This work aimed to examine the effects of drinking an "energy drink" upon (i) short-term maximal performance, (ii) reaction times and (iii) psychological factors (i.e., mood state, ratings of perceived exertion (RPE) and affective load) and on physiological parameters such as blood pressure, blood glucose, hematological parameters and other biochemical parameters.

A total of 22 healthy and regularly active physical-education male students from various sports disciplines volunteered to participate in this study. Potential participants were initially screened through telephone interviews based on the inclusion criteria: 18-40 years of age, BMI less than 25 kg/m2, and low (<1.5g/month ) and not regular caffeine users. Exclusion criteria included diagnosis of any chronic metabolic disease such as type 2 diabetes or cardiovascular disease, an auto-immune disease such as rheumatoid arthritis, lupus or type 1 diabetes, liver disease and the intake of any medications/ or dietary supplements known to influence blood glucose concentrations and/or blood pressures.

The study was conducted according to the declaration of Helsinki and the protocol was fully approved (identification code: 8/16) by the review board "Local committee of the Laboratory of Biochemistry, CHU Habib Bourguiba, Sfax, Tunisia." After a throughout explanation of the protocol and responses to all questions, participants signed a written informed consent.

Subjects were instructed to avoid nicotine, alcohol, dietary supplements, medications and all other stimulants and to maintain their normal dietary, sleep and physical activity patterns before test sessions. Caffeine and other caffeinated products (e.g., chocolate, caffeinated gums, caffeine containing beverages) were avoided for 48 hours and food for at least 4 hours before testing.

A double blind, placebo-controlled, counterbalanced, crossover design was adopted for this study. The randomized order of testing was determined using free online software (www.randomization.com). Neither staff nor participants were informed about the names of the two drinks, and blinding was strictly maintained by emphasizing to both staff and participants that both drinks adhered to healthy principles, and that each was advocated by certain sports medicine experts. Two familiarization sessions were completed before definitive test sessions in order to eliminate any learning effects on physical performance and reaction time measurements. During the second familiarization session, body mass and height were recorded. Each participant visited the laboratory for two formal test sessions, drinking a caffeine-containing ED (RB) and a caffeine and taurine free beverage drink (PL). All sessions were arranged in the early evening hours to avoid any time of day effects. The two definitive test sessions were separated by an interval of seven days to allow sufficient recovery between tests and to ensure caffeine washout. To avoid identification, two opaque and unmarked cans of RB or PL were ingested by each participant (i.e., 500 mL) in the presence of a researcher. The two drinks were similar in volume, texture, and appearance. One can of RB drink (i.e., 250ml) contained 80 mg of caffeine, 1 g of taurine, 27 g of carbohydrates, 0.6 g of protein, 5 mg vitamin B6 and 487 kJ of energy. The PL drink was prepared by an agri-food engineer; it did not contain any caffeine or taurine, but comprised carbonated water, carbohydrates, citric acid lemon juice reconstituted from concentrate (1 %), supplemented by flavorings, of sodium citrate, acesulfame K, sucralose, potassium sorbate and RB (a flavoring that contains propylene glycol E1520 (0.23 mL).

Beverages were prepared, shaken and chilled in a refrigerator at 14h00 by an investigator who took no part in the test sessions or data analysis, but prepared the alphanumeric code identifying the tested drink. At 17h00, the cooled beverages were served in sealed plastic opaque water bottles and consumed using an opaque straw. Participants were instructed to drink the fluid quickly (within 1±0.5 min) 60 minutes before their test session and not to discuss or compare tastes or to make any assumptions about what they had ingested. The interval of 60 minutes was chosen as optimal for a complete caffeine absorption and thus a peaking of a caffeine concentration .

Subjects were supervised by staff to ensure that they drank the entire quantity of fluid, and no exchange of bottles was allowed. The last standardized meal (i.e., lunch) before the beginning of the test session was taken at 13h00. Temperature and relative humidity of the laboratory were similar over the test sessions, with a temperature of around 22°C and a relative humidity between 45 and 55 %. During each test session (from 18h00), resting blood glucose and blood pressures were measured and the participants completed the POMS questionnaire.

A 5-min treadmill warm-up was performed. After that, the reaction time, and handgrip force were determined, and the 30-s Wingate test was performed. RPE scores, blood glucose and blood pressures were then measured, and the affective load was calculated. To investigate the effects of RB on the acute physiological and psychological responses to exercise, blood measures and psychological test measures were collected immediately pre- and post-exercise.

Inclusion Criteria:

• 18-40 years of age,
• BMI less than 25 kg/m2,
• low (<1.5g/month [20]) caffeine users
• not regular caffeine users.

Exclusion Criteria:

• chronic metabolic disease such as type 2 diabetes and cardiovascular disease
• auto-immune disease such as rheumatoid arthritis, lupus or type 1 diabetes,
• liver disease
• intake of any medications/ or dietary supplements known to influence blood glucose and/or blood pressures.

To avoid identification, two opaque and unmarked cans [24-26] of Red Bull or Placebo were ingested by each participant (i.e., 500 mL) in the presence of a researcher.