Caffeinated Chewing Gum Improves Sympathetic Nerve Activity and Simulates Wrestling Performance: a Double-blind Crossover Trial
Phase: NA
Conditions: Caffeine, Control
Sponsor: National Taiwan Sport University

Caffeinated chewing gum improves sympathetic nerve activity and simulates 
wrestling performance: a double-blind crossover trial 
 
Document Date: Dec 25, 2024 
IRB approved date: March 3, 2024 
 
Project summary 
The purpose of this study was to investigated whether caffeine intake is effective in 
improving the performance of wrestlers in a simulated competition. The study was 
designed to involve 16 healthy adult male athletes who had been trained in professional 
wrestling and were divided into caffeine trial (CAF trial) and placebo trial (PL trial) in 
a randomized crossover double-blind study design. After three days of dietary control, 
participants arrived at the laboratory in the afternoon of the day of the trial to collect 
saliva samples in a quiet state and were fitted with a heart rate monitor. Participants 
chewed either caffeine-containing gum at 3 mg/kg body weight (CAF trial) or placebo-
containing gum without caffeine (PL trial) for 10 minutes and then spit it out. 
Participants then underwent a 15-minute dynamic warm-up. At the end of the warm-up, 
participants performed 2 rounds of 3-minute wrestling-specific simulated matches, and 
the number of falls in the simulated matches was recorded. After the simulation match, 
saliva samples were collected from the participants again. The saliva samples were 
analyzed for caffeine alpha-amylase concentration. Differences between the two trials 
were analyzed using a paired sample t-test and significance was set at α<0.05. 
 
 
Introduction 
At the Olympic Games and the World Athletics Championships, wrestling is one of the 
most popular event. In addition to specialized skills, the key to winning a wrestling 
match is muscle strength and anaerobic power (1, 2). In a previous study, 92 male 
wrestlers were divided into elite and non-elite groups according to their performance, 
and it was found that absolute muscular strength and anaerobic power were the key 
factors that differentiated elite wrestlers from non-elite wrestlers(2). From these studies, 
it appears that improving strength as well as anaerobic power of a wrestler is an 
important factor in winning a competition. 
 
Many studies have suggested that caffeine (1,3,7-trimethylxanthine) supplementation 
prior to exercise is effective in improving maximal muscle strength and anaerobic 
capacity, with approximately small to moderate improvements (3). It has been 
established that the mechanisms in question consist of antagonised adenosine receptors. 

The resulting effect of this is a reduction in the level of fatigue experienced during 
periods of physical exertion. Furthermore, it has been demonstrated that the release of 
calcium ions from the sarcoplasmic reticulum is increased, while at the same time the 
activity of sodium-potassium ATPase (Na+/K±ATPase) is maintained. In addition to 
the abovementioned effects, it has been observed that the aforementioned mechanisms 
stimulate glycolysis (4, 5). A dose of 3-6 milligrams of caffeine per kilogram of body 
weight is recommended before exercise. This can be achieved through energy drinks or 
capsules(3). 
 
Another physiological mechanism that may affect maximal muscle strength and 
anaerobic power may be related to the activation of sympathetic nerves. Recent studies 
have demonstrated that enhanced sympathetic activation during exercise can be an 
effective strategy for reducing fatigue and increasing energy output at maximum 
intensity (6, 7). In order to comprehend the status of sympathetic nerve activation, it is 
possible to quantify the electrocardiographic signal from the heart, the α-amylase levels 
in saliva, or the plasma catecholamine concentration (8). Caffeine supplementation has 
been demonstrated in previous studies to enhance sympathetic nerve activity(9). In 
addition to the ingestion of capsules, the chewing of caffeinated gum has been observed 
to enhance sympathetic nerve activity and significantly reduce fatigue during maximal 
sprinting(7). 
 
It has been demonstrated that chewing caffeinated gum can enhance the rate of caffeine 
absorption, leading to a more expeditious attainment of the maximum blood 
concentration of caffeine(10). The absorption of caffeine from chewing caffeine gum is 
a process that occurs through the mucous membrane of the oral cavity. The rate of 
absorption is such that the highest concentrations of caffeine in the blood can be reached 
within approximately fifteen minutes after chewing (11). With regard to the effects of 
caffeine on exercise performance, studies based on systematic reviews and meta-
analyses have found that the ingestion of caffeine chewing gum at a dose greater than 
3 mg/kg of body weight prior to exercise can effectively improve maximal muscular 
strength and anaerobic power (12). In the case of weight classes being required for 
events such as wrestling, the caffeinated chewing gum has been demonstrated to reduce 
water intake and may decrease the risk of being overweight. This may serve as an 
additional effective method of supplementing caffeine intake. The purpose of this study 
was to investigate the effect of caffeinated chewing gum on sympathetic nerve activity 
on wrestling simulation match. 
 
 

Methods 
Study design 
In this study, a double-blind crossover experimental design was used, in which 
participants were randomised to either the caffeinated chewing gum trial (CAF) or the 
placebo chewing gum trial (PL). After a seven-day washout period, the participants 
were switched to another trial of experiments (Figure 1). All tests were conducted after 
a regular 6-day training period with a day off to ensure that participants were in a similar 
physiological state prior to the experiment. Prior to the experiment, participants were 
asked to perform at least 2 simulated matches to familiarise with the simulated matches 
before the main experiment. The primary outcome measure was the number of falls in 
the simulated match, while the secondary outcome was α-amylase concentration in 
saliva, as an indicators of sympathetic nerve activity. The study commenced on 11 June 
2024 and concluded on 23 December 2024. 
 
Participants 
Seventeen professionally wrestling male athletes were recruited for this study. After one 
participants withdrew due to personal reasons, the final results of 16 participants (age: 
21.8 ± 1.0 years; height: 168.4 ± 4.5 cm; mass: 68.2 ± 8.7 kg) were analyzed. The 
inclusion criteria for this study were: (i) more than 6years of professional wrestling 
training; (ii) more than 6 months of continuous training; (iii) recovering from sports 
injuries such as strains and contusions for at least 3 months. The exclusion criteria were: 
(i) non-professionally wrestling athlete; (ii) none regular training in the past 6 months; 
(iii) less than 3 months of recovery from a sports injury, or present epilepsy, 
hypertension, hyperlipidemia, heart disease, arthritis, osteoporosis, or brain injury; (iv) 
a history of caffeine allergy. The consent form is completed after the participant has 
given consent. This study was conducted following the Declaration of Helsinki. 
 
Sample size calculation 
In this study, the number of participants in request was analysed using the G*Power 
software (version 3.1.9.4, Universität Düsseldorf, Germany) (13). In a previous study, 
12 wrestling athletes were given the Pittsburgh Wrestling Performance Test (PWPT) 
after a caffeine intake and found that supplementation with a high dose of caffeine 
significantly reduced the completion time (56.8 ± 2.0 vs. 52.9 ± 1.8 s; p < .05) of the 
PWPT. Based on this data, we calculated that the effect size of this study reaches 2. The 
calculation was based on an alpha level of 0.05and a correlation coefficient of 0.80, it 
was calculated that 6 participants would be sufficient to meet the requirement of 
participants.  
 

Protocol 
Prior to the first official test, the participant must record the diet and training for the 3 
days prior to the test, and consume the same diet and training before the next test. On 
the day of the test, the same breakfast and lunch will be provided to the participant to 
ensure the same energy intake before the test. 
 
All the experiments were conducted at 3pm. Upon arrival at the laboratory, the 
participants were required to undergo saliva collection. Three milliliters of saliva were 
spat into a saliva tube for later analysis. After the participants putted on heart rate 
monitor, participants were instructed to chew either caffeine-containing gum (3 mg/kg 
body weight) or a caffeine-free placebo gum for a period of 10 minutes. Thereafter, the 
gum was to be expectorated. Following this, the participants underwent a dynamic 
warm-up for a period of 15 minutes. It has been established that the highest blood 
concentration of caffeine is achieved 15 minutes after the chewing of the gum and 
subsequent expulsion(11). After warming up, participants are permitted to engage in 
any form of self-stimulation they deem appropriate, including yelling, punching the 
dummy, and other similar actions. The present action has been meticulously designed 
to enhance sympathetic nerve activity. This finding serves to corroborate the hypothesis 
that caffeine supplementation exerts a stimulant effect on sympathetic nerve activity. 
 
After warming up, the grip strength of the participant's hands was measured with a grip 
strength meter (Smedlay’s Hand Grip Dynamometer TTM, Tokyo, Japan). 
Subsequently, participants engaged in a 3-minute, 2-round simulated wrestling match. 
This simulation has been shown to have good reliability and validity(14). In each 3-
minute round of the simulation, one dummy fall was performed in the first 30 seconds, 
with an interval of 10 seconds each time. The participants then try to complete the 
maximum number of falls in 20 seconds. After the fall, the participants were given a 
10-second break for a total of 1 minute. The same pattern was followed in the 2nd and 
3rd minutes. During the 3rd minute, the duration of continuous wrestling was extended 
to 30 seconds. After a 30-second break, the participant repeated the same simulation 
and record the total number of falls. After the simulation competition, saliva was 
collected again and caffeine concentration and α-amylase concentration were measured. 
 
Caffeine and placebo gum 
The caffeinated chewing gum employed in the present study (Military Energy Gum, 
Arctic Mint flavour; Stay Alert, Chicago, USA) has been utilised in previous studies(7, 
10). It is estimated that a single piece of gum contains approximately 5 grams and 100 
milligrams of caffeine. The placebo gum was a commercially available blue mint gum 

(Lotte Gum Mint., Ltd., Tokyo,Japan). In order to administer a dose of 3 mg/kg of 
caffeine per unit of body weight, it was necessary to crush, grind, blend, reshape and 
flavor all chewing gums with 0.3 g of peppermint-flavored powder. The process was 
designed to ensure that the new chewing gums were comparable to the original in terms 
of appearance, color, flavor, weight and size. Previous studies have demonstrated that 
this method renders it impossible for the participant to distinguish the type of gum (7, 
15). 
 
 
Saliva sample collection and analysis 
Saliva samples were stored in a refrigerator at -80 degrees immediately after collection. 
For analysis, samples were thawed and centrifuged at 4000 rpm for 5 min. 500 μL of 
saliva samples were transferred to glass tubes for mass spectrometry analysis and 
Enzyme-linked immunosorbent assay (ELISA) analysis respectivelyThe analysis of 
saliva α-amylase concentration was conducted utilising an enzyme-linked 
immunosorbent assay (ELISA) with commercially available reagents (Neogen 
Corporation, Kentucky, USA; Salimetrics LLC, State College, PA, USA).  
 
Statistical analysis 
All data are presented as means ± standard deviations. The Shapiro–Wilk test was 
utilised to evaluate the normality of the data. The grip strength and number of fall on 
each rounds between the two trials were analyzed using paired t-tests. The saliva 
caffeine and α-amylase concentration and RPE were analyzed using two-way ANOVA 
with repeated measures. If the interaction effect (trial × time) were significant, the 
Bonferroni method was used to perform post hoc comparisons. Effect sizes were 
calculated using Cohen's d to quantify the magnitude of observed ef-fects and defined 
as trivial (<0.20), small (0.20–0.40), moderate (0.40–0.80), and large (>0.8), 
respectively. The power value of each data was conducted using G*Power 3.1.9.6 
software (13) 
 
 
 

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