The use of nutritionally supplemented drinks with thermogenic ingredients for weight loss and related improvements while following a diet and exercise program has increased significantly. Specifically, the thermogenic “energy” drink (thermogenic drink [TD]) market was estimated to be a $5.7 billion industry in 2006 with over 500 TDs being marketed worldwide (7). As a result, the need to investigate the efficacy of TDs has increased in an attempt to explain associated efficacy and mechanisms of action (10,15,29).
Common ingredients in TDs include caffeine, epigallocatechin gallate (EGCG), and taurine. Caffeine increases lipolysis through stimulation of adenosine and β-adrenergic receptors resulting in increases in circulating epinephrine (13,26) and serum free fatty acids (26). Epigallocatechin gallate enhances fat oxidation (6), alters food digestibility (20,27), and can downregulate stearoyl-CoA desaturase gene expression (20); an effect linked to a reduction of adiposity and lipid synthesis. Consequently, caffeine and EGCG should work synergistically because epinephrine secretion (from caffeine ingestion) enhances metabolism, whereas EGCG reduces net energy intake and offsets further increases in adiposity. Taurine ingestion may facilitate lipolysis by increasing peroxisome proliferator-activated receptor-γ coactivator-1α in white adipose tissue (37), an effect that warrants the addition of taurine to the weight-loss products.
Recent research has reported that products containing caffeine and EGCG as primary ingredients (or in combination) increases metabolism and promotes lipolysis (6,10-12,29,31). Although less research is available on taurine, studies have suggested it to positively influence mood, lipid metabolism, and weight loss and when combined with caffeine to influence memory, mood, and information processing (1,34,39,42). Our laboratory has recently found a TD containing caffeine, EGCG, and taurine to significantly increase metabolism and promote lipolysis (10,29). Specifically, acute consumption of this TD significantly increased resting energy expenditure (REE) and serum markers of lipolysis assessed by free fatty acid and glycerol appearance (10). In a follow-up investigation, our laboratory reported on the effects of chronic TD consumption. Free fatty acid area under the curve (AUC) values were significantly greater after TD ingestion compared with a noncaffeinated, noncaloric placebo after 28 days of supplementation, and percent body fat and fat mass were significantly decreased in the TD group when compared with the placebo group independent of exercise (29).
The majority of research conducted on TDs has examined the efficacy of the product compared with a noncaffeinated, noncaloric placebo (10,23,30), but no studies to our knowledge have directly examined the physiological differences that may occur between genders. This is important as current formulations of TDs do not standardize the dosage of active ingredients relative to body mass; thus making it likely that divergent responses may occur between genders given that men generally possess more body mass relative to women (23). Further support to this hypothesis has shown that women tend to oxidize less fat for energy at rest and during exercise compared with men (25,31). The present investigation involves a retrospective analysis to a previously published data set (29) in which no gender effect was investigated. The initially published report examined the overall efficacy of the TD without a gender-specific analysis. The purpose of this investigation was to examine if gender differences were evident between a group of men and women who consumed a commercially available TD for a period of 4 weeks for changes in body mass, body composition, energy expenditure, and markers of lipolysis.
Experimental Approach to the Problem
This study was conducted as a single-blind, parallel, matched-pair, placebo-controlled investigation. Upon recruitment, subjects were matched into clusters according to age and body mass before beginning the study. In a balanced format, all subjects (N = 60) were assigned to ingest on a daily basis either a TD (men: n = 15 and women: n = 15) or a placebo drink (men: n = 15 and women: n = 15) for the duration of the study. All participants were tested at baseline (day 0) and after 28 days (day 28) for changes in free fatty acids, glycerol, REE, respiratory exchange ratio, body mass, and body composition. To determine metabolic changes in all groups, the response to ingestion of a single 336 mL drink on days 0 and 28 was measured by repeated blood samples, energy expenditure, and substrate oxidation measurements before (0 minutes) and 60, 120, and 180 minutes after drink ingestion. Primary outcome variables were gender-specific responses to metabolic indicators (e.g., free fatty acid, glycerol, respiratory exchange ratio, and REE), whereas secondary outcomes were changes in body mass and body composition. It is hypothesized from these data that daily ingestion of a TD for 4 weeks will result in gender-dependent outcomes relative to changes in biochemical markers of lipolysis (e.g., free fatty acids and glycerol), substrate oxidation, energy expenditure, and body mass and body composition changes.
Healthy college-aged men (mean ± SE; 23.2 ± 0.74 years; 177.2 ± 1.12 cm; 81.7 ± 2.07 kg; 22.8 ± 1.33%; n = 30) and women (23 ± 1 years, 23.4 ± 0.56 years; 165.6 ± 1.59 cm; 62.1 ± 1.18 kg; 28.3 ± 1.36%; n = 30) chose to participate in this study. All testing was conducted after the participant completed the informed consent and comprehensive medical history questionnaires in compliance with the Institutional Review Board at the University of Oklahoma. Participants were excluded if they (a) had any history of metabolic, hypertension, hepatorenal, musculoskeletal, autoimmune, or neurological disease; (b) were currently taking thyroid, antihyperlipidemic, hypoglycemic, antihypertensive, or androgenic medications; and (c) had taken nutritional supplements or regularly consumed foods that may contain ingredients or bioactive compounds that may affect metabolism (i.e., over 100 mg·d−1 of caffeine, ephedrine alkaloids, guggulsterones, etc.) or muscle mass (i.e., creatine, protein, amino acids, androstenedione, dihydroepiandrosterone, etc.) within 3 months of starting the study.
In a single-blind fashion, participants were randomly assigned to ingest 336 mL of a TD that was provided by the manufacturer (CELSIUS, Celsius, Inc., Delray Beach, FL, USA) or a noncaffeinated noncaloric PLA (Caffeine-Free Diet Coke, Coca Cola) after baseline testing at any point during the day for the duration of the investigation. The TD contained a proprietary blend of guarana extract (seed), green tea leaf extract (leaf) standardized to 10% EGCG, 200 mg caffeine, taurine, glucuronolactone, and ginger extract (root) (Table 1). The randomly assigned drinks were stripped of all identifying information before administration and compliance was monitored by having participants pick up drinks from the laboratory on a weekly basis and completing an ingestion log throughout the duration of the study. Additionally, participants were instructed to avoid regular consumption of foods known to be high in caffeine such as caffeinated soft drinks and coffee. Dietary intake was monitored with 3 day dietary recalls (2) (2 week days and 1 weekend day) completed before day 0 and day 28 testing sessions to ensure between-group homogeneity for energy and macronutrient consumption and were assessed using the Food Processor III Nutrition Software version 8.6 (ESHA Nutrition Research, Salem, OR, USA).
During days 0 and 28, each subject reported to the laboratory after a 12-hour fast. Upon arrival, participants were asked to void their bladder and change into a tight fitting swimsuit. Participants were barefoot for measurement of height that was conducted using a standiometer (Detecto, Webb City, MO, USA). Assessment of body mass, fat mass, fat-free mass, and percent body fat was done using air plethysmography (BOD POD®, Life Measurement Inc. Concord, CA, USA). Before each test, the device was calibrated according to the manufacturer's instructions, and percent body fat was estimated using the Brozek equation (8). Our laboratory colleagues have previously demonstrated total error of measurement values of 0.66% body fat using this technique (29). After body composition assessment, participants rested in a quiet room for 5 minutes where heart rate and blood pressure were determined using an electronic sphygmomanometer (HEM-757, Omron HealthCare Inc., Vernon Hills, IL, USA), which has been reported to have an SD of ±3 mm Hg for blood pressure and to be within ±5% of pulse rate.
Resting energy expenditure and respiratory exchange ratio were determined using indirect calorimetry (True One 2400® Metabolic Measurement System, ParvoMedics Inc., Sandy, UT, USA). Before each testing session, the metabolic cart was calibrated according to the manufacturer's specification. For testing, a clear, hard plastic breathing hood with a plastic drape was attached to a metabolic cart and was placed over the participant's head and upper torso. To ensure accuracy of REE assessment, percent CO2 values were maintained within 1.0-1.2% by manual adjustment of flow rate. Mean oxygen uptake (V̇o2) and carbon dioxide output (V̇co2) were measured for each breath and averaged over 15-second intervals. Resting energy expenditure and respiratory exchange ratio were measured for 20 minutes, but the data reported represent a 5-minute window at the end of the collection period in which a criterion variable (V̇o2) deviated by less than 5%. Test-retest correlations (r) using this device were in the range of 0.550-0.747 with a mean intraclass coefficient of 0.893, p < 0.001. Before drink ingestion, an initial measurement (0 minute) was made, and after drink ingestion, participants had their REE and respiratory exchange ratio determined at 60, 120, and 180 minutes after drink ingestion.
Participants then donated approximately 6 mL of fasting blood from an antecubital vein by using standard venipuncture techniques. All blood samples were collected in 6-mL serum separation vacutainer tubes (BD Vacutainer®, Franklin Lakes, NJ, USA). Each vacutainer was inverted several times and immediately centrifuged at 3,500 rpm for 15 minutes at room temperature. The resulting supernatant was then aliquoted into 2 microcentrifuge tubes and stored at −20°C for subsequent analysis of glycerol and free fatty acids. Before drink ingestion, an initial blood sample (0 minutes) was collected, and after drink ingestion, subjects donated additional blood samples 60, 120, and 180 minutes after drink ingestion. After day 0 testing, participants ingested 1 daily dose of the randomly assigned drink per day for 27 days before returning to the laboratory on day 28 at the same time of the day as baseline testing and performed identical testing procedures as was conducted on day 0.
Lipolysis was determined in duplicate by the assessment of serum concentrations of glycerol and free fatty acids. Glycerol was assessed using a calibrated commercial oxidase enzyme reaction analyzer and reagents (Analox GM7, Analox Instruments, London, United Kingdom). Free fatty acids were assessed using a commercial colorimetric assay (Roche, Penzeberg, Germany) and read with a spectrophotometer (SmartSpec™Plus, Bio-Rad, Hercules, CA, USA) at a wavelength of 546 nm. Assay precision (coefficient of variation [C V]) and accuracy (percent of nonrecovery) were calculated using control samples for glycerol and free fatty acids by 8 replicate determinations. The reported C V and percent of nonrecovery for glycerol (0.24 mM control serum: 2.5 and 1.2%) and free fatty acids control serums (0.35 mM control serum: 8.9 and 3.6%) were found to be within acceptable ranges (4,14).
Separate AUC analyses using the trapezoidal method were performed for free fatty acids, glycerol, and REE on days 0 and 28. Respiratory exchange ratio analysis data obtained from each time point (pre, 60, 120, and 180 minutes) were averaged to attain a 3-hour value for each group of participants on both days 0 and 28. Thus, all nutritional data, body composition, respiratory exchange ratio, and all AUC data were analyzed using separate 4 × 2 (group × time point [days 0 and 28]) repeated-measures analysis of variance, Bonferroni post hoc comparisons were performed to delineate within group changes, and Tukey post hoc comparisons were used to delineate between-group changes. To clearly delineate between-gender responses according to our hypothesis, free fatty acid, glycerol, REE, and respiratory exchange ratio data were compared between genders on days 0 and 28 using separate independent t-tests. Within-gender responses were evaluated using separate dependent t-test. All statistical analyses were performed using SPSS (version 16.0, SPSS Inc., Chicago, IL, USA). Data are presented as mean ± SE, and significance for all statistical analyses was determined using an alpha level of ≤0.05.
Nutrient consumption was normalized for body mass (in kilograms) and is presented in Table 2. No significant interactions (p > 0.05) were found for energy, protein, carbohydrate, or fat consumption. There was a main effect for time because there was a significant decrease in carbohydrate consumption (p < 0.05) from days 0 to 28. Within-group analyses revealed a significant decrease in carbohydrate consumption in the female PLA group. A significant interaction was found for caffeine consumption (p < 0.001). There was a main effect for group (p < 0.001), because the TD groups consumed significantly more caffeine than did the PLA groups and time (p < 0.001) because caffeine consumption was significantly greater on day 28 compared to on day 0 in the male and female TD groups.
Day 0 and Day 28 Thermogenic and Lipolytic Response
Acute and chronic thermogenic and lipolytic data using AUC analysis with day 0 and day 28 response graphs for free fatty acids, glycerol, and REE are presented in Figures 1-3, respectively. There was no significant interaction (p = 0.11) for free fatty acid AUC analysis, but there was a significant group effect (p = 0.002) as the male PLA group mobilized significantly less free fatty acids than every other group on day 0 and the female TD group mobilized significantly more free fatty acids than every other group on day 28. The response graph indicated that the male TD group had significantly greater serum free fatty acid concentrations at 60, 120, and 180 minutes post-drink consumption compared to on day 28. Additionally, the female TD group had significantly greater serum free fatty acid concentrations than did the male TD group at 120 and 180 minutes post-drink consumption on day 28. There was a significant interaction for glycerol AUC (p = 0.02), because the male PLA group experienced a significant increase (p = 0.04) in serum glycerol appearance from days 0 to 28. There was no main effect for group (p = 0.82) or time (p = 0.43). No interaction was present for REE AUC (p = 0.09), but there was a main effect for group (p < 0.001). On day 0, the female TD group had a lower REE than did the male PLA and male TD groups, whereas the female PLA group had a lower REE than did the male PLA, male TD, and female TD groups. On day 28, the female TD and female PLA groups had significantly lower REE than did the male TD and male PLA groups. Resting energy expenditure was significantly greater in the male TD and male PLA groups compared with the female TD and female PLA groups at each time point on days 0 and 28. Acute and chronic respiratory exchange ratio data along with day 0 and 28 response graphs are presented in Figure 4. There was no significant interaction (p = 0.08), group (p = 0.06), or time effect (p = 0.27). The response graph indicated that the female TD group had a significantly higher respiratory exchange ratio on day 28 compared to on day 0 at 120 (p = 0.03) and 180 (p = 0.03) minutes post-drink consumption. Additionally, the female TD group had significantly greater respiratory exchange ratio values 120 minutes (p = 0.03) post-drink consumption on day 0 and 60 minutes (p = 0.04) post-drink consumption on day 28 compared to the male TD group.
No significant interaction was present for body mass (p = 0.10) or fat-free mass (p = 0.13), but there was a main effect for group in each condition (p < 0.001) as men were significantly heavier and had more fat-free mass than did women. There was a significant interaction for fat mass (p < 0.001) and percent body fat (p = 0.002) as the male TD group lost a significant amount of fat mass and the female PLA group gained a significant amount of fat mass over the duration of the investigation. A group effect was present for percent body fat (p = 0.02) as women had significantly more percent body fat than men (Table 3). Figure 5A, B display the delta change (prevalue − postvalue) for percent body fat and an individual response graph, respectively.
The purpose of the present study was to investigate gender-specific metabolic responses to prolonged of a commercially available TD. The primary findings from this study suggested that women tended to mobilize greater amounts of fat (Figure 2), whereas men tended to oxidize more fat (Figure 4) and experienced improvements in body composition (Figure 5). Previous investigations have examined the metabolic response of TD ingestion; however, none of these studies examined any gender-related outcome (10,16,19). Our laboratory has reported increases in energy expenditure and fat mobilization after acute and prolonged ingestion of a TD containing caffeine, EGCG, and taurine (10). The prolonged study of this data set investigated mixed-gender effects (29) but failed to investigate any gender-specific outcomes; a potential effect that was brought to light by a recent report of Lockwood et al. (23). This study indicated that with exercise over a 12-week period, men may experience greater changes in body composition than do women (23). Thus, the decision was made to perform a retroactive analysis of the Roberts et al. data set (29) to determine the immediate metabolic changes for each gender that occur after acute ingestion of a TD before and after 28 days of daily ingestion. It was hypothesized that women would experience a greater metabolic effect because of greater relative dosing (milligrams per kilogram body mass) of active ingredients and gender differences in substrate use at rest and during exercise (17,25). As expected, men in the TD group received a significantly lower relative dose of caffeine compared to the women in the TD group (male TD = 2.5 ± 0.39, female TD = 3.3 ± 0.59, p = 0.001). Specific delivered dosages of EGCG and taurine could not be calculated because of their inclusion in a proprietary blend of the ingredients; however, popular over-the-counter thermogenic drinks contain around 90 mg of EGCG (28) and 1-2 g of taurine (3). Relative dosing values are important because of the commonly reported dose-related outcomes associated with these ingredients (9,22).
Caffeine is thought to be the likely candidate for the metabolic action seen between the genders. Briefly, the lipolytic mechanisms associated with caffeine ingestion include the following: (a) an enhanced secretion of epinephrine (5) and greater production of cAMP-mediated activation of hormone-sensitive lipase and the subsequent liberation of glycerol and free fatty acids into the plasma (38), (b) phosphodiesterase inhibition, leading to further sustainment of cAMP and subsequent activation of lipolysis, and (c) antagonism of peripheral adenosine (A1) receptors, yet further increasing intraadipocyte cAMP levels. It is likely that caffeine is the primary and only agent resulting in lipolytic actions; however, firm conclusions to this point cannot be drawn from our study design. It is also possible that greater values in serum estrogen in women may work to further sustain caffeine levels through its inhibition of cytochrome P450 1A2 (21,35). Therefore, although plasma caffeine concentrations were not assessed, women consuming the TD may have had greater circulating concentrations of caffeine than did men allowing for greater fat mobilization via the aforementioned lipolytic mechanisms. In support of this, day-28 free fatty acid AUC analysis revealed that the women who ingested a TD had significantly more serum free fatty acids than did the other 3 cohorts. However, the lack of plasma caffeine, epinephrine, and estrogen assessment is a limitation in the present investigation and limits our ability to further explore these possibilities. While taurine has been used as a lipolytic agent that may improve mood (1) and decrease risk of heart disease (41,42), although it is unknown whether the dose received in either gender was sufficient to elicit a physiological response. Although Zhang et al. reported that a daily 3-g dose for 7 weeks reduced triglycerides and reduced body mass (42), a post hoc analysis revealed no such effect from the present study (data not shown).
In contrast to the initial hypothesis, the man who ingested the TD experienced significant reductions in fat mass and percent body fat in comparison to men who ingested PLA and women who ingested the TD. Although contradictory to greater liberation of free fatty acids in women who ingested a TD on days 0 and 28, men who ingested the TD lost more fat mass. Respiratory exchange ratio data (Figure 4) suggest a greater oxidation of fat by men who ingested the TD in comparison to women who also ingested the TD on days 0 and 28. This is supported previously where women were found to oxidize less fat for energy than men at rest (25,36). Hence, this between-gender endocrinological phenomenon would again help explain the significant changes in percent body fat and fat mass for the male TD group and the lack of response in the female TD group.
A lack of internal stability in our body composition measurement or changes in normal dietary intake are potential confounding influences; however, we have previously reported a total error of measurement for body composition using the BOD POD to be approximately 0.66% with a test-retest intraclass coefficient of >0.99 (29). Using minimum difference standards (40), the 1.18% gender difference (Figure 5) seen in body composition exceeded the SEM of this device of 0.48% (24), thus suggesting that this effect was a real difference. Furthermore, a significant decrease in carbohydrate intake occurred in the female PLA group; however, no other changes in dietary variables were found (Table 2). It is likely that self-reporting errors may have occurred in the dietary analysis (33), but previous studies have also reported validity with 3-day food records (32).
In conclusion, findings from this study provide preliminary biochemical and metabolic evidence of greater increases in circulating free fatty acids in women (Figure 1), whereas greater levels of fat oxidation and overall improvements in body composition were seen in men (Figure 5). Although supportive of other literature (23), the mechanism(s) associated with these findings are not well understood because previous research has suggested an inhibitory effect of estrogen over cellular respiration components (35), whereas the rates of reesterification (18) between genders may have also impacted the findings from the present investigation.
Fitness and exercise professionals are faced with a barrage of efficacy-based questions in their practice regarding exercise and nutrition. The use of thermogenic ingredients and especially TDs in the past several years has increased by several degrees of magnitude. Completed studies have shown that immediate effects of these drinks results in an increase in lipolysis and energy expenditure. Outcomes from this study help fitness enthusiasts and professionals understand the gender-specific responses that may occur in response to ingesting a commercially available TD. As such, it appears that greater increases in free fatty acids (a marker of lipolysis) result in women, whereas men may experience a greater decrease in body composition after daily ingestion of a TD for 28 days.
The authors would like to thank the study participants and additional graduate students and faculty in the Department of Health and Exercise Science who assisted in part with this study investigation. Celsius Inc. (Delray Beach, FL) provided funding for this project through an unrestricted research grant to the University of Oklahoma. All researchers involved independently collected, analyzed, and interpreted the results from this study and have no financial interests concerning the outcome of this investigation; thus, no conflicts of interest are evident in this report. The results from this study do not constitute endorsement of the product by the authors, their institution or the National Strength and Conditioning Association.
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Keywords:Copyright © 2010 by the National Strength & Conditioning Association.
metabolism; thermogenesis; caffeine; EGCG; energy drink