RED BULL INDUCES BIOCHEMICAL CHANGES IN WISTAR RAT LIVER

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1 Annals of RSCB Vol. XVIII, Issue 2/213 RED BULL INDUCES BIOCHEMICAL CHANGES IN WISTAR RAT LIVER Mihaela Crişan, Camelia Munteanu, Corina Roşioru, Camelia Lang FACULTY OF BIOLOGY AND GEOLOGY, BABEŞ-BOLYAI UNIVERSITY, CLUJ-NAPOCA, ROMANIA Summary Despite the increasing popularity of energy drinks consumption, there are concerns regarding the safety and effectiveness of this type of drink. The aim of the present study was to evaluate the changes in blood and liver biochemical parameters in male Wistar rats, after 15 days of ad libitum Red Bull energy drink administration. Increased serum transaminases and LDH activities, and a high glycaemia were observed. An alarmingly high liver glycogen deposition was also recorded. These results showed that Red Bull consumption may lead to hepatic injuries and to unexpected metabolic modifications. Further studies are required to elucidate whether energy drinks are a safe choice for athletic and cognitive performance enhancement. Key words: Red Bull, energy drink, liver camelia.lang@hasdeu.ubbcluj.ro Introduction The raising interest in improvement of sport performance led to an increasing demand for ergogenic supplements. Energy drinks, promoted as cognitive and athletic performance enhancers, are some of the most popular supplements worldwide. Red Bull is the best sold energy drink in the world, with 4.5 billion cans sold per year, and the number is ever increasing ( According to a recent EFSA (European Food Safety Authority) report, 12% of european adults (18-65 years) and 12% of teenagers (1-18 years) are high chronic energy drinks consumers, with 7 liters of energy drinks consumed in one month (aproximately a can of 25 ml/day). Despite the popularity of these drinks, their effects on consumers health are quite controversial and not sufficient research on energy drinks safety has been conducted yet. There are several studies recording a modest improvement of energy drinks consumption on physical endurance (Alford et al., 21, Baumm and Weiss, 21, Seidl 118 et al., 2), but also studies that showed no significant enhancement of endurance related to the consecutive energy drinks consumption (Carvajal-Sancho and Moncada-Jiménez, 25; Eckerson et al., 213, etc). Some of the compounds found in energy drinks are used in therapy for treating certain disorders, like taurine and niacin for dyslipidemias. Other compounds, like glucuronolactone, are not well studied. An alarming number of side effects and even deaths were reported, as a consequence of energy drinks consumption. Arrythmia, cardiac arrest and hepatitis are some of the quoted side effects (Ballard et al., 21; Clauson et al., 28; Vivekanandarajah et al., 211). The present study sought to observe changes of some biochemical parameters in liver and serum in Wistar rats, following Red Bull ingestion ad libitum for 15 days. This study was designed as the pilot phase of a more complex project concerning the effects of Red Bull consumption on biochemical parameters in liver, brain, skeletal and cardiac muscle.

2 Annals of RSCB Vol. XVIII, Issue 2/213 Material and methods All chemicals used in this study were of analytical grade, purchased from Sigma-Aldrich Chemie GmbH, Germany, Nordic Invest S.R.L., Romania and S.C. BioZyme S.R.L, Romania. Red Bull energy drink was purchased from the local market. 16 male Wistar albino rats, weighing 186.6±3.15 g were used. Animals were housed in the zoobase of Molecular Biology & Biotechnology Department, School of Biology and Geology, Babeş-Bolyai University, Cluj- Napoca. They were kept in hygienic conditions and received a standard diet. Animals were organized in two groups: the control group (n=6), received tap water and the Red Bull group (n=1), received Red Bull energy drink ad libitum. After 15 days, the animals were killed and blood and liver samples were harvested for biochemical assays: glycaemia, glucose and glycogen hepatic content, total serum and liver proteins, ALT (alanine aminotransferase), AST (aspartate aminotransferase), LDH (lactate dehydrogenase) enzymatic activities in serum. Blood and liver glucose was determined using the Somogy-Nelson method (Somogy, 1945; Nelson, 1944). Glycogen content was measured using Lo s method (Lo et al., 197). Total protein concentration was assayed with Bradford reagent (Bradford, 1976). ALT and AST activities were assessed according to the photocolorimetric method described by Reitman and Frankel (1957), and for LDH activity, the method of Bergmeyer and Bernt (1974) was used. Results were analyzed using two tailed t test and considered statistically significant at p.5. Bull has 27 g carbohydrates, and other compounds that may affect carbohydrates metabolism, such as niacin. According to Zhou et al. (21) and Li et al. (29), niacin supplementation in high carbohydrate diets may be one of the causes of diabetes epidemic. Moreover, niacin can induce insulin resistance (Chang et al., 26; Greenbaum et al., 1996). A can of Red Bull assures 1% of niacin RDA (Recommended Daily Allowance). Even if this quantity is not usually seen as a risk, it should be considered that high carbohydrate content of Red Bull may amplify niacin s side effects. A normal diet assures niacin RDA and there is no need for external supplementation in healthy subjects. mg/dl Glycaemia Fig. 1. Glycaemia in rats treated with Red Bull energy drink ad libitum (n=1) compared with C group (n=6). The results are expressed as mean value ± standard error; significant difference by t test ( p.5). Significant liver glucose rise seen in this experiment (Fig. 2) is also a consequence of the high carbohydrate content in Red Bull energy drink. It was demonstrated that in conditions such as type 2 diabetes, the depletion of skeletal muscle glycogen is associated with a rise in liver carbohydrates (Eldar-Finkelman and Yarkoni, 22). Results and discussion As expected, Red Bull administration led to a significant increase in glycaemia (Fig. 1). A 25 ml can of Red 119

3 Annals of RSCB Vol. XVIII, Issue 2/ Liver glucose Fig. 2. Liver glucose in rats treated with Red Bull ( p.1). The glycogen content in liver was tripled by Red Bull ad libitum administration (Fig. 3). Abnormal glycogen deposition in liver due to Red Bull administration has been noted for the first time in the present study. Hepatic glycogen deposition is seen in 8 % of chronic diabetic patients. It is believed that this condition is closely related to liver enzymes abnormalities (Levinthal and Tavill, 1999). In a study conducted by Clore et al. (1992) in humans, after 3 days of fasting, hepatic glycogen content of type 2 diabetic patients was 2-fold higher than that of controls Liver glycogen Fig. 3. Liver glycogen in rats treated with Red Bull ( p.5). In the present study, serum and liver protein concentrations were not affected by Red Bull administration (Figs. 4 and 5). mg/dl Serum protein Fig. 4. Serum protein in rats treated with Red Bull Liver protein Fig. 5. Liver protein in rats treated with Red Bull As seen below, ingestion of Red Bull ad libitum in rats led to a significant rise of serum AST, ALT and LDH activities (Figs 6, 7, and 8). µ g pyruvate/ ml serum/ hour AST activity in serum Fig. 6. Serum AST activity in rats treated with with Red Bull energy drink ( p.5). Ebuehi et al. (211) reported an increase in serum protein after administration of 5 ml Red Bull daily, for 36 days, in rabbits. 12

4 Annals of RSCB Vol. XVIII, Issue 2/213 µg pyruvate/ g tissue/ hour ALT activity in serum Fig. 7. Serum ALT activity in rats treated with with Red Bull energy drink ( p.5). µmoles/ pyr/ml serum/min LDH activity in serum Red Bull Fig. 8. Serum LDH activity in rats treated with with Red Bull energy drink ( p.5). Our results are supported by other studies, which reported hepatic injuries related with energy drink consumption. Khayat et al. (212) studied the effects of three types of energy drinks including Red Bull, on Wistar rat liver. Animals received 1.5 ml energy drink daily, for 4 weeks. Results showed mild hepatotoxicity, with increased transaminase activities in serum, and hepatic tissue alterations. Akande and Bankojo (211) conducted a study in which Power Horse energy drink administration in rats led to high AST, ALT and ALP (alkaline phosphatase) activities in serum. Conclusions Ad libitum Red Bull administration in Wistar rats is proven to increase blood glucose and to affect liver carbohydrate metabolism. Excess glycogen deposition in liver due to Red Bull administration has been noted for the first time in the present study. Constant Red Bull consumption may lead to hepatic injuries. 121 References Akande I.S., Banjoko O.A., Assessment of Biochemical Effect of Power Horse Energy Drink on Hepatic, Renal and Histological Functions in Sprague Dawley Rats, Annu. Rev. & Res. Biol., 1(3): 4556, 211 Alford C., Cox H., Wescott R., The effects of red bull energy drink on human performance and mood, Amino Acids, 21(2): , 21 Baum M., Weiss M., The influence of a taurine containing drink on cardiac parameters before and after exercise measured by echocardiography, Amino Acids, 2: 75-82, 21 Bergmeyer, H. U., Brent, E., UV-assay with pyruvate and NADH, in: Bergmeyer, H. U., ed., Methods of enzymatic analysis, Academic Press, New York, , 1974 Bradford, M.M., A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of proteindze binding, Analit. Biochem., 72: , 1976 Carvajal-Sancho A., Moncada-Jiménez J., The acute effect of an energy drink on physical and cognitive performance of male athletes, Kinesiologia Slovenica, 11(2): 5 16, 25 Chang A., Smith J., Galecki A., Bloem C., Halter J., Impaired beta-cell function in human aging: response to nicotinic acidinduced insulin resistance, J. Clin. Endocrinol. Metab.: 91: , 26 Clauson K.A., Shields K.M., McQueen C.E., Persad N., Safety issues associated with commercially available energy drinks, J. Am. Pharm. Assoc., 48(3): e55-63, 28 Clore J., Post E., Bailey, D., Nestler J., Blackard W., Evidence for increased liver glycogen in patients with noninsulindependent diabetes mellitus after a 3-day fast., J. Clin. Endocrinol. Metab., 74: , 1992 Ebuehi O.A., Ajayl O.E., Onyeulor A.L., Awelimobor D., Effects of oral administration of energy drinks on blood chemistry, tissue histology and brain acetylcholine in rabbits, Nig. Q. J. Hosp. Med., 21(1): 29-34, 21 Eldar-Finkelman H., Yarkoni M., Insulin Resistance and Glycogen Synthesis: Roles in Liver, Muscle and Adipose Tissue, in:

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