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Journal of Dietary Supplements, 11(1):64–79, 2014


C 2014 by Informa Healthcare USA, Inc.
Available online at www.informahealthcare.com/jds
DOI: 10.3109/19390211.2014.887602

ARTICLE

A Systematic Review on the Herbal Extract


Tribulus terrestris and the Roots of its Putative
Aphrodisiac and Performance Enhancing Effect

Ahmed Qureshi, Declan P Naughton, & Andrea Petroczi


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School of Life Sciences, Kingston University, Kingston upon Thames, Surrey,


London KT1 2EE, UK

ABSTRACT. Tribulus terrestris (TT) is a dicotyledonous herbal plant of the Zygophyl-


laceae family. In ancient medicine, extracts of the aerial parts and fruits have been used
for its diuretic, tonic, and aphrodisiac properties. Today, TT is widely used by athletes
and bodybuilders based on the belief, fueled by claims in marketing information, that
it can enhance testosterone concentrations. To assess TT’s effect on testosterone levels
For personal use only.

in human and animals, an electronic literature search out using seven databases and the
patent database up to August 2013 was carried out. Randomized control trials, which in-
cluded healthy human subjects ingesting TT as sole or combined supplement, along with
animal studies with TT as a sole treatment across a number of species were included.
Eleven studies met the inclusion criteria, including one patent application. The results
showed that trials varied in duration, dosage and supplementation with TT as sole or
combined treatment, rendering meta-analysis impossible. A limited number of animal
studies displayed a significant increase in serum testosterone levels after TT adminis-
tration, but this effect was only noted in humans when TT was part of a combined sup-
plement administration. Literature available for the effectiveness of TT on enhancing
testosterone concentrations is limited. Evidence to date suggests that TT is ineffective
for increasing testosterone levels in humans, thus marketing claims are unsubstanti-
ated. The nitric oxide release effect of TT may offer a plausible explanation for the ob-
served physiological responses to TT supplementation, independent of the testosterone
level.

KEYWORDS. androgen, health claim, herbal supplement, nitric oxide, saponin,


sport, testosterone

BACKGROUND
Individuals seek testosterone enhancement via synthetic and nonsynthetic routes in
order to enhance athletic performance and/or physical appearance (Sjöqvist, Garle,
& Rane, 2008). Pharmaceutical advances over time have allowed the production of

Address correspondence to: Andrea Petroczi, School of Life Sciences, Kingston University, Kingston upon
Thames, Surrey, London KT1 2EE, UK; (E-mail: K A.Petroczi@kingston.ac.uk).
(Received 8 October 2013; accepted 21 January 2014)

64
Systematic Review on Tribulus terrestris 65

potent androgen elevating drugs with fewer side effects, but the moral and health
issues still remain. The thriving dietary supplement market offers a wide variety of
natural sources for sports people with proven and putative effects on athletic per-
formance (Maughan, 2005; Maughan, Greenhaff, & Hespel, 2011). Herbal Tribulus
terrestris (TT) supplements are widely available in high street dietary supplement
retailers and on the Internet with prices typically under £0.1 per capsule. Owing
to its putative effect on increasing serum testosterone levels, TT, is widely used by
bodybuilders and gaining popularity among male athletes for muscular hypertro-
phy, skeletal muscle strength development, boosting libido, promoting testicular de-
velopment as well as a treatment for hypertension, relieving kidney disorders and
colic while remaining “natural” (Antonio, Uelmen, Rodriguez, & Earnest, 2000;
Arcasoy, Erenmemisoglu, Tekol, Kurucu, & Kartal, 1998; Bashir, Tahir, Samee
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& Munir, 2009; Gauthaman, Adaikan, & Prasad, 2002). Supplement producers
are capitalizing on this trend, including patents for performance-enhancing food
supplements containing TT extract (Golini, 2011; Rodriquez, 2009). Evidence
emerging from the scientific literature is less convincing. Although animal studies
have shown TT mediated increases in levels of testosterone, dihydrotestosterone,
and dehydroepiandrosterone, clinical experiments failed to confirm this effect in
humans (Antonio et al., 2000; Brown et al., 2000; Bucci, 2000; Gauthaman &
Ganesan, 2008; Rogerson, Riches, Jennings, Mier, & Marshall-Graidisnik, 2007;
Saudan, Baume, Emery, Strahm, & Saugy, 2008). Caution is called for against using
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TT until further convincing evidence become available (Borrione, Di Luigi, Maf-


fulli, & Pigozzi, 2008; Brown, Vukovics, & King, 2006; Kreider et al., 2010; Tamler &
Mechanick, 2007).
In the European Union, the use of nutrition and health claims in commercial
communications that directly inform customers or create a perception of specific
health benefits is regulated by the Regulation (EC) No 1924/2006 on nutrition
and health claims made on foods, with scientific substantiation of specific health
claims being the cornerstone of the regulation (Regulation (EC) No 1924/2006).
The Regulation specifically states that “a claim should be scientifically substan-
tiated by taking into account the totality of the available scientific data, and by
weighing the evidence” (Regulation (EC) No 1924/2006, point 17, p. 5). At the
time of this review (2013), the EU Register of Nutrition and Health Claims
(http://ec.europa.eu/nuhclaims/), which contains authorized and nonauthorized
health claims, the conditions of use and applicable restrictions (if any), or the
reasons for nonauthorization, shows no record for Tribulus terrestris. Similar
regulations outside Europe found around the word, calling for a harmonized
approach to claim substantiation (Richardson et al., 2003).
Despite the questionable scientific evidence and regulations regarding health
claims around the world, the common themes emerging from information available
from unregulated sites on the Internet on TT are centered on the putative effect
comparable to steroids while being natural and legal. Specifically, claims include
increasing luteinizing hormone responsible for testosterone production in males:
“Can help increase growth compound levels”; helping natural testosterone produc-
tion: “Natural herb helps increase the natural production of testosterone”, “Tribulus
is used to improve fertility and boost male test production” and some even include a
66 Qureshi et al.

figure by which increase in testosterone can be expected: “Research has shown that
when taking Tribulus terrestris it can increase your testosterone levels by up to 50%.
Taking Tribulus may help you improve your stamina and endurance as well as help
speed up muscle growth.”
Current understanding on the Internet is mixed. This comprehensive open
source of information available to consumers suggests TT has aphrodisiac prop-
erties, a common understanding shared by the majority. However, a split in belief
occurs when questioning its role in testosterone boosting. Forum users over the age
of 30 have claimed to have seen a pronounced effect on muscle gain and increased
libido due to an increase in testosterone when ingesting a product containing TT.
These claims cannot be verified as no measurements were taken, but efficacy may
be seen in this age group due to a decrease in testosterone in the aging process,
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suggesting TT may be effective when testosterone concentrations are significantly


lower than the normal range.
Tribulus terrestris is a dicotyledonous herb that belongs to the Zygophyllaceae
family and is widely grown in Africa, Australia, Asia, and Europe. Chinese tra-
ditional medicine utilizes TT to improve visual acuity and as an anticonvulsant,
while Indian herbal medicine takes advantage of its diuretic, tonic, and its aphro-
disiac properties (Chopra, Chopra, Handa, & Kapur, 1958, Gauthaman, Adaikan,
& Prasad, 2003; Gauthaman & Ganesan, 2008). Its popularity grew in Eastern Eu-
rope in the 1970s once its properties were discovered by the Bulgarian Olympic
For personal use only.

weight lifting team and has continued to flourish as it is regarded as a safe, legal,
and herbal alternative to anabolic steroids. It is most commonly ingested in extract
form, which is largely composed of the aerial portions of the plant or the fruits
that mainly contain steroidal glycosides (saponins) of the furasatanol type found
in TT (Kostova & Dinchev, 2005; Koumanov, Bozadjieva, Andreeva, Platonva, &
Ankov, 1982; Liu et al., 2010; Tomova et al., 1981; Su et al., 2009). Specifically, a po-
tent saponin Protodioscin has been isolated in TT (Ganzera, Bedir, & Khan, 2001).
Protodioscin is assumed to be the efficacious compound which can allegedly cause
an increase in testosterone and its androgenic derivatives, dehydroepiandrosterone
and dihydrotestosterone (Dikova & Ognyanova, 1983).
The proposed mechanism by which TT can theoretically raise serum testos-
terone levels is by stimulating androgen receptors within the brain, which causes
the posterior pituitary gland to secrete more luteinising hormone, thus stimulat-
ing the testes to synthesize more testosterone (Koumanov et al., 1982). In turn,
this androgen receptor stimulating effect in the brain leads to a misinterpretation
of the body’s sex hormone levels, assuming the levels are lower than they really
are.
TT is commonly used by athletes and bodybuilders, based on the belief it
has testosterone enhancing capabilities. While there is a plethora of ‘‘reviews”
on TT posted on the internet, they are mostly available via or linked to body-
building websites and online retailers. Despite the health claims and popularity,
no scientific and unbiased review is available on TT. This review aims to ad-
dress this gap by critically assessing and summarizing the evidence from trials on
the effect of TT on humans and animals for or against its role as a testosterone
booster.
Systematic Review on Tribulus terrestris 67

METHODS
This systematic review was conducted according to the PRISMA protocol
(Moher, Liberati, Tetzlaff, & Altman, 2009). Furthermore, in line with the guid-
ance for evaluating health claims, the review was focused on the strongest evidence
available from intervention studies (Ellwood, Trumbo, & Kavanaugh, 2010; Verha-
gen, Vos, Francl, Heinonen, & van Loveren, 2010).

Data Sources
In order to form a comprehensive view of TT efficacy, a total of eight electronic
databases, namely Pubmed, Medline, Science Direct, Web of Knowledge, Google
Scholar, Proquest Science Journals, SAGE Journals Online, and Patents were
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searched up to August 2013 using the search terms for which data were sought:
‘‘Tribulus terrestris” AND supplementation AND testosterone AND human OR
animal. Reference lists of the included articles were also searched for further rele-
vant articles. Included articles were limited to in vivo trials with humans or animals,
published in English.

Study Selection
An article was included if it met the following criteria: (1) randomized controlled
clinical trial (RCT); (2) involved healthy human volunteers with testosterone lev-
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els in the normal range prior to treatment or intact animals; (3) in which subjects
were administered with any type of Tribulus terrestris, regardless of country of ori-
gin, administration route, and preparation of TT. Trials that varied in duration,
dosage, and used TT as a combined treatment were also included with either or
both male and female subjects. Trials that stated changes in testosterone levels but
presented no clinical data within the papers were excluded. The potential risk of
bias was assessed at study level using the Cochrane Collaboration’s tool.
For the identification stage, two reviewers (AQ and AP) set and refined the
search criteria for selection. Based on the exclusion and inclusion criteria, one re-
viewer (AQ) assessed each potential output for inclusion and extracted the infor-
mation from the final set of included articles. Further selection based on the quality
was not available owing to the limited number of intervention studies met the cri-
teria, thus all eligible studies were included with their limitations discussed. Results
based on the extracted information were reviewed by two reviewers (AP and DPN).
The literature search identified 667 articles for possible inclusion, which was re-
duced to 34 after screening the abstract and title leaving 34 full articles to be re-
trieved and scrutinized. Twenty-three of these articles were further excluded due
to reasons summarized in Figure 1. The 11 studies remaining were split between an-
imal and human studies, 4 and 7, respectively. Studies with insufficient details for the
intervention and/or the key outcome measure were excluded, which may present
bias across the studies.

RESULTS
The selected studies are summarized in Tables 1 and 2 according to the following
key information: description of the study (e.g., whether human or animal study,
68 Qureshi et al.
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FIGURE 1. Flow diagram of the PRISMA literature search process.

age, number of participants, and study duration), characteristics of treatment and


control (e.g., type of Tribulus, dose, route of administration and frequency of treat-
ment), and outcome of treatment and consequences, if any. Increase in serum
testosterone concentration was considered as a key outcome.
The most comprehensive investigation was an animal study was on a number
of different subjects at varied doses, totaling 84 animals (Gauthaman & Ganesan,
2008). A two-armed group design with primates, rabbits and rats were conducted
over 8 weeks in which the sole intervention regime was TT. The remaining three
animal studies were conducted on Wistar rats, where studies varied from each other
by dosage, duration of treatment, and number of animals (Ghosian Moghaddam,
Khalili, Maleki, & Ahmad Abadi, 2013; Martino-Andrade et al., 2010; Singh, Nair,
& Gupta, 2012). All animal studies employed a placebo treatment.
The absence of clear evidence for the efficacy of TT in elevating testosterone in
humans warrants a critical analysis of the trials. Two studies were notable as the
sample sizes were proportionally greater (n = 22 and n = 21) and the duration of
the studies were considerably longer at 5 and 4 weeks, respectively (Neychev &
Mitev, 2005; Rogerson et al., 2007). One study adopted a two-armed parallel group
double-blind design with a placebo regimen, where blind questionnaire was carried
out at the end of the treatment to determine whether the blinding had been effec-
tive (Rogerson et al., 2007). The second study employed a three-armed parallel
group that was a single blind study also with a placebo regimen (Neychev & Mitev,
2005). An additional trial was a nonrandomized trial without a placebo very small
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TABLE 1. TT Supplementation Studies Using Animal Models

Sample Size Age Form of


(years) Duration of Placebo Testosterone Key
Reference Study Herbal Intervention Regimen Measured Out-come Other Outcomes

Gauthaman and Primate n=5 n=5 Serum (+) Dihydrotestosterone and


Ganesan 2008 n = 10, no age Each primate was with Blood sample Testosterone Dehydroepiandrosterone
Singapore stated concentrations of 7.5,15 taken 15 mins had short lived increases
180 mins and 30 mg/kg Intravenously before
treatment
acted as
control
White Rabbit n = n = 6 (TT1 = 2.5 mg/kg) n=6 Serum (−) Serum Dihydrotestosterone
24, no age stated n = 6 (TT2 = 5 mg/kg) Testosterone increase was statistically
8 weeks n = 6 (TT3 = 10 mg/kg) significant
Once daily for all
concentrations
Normal n = 10 (TT1 = 2.5 mg/kg) n = 10 Serum (−) Serum Dihydrotestosterone
Sprague–Dawley n = 10 (TT2 = 5 mg/kg) Testosterone increase not statistically
rat n = 40, no age n = 10 (TT3 = 10 mg/kg) significant
stated Once daily for all
8 weeks concentrations
Singh et al. 2012 Wistar rat n = 18 n = 6 (LAET = 50 mg/kg) n = 6 (Gum Serum (+) Increase in sexual behaviour
India 13 days n = 6 (LAET = 100 mg/kg) Acacia = Testosterone noted
Once daily, orally 2 mL/kg)
Martino-Andrale Wistar rat n = 55 13 n = 14 (TT1 = 11 mg/kg) n = 13 (distilled Serum (−) nil
et al. 2010 weeks old n = 14 (TT2 = 42 mg/kg) water) Testosterone
Brazil 28 days n = 14 (TT3 = 110 mg/kg)
Once daily via oral gavage
Moghaddan et al. Wistar rat n = 48, no n = 12 (TT1 = 6.25% plant n = 12 Serum (+) Groups treated with TT had
2013 Iran age stated food) Testosterone higher concentrations of LH,
21 days n = 12 (Mph = 70 mg/mL/day) but no significant change in
n = 12 (TT2 + Mph = 6.25% FSH levels.
+ 70 mg/mL/day)

LAET: Lyophilized aqueous extract of the dried fruits of Tribulus terrestris.


TT: Tribulus terrestris.

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70
TABLE 2. TT Supplementation Studies With Human Participants

Sample Size Age Form of


(years) Duration of Testosterone
Reference Study Herbal Intervention Placebo Regimen measured Key Out-come Other Outcomes

Rogerson et al., n = 22 n = 11 n = 11 Measured in T/E (–) Strength and fat


2007 19.8 ± 2.9 Standardized 450 mg TT ratio free mass
Australia 5 weeks (60% saponins) increased in both
One capsule daily, orally groups
Neychev & Mitev, n = 21 200mg Bulgarian TT n=7 Measured in Total (–) Androstenedione
2005 20–36 (60% saponins) Testosterone and luteinizing
Bulgaria 4 weeks n = 7 (TT1 = 20 mg/kg) hormone not
n = 7 (TT2 = 10 mg/kg) significantly
One capsule three times a altered
day, orally
Saudan et al., 2008 n=2 n=2 n=0 Measured in T/E (–) Concentrations of
Switzerland Female∗ 26 and 40 250 mg Tribestan R two ratio DHEA and LH did
2 days capsules three times a not show
day, orally significant
variation
Brown et al., 2000 n = 20 n = 10 consumed n = 10 Measured free and (–) Serum Estradiol
USA 19–29 ‘ANDRO-6’: 750 mg TT 250 mg rice flour total testosterone and serum
8 weeks (2 weeks 300 mg Androstenedione three times a day, estrone elevated
on, 1 week off) 150 mg DHEA 625 mg orally on weeks 2,5,8
Chrysin 300 mg and weeks 5 and
Indole-3-carbinol 540 mg 8, respectively.
Saw Palmetto One Increase in
capsule three times a day, muscle strength
orally in both groups
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Brown et al., 2001 n = 55 in total n = 28 consumed ‘ n = 27 Measured free and (–) Basal serum free
USA 30 years n = 20 AND-HB’: Rice flour total testosterone testosterone
40 years n = 20 1350 mg TT concentration in
50 years n = 15 300 mg Androstenediol 30years 0 higher
30–58 450 mg Saw Palmetto than 50 years.
4 weeks 450 mg Indole-3-carbinol Basal serum
300 mg Chrysin androstenedione
1500 mg yLA and DHT
One capsule three times concentrations
a day, orally higher in
30 years,
compared to
40 years
Kohut et al., 2003 n = 16 n=8 n=8 Measured serum (–) Increased levels of
USA 50–59 750 mg TT Rice flour free testosterone Free
4 weeks 150 mg DHEA testosterone,
300 mg Androstenedione androstendione,
625 mg Chrysin estradiol and
300 mg Indole-3-carbinol DHT from week 1
540 mg Saw Palmetto to week 4
One capsule three times
a day, orally
Chaudhary et al., n = 148 n = 38 (E-MA-H) Low n = 36 Measured serum (–) Statistically
2011 no ages stated Dose n = 37 (E-MA-H) 2 capsules daily testosterone significant
India 8 weeks High Dose n = 37 increase in
(E-MA-HP) Two capsules IIEF-A, IIEF-B
daily and IPE in all
groups ingesting
supplement

T/E: Testosterone/Epitestosterone ratio.


yLA: gamma-linolenic acid.
DHEA: Dehydroepiandrosterone.
TT: Tribulus terrestris.
E-MA-H: Seven drug formulation comprising of herbal extracts.
E-MA-HP: Nine drug formulation comprising of herbal extract.

71
72 Qureshi et al.

(n = 2) and lasted only 2 days (Saudan et al., 2008). All three trials used standard-
ized versions of TT in capsule form, ingested orally (Neychev & Mitev, 2005; Roger-
son et al., 2007; Saudan et al., 2008). Variations in the dose administered across the
RCTs add to the complexity of harmonizing results from the reports.
The remaining four studies employed a combination of herbal drugs, including
TT, in their intervention regime, and again a variety of doses were used across the
studies (Brown et al., 2000, 2001; Chaudhary, Patel, Kulkari, & Sonawane, 2012;
Kohut et al., 2003). Three of these employed a two-armed design, with one arm
being the placebo, where the subjects were randomly assigned in a double-blind
manner and ingested the drugs orally in capsule form (Brown et al., 2000, 2001;
Chaudhary et al., 2012). One employed a four-armed design (Kohut et al., 2003).
The duration of these studies ranged between 4 and 8 weeks.
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Affect of Tribulus Terrestris as a Sole Treatment on Testosterone Levels in Young


Men
Two RCTs tested the effects of TT versus placebo on males aged between 19 and
36 (Neychev & Mitev, 2005; Rogerson et al., 2007). One study researched the effect
of TT supplementation on elite male rugby players over a five-week period using
a single dose of TT over the duration (Rogerson et al., 2007). This study showed
no significant change in urinary T/E ratio thus TT showed negative effect. Mean
urinary T/E data were used from the chart as no statistical figures were given, from
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week 0 to week 6 the placebo T/E ratio increased by 7.1%, the group ingesting
TT showed a decrease in T/E ratio of 5.8%. The second study was conducted to
ascertain whether TT influences androgen synthesis in men between the ages of
20–36 (Neychev & Mitev, 2005). Experimental subjects were given two different
doses of TT extract, 20 mg/kg (TT1) and 10 mg/kg (TT2). This study produced
results where the concentration of total testosterone was not significantly altered by
the ingestion of TT extract in both groups. The mean value for the group TT1 was
15.75 ± 1.75 nmol/L, TT2 16.32 ±1.57 nmol/L and the mean for the control group
was 17.74 ± 1.09 nmol/L (p > .05).

Affect of Tribulus Terrestris as a Sole Treatment on Testosterone Levels in


Women
Only one RCT has been conducted with female subjects. This study was based on
a very small-sample size of two female volunteers and measured the effect of TT
(Tribestan) on testosterone production (Saudan et al., 2008). This short-term study
showed no significant change in T/E ratio when ingesting 500 mg of Tribestan. Sub-
ject 1 had an increase in T/E ratio of 10%, subject 2 had in increase in T/E ratio of
5.5% when compared to baseline readings over 2 days.

Affect of Tribulus Terrestris Utilized in a Combined Supplement and Its Effect


on Human Testosterone Levels
Four RCTs measured the effects of a herbal supplement combined with androgen
precursors versus placebo on the serum testosterone levels in young men carry-
ing out resistance training, endocrine-lipid responses in older men when ingest-
ing the supplement chronically, the effect of the supplement on immune func-
tion in middle-aged men and treating male sexual dysfunction with a combined
Systematic Review on Tribulus terrestris 73

supplement (Brown et al., 2000, 2001; Chaudhary et al, 2012; Kohut et al., 2003). No
significant changes in total testosterone levels were observed in either of the studies,
however free testosterone levels were increased in two RCTs (Brown et al., 2000,
2001). The first RCT reported no effects of the supplement on free and total serum
testosterone levels, while the second RCT had free testosterone concentration in-
crease of 31.88% in 30-year olds, 19.04% in 40-year olds, and 19.23% in 50-year
olds when ingesting the herbal supplement and placebo control free testosterone
concentrations had only slight increases at 1.38%, 0%, and a 2.08% decrease, re-
spectively, when compared to baseline readings taken at week 0 for both arms of the
study (Brown et al., 2000, 2001). The third RCT only measured free testosterone
levels in subjects and reported that there was an increase of 36.33% in the concen-
tration of free testosterone in the supplement group and a decrease of 5.1% in the
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placebo group after 4 weeks (Kohut et al., 2003). The fourth trial conducted in a
patent study showed that all serum testosterone concentrations dropping below the
baseline readings after 60 days of supplementation producing no significant results
(Chaudhary et al., 2012).

Affect of Tribulus Terrestris on Animal Testosterone Levels


Four large studies conducted on a number of species of animal produced varying
results when administered with TT as a sole intervention regimen (Gauthaman
& Ganesan, 2008; Ghosian Moghaddam et al., 2013; Martino-Andrade et al.,
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2010; Singh et al., 2012). The comparative study of the hormonal effects of TT on
primates, rabbit, and rat only produced significant increase in testosterone levels
in all dosages when tested on primates when being compared to baseline for the
acute 180-min duration, but no significant increase was visible in white rabbit or
Sprague–Dawley rat during the 8-week study (Gauthaman & Ganesan, 2008). The
second study analyzed the aphrodisiac activity of Albino Wistar rat when being
treated with aqueous extract of TT and the third and fourth study focused on
how endocrine sensitive organs are affected by administration of TT in Wistar rats
(Martino-Andrade et al., 2010; Singh et al., 2012). Of the these two studies, one
found a significant increase in serum testosterone levels when compared to the
control group, with 50 mg/kg of LAET (lyophilized aqueous extract of Tribulus ter-
restris) producing a mean increase of 30% and 100 mg/kg producing a 56.6% mean
increase when compared to the control after 13 days (Singh et al., 2012). On the
contrary, the other study showed no significant change in testosterone concentra-
tions in rats that were treated with 11, 42, and 110 mg/kg/day when compared to the
control group, with mean increases being 15.78%, 42.10%, and 2.7%, respectively
(Martino-Andrade et al., 2010). The last animal study administered TT as a com-
bined treatment with morphine to evaluate the role of TT on treating hormonal
imbalances caused by morphine addiction in Wistar rats (Ghosian Moghaddam
et al., 2013). A significant, two-fold increase in the serum testosterone level was
noted in the group that was treated with TT compared to the morphine-only group.

DISCUSSION
Critical evaluation of the findings in the 11 included studies suggests that TT is in-
effective at increasing testosterone levels in both humans and animals. This finding
74 Qureshi et al.

however may be limited due to a number of factors that could bias the results at the
study level. One RCT did not employ a placebo control and did not have a stan-
dardized time when TT was ingested by the two subjects. Although the majority of
the studies used placebos, only one RCT reported successful blinding of subjects
after carrying out a questionnaire. The remaining RCTs were not fully successful in
minimizing bias. Sample sizes ranged from 2 to 56, which are considered compara-
tively small, with no studies carrying out power calculations. Given the widespread
sale and advertisement of TT in bodybuilder websites and gyms, it is surprising that
such population is largely absent. It is notable that no pharmaceutical type dosage
studies have been conducted and that a variety of doses were adopted across the
varied trials. A further complication arises with the use of a natural product or ex-
tracts in that doses of putative active components may vary considerably depending
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on the extraction, preparation, and storage history of the product.


Only three studies found an increase in testosterone after administration of TT;
all three studies were animal studies, with one trial on primates and the remain-
ing two on Wistar rats (Gauthaman & Ganesan, 2008; Ghosian Moghaddam et al.,
2013; Singh et al., 2012). The study involving sexually sluggish male rats may not be
entirely compelling as these rats may had preexisting low levels of testosterone and
its precursors, so any increase witnessed may be testosterone levels returning to a
normal range. The second study employing Wistar rat may also not be entirely valid
as morphine causes androgen levels to decrease when being administered chroni-
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cally. The increase in serum testosterone level shown may also be assumed to levels
returning to a normal range. These results could suggest TT to be used as a treat-
ment for hormone replacement therapy, but further studies are required to confirm
this effect. The results from the trial with primates showing an increase in serum
testosterone levels are limited to showing a short-lived initial rise in the testos-
terone level which returned to baseline after just 90 minutes. The study did not
state whether baseline readings, which acted as control, were within the normal
range of primate testosterone levels.
Currently, the dose that supposedly causes an increase in testosterone is un-
known. Human sole TT treatment varied from 450 mg to 2400 mg daily, and human
combined treatment varied from 2250 mg to 4050 mg daily of TT extract within
combined substances. A greater extent of variation was noted in animal trials, from
2.5 mg/kg up to 110 mg/kg daily. Further studies would be needed to demonstrate an
alleged optimal dose. Variation was also seen over the majority of experiments in
duration of supplementation, age of subjects in human trials, age of animal subjects
and training history, and status. All these factors can play a role in the efficacy of TT
and future repeats of experiments may benefit from standardizing such variables so
that a true comparison can be undertaken. A meta-analysis is favored when carry-
ing out a review such as this, but could not be performed as no statistical correlation
could be made between the varying data.
To further improve trials conducted in the future, the form of testos-
terone measured also needs to be standardized. Serum testosterone, testos-
terone/epitestosterone from urine, and total and free testosterone forms were a
number of the forms measured in the studies analyzed. Two human RCTs showed
increased levels of free testosterone with unchanged total testosterone levels,
but were classed as flawed studies due to TT being combined with DHEA and
Systematic Review on Tribulus terrestris 75

androstenedione, which are known to increase the free testosterone fraction by


reducing sex hormone-binding globulin (Beckham & Earnest, 2003). Changes in
total testosterone did not occur in any of the studies, but would have been a key
indicator that TT had been effective.
Given the low number of trials of TT and the lack of clinical control over factors
such as diet, one key confounding factor remains in the potential role of diet and
lifestyle in altering steroid levels. Recent studies reveal that dietary components
such as catechins found in green and white teas or quercetin abundant in red wine
are effective inhibitors of key enzymes involved in glucuronidation of testosterone
for urinary clearance (Jenkinson, Petroczi, & Naughton, 2013). Future human tri-
als should carefully control diet as well as other drugs which are known to affect
glucuronidation rates, at least in vivo.
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Furthermore, no studies were found that considered or tested alternative expla-


nations for the observed physiological responses in the absence of any increase in
the testosterone level. TT may have a performance-enhancing effect via having pos-
itive effect on aerobic capacity (Milašius, Pečiukonienė, Dadelienė, & Skernevičius,
2010), however, this effect might be due to TT having strong inhibitory activ-
ity on COX-2, thus effecting the release of nitric oxide (Hong et al., 2002). Al-
though at this stage the link is only speculative, nitric oxide supplying formulations
are another popular substance among those seeking performance enhancing ef-
fect (Petroczi & Naughton, 2010). Nitric oxide release can also offer an explana-
For personal use only.

tion for the ‘‘aphrodisiac effect” of TT alternative to testosterone increase. It has


been shown that protodioscin triggers nitric oxide release in the corpora cavernosa
and corpus spongiosum, thus having a pro-erectile effect (Adaikan, Gauthaman,
Prasad, & Ng, 2001) independent of testosterone. Finally, nitric oxide release may
also explain the traditional use of TT in treating cardiac problems (Bowen, 1990;
Ojha et al., 2008).
A variety of dosages and frequency of doses were given within and across tri-
als to assess the therapeutic efficacy of TT. The understanding of which TT is
used by bodybuilders after a cycle of steroids is, that it reduces the incidence of
adverse effects related with steroid use, however no studies assessed adverse ef-
fects of TT. As all the studies were relatively short term, long-term supplement-
ing effects may be useful to evaluate efficacy and toxicity in human and animal
subjects.
The mechanism of action of TT is still to be proved, as only two human stud-
ies reviewed measured and demonstrated that no significant increase in the neces-
sary key hormone, luteinizing hormone, had occurred. One animal study showed
significant increases in LH, but no change in testosterone concentration. TT al-
legedly increases luteinizing hormone in the process of increasing testosterone
concentrations.
As the popularity of TT has grown over the past few decades, a large number of
supplement manufacturers have begun to meet demand by producing and market-
ing TT as a testosterone booster. When questioned over its mechanism of action,
many are elusive when stating the pathway. Patent applications for herbal supple-
ments containing TT claims to decrease body fat, increase strength and lean muscle
mass, some linking the relationship between testosterone deficiency and sexual dis-
orders. This has also caused TT to be marketed as an aid to reduce male sexual
76 Qureshi et al.

dysfunction, but evidence or exemplification that TT works is still to be found by


these inventors applying for patents.
Continued use of the herb has been seen across the world in many gyms, where
the anecdotally reported performance enhancing effects could be through a dif-
ferent mechanism or a placebo effect or through an entirely different substance
ingested alongside TT. The latter may knowingly come from a combination of sup-
plements used by the individual or through contamination. Contamination of di-
etary supplements owing to poor quality control or deliberate spiking is a grow-
ing concern in food safety as well as anti-doping (Maughan, 2005; Petroczi, Taylor,
& Naughton, 2011). Research shows that a congruent pattern exist between the
putative health claim or effect of the supplement and its respective contaminants
(Petroczi et al., 2011). Thus, outside controlled clinical trials with TT as a sole sup-
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plement, performance enhancing experiences of TT users cannot be credited to TT


alone.

Limitations
The main limitation of this review arises from the small number of studies found
eligible for inclusion and the quality of the findings presented in these papers. Doses
and durations varied widely across the studies, with very few trials specifically aimed
to test TT as a testosterone booster. One particularly limiting factor was the use
of TT as part of a supplementation combination, making linking any effect to TT
For personal use only.

alone impossible. The review also revealed a hiatus in research into the mechanism
of and effectiveness of TT supplement as only two human studies demonstrated
that no significant increase in the necessary key hormone, luteinizing hormone, had
occurred. Finally, the review is limited to research outputs published in English.

CONCLUSION
Despite the popularity of TT supplements, the literature available on the effective-
ness of TT supplement is surprisingly scarce. On the basis of the results summarized
in this review, TT appears to be ineffective at increasing testosterone levels. While
people in need for boosting testosterone levels for health reasons would benefit
from a natural dietary supplement, further research is required to elucidate the
efficacy, proposed mechanism of action, optimal dose and frequency, and any po-
tential side-effects to establish any health claims relating to TT supplementation
in humans. Investigating a potential performance enhancing effects via nitric oxide
release could make an important contribution to anti-doping research.

Declaration of interest: The authors report no conflicts of interest. The authors


alone are responsible for the content and writing of this paper.

ABOUT THE AUTHORS


Ahmed Qureshi, Declan P Naughton, and Andrea Petroczi are affiliated with
School of Life Sciences, Kingston University, Kingston upon Thames, Surrey,
London KT1 2EE, UK.
Systematic Review on Tribulus terrestris 77

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