Pe o mance
Ou comes: A Sys ema ic Re iew and
Me a-Analysis. Nu ien s 2022,14,
3479. h ps://doi.o g/10.3390/
nu14173479
Academic Edi o s: Isabel Sosped a
López and José
Miguel Ma ínez-Sanz
Recei ed: 1 Augus 2022
Accep ed: 21 Augus 2022
Published: 24 Augus 2022
Publishe ’s No e: MDPI s ays neu al
wi h ega d o ju isdic ional claims in
published maps and ins i u ional a il-
ia ions.
Copy igh : © 2022 by he au ho s.
Licensee MDPI, Basel, Swi ze land.
This a icle is an open access a icle
dis ibu ed unde he e ms and
condi ions o he C ea i e Commons
A ibu ion (CC BY) license (h ps://
c ea i ecommons.o g/licenses/by/
4.0/).
nu ien s
Sys ema ic Re iew
E ec s o Ci ulline Supplemen a ion on Di e en Ae obic
Exe cise Pe o mance Ou comes: A Sys ema ic Re iew and
Me a-Analysis
Ai o Vi ibay 1,2, Julen Fe nández-Landa 3, A kai z Cas añeda-Baba o 4,* , Pila S. Collado 2,
Diego Fe nández-Láza o 5,6 and Juan Mielgo-Ayuso 7,*
1Glu 4Science, Physiology, Nu i ion and Spo , 01004 Vi o ia-Gas eiz, Spain
2Ins i u e o Biomedicine (IBIOMED), Uni e si y o Leon, 24071 Leon, Spain
3Physical Educa ion and Spo s Depa men , Facul y o Educa ion and Spo , Uni e si y o he Basque
Coun y (UPV/EHU), 01007 Vi o ia-Gas eiz, Spain
4Heal h, Physical Ac i i y and Spo s Science Labo a o y, Depa men o Physical Ac i i y and Spo s,
Facul y o Educa ion and Spo , Uni e si y o Deus o, 48007 Bilbao, Spain
5Depa men o Cellula Biology, Gene ic, His ology and Pha macology, Facul y o Heal h Sciences,
Uni e si y o Valladolid, Campus o So ia, 42003 So ia, Spain
6Neu obiology Resea ch G oup, Facul y o Medicine, Uni e si y o Valladolid, 47005 Valladolid, Spain
7Depa men o Heal h Sciences, Facul y o Heal h Sciences, Uni e si y o Bu gos, 09001 Bu gos, Spain
*Co espondence: [email p o ec ed] (A.C.-B.); [email p o ec ed] (J.M.-A.)
Abs ac :
Supplemen a ion wi h Ci ulline (Ci ) has been shown o ha e a posi i e impac on ae obic
exe cise pe o mance and ela ed ou comes such as lac a e, oxygen up ake (VO
2
) kine ics, and he a e
o pe cei ed exe ion (RPE), p obably due o i s ela ionship o endogenous ni ic oxide p oduc ion.
Howe e , cu en esea ch has shown his o be con o e sial. The main objec i e o his sys ema ic
e iew and me a-analysis was o analyze and assess he e ec s o Ci supplemen a ion on ae obic
exe cise pe o mance and ela ed ou comes, as well as o show he mos sui able doses and iming
o inges ion. A s uc u ed li e a u e sea ch was ca ied ou by he PRISMA
®
(P e e ed Repo ing
I ems o Sys ema ic Re iews and Me a-Analyses) and PICOS guidelines in he ollowing da abases:
Pubmed/Medline, Scopus, and Web o Science (WOS). A o al o 10 s udies we e included in he
analysis, all o which exclusi ely compa ed he e ec s o Ci supplemen a ion wi h hose o a placebo
g oup on ae obic pe o mance, lac a e, VO
2
, and he RPE. Those a icles ha used o he supplemen s
and measu ed o he ou comes we e excluded. The me a-analysis was ca ied ou using Hedges’ g
andom e ec s model and pooled s anda dized mean di e ences (SMD). The esul s showed no
posi i e e ec s o Ci supplemen a ion on ae obic pe o mance (pooled SMD = 0.15; 95% CI (
−
0.02 o
0.32); I
2
, 0%; p= 0.08), he RPE (pooled SMD =
−
0.03; 95% CI (
−
0.43 o 0.38); I
2
, 49%; p= 0.9), VO
2
kine ics (pooled SMD = 0.01; 95% CI (
−
0.16 o 0.17); I
2
, 0%; p= 0.94), and lac a e (pooled SMD = 0.25;
95% CI (
−
0.10 o 0.59); I
2
, 0%; p= 0.16). In conclusion, Ci supplemen a ion did no p o e o ha e
any bene i s o ae obic exe cise pe o mance and ela ed ou comes. Whe e ch onic p o ocols seemed
o show a posi i e endency, mo e s udies in he ield a e needed o be e unde s and he e ec s.
Keywo ds: Ci ulline; e gogenic aids; physical pe o mance; ni ic oxide; ae obic; endu ance
1. In oduc ion
Ae obic o ca dio espi a o y endu ance exe cise places high physiological and me abolic
demands on he human body’s o ganic egula o y sys ems [
1
]. F om a physiological poin
o iew, pe o mance in ca dio espi a o y endu ance spo s, whe e he e o gene ally las s
longe han 5 min, depends p ima ily on h ee ela ed ac o s: maximal oxygen up ake
(VO
2
max), which can be de ined as he maximum oxygen-ca ying capaci y o a subjec ;
he indi idual lac a e h eshold (ILT), which has many names and is ela ed o di e en
physiological concep s bu which, in sho , e e s o he in ensi y o exe cise a which he
Nu ien s 2022,14, 3479. h ps://doi.o g/10.3390/nu14173479 h ps://www.mdpi.com/jou nal/nu ien s
Nu ien s 2022,14, 3479 2 o 28
concen a ion o lac a e in he blood inc eases exponen ially compa ed o basal le els due
o he inabili y o be oxidized in he mi ochond ia; and he e iciency o unning economy,
de ined as he oxygen consump ion o ene gy equi ed by an indi idual o main ain a
gi en unning pace [
2
–
6
]. In addi ion, he impo ance o psychological sensa ion is o
he u mos impo ance in spo , as i is conside ed a o m o bio eedback in eg a ed in o
he in o ma ion and e o egula ion sys em be ween he cen al command and he local
pe iphe al mechanisms [7,8].
In compe i i e spo , a 1–2% imp o emen in pe o mance can make he di e ence
be ween winning and losing a compe i ion and is, he e o e, a c i ical di e ence [
9
]. To
imp o e spo s pe o mance, a hle es wo k wi h complex, ca e ully planned aining and
nu i ion sys ems [
10
,
11
]. Nu i ion and supplemen a ion a e he e o e impo an ools in
op imizing pe o mance, especially in endu ance and ae obic disciplines [
6
,
12
]. In hei
sea ch o he ul ima e pe o mance imp o emen , a hle es o en make use o a a ie y o
spo s supplemen s [
13
]. Among he mos -used supplemen s, p o eins and amino acids
a e he mos consumed, wi h a equency o use o 35–40% [
14
]. Howe e , he use o
nu i ional supplemen s wi h unc ions ela ed o Ni ic Oxide (NO) syn hase has inc eased
conside ably in he ield o spo , due o e idence linking hei in ake o posi i e e ec s on
spo s pe o mance [12,15,16].
The e gogenic in e es in NO- ela ed supplemen s and hei di e en ways o ob ain-
ing hem ocuses on he po en ial e ec s ha his bioac i e compound can ha e on he
body. These include heal h-enhancing e ec s such as he egula ion o blood low and
blood p essu e, he main enance o gas ic in eg i y, and p o ec ion agains ischemic issue
damage [
17
–
19
]. They ha e also been ound o be ela ed o imp o ed pe o mance in
di e en ypes o disciplines and h ough di e en supplemen s, imp o ing asodila ion
and angiogenesis, causing an inc ease in mi ochond ial espi a ion and biogenesis, pos-
i i ely impac ing glucose up ake, op imizing oxygen egula ion, and imp o ing muscle
con ac ion [20–26].
Among he supplemen s linked o hese e ec s a e wo non-essen ial amino acids,
namely A ginine (A g) and Ci ulline (Ci ) [
16
,
27
]. Ci is an amino acid abundan ly
ound in wa e melon (Ci ullus ulga is) and can be syn hesized om A g and ano he
amino acid called O ni hine (O n), wi h plasma glu amine and A g i sel being he main
p ecu so s [27,28]
. Likewise, Ci is a p ecu so o A g [
29
,
30
]. The e o e, Ci plays an
essen ial ole in he bioa ailabili y o A g and also in he subsequen syn hesis o NO [
29
].
Howe e , unlike A g, Ci passes di ec ly in o he kidneys wi hou being ca abolized by he
enzyme a ginase [
31
]. Ci i sel can inhibi a ginase ac ion and limi he ca abolism o A g
o O n [
32
]. I is he e o e hypo hesized ha die a y Ci in ake migh be mo e e icien in
inc easing A g bioa ailabili y and NO syn hesis han A g supplemen a ion i sel [28,32].
Fo his eason, A g and Ci supplemen a ion has been used in a hle es om di e en
disciplines and condi ions o imp o e a hle ic pe o mance [
20
,
33
–
37
]. A ecen sys ema ic
e iew and me a-analysis [
24
] showed a posi i e e ec o A g supplemen a ion in di e en
spo s disciplines, depending on he ene gy me abolism used. Howe e , e idence o he
e ec o Ci on ae obic and ca dio espi a o y endu ance exe cise is inconsis en o da e.
The e a e insu icien s udies ha ha e quali a i ely and quan i a i ely analyzed i s e ec
on physical pe o mance and he di e en a iables in ol ed.
The physiological demands o endu ance spo s disciplines a e di ec ly ela ed o
he abili y o he ca dio ascula sys em o p o ide oxygen and nu ien s o he muscles,
and he unc ionali y o he pe iphe al (muscula ) s uc u es o ob ain he equi ed ene gy,
ha is, adenosine iphospha e (ATP), as e icien ly as possible. The e o e, he aining,
nu i ion, and supplemen a ion me hods used a e o pa icula impo ance o a hle es in
hese disciplines. They a e he e o e o g ea in e es conside ing he po en ial bene icial
e ec s ha he in ake o supplemen s ha exogenously and endogenously s imula e NO
syn hesis ha e in ela ion o he demands equi ed by long-du a ion exe cise [
20
]. The
inc eased syn hesis o his bioac i e compound could ul ima ely esul in mul iple bene icial
e ec s [17,20,38–40].
Nu ien s 2022,14, 3479 3 o 28
Despi e he p omising e idence o Ci supplemen a ion being used in bo h high
in ensi y, powe , and s eng h exe cise [
41
,
42
], as well as in long- e m exe cise [
43
–
45
],
he e a e no conclusi e esul s abou i s possible e ec s. Al hough acu e supplemen a ion
wi h Ci (single o mul iple doses wi hin a single day) has been shown o signi ican ly
inc ease plasma le els o A g and NO, no imp o emen in a hle ic pe o mance has been
obse ed [
46
,
47
]. Simila ly, no e ec was shown on VO
2
max, anae obic h eshold, ime
o exhaus ion, and he pe o mance pa ame e s analyzed a e doses o Ci o be ween
6 and 12 g
adminis e ed 1–2 h be o e exe cise [
43
,
44
,
48
]. In con as , supplemen a ion using
ch onic p o ocols has shown mo e p omising esul s in ela ion o endu ance pe o mance
a iables. Fo example, 7-day Ci supplemen a ion o 2.4 g/day and 6 g/day o Ci
imp o ed pe o mance in a 4 km ime ial [
49
] and VO
2
kine ics and a igue ole ance in
s enuous exe cise on a cycle-e gome e [
45
], espec i ely. Howe e , one aspec o conside
in he in e p e a ion o hese esul s is he le el o he pa icipan s in he s udies. The bes
esul s ega ding supplemen a ion wi h e gogenic aids ollowing he NO ou e ha e been
mainly ela ed o un ained subjec s o o pa icipan s who a e a low compe i i e le els,
whe eas no majo di e ences ha e been ound in a hle es wi h high aining le els [20].
Wi h his in mind, his sys ema ic e iew and me a-analysis aim o quali a i ely and
quan i a i ely analyze he impac o Ci supplemen a ion on ae obic pe o mance and
ela ed a iables such as lac a e, VO
2
kine ics, and he RPE o analyze he cu en e idence
in an o de ly and sys ema ic way, o ex end he cu en unde s anding o he e icacy,
e ec s, and mechanisms behind spo s supplemen a ion.
2. Me hods
2.1. Li e a u e Sea ch S a egies
This sys ema ic e iew and me a-analysis we e conduc ed by he PRISMA (P e e ed
Repo ing I ems o Sys ema ic Re iews and Me a-Analyses) guidelines [
50
] and he PICOS
inclusion c i e ia de ini ion model: “P” (sample o popula ion): “ac i e”; I (in e en ion):
“impac o Ci ulline on spo s pe o mance”; C (compa ison): “Ci ulline supplemen a ion
compa ed o Placebo”; O (ou comes): “ae obic spo s pe o mance and ela ed a iables”;
S (s udy design): “double-blind, c osso e , andomized, clinical ial” [
51
]. A e iew o he
scien i ic li e a u e was ca ied ou , including hose s udies ha analyzed he e ec o Ci
supplemen a ion on ae obic spo s pe o mance and i s ela ed a iables du ing exe cise,
namely, lac a e, he a e o pe cei ed exe ion (RPE), and oxygen up ake kine ics (VO
2
).
The sea ch was conduc ed using he espec i e sea ch engines o he di e en scien i ic
da abases, wi hou applying any empo al il e , hus co e ing all en ies om he i s
s udies on he subjec in 2006 un il he la es published o da e in 2021. Speci ically, he
PubMed/MEDLINE, Web o Science (WOS), and Scopus da abases we e used, using he
ollowing Boolean sea ch equa ion: L-Ci ulline [All Fields] OR “Ci ulline” [All Fields]
OR “wa e melon juice” [All Fields] AND supplemen a ion [All Fields] AND ((“spo s”
[MeSH Te ms] OR “spo s” [All Fields] OR “spo ” [All Fields] OR “spo ” [All Fields])
OR (“exe cise” [MeSH Te ms] OR “exe cise” [All Fields])) AND (“endu ance” [All Fields]
OR “pe o mance” [All Fields] OR “ae obic” [All Fields] OR “lac a e” [All Fields] OR
“pe cei ed exe ion” [All Fields] OR “oxygen kine ics” [All Fields]).
Using his equa ion, ele an a icles in he ield we e selec ed by applying a “snowball”
sea ch s a egy h ough he e e ences ob ained in he ini ial sea ch. All i les and abs ac s
we e c oss-checked and analyzed o iden i y duplica es o any po en ial e o s, as shown in
he low cha . To do his, he “duplica e sea ch” op ion in Mendeley Re e ence Manage
was used. All i les and abs ac s we e also sc eened o conside whe he eading he ull
a icles would be necessa y. The sea ch, sc eening, and selec ion o published s udies we e
ca ied ou independen ly by wo au ho s (A.V.M. and J.M.-A.), acco ding o he p oposed
inclusion and exclusion c i e ia.
Nu ien s 2022,14, 3479 4 o 28
2.2. Inclusion and Exclusion C i e ia
The inclusion c i e ia applied in his sys ema ic e iew and me a-analysis o he
selec ion o he a icles o be conside ed in he s udy we e as ollows:
- S udies pe o med on humans.
-
S udies wi h an app op ia e design, using a double-blind, andomized, c osso e
me hodology.
-
S udies wi h 2 well-di e en ia ed g oups compa ing he e ec s o Ci ulline supple-
men a ion (expe imen al g oup) e sus a placebo (con ol g oup).
-
S udies we e conduc ed on ac i e people, ec ea ional a hle es, well- ained a hle es,
and eli e o heal hy p o essional a hle es, bo h men and women.
-
S udies in which he compa ison o placebo e sus supplemen a ion was only wi h
Ci ulline and o he possible masking compounds, a oiding he in ake o di e en
e gonu i ional aids a he same ime.
- S udies speci ying he amoun and ype o supplemen a ion used.
-
S udies measu ing ae obic physical pe o mance using con inuous es s las ing 5 min
o mo e, ollowing he c i e ia ela ing o he measu emen o VO2max.
- S udies measu ing blood lac a e, he RPE, and di e en a iables o VO2kine ics.
- S udies published in any language.
- S udies o which he ull manusc ip was a ailable.
The ollowing exclusion c i e ia we e used o disca d a icles ound in he ini ial sea ch:
-
S udies in which supplemen a ion did no in ol e Ci ulline alone, including hose
using a combina ion o di e en ni ic oxide pa hway supplemen s.
-
S udies in which he e was no eal p esence o a con ol g oup inges ing a subs ance
conside ed o be a placebo.
- S udies in which o he supplemen s o e gonu i ional aids ha migh in e e e wi h
pe o mance we e used in he in e en ions, conside ing he scien i ic e idence in he
li e a u e, such as Ca eine, Be a-Alanine, C ea ine, e c.
-
S udies in which pe o mance was measu ed using es s las ing less han 5 min and/o
measu ing a iables ela ed o anae obic pe o mance.
- S udies conduc ed on an unheal hy, sick, o inju ed popula ion.
Finally, no il e s we e applied conce ning he gende , le el, o spo ing discipline
o he subjec s pa icipa ing in he s udies, and any esea ch ha analyzed he p oposed
a iables in any si ua ion ha me he inclusion c i e ia was conside ed o be alid.
2.3. S udy Selec ion
A e sea ching, he a ious s udies we e sc eened and unila e ally selec ed o analysis
by he au ho . A e applying he inclusion and exclusion c i e ia, da a we e ex ac ed om
each s udy as shown in he “Resul s” sec ion. The sou ce o he s udies, including au ho s,
yea o publica ion, and ci a ion in o ma ion, he sample size and cha ac e is ics o he
pa icipan s (le el, age, and gende ), he adminis a ion o he supplemen o be analyzed
and he placebo o be con olled o (dose, iming), he inal a iables o be e iewed
( es , in ensi y, and cha ac e is ics), and, inally, he main conclusions o each s udy we e
ex ac ed by he au ho wi h he help o a sp eadshee (Mic oso Inc., Sea le, WA, USA).
Subsequen ly, he da a p esen ed we e analyzed and se e al s udies we e excluded o
ele an easons un il he s udies o be included in his sys ema ic e iew and me a-analysis
we e ob ained.
2.4. Measu ing Va iables
The a ailable scien i ic li e a u e on Ci supplemen a ion and i s e ec on ae obic
physical exe cise was examined in e ms o pe o mance in he physical es s conduc ed,
he analysis o lac a e as a de e mining ene gy me aboli e, he RPE, and VO
2
kine ics as a
physiological ac o limi ing ae obic pe o mance. To quan i y ae obic spo s pe o mance,
measu emen a iables classi ied acco ding o he du a ion o he es s we e used. The
Nu ien s 2022,14, 3479 5 o 28
5 min es appea ed o be a eliable es o de e mining maximal ae obic speed, ela ed
o VO
2
max, and he e o e sugges ed he ime ha a subjec can main ain he lowes
in ensi y ela ed o his physiological equi emen [
52
,
53
]. Taking in o accoun ha VO
2
max ma ks a subjec ’s maximum ae obic powe , he au ho es ablished es s
las ing ≤5 min
as ime c i e ia o analyzing ae obic spo s pe o mance in his sys ema ic e iew and
me a-analysis.
Mo e speci ically, se e al es s we e included in he pe o mance analysis o de e mine
ae obic pe o mance, bo h on he cyclo-e gome e and eadmill, acco ding o he p oposed
c i e ia: inc emen al and sus ained es s o exhaus ion; B uce p o ocol; eadmill es a
sub-maximal in ensi y (40% o HHR); cons an wo k es a 70% o VO
2
max; mode a e
in ensi y es (90% o HHR); cons an wo k es a 70% o VO
2
max; mode a e in ensi y es
(90% o VO
2
max); mode a e in ensi y es (90% o GET) ollowed by a high-in ensi y es on
he day (GET + di e ence be ween GET and VO
2
max wo k); 6 min cons an wo k es (70%
VO
2
max) ollowed by a 30 s all-ou es a 70% o VO
2
max; a 30 s all-ou es ; 10 epe i ions
o 15 s a maximum in ensi y (sp in ); 5 min es and an inc emen al es o exhaus ion a
100% o indi idual peak powe ; a hal -ma a hon ace; a 4 km and 40 km indi idual ime
ial; and an SRT es comp ising a 6 ×1-min sp in a 120% o maximum powe .
Likewise, o he analysis o lac a e, he measu emen o his bioma ke was analyzed
in he di e en s udies in he same way as o he RPE, using he Modi ied Bo g Scale (0–10)
o Bo g Ra ing o Pe cei ed Exe ion Scale (6–20). In u n, di e en ela ed a iables we e
used o analyze he esponses o VO
2
kine ics. These included: ne oxygen expendi u e,
mean esponse ime (MRT), oxygen de ici , mean pulmona y VO
2
esponse ime, mean
o e all pulmona y gas esponse ime, issue oxygena ion index, muscle oxygena ion,
pulmona y VO2, VO2max, gas exchange de ined h eshold, and peak VO2.
Fo he s a is ical analysis o he quan i a i e syn hesis (me a-analysis), he sample
size, means, and s anda d de ia ions o he di e en a iables we e ex ac ed o he wo
g oups compa ed: he Ci supplemen a ion g oup and he placebo g oup, espec i ely. In
hose cases whe e da a we e no ound in a nume ical o m, he esul s we e ex ac ed by
analyzing he igu es and g aphs using he Re iew Manage (Re man) 5.4 (Copenhagen:
The No dic Coch ane Cen e, The Coch ane Collabo a ion, Copenhagen, Denma k), which
es ima es he esul s based on he p ocess o coding he pixels o he espec i e images.
2.5. Publica ion Bias
Publica ion biases we e assessed using Egge ’s s a is ical es , whe eby publica ion
bias occu s when p< 0.05 [
54
]. To isualize he calcula ion and in e p e he esul s, he
ollowing unnel plo s we e pe o med, ollowed by Egge ’s s a is ic o con i m o e u e
publica ion bias o each a icle included in his sys ema ic e iew (Figu e 1). Egge ’s
analysis and he unnel plo s sugges low publica ion bias in he 4 ca ego ies analyzed.
2.6. Assessing he Quali y o In es iga ions
This sys ema ic e iew was p epa ed by he Coch ane Collabo a ion Guidelines [
55
].
The quali y assessmen o he di e en a icles was ca ied ou independen ly by wo
au ho s (A.V.M. and J.M.-A.). The isk o bias analysis was sepa a ed in o six dis inc
domains, as shown in Figu e 2: selec ion biases ( andom sequence gene a ion, alloca ion
concealmen ); biased elabo a ion (blinding o pa icipan s and esea che s); de ec ion bias
(blinding o ou come assessmen ); a i ion bias (incomple e ou come da a); epo ing bias
(selec i e epo ing); and o he ypes o biases. Rega ding he assessmen o he domains,
i was conside ed o be: “low” i he c i e ion had a low isk o bias and i was unlikely
o al e he esul s, o “high” i he c i e ion had a high isk o bias and i was likely o
se iously unde mine he esul s. I he isks o bias we e no known, hey we e ca ego ized
by labeling hem as “unknown”, which signi ies doub s as o he esul s.
Nu ien s 2022,14, 3479 6 o 28
Nu ien s 2022, 14, x FOR PEER REVIEW 6 o 30
(a) (b)
(c) (d)
Figu e 1. Funnel plo s o he s anda d e o o ae obic (a) he RPE, (b) VO2 kine ics, (c) lac a e, and
(d) spo s pe o mance da a, using Hedges’ g. SE: s anda d e o ; SMD: s anda d mean di e ence.
2.6. Assessing he Quali y o In es iga ions
This sys ema ic e iew was p epa ed by he Coch ane Collabo a ion Guidelines [55].
The quali y assessmen o he di e en a icles was ca ied ou independen ly by wo au-
ho s (A.V.M. and J.M.-A.). The isk o bias analysis was sepa a ed in o six dis inc do-
mains, as shown in Figu e 2: selec ion biases ( andom sequence gene a ion, alloca ion con-
cealmen ); biased elabo a ion (blinding o pa icipan s and esea che s); de ec ion bias
(blinding o ou come assessmen ); a i ion bias (incomple e ou come da a); epo ing bias
(selec i e epo ing); and o he ypes o biases. Rega ding he assessmen o he domains,
i was conside ed o be: “low” i he c i e ion had a low isk o bias and i was unlikely o
al e he esul s, o “high” i he c i e ion had a high isk o bias and i was likely o se i-
ously unde mine he esul s. I he isks o bias we e no known, hey we e ca ego ized
by labeling hem as “unknown”, which signi ies doub s as o he esul s.
Figu e 1.
Funnel plo s o he s anda d e o o ae obic (
a
) he RPE, (
b
) VO
2
kine ics, (
c
) lac a e, and
(d) spo s pe o mance da a, using Hedges’ g. SE: s anda d e o ; SMD: s anda d mean di e ence.
Nu ien s 2022, 14, x FOR PEER REVIEW 6 o 30
(a) (b)
(c) (d)
Figu e 1. Funnel plo s o he s anda d e o o ae obic (a) he RPE, (b) VO2 kine ics, (c) lac a e, and
(d) spo s pe o mance da a, using Hedges’ g. SE: s anda d e o ; SMD: s anda d mean di e ence.
2.6. Assessing he Quali y o In es iga ions
This sys ema ic e iew was p epa ed by he Coch ane Collabo a ion Guidelines [55].
The quali y assessmen o he di e en a icles was ca ied ou independen ly by wo au-
ho s (A.V.M. and J.M.-A.). The isk o bias analysis was sepa a ed in o six dis inc do-
mains, as shown in Figu e 2: selec ion biases ( andom sequence gene a ion, alloca ion con-
cealmen ); biased elabo a ion (blinding o pa icipan s and esea che s); de ec ion bias
(blinding o ou come assessmen ); a i ion bias (incomple e ou come da a); epo ing bias
(selec i e epo ing); and o he ypes o biases. Rega ding he assessmen o he domains,
i was conside ed o be: “low” i he c i e ion had a low isk o bias and i was unlikely o
al e he esul s, o “high” i he c i e ion had a high isk o bias and i was likely o se i-
ously unde mine he esul s. I he isks o bias we e no known, hey we e ca ego ized
by labeling hem as “unknown”, which signi ies doub s as o he esul s.
Figu e 2. Risk-o -bias g aph exp essed as pe cen ages.
Figu e 3, below, is a clea isual ep esen a ion o he analysis bias o each a icle
included in his sys ema ic e iew and me a-analysis.
Nu ien s 2022,14, 3479 7 o 28
Nu ien s 2022, 14, x FOR PEER REVIEW 7 o 30
Figu e 2. Risk-o -bias g aph exp essed as pe cen ages.
Figu e 3, below, is a clea isual ep esen a ion o he analysis bias o each a icle
included in his sys ema ic e iew and me a-analysis.
Figu e 3. Summa y o he isk o bias. Re iew o he au ho s’ j o he isk o bias in he di e en
i ems selec ed om each a icle included in he sys ema ic e iew and me a-analysis. indica es a
low isk o bias; indica es an unknown isk o bias [43–45,48,49,56–60].
3. Resul s
3.1. Main Sea ch
The ini ial li e a u e sea ch using he elec onic da abases showed a o al o 94 ec-
o ds ma ching he Boolean sea ch engines p oposed in he sea ch equa ion. Addi ionally,
eigh eco ds we e ound using o he e e ence lis s, yielding an ini ial o al numbe o
102 a icles. Howe e , only 10 me all o he inclusion and exclusion c i e ia and we e
he e o e included in he sys ema ic e iew and me a-analysis. Following he p ocess o
duplica e analysis, 17 a icles we e excluded, lea ing 85 e e ences o sc eening. O hese,
28 we e excluded o di e en easons: 21 e e ences used a non-human sample and se en
we e ei he na a i e o sys ema ic e iews and/o me a-analyses. The e o e, 57 eco ds
we e sc eened o ull- ex analysis and assessed o eligibili y. A e he assessmen , a
o al o 48 eco ds we e excluded o a ious easons ela ed o he inclusion and exclusion
c i e ia. O hese, 19 s udies we e ejec ed because hey had used mo e han one
Figu e 3.
Summa y o he isk o bias. Re iew o he au ho s’ j o he isk o bias in he di e en i ems
selec ed om each a icle included in he sys ema ic e iew and me a-analysis.
Nu ien s 2022, 14, x FOR PEER REVIEW 7 o 30
Figu e 2. Risk-o -bias g aph exp essed as pe cen ages.
Figu e 3, below, is a clea isual ep esen a ion o he analysis bias o each a icle
included in his sys ema ic e iew and me a-analysis.
Figu e 3. Summa y o he isk o bias. Re iew o he au ho s’ j o he isk o bias in he di e en
i ems selec ed om each a icle included in he sys ema ic e iew and me a-analysis. indica es a
low isk o bias; indica es an unknown isk o bias [43–45,48,49,56–60].
3. Resul s
3.1. Main Sea ch
The ini ial li e a u e sea ch using he elec onic da abases showed a o al o 94 ec-
o ds ma ching he Boolean sea ch engines p oposed in he sea ch equa ion. Addi ionally,
eigh eco ds we e ound using o he e e ence lis s, yielding an ini ial o al numbe o
102 a icles. Howe e , only 10 me all o he inclusion and exclusion c i e ia and we e
he e o e included in he sys ema ic e iew and me a-analysis. Following he p ocess o
duplica e analysis, 17 a icles we e excluded, lea ing 85 e e ences o sc eening. O hese,
28 we e excluded o di e en easons: 21 e e ences used a non-human sample and se en
we e ei he na a i e o sys ema ic e iews and/o me a-analyses. The e o e, 57 eco ds
we e sc eened o ull- ex analysis and assessed o eligibili y. A e he assessmen , a
o al o 48 eco ds we e excluded o a ious easons ela ed o he inclusion and exclusion
c i e ia. O hese, 19 s udies we e ejec ed because hey had used mo e han one
indica es a low
isk o bias;
Nu ien s 2022, 14, x FOR PEER REVIEW 7 o 30
Figu e 2. Risk-o -bias g aph exp essed as pe cen ages.
Figu e 3, below, is a clea isual ep esen a ion o he analysis bias o each a icle
included in his sys ema ic e iew and me a-analysis.
Figu e 3. Summa y o he isk o bias. Re iew o he au ho s’ j o he isk o bias in he di e en
i ems selec ed om each a icle included in he sys ema ic e iew and me a-analysis. indica es a
low isk o bias; indica es an unknown isk o bias [43–45,48,49,56–60].
3. Resul s
3.1. Main Sea ch
The ini ial li e a u e sea ch using he elec onic da abases showed a o al o 94 ec-
o ds ma ching he Boolean sea ch engines p oposed in he sea ch equa ion. Addi ionally,
eigh eco ds we e ound using o he e e ence lis s, yielding an ini ial o al numbe o
102 a icles. Howe e , only 10 me all o he inclusion and exclusion c i e ia and we e
he e o e included in he sys ema ic e iew and me a-analysis. Following he p ocess o
duplica e analysis, 17 a icles we e excluded, lea ing 85 e e ences o sc eening. O hese,
28 we e excluded o di e en easons: 21 e e ences used a non-human sample and se en
we e ei he na a i e o sys ema ic e iews and/o me a-analyses. The e o e, 57 eco ds
we e sc eened o ull- ex analysis and assessed o eligibili y. A e he assessmen , a
o al o 48 eco ds we e excluded o a ious easons ela ed o he inclusion and exclusion
c i e ia. O hese, 19 s udies we e ejec ed because hey had used mo e han one
indica es an unknown isk o bias [43–45,48,49,56–60].
3. Resul s
3.1. Main Sea ch
The ini ial li e a u e sea ch using he elec onic da abases showed a o al o 94 eco ds
ma ching he Boolean sea ch engines p oposed in he sea ch equa ion. Addi ionally,
eigh eco ds we e ound using o he e e ence lis s, yielding an ini ial o al numbe o
102 a icles. Howe e , only 10 me all o he inclusion and exclusion c i e ia and we e
he e o e included in he sys ema ic e iew and me a-analysis. Following he p ocess o
duplica e analysis, 17 a icles we e excluded, lea ing 85 e e ences o sc eening. O hese,
28 we e excluded o di e en easons: 21 e e ences used a non-human sample and se en
we e ei he na a i e o sys ema ic e iews and/o me a-analyses. The e o e, 57 eco ds
we e sc eened o ull- ex analysis and assessed o eligibili y. A e he assessmen , a
o al o 48 eco ds we e excluded o a ious easons ela ed o he inclusion and exclusion
c i e ia. O hese, 19 s udies we e ejec ed because hey had used mo e han one supplemen
in conjunc ion wi h he ILC, nine s udies we e excluded because hey employed o he
un ela ed supplemen s, six we e elimina ed because hey used subjec s un ela ed o he
inclusion c i e ia, and 14 we e excluded because hey analyzed a iables un ela ed o
hose p oposed in he me hodology o his pape . Following his sui abili y analysis,
Nu ien s 2022,14, 3479 8 o 28
10 a icles we e chosen o quali a i e syn hesis (sys ema ic e iew). Subsequen ly, a e
analyzing and ex ac ing he nume ical and/o image da a, hese 10 we e included in
he inal quan i a i e syn hesis (me a-analysis). The e o e, he 10 a icles ha me all he
p oposed inclusion and exclusion c i e ia we e included in bo h he sys ema ic e iew and
he me a-analysis [43–45,48,49,56–60].
The low cha in Figu e 4, below, shows he sequence o ac ions ha we e ca ied ou
du ing he main sea ch, ollowing he PRISMA guidelines [
50
] and jus i ying he easons
o he exclusion o he a icles emo ed om he syn hesis.
Nu ien s 2022, 14, x FOR PEER REVIEW 8 o 30
supplemen in conjunc ion wi h he ILC, nine s udies we e excluded because hey em-
ployed o he un ela ed supplemen s, six we e elimina ed because hey used subjec s un-
ela ed o he inclusion c i e ia, and 14 we e excluded because hey analyzed a iables
un ela ed o hose p oposed in he me hodology o his pape . Following his sui abili y
analysis, 10 a icles we e chosen o quali a i e syn hesis (sys ema ic e iew). Subse-
quen ly, a e analyzing and ex ac ing he nume ical and/o image da a, hese 10 we e
included in he inal quan i a i e syn hesis (me a-analysis). The e o e, he 10 a icles ha
me all he p oposed inclusion and exclusion c i e ia we e included in bo h he sys ema ic
e iew and he me a-analysis [43–45,48,49,56–60].
The low cha in Figu e 4, below, shows he sequence o ac ions ha we e ca ied
ou du ing he main sea ch, ollowing he PRISMA guidelines [50] and jus i ying he ea-
sons o he exclusion o he a icles emo ed om he syn hesis.
Figu e 4. Flowcha o he p ocess o selec ion, sc eening, sui abili y, and inclusion o a icles in-
cluded in he sys ema ic e iew and me a-analysis. Adap ed om PRISMA guidelines [50].
3.2. S udy Cha ac e is ics
Table 1 shows he s udy cha ac e is ics (design, gende o he subjec s, as well as he
ype, dose, and ime o supplemen a ion) o he s udies included in he sys ema ic e iew
and me a-analysis. The o al sample analyzed comp ised 10 a icles o aling 173 subjec s.
All 10 a icles used a double-blind, andomized, placebo-con olled, c osso e me hodol-
ogy [43–45,48,49,56–60]. None o he s udies p esen ed any con lic o in e es conce ning
hei in e en ion and he supplemen a ion used. O he 10 s udies analyzed, ou o hem
in ol ed ec ea ional a hle es [45,57–59], h ee we e well- ained and/o eli e spo speo-
ple [44,49,60], wo s udies in ol ed ac i e adul s [43,56], and he emaining a icle was
conce ned wi h in e collegia e a hle es [48]. In he 10 a icles [43–45,48,49,56–60], wo com-
pa a i e g oups (Ci and placebo) we e sepa a ed, while in wo o hem, an addi ional
Figu e 4.
Flowcha o he p ocess o selec ion, sc eening, sui abili y, and inclusion o a icles included
in he sys ema ic e iew and me a-analysis. Adap ed om PRISMA guidelines [50].
3.2. S udy Cha ac e is ics
Table 1shows he s udy cha ac e is ics (design, gende o he subjec s, as well as he
ype, dose, and ime o supplemen a ion) o he s udies included in he sys ema ic e iew
and me a-analysis. The o al sample analyzed comp ised 10 a icles o aling 173 subjec s.
All 10 a icles used a double-blind, andomized, placebo-con olled, c osso e me hodol-
ogy [
43
–
45
,
48
,
49
,
56
–
60
]. None o he s udies p esen ed any con lic o in e es conce ning
hei in e en ion and he supplemen a ion used. O he 10 s udies analyzed, ou o hem
in ol ed ec ea ional a hle es [
45
,
57
–
59
], h ee we e well- ained and/o eli e spo speo-
ple [
44
,
49
,
60
], wo s udies in ol ed ac i e adul s [
43
,
56
], and he emaining a icle was
conce ned wi h in e collegia e a hle es [
48
]. In he 10 a icles [
43
–
45
,
48
,
49
,
56
–
60
], wo
compa a i e g oups (Ci and placebo) we e sepa a ed, while in wo o hem, an addi ional
g oup was used o compa e di e en dosages [
43
,
48
]. A g oup was used in one o hem o
compa e he iming o Ci in ake and he ime o inges ion [43].
Nu ien s 2022,14, 3479 9 o 28
Table 1.
Design, cha ac e is ics, and gende o subjec s. Type, dose, and iming o supplemen a ion
o he s udies ha we e included in he sys ema ic e iew and me a-analysis.
S udy design Randomized, double-blind, c osso e , and
placebo-con olled
10 s udies
[43–45,48,49,56–60]
Cha ac e is ics o he
subjec s
Rec ea ional 4 s udies [45,57–59]
In e collegia e 1 s udy [48]
Well- ained 3 s udies [44,49,60]
Ac i e 2 s udies [43,56]
Gende o subjec s
Male 7 s udies
[44,45,49,57–60]
Female 0 s udies
Bo h 3 s udies [43,48,56]
Type o Ci ulline
supplemen a ion
L-Ci ulline 6 s udies
[44,45,49,57,59,60]
Wa e melon Juice 1 s udy [48]
Wa e melon Juice + L-Ci ulline 2 s udies [57,59]
Ci ulline Mala e 2 s udies [44,58]
Dose used
1 g o supplemen 1 s udy [48]
2.4 g o supplemen 1 s udy [49]
3 g o supplemen 1 s udy [43]
3.45 g o supplemen 2 s udies [57,59]
6 g o supplemen 4 s udies [45,48,56,60]
8 g o supplemen 1 s udy [58]
9 g o supplemen 1 s udy [43]
12 g o supplemen 1 s udy [44]
In ake ime
Acu e
1 h be o e he es 2 s udy [44,58]
1–2 h be o e he es 1 s udy [48]
2 h be o e 1 s udy [59]
3 h be o e 1 s udy [43]
Ch onic
1 day 1 s udy [43]
6 days 1 s udy [45]
7 days 3 s udies [49,56,60]
16 days 1 s udy [57]
A o al o i e s udies used an acu e dose o supplemen a ion [
43
,
44
,
48
,
58
,
59
], o which
wo used Ci 1 h be o e he es [
44
,
61
], one s udy 1 h o 2 h be o e he expe imen al
es [
48
], ano he s udy 2 h be o e, [
59
] and he emaining s udy 3 h be o e he es [
43
]. In
con as , six s udies used a ch onic supplemen a ion me hodology anging om 1 day o
16 days [
43
,
45
,
56
,
57
,
59
,
62
]. O hese, Hickne e al. [
43
] used 1-day supplemen a ion, while
Bailey e al. [
45
] used 6 days o supplemen a ion. Th ee s udies [
49
,
56
,
60
] supplemen ed
he Ci o 7 days, and he emaining [57] o a longe pe iod o 16 days.
In all he s udies e iewed, he dose o Ci supplemen a ion was absolu e, wi h no
s udies using doses ela i e o body mass o o he a iables, as is he case wi h o he
supplemen s [
63
]. L-Ci ulline was used as a supplemen in six s udies [
43
,
45
,
48
,
49
,
56
,
60
],
while in one [
63
], pa icipan s inges ed wa e melon juice. In wo o he
s udies [57,59]
,
a mix u e o bo h supplemen s (wa e melon juice en iched wi h L-Ci ulline) was used,
whe eas he wo emaining s udies [
44
,
58
] in ol ed an in ake o Ci ulline Mala e. Rega d-
Nu ien s 2022,14, 3479 16 o 28
Table 4. Summa y o s udies included in he sys ema ic e iew and me a-analysis in es iga ing he e ec o IAC on oxygen up ake kine ics (VO2).
Au ho /s Popula ion In e en ion Tes Va iables Analyzed Main Conclusions
Ashley e al., 2018 [56]
Fi een ac i e adul s
(8 women, 7 men)
(22 ±2 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 6 g/day L-Ci ulline
o 7 days.
T eadmill unning a 40%
HRR. Measu emen o
oxygen consump ion by
indi ec calo ime y.
•Ne oxygen expendi u e
g oup 1
•
Mean Response Time (MRT)
g oup 1
•Oxygen de iciency g oup 1
↔
↑
↑
Ele en ac i e adul s
(7 women, 4 men)
(74 ±7 yea s).
•Ne oxygen expendi u e
g oup 2
•
Mean Response Time (MRT)
g oup 2
•Oxygen de iciency g oup 2
↔
↔
↔
Bailey e al., 2015 [45]
Ten heal hy, ac i e,
ec ea ional subjec s. Men
(19 ±1 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 6 g L-Ci ulline +
4.3 g mal odex in (wi h 500 mL wa e )
o 6 days + 90 min be o e he es .
Two cycle-e gome e es s:
1 mode a e in ensi y es on
day 6 (90% o GET) + 1
high-in ensi y es on day 7
(GET + di e ence be ween
GET and VO2max wo k).
•Pulmona y VO2Mean
Response Time (s)
•
Mean Response Time (MRT)
g oup 2.
•Tissue oxygena ion
index (%)
↑
↑
↑
Bailey e al., 2016 [57]
Eigh heal hy, ac i e,
ec ea ional subjec s. Men (22
±2 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled. ~3.4 g/day o
L-Ci ulline in 300 mL o Wa e melon
Juice Concen a e o 16 days.
Cons an cycle-e gome e
wo ks a 70% o VO2max o
exhaus ion.
•Muscle oxygena ion
•Pulmona y VO2
↑
↔
Six min o cons an wo k
(70% VO2max) + 300all-ou
on cycle-e gome e .
•Muscle oxygena ion
•Pulmona yVO2
↑
↔
Nu ien s 2022,14, 3479 17 o 28
Table 4. Con .
Au ho /s Popula ion In e en ion Tes Va iables Analyzed Main Conclusions
Cu u ello e al., 2014
[48]
Ele en heal hy men
(20.6 ±1.2 yea s) and ele en
heal hy women
(21.0 ±1.3 yea s);
in e collegia e eam playe s.
Randomized, double-blind, c osso e ,
placebo-con olled; 6 g L-Ci ulline in
710 mL suc ose solu ion inges ed 1 o 2 h
be o e he es .
B uce’s eadmill p o ocol o
exhaus ion.
•VO2max. g oup 1
•Vasodila a ion g oup 1
•Anae obic h eshold
de ined by gas exchange
g oup 1
↔
↔
↔
Randomized, double-blind, c osso e ,
placebo-con olled; 710 mL o
Wa e melon Juice (1 g L-Ci ulline)
inges ed 1 o 2 h be o e he es .
B uce’s eadmill p o ocol o
exhaus ion.
•VO2max. g oup 2
•Vasodila a ion g oup 2
•Anae obic h eshold
de ined by gas exchange
g oup 2
↔
↔
↔
Hickne e al., 2006 [43]
Se en physically ac i e men
and women (18–40 yea s).
Randomized, double-blind, c osso e ,
and placebo-con olled; 3 x 3 g o
L-Ci ulline o 24 h s.
Inc emen al eadmill es o
exhaus ion.
•VO2peak g oup 1 ↔
Ten physically ac i e men
and women (18–40 yea s).
Randomized, double-blind, c osso e ,
and placebo-con olled; 3 g L-Ci ulline
3 h be o e he es .
Inc emen al eadmill es o
exhaus ion.
•VO2peak g oup 2 ↔
Suzuki e al., 2016 [49]
Twen y- wo heal hy and
well- ained men
(29 ±8.4 yea s).
Randomized, double-blind, c osso e ,
and placebo-con olled; 2.4 g/day o
L-Ci ulline (9 capsules) o 1 week
be o e he es (be o e bed ime) + 1 h
be o e he es .
Fou km indi idual ime ial
on cycle-e gome e .
•VO2kine ics ↔
↑
: S a is ically signi ican imp o emen o Ci ulline ea men ;
↔
no s a is ically signi ican imp o emen o Ci ulline ea men . VO
2
: Oxygen up ake. VO
2
max: Maximal Oxygen
up ake. HRR: Hea Ra e Rese e. MRT: Mean Response Time. GET Gas Exchange Th eshold.
Nu ien s 2022,14, 3479 18 o 28
O he six s udies ha looked a he impac o Ci on VO
2
esponse, wo [
45
,
56
]
analyzed pulmona y O
2
up ake (VO
2
) mean esponse ime (MRT) and ound s a is ically
signi ican e ec s ha showed an imp o emen in his a iable. Howe e , while hey
ound s a is ically signi ican e ec s ha showed an imp o emen in his a iable, Ashley
e al. [
56
] only ound hese esul s in one o he g oups, wi h no bene i s ha ing been
epo ed in he g oup o people o e 60 yea s o age. Addi ionally, wo s udies looked a
muscle oxygena ion [
57
] and issue [
45
] in submaximal and maximal exe cises and ound
signi ican imp o emen s ela ed o hese a iables. In con as , Cu u ello e al. [
48
] did
no ind signi ican esul s in VO
2
max. o anae obic h eshold, no did Bailey e al. [
57
] in
pulmona y VO
2
, o Hickne e al. [
43
] in VO
2
peak. Likewise, Ashley e al. [
56
] epo ed
no signi ican di e ences in ne oxygen expendi u e, no did he emaining s udy [
49
] in
VO2kine ics.
Figu e 7shows he esul s o he me a-analysis abou VO
2
kine ics a e Ci supplemen-
a ion, whe e no signi ican e ec was ound o his a iable (p= 0.94). A me a-analysis o
he selec ed s udies ound low inconsis ency le els (I = 0%; p= 0.91). One s udy showed a
la ge e ec on a single measu ed a iable, namely, he pulmona y gas exchange mean e-
sponse ime a e he high-in ensi y cycle-e gome e exe cise [
45
]. In addi ion, wo a iables
showed a small e ec [
56
], while he emaining 17 we e ela ed o a negligible e ec . The e-
o e, he me a-analysis showed a negligible e ec on VO
2
kine ics by Ci supplemen a ion
(pooled SMD = 0.01; 95% CI (−0.16 o 0.17).
Nu ien s 2022, 14, x FOR PEER REVIEW 19 o 30
Figu e 7 shows he esul s o he me a-analysis abou VO2 kine ics a e Ci supple-
men a ion, whe e no signi ican e ec was ound o his a iable (p = 0.94). A me a-anal-
ysis o he selec ed s udies ound low inconsis ency le els (I = 0%; p = 0.91). One s udy
showed a la ge e ec on a single measu ed a iable, namely, he pulmona y gas exchange
mean esponse ime a e he high-in ensi y cycle-e gome e exe cise [45]. In addi ion, wo
a iables showed a small e ec [56], while he emaining 17 we e ela ed o a negligible
e ec . The e o e, he me a-analysis showed a negligible e ec on VO2 kine ics by Ci sup-
plemen a ion (pooled SMD = 0.01; 95% CI (−0.16 o 0.17).
Figu e 7. Fo es plo compa a i e s udy o he e ec s o Ci supplemen a ion on VO2 kine ics
[43,45,48,49,56,57].
3.6. E ec o Ci ulline on Lac a e
Fou o he nine s udies included in he sys ema ic e iew and me a-analysis looked
a lac a e as he main me aboli e o glycolysis [64] a e he inges ion o Ci and placebo.
Table 5 shows hese s udies, including hei me hodology and hei main conclusions e-
ga ding lac a e measu emen .
In h ee s udies [44,57,59], an inc ease in blood lac a e concen a ion was shown in
he expe imen al g oup compa ed o he con ol g oup. Ma ínez e al. [59] analyzed lac-
a e a e a submaximal e o o 40 km, whe eas Bailey e al. [57] did so a e sub-maximal
and maximal e o , and Cunni e e al. [44] a e a sp in e o . The emaining s udy,
ca ied ou by Hickne e al. [43], did no show a signi ican inc ease in lac a e in ei he o
he wo expe imen al g oups in which di e en doses and imings o Ci supplemen a ion
we e used.
The me a-analysis did no show any signi ican posi i e e ec on lac a e compa ed
o placebo (p = 0.16). The s udies analyzed ound low inconsis ency be ween hem, as
shown by he me a-analysis (I = 0%; p = 0.94). Figu e 8 shows he esul s ob ained. I indi-
ca es he mode a e e ec o lac a e ound in he s udy by Ma ínez e al. [59] a e pa ici-
pan s comple ed a hal -ma a hon ace. Howe e , wo a iables om one s udy [57]
showed a small e ec , and he emaining h ee, ob ained om wo s udies, showed a
small e ec [43,44] and a negligible e ec . The e o e, he me a-analysis showed a small
e ec o Ci supplemen a ion on lac a e (SMD g ouped = 0.25; 95% CI (−0.10 o 0.59)).
Figu e 7.
Fo es plo compa a i e s udy o he e ec s o Ci supplemen a ion on VO
2
kine ics [
43
,
45
,
48,49,56,57].
3.6. E ec o Ci ulline on Lac a e
Fou o he nine s udies included in he sys ema ic e iew and me a-analysis looked
a lac a e as he main me aboli e o glycolysis [
64
] a e he inges ion o Ci and placebo.
Table 5shows hese s udies, including hei me hodology and hei main conclusions
ega ding lac a e measu emen .
Nu ien s 2022,14, 3479 19 o 28
Table 5. Summa y o s udies included in he sys ema ic e iew and me a-analysis in es iga ing he e ec o Ci on lac a e.
Au ho /s Popula ion In e en ion Tes Va iables Analyzed Main Conclusions
Bailey e al., 2016 [57]
Eigh heal hy, ac i e,
ec ea ional subjec s. Men
(22 ±2 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; ~3.4 g/day o
L-Ci ulline in 300 mL o Wa e melon
Juice Concen a e o 16 days.
Cons an cycle-e gome e wo ks a
70% o VO2max o exhaus ion.
•Lac a e ↑
Six min o cons an wo k (70% VO
2
max) + 30” all-ou on
cycle-e gome e .
•Lac a e ↑
Cunni e e al.,2016 [44]
Ten well- ained heal hy men
(23.5 ±3.7 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 12 g Ci ulline
Mala e (dissol ed in 400 mL wa e )
60 min be o e comple ing he es s.
Two exe cises o 10 epe i ions o
15 s all-ou (sp in ) on a
cycle-e gome e , ollowed by 5 min
o es and an inc emen al es o
exhaus ion a 100% o indi idual
peak powe .
•Lac a e ↑
Hickne e al., 2006 [43]
Se en physically ac i e men
and women (18–40 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 3 ×3 g o
L-Ci ulline o 24 h
Inc emen al eadmill es o
exhaus ion.
•Lac a e ↔
Ten physically ac i e men
and women (18–40 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 3 g L-Ci ulline 3 h
be o e he es .
Inc emen al eadmill es o
exhaus ion.
•Lac a e ↔
Ma ínez-Sánchez e al., 2017
[59]
Twen y- wo heal hy male
ama eu unne s
(35.3 ±11.4 yea s).
Randomized, double-blind, c osso e ,
placebo-con olled; 500 mL o Fashion
Wa e melon Juice en iched wi h
L-Ci ulline (3.45 g pe 500 mL) aken
2 h be o e he ace.
Hal -Ma a hon Race •Lac a e ↑
↑: S a is ically signi ican imp o emen o Ci ulline ea men ; ↔no s a is ically signi ican imp o emen o Ci ulline ea men .
Nu ien s 2022,14, 3479 20 o 28
In h ee s udies [
44
,
57
,
59
], an inc ease in blood lac a e concen a ion was shown in he
expe imen al g oup compa ed o he con ol g oup. Ma ínez e al. [
59
] analyzed lac a e
a e a submaximal e o o 40 km, whe eas Bailey e al. [
57
] did so a e sub-maximal and
maximal e o , and Cunni e e al. [
44
] a e a sp in e o . The emaining s udy, ca ied
ou by Hickne e al. [
43
], did no show a signi ican inc ease in lac a e in ei he o he
wo expe imen al g oups in which di e en doses and imings o Ci supplemen a ion
we e used.
The me a-analysis did no show any signi ican posi i e e ec on lac a e compa ed
o placebo (p= 0.16). The s udies analyzed ound low inconsis ency be ween hem, as
shown by he me a-analysis (I = 0%; p= 0.94). Figu e 8shows he esul s ob ained. I
indica es he mode a e e ec o lac a e ound in he s udy by Ma ínez e al. [
59
] a e
pa icipan s comple ed a hal -ma a hon ace. Howe e , wo a iables om one s udy [
57
]
showed a small e ec , and he emaining h ee, ob ained om wo s udies, showed a small
e ec [
43
,
44
] and a negligible e ec . The e o e, he me a-analysis showed a small e ec o
Ci supplemen a ion on lac a e (SMD g ouped = 0.25; 95% CI (−0.10 o 0.59)).
Nu ien s 2022, 14, x FOR PEER REVIEW 20 o 30
Figu e 8. Fo es plo compa a i e s udy o he e ec s o Ci supplemen a ion on lac a e
[43,48,57,59].
Figu e 8.
Fo es plo compa a i e s udy o he e ec s o Ci supplemen a ion on lac a e [
43
,
48
,
57
,
59
].
4. Discussion
The main objec i e o his sys ema ic e iew and me a-analysis was o analyze and
summa ize he cu en e idence on he e ec s o Ci supplemen a ion on ae obic spo s
pe o mance and ela ed a iables such as lac a e, VO
2
kine ics, and he RPE. O e all,
he esul s ob ained indica e ha Ci had no bene icial e ec s on ae obic pe o mance,
lac a e concen a ion, VO
2
kine ics, and/o he RPE in he es s pe o med. The doses
and iming o in ake di e ed be ween he s udies analyzed, sugges ing ha he e may be
di e en mechanisms a play depending on hese a iables. Rega ding iming, ch onic
supplemen a ion o 6–16 days seemed o ha e posi i e, albei s a is ically non-signi ican
e ec s, while acu e supplemen a ion (1 h–3 h be o e he es ) did no ha e a posi i e impac .
In his sense, bo h a ch onic dose o Ci be ween 3.4 and 6 g/day o 6–16 days and an
acu e dose (1 h–3 h be o e) o be ween 1 and 12 g did no signi ican ly imp o e ae obic
pe o mance. Simila ly, Ci did no imp o e he RPE sco e unde he same supplemen a ion
condi ions, and VO
2
kine ics we e no signi ican ly al e ed. Supplemen a ion wi h 3–12 g
o acu ely and ch onically inges ed Ci showed a clea end o inc eased lac a e, al hough
he o e all e ec was only small (p= 0.16).
Conside ing he al eady known ole o Ci in he syn hesis and p oduc ion o NO
and he possible e ec s associa ed wi h his compound, he po en ial impac o i s supple-
men a ion on physical pe o mance and associa ed a iables in di e en ypes o exe cise
and using di e en es s has been s udied in ecen yea s [
59
,
65
]. A posi i e impac on
a ious aining adap a ions ela ed o o he NO pa hway supplemen s has also been
sugges ed [
24
,
26
]. Howe e , s udies examining such e ec s a e sca ce. As a esul , he
e idence o Ci supplemen a ion is con o e sial and, he e o e, i is no conside ed o
be an e gogenic aid backed by s ong scien i ic e idence [
12
,
15
]. The e o e, al hough he
mos commonly used dose was be ween 3.4 and 6 g, he e was no consensus on doses and
p o ocols o use o imp o e spo s pe o mance. The esul s ob ained in his sys ema ic
Nu ien s 2022,14, 3479 21 o 28
e iew and me a-analysis a e simila ly con o e sial, as hey showed no uni o m impac
on hose a iables measu ed a e he inges ion o his amoun . Likewise, g ea e in akes
(9–12 g) ha ha e shown posi i e e ec s o o he ela ed supplemen s such as A g [
24
]
ha e no shown such bene i s in his s udy. Simila ly, doses o less han 3.4 g ha e no been
ound o ha e a posi i e impac on pe o mance o associa ed a iables. Howe e , i is
wo h highligh ing he possible di e ences associa ed wi h he in ake iming, as a sligh
addi ional bene i seems o be shown in hose p o ocols based on ch onic supplemen a-
ion compa ed o acu e supplemen a ion. This could be explained by he esul s al eady
ob ained in s udies wi h o he supplemen s o he NO pa hway, such as bee oo juice o
ni a e, whe e ch onic supplemen a ion seems o be necessa y o gene a e adap a ions a
a highe le el ela ed o NO [
25
]. Abou acu e supplemen a ion, i is well known in he
li e a u e ha ni a e and ni i e le els peaked a e 2.5–3 h [
39
] and A g a ound 30–90 min
a e inges ion [
66
]. Taking his in o accoun , and conside ing he ole played by Ci in
he syn hesis o A g oge he wi h NO om he ni a e–ni i e–NO pa hway, acu e supple-
men a ion should p opose p o ocols by hese p o en e ec s. The p esen s udy sugges s
di e en e ec s be ween one p o ocol and he o he ; he e o e, he e is a need o s udy bo h
ypes o supplemen a ion o unde s and which me hodology could be mo e app op ia e
o he op imiza ion o spo s pe o mance.
The long- e m pe o mance o endu ance depends on he abili y o e icien ly syn-
hesize ATP p ima ily h ough oxida i e pa hways, as well as o p oduce and oxidize
lac a e h ough oxida i e phospho yla ion [
1
,
67
,
68
]. I is p ima ily in luenced by di e en
a iables such as lac a e concen a ion, which is an indi ec measu e o cellula me abolism;
VO
2
kine ics, which de e mines physiological demands; and he RPE, as a subjec i e pe -
cep ion o he e o made. The use o di e en nu i ional and supplemen a ion me hods
can he e o e make a signi ican di e ence in spo ing pe o mance [
6
,
12
]. Mo eo e , ak-
ing in o accoun he physiological e ec s ela ed o NO a he en ila o y le el, as well
as ene gy e iciency and muscle con ac ion, using supplemen s such as Ci o enhance
i s endogenous syn hesis could be an in e es ing s a egy [
25
]. While he e a e s udies
ha ha e shown signi ican imp o emen s in hei pa icipan s [
43
,
45
,
57
,
60
], he e a e
se e al o he pape s whose esul s ha e no shown any posi i e e ec [
48
,
49
,
59
]. Con-
sequen ly, Ci supplemen a ion, in isola ion, is no cu en ly ecommended o ae obic
pe o mance enhancemen .
4.1. E ec on Ae obic Spo s Pe o mance
Long- e m spo ing pe o mance depends p ima ily on high ae obic capaci y, which
allows he ca dio ascula sys em o supply su icien oxygen o mee he needs o exe cise,
and high me abolic lexibili y ha acili a es he use o ene gy subs a es acco ding o he
demands o he momen [
1
,
69
]. Fo example, he use o a nu i ional s a egy based on
highe o lowe ca bohyd a e in ake can make a signi ican di e ence in winning o losing
a compe i ion such as an asphal ma a hon [
6
]. The use o an e ec i e e go-nu i ional
aid can also make an impo an di e ence [
6
]. In his ega d, Ci supplemen a ion has
been shown o ha e bene icial e ec s associa ed wi h ae obic capaci y. Bailey e al. [
45
]
ound a signi ican imp o emen in exe cise ole ance ime and wo k comple ed in wo
cycle-e gome e es s a di e en in ensi ies (mode a e and se e e) a e ch onic supple-
men a ion o 6 days, wi h he las dose aken 90 min be o e exe cise (6 g o Ci ). Simila ly,
S anelle e al. [60]
epo ed clea bene i s in he ime o comple e a 40 km indi idual ime
ial, as well as in a e age powe ou pu , ollowing a simila supplemen a ion p o ocol
bu wi h a 1-day longe du a ion. Howe e , he same s udy showed no posi i e e ec s in
a se ies o six sp in s pe o med immedia ely a e comple ing he ime ial. In ano he
simila s udy, in which he iming o he dose was longe (16 days) bu smalle (3.4 g/day
o Ci ), no imp o emen in ime o exhaus ion was ound in a cons an wo k es a 70%
o VO
2
max [
57
]. These esul s could be explained by he lowe dose inges ed in he la e
s udy, which was almos hal o he dose wi h posi i e e ec s.
Nu ien s 2022,14, 3479 22 o 28
Using an acu e supplemen a ion me hodology, Hickne e al. [
43
] ound a signi ican
imp o emen in ime o exhaus ion in wo g oups wi h di e en doses o Ci : 9 g/day
inges ed o e 24 h wi h a inal in ake 3 h be o e he es , and a single dose o 3 g/day
aken 3 h be o e he es . Ma ínez e al. [
59
] ound no posi i e esul s in he inal ime o
a hal -ma a hon a e he use o a simila dose o 3.45 g (in he o m o wa e melon) was
inges ed 2 h be o e exe cise, and a single dose o 3 g/day was aken 3 h be o e he es .
The same esul s we e ob ained in ano he s udy ha compa ed wo di e en doses (6 g
and 1 g) inges ed 1–2 h be o e an inc emen al p o ocol o exhaus ion among in e collegia e
playe s, wi hou showing any imp o emen associa ed wi h ime o exhaus ion [
48
]. While
low doses could be he eason o hese esul s, a s udy conduc ed by Cunni e e al. [
44
]
showed ha e en a dose o 12 g inges ed 60 min be o e an in e mi en es ollowed by an
exhaus ion es did no p o ide any bene i s in ime o exhaus ion, a e age powe , wo k
done, dis ance co e ed, o a e age speed. I seems ha acu e supplemen a ion wi h Ci
does no yield any imp o emen in ae obic spo s pe o mance, ega dless o he dose used
and he le el o he sample. In he same ein, i is pa icula ly in e es ing o conside he
pha macokine ics o Ci o s udy he bes iming me hodology o op imiza ion in he same
way as o o he supplemen s such as A g.
Acu e and ch onic supplemen a ion may ha e di e en physiological mechanisms o
ac ion and may also be ela ed o he doses o Ci supplemen ed. The esul s o his sys em-
a ic e iew and me a-analysis show no clea bene icial e ec s o ei he s a egy
(p= 0.08)
,
al hough ch onic supplemen a ion wi h an amoun close o and/o abo e
6 g/day
may
show a end owa ds imp o emen , in con as o he acu e app oach.
4.2. E ec on he RPE
The a e o pe cei ed exe ion is a powe ul indica o o he in e nal load ha a subjec
endu es when exe cising. I is also ela ed o mul iple o he a iables ha a y along wi h
he du a ion and in ensi y o exe ion, such as di e en physiological a iables ela ed o
oxygen consump ion and hea a e, me abolic a iables such as lac a e concen a ion o
glycogen deple ion, and o he speci ic phenomena such as muscle damage, empe a u e,
o dehyd a ion [
70
,
71
]. I s quan i ica ion by means o a subjec i e a ing scale such as
he Bo g scale [
72
] has been shown o be alid o de ec ing physiological and me abolic
changes du ing exe cise, as well as o moni o ing in e nal load du ing aining [
73
,
74
].
The sensa ion o exe ion o an exe cise can be modi ied by di e en ools ela ed o
nu i ion and supplemen a ion, such as ca bohyd a e in ake, luid and elec oly e in ake,
o supplemen a ion wi h subs ances such as ca eine o c ea ine [
75
]. In he same ein,
NO- ela ed supplemen s such as ni a e o bee oo juice ha e al eady been shown o ha e
di ec and bene icial e ec s on he RPE du ing and a e exe cise [61,76].
Rega ding Ci supplemen a ion, Hickne e al. [
43
] showed an inc ease in he sub-
maximal RPE o wo g oups o en and se en physically ac i e men and women du ing
an inc emen al es o exhaus ion a e inges ing 9 g/day and 3 g/day, espec i ely. The
same esul s we e epo ed by ano he s udy using an in e media e dose o 6 g/day in
nine well- ained male ia hle es and compe i o s compe ing in a 40 km indi idual ime
ial [
60
]. These esul s in he RPE co espond wi h an imp o emen in ae obic capaci y
in bo h s udies, demons a ing ha , along wi h an imp o emen in spo s pe o mance,
he e could be a posi i e e ec o Ci on he RPE. On he con a y, wo o he s udies did
no ind signi ican di e ences be ween hei esul s. Ma ínez e al. eco ded no change
in he RPE o 22 male ama eu unne s a e he acu e supplemen a ion o 3.4 g o Ci in
a hal -ma a hon ace. Wi h simila esul s, Suzuki e al. [
49
] ound no imp o emen in
he RPE o 22 well- ained men who ch onically inges ed 2.4 g o Ci o 7 days p io o
compe ing in a 4 km indi idual ime ial.
While a lowe sco e on he RPE scale may lead o an imp o emen in spo s pe o -
mance due o a lowe pe cep ion o a igue, an inc ease in he RPE scale may a he same
ime lead o an inc ease in exe cise in ensi y and hus o highe sel -demand in exe cise,
which could lead o be e pe o mance. Howe e , such an inc ease should be ela ed
Nu ien s 2022,14, 3479 23 o 28
o an objec i e imp o emen in pe o mance measu ed by o he a iables, as was he
case in he s udies analyzed in his sys ema ic e iew and me a-analysis [
43
,
60
]. These
esul s could be explained by a mul i ude o ac o s ela ed o he e ec s o Ci and NO
on pe o mance, including inc eased muscle oxygena ion, educed oxygen consump ion,
and imp o ed muscle con ac ion [
20
]. Despi e he posi i e esul s, i canno be concluded
ha Ci supplemen a ion imp o es he RPE du ing endu ance exe cise due o he limi ed
li e a u e a ailable and he esul s ob ained in his me a-analysis (p= 0.90).
4.3. E ec on VO2Kine ics
A subjec ’s oxygen consump ion is di ec ly ela ed o he equi emen s o di e en
exe cise in ensi ies and esponds di ec ly o he physiological and me abolic demands
o pe iphe al issues [
8
,
77
]. Thus, he a io o VO
2
o CO
2
inc eases esponds ia he
Respi a o y Exchange Ra io (RER) o he change in ene gy subs a es in a o o he
p io i y use o ca bohyd a es and co esponds (like lac a e p oduc ion) o inc eased exe cise
in ensi y [
78
]. Di e en changes in VO
2
kine ics ela ed o eac ion ime, en ila o y
capaci y, o dec eased oxygen deb could lead o imp o ed en ila o y e iciency, inc eased
issue oxygena ion a a gi en in ensi y, and he e o e inc eased a hle ic pe o mance [
77
].
Supplemen s ha induce inc eased NO p oduc ion such as bee oo juice may be associa ed
wi h posi i e changes in VO
2
kine ics h ough mul iple mechanisms, including inc eased
asodila ion and a co esponding inc ease in blood low o pe iphe al issues [25,79].
Along he same lines, Ci could exe i s e ec on VO
2
kine ics. In ac , a s udy ha
analyzed he impac o i s supplemen a ion, using a dose o 6 g/day o 6 days, ound
signi ican imp o emen s in MRT and he issue oxygena ion index, which we e ela ed o
di ec imp o emen s in VO
2
kine ics i sel [
45
]. Simila ly, ano he s udy by he same g oup
o scien is s epo ed a signi ican inc ease in muscle oxygena ion a bo h submaximal and
maximal exe cise a e 16 days o long- e m supplemen a ion wi h 3.4 g/day o Ci [
57
].
In he same s udy, howe e , he imp o emen in VO
2
was no signi ican . Simila esul s
we e ound by Ashley e al. [
56
] in MRT and oxygen de ici in a g oup o 15 ac i e adul s
who inges ed 6 g/day o Ci o 7 days be o e walking a he submaximal in ensi y on
a eadmill. In he same s udy, howe e , he esul s we e no posi i e o a g oup o
people wi h an a e age age o o e 70 yea s old. In con as , Hickne e al. [
43
] showed
no bene i in VO
2
peak in hei s udy o physically ac i e men and women. Cu u ello
e al. [
48
] also epo ed no bene i wi h espec o VO
2
max, asodila ion, and VO
2
a he
anae obic h eshold. Along simila lines, ano he s udy wi h simila doses o Ci showed
no imp o emen s in VO
2
max kine ics [
49
]. The di e ences in he esul s shown be ween
he di e en s udies could be explained by he dose and iming o Ci supplemen a ion.
As wi h o he NO pa hway supplemen s, ch onic in akes o Ci appea o be necessa y o
ob ain g ea e imp o emen s in he ca dio ascula sys em.
While acu e supplemen a ion showed no bene i s o VO
2
kine ic a iables, hose
s udies in which ch onic supplemen a ion was used (7–16 days) a a dose o app oxima ely
3.4–6 g/day appea ed o imp o e hese a iables. Howe e , in iew o he esul s ob ained
in his me a-analysis (p= 0.94), i canno be de e mined ha Ci supplemen a ion has a
posi i e impac on VO
2
. The e o e, u he s udies should be ca ied ou wi h ch onic doses
and wi h he combina ion o o he po en ially syne gis ic supplemen s ha could show
bene icial e ec s, such as hose p e iously documen ed in he li e a u e [62].
4.4. E ec on Lac a e
Lac a e is he main me aboli e o glycolysis du ing exe cise and i s p oduc ion inc eases
p opo ionally o he in ensi y o exe cise un il i exceeds he mi ochond ial clea ance ca-
paci y and accumula es in he bloods eam [
68
]. While in ensi y is he main ac o in i s
inc ease, he a ailabili y o muscle and li e glycogen, exogenous glucose, and he subjec ’s
ibe ype, among o he ac o s, de e mine i s p oduc ion [
68
]. Howe e , i s oxida ion is lim-
i ed by he unc ionali y o he mi ochond ion and he associa ed me abolic lexibili y [
69
].
Al hough i s inc ease in blood is conside ed a nega i e e ec , especially a submaximal
Nu ien s 2022,14, 3479 24 o 28
in ensi ies, he inc ease in i s p oduc ion is synonymous wi h inc eased glycoly ic ac i a ion
and, he e o e, mo e e icien uel use a highe in ensi ies equi ing highe physiological
demands [
6
,
69
]. Consequen ly, i s inc ease a e a submaximal o maximal es , especially
when accompanied by an objec i e imp o emen in pe o mance, can mean an inc ease in
exe cise in ensi y and, he e o e, in he abili y o exe cise a a highe in ensi y [
69
,
80
,
81
].
Va ious e go-nu i ional aids such as ca eine, be a-alanine, o bica bona e ha e been shown
o signi ican ly inc ease lac a e le els, accompanied by an imp o emen in spo s pe o -
mance [
82
], as well as o he NO- ela ed supplemen s such as bee oo juice [
83
,
84
]. Ch onic
supplemen a ion o 16 days a a dose o 3.4 g/day showed a signi ican inc ease in lac a e
a submaximal and maximal exe cise in ec ea ional spo speople [
57
]. The same esul s
we e obse ed wi h o he acu e supplemen a ion me hodologies. Ma ínez-Sánchez A.
e al. [
59
] ound signi ican inc eases in lac a e a e a maximal in e mi en es ollowed by
a con inuous es wi h a high in ake o Ci (
12 g 60 min
be o e he es ). Along he same
lines, one s udy showed highe lac a e le els a e a hal -ma a hon ace compa ed o a
placebo, al hough pa icipan s had inges ed a much lowe dose (3.45 g inges ed 2 h be o e).
Howe e , Hickne e al. [
43
] obse ed no di e ences om he con ol g oup and be ween
g oups when compa ing wo di e en doses o Ci (9 g s. 3 g) in an inc emen al eadmill
es o exhaus ion. Al hough he la e esul s we e no s a is ically signi ican , he aw da a
om he s udy showed a posi i e end, sugges ing he e ec p e iously obse ed in he
o he s udies.
Ci supplemen a ion could inc ease lac a e concen a ion h ough an inc ease in exe -
cise in ensi y ha could be explained by an inc eased abili y o exe cise a a highe in ensi y,
wi h highe bioene ge ic e iciency, and wi h inc eased ole ance o lac a e [
20
,
44
]. Al hough
he esul s o he s udies analyzed could be conside ed posi i e, he me a-analysis ca ied
ou has no shown signi ican esul s (p= 0.16). These esul s could be jus i ied by he small
numbe o s udies examined. Howe e , he e is no consensus om which o conclude ha
Ci has a posi i e e ec on lac a e inc ease du ing endu ance exe cise.
4.5. S eng hs and Limi a ions
This sys ema ic e iew has se e al s eng hs and weaknesses. The o al numbe o
selec ed a icles ep esen s an impo an limi a ion due o he sca ci y o s udies ha ha e
analyzed Ci supplemen a ion on ae obic pe o mance and i s a iables. Addi ionally, he e
has been a b oad a ie y in supplemen a ion p o ocols, bo h in dosage and iming, which
means ha he likelihood o ob aining accu a e conclusions is commensu a ely lowe . This
may be due o he con o e sial esul s o he scien i ic li e a u e which, o he momen ,
do no allow o he mos app op ia e doses and p o ocols o use o be es ablished. The
oppo uni ies o ad ancemen in his a ea a e hus hinde ed acco dingly. Addi ionally, he
a iables analyzed and he es s ca ied ou widely di e ed om one ano he . Ne e heless,
hey we e all ela ed o he demands o ae obic exe cise, and he e o e he conclusions ha e
been d awn igo ously.
On a di e en no e, he exhaus i e me hodology ca ied ou in his sys ema ic e iew
and me a-analysis, in e ms o s udy selec ion, he analysis o possible biases, he analysis
o a iables, and he in e p e a ion and quan i ica ion o esul s, is an impo an s eng h o
he s udy.
4.6. Fu u e Resea ch Lines
Acco ding o he esul s ob ained in his s udy, and conside ing i s s eng hs and
weaknesses, u u e lines o esea ch should ocus on he homogeneous s udy o Ci supple-
men a ion, ying o homogenize doses, iming, and he mos ep esen a i e a iables o
he e ec s o supplemen a ion, as is he case wi h o he supplemen s o he NO pa hway.
In iew o he p omising esul s, u he s udies should be p oposed ha use a ch onic
supplemen a ion me hodology and a highe doses han he usual ones (6 g/min). This
would make i possible o measu e no only he acu e e ec s on pe o mance bu also
he possible physiological and me abolic adap a ions al eady analyzed o o he sup-
Nu ien s 2022,14, 3479 25 o 28
plemen s [
26
]. Al hough his sys ema ic e iew and me a-analysis has no been able o
es ablish ela ionships in e ms o he sample o s udies, i may be o in e es o analyze he
di e ence in e ec s be ween pa icipan s o di e en le els, due o he di e ences al eady
epo ed in simila supplemen s be ween eli e and ama eu a hle es [
24
]. Addi ionally, he
pha macokine ics o CWB and he abso p ion mechanisms o exogenous supplemen a ion
a e wo a eas o be s udied in dep h ha could p o ide new and mo e speci ic knowledge
o ex end he unde s anding o he e ec s o Ci supplemen a ion.
Finally, and conside ing he ole o Ci in he NO me abolic pa hways and he close
ela ionship wi h he A g on he one hand, and ni a e and ni i e on he o he , i could be o
g ea in e es o analyze he join e ec s o di e en supplemen s o his pa hway on spo s
pe o mance, as well as on di e en associa ed physiological and me abolic a iables.
5. Conclusions
This sys ema ic e iew and me a-analysis has shown no signi ican posi i e e ec s o
Ci supplemen a ion on he di e en a iables analyzed in ela ion o ae obic spo s pe -
o mance. Ch onic supplemen a ion (6–16 days) wi h an amoun be ween
3.4 and 6 g/day
showed a posi i e endency o imp o e ae obic pe o mance, bu wi hou s a is ical sig-
ni icance. Acu e supplemen a ion (1–3 h) wi h 1–12 g o Ci did no signi ican ly imp o e
ae obic pe o mance o any ela ed a iables. No imp o emen in VO
2
kine ics o he
RPE was obse ed a e Ci supplemen a ion, i espec i e o doses and iming. In con as ,
Ci showed a posi i e endency o inc ease lac a e bu wi h no signi ican esul s. Ci
supplemen a ion does no appea o be e ec i e in inc easing ae obic a hle ic pe o mance
and is he e o e no ecommended o his pu pose. Due o he pauci y o s udies on his
subjec , as well as he he e ogenei y o he p o ocols used, u he s udies a e needed o
unde s and he impac o such supplemen a ion on a hle ic pe o mance.
Au ho Con ibu ions:
A.V. and J.M.-A.: Concep ion and design, analysis and in e p e a ion o
he da a, d a ing o he pape , and c i ical e iew and app o al o he inal e sion submi ed o
publica ion. A.C.-B. and J.F.-L.: Analysis and in e p e a ion o he da a, d a ing o he pape , and
c i ical e iew and app o al o he inal e sion submi ed o publica ion. P.S.C. and D.F.-L.: D a ing
o he pape and c i ical e iew and app o al o he inal e sion submi ed o publica ion. All
au ho s ha e ead and ag eed o he published e sion o he manusc ip .
Funding: This esea ch has no ecei ed any ex e nal unding.
Ins i u ional Re iew Boa d S a emen : No applicable.
In o med Consen S a emen : No applicable.
Da a A ailabili y S a emen : No applicable.
Con lic s o In e es : The au ho s a i m no con lic o in e es .
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