Experimental approach to the problem

A randomized, placebo controlled, double-blind study was used to compare the effects of consuming a low dose of Teacrine® (TC-LD), a high dose of Teacrine® (TC-HD) or a placebo (PLA) for 8 weeks on apparently healthy, recreationally active, regular caffeine consuming, males and females. The following dependent variables were assessed to determine supplementation-induced differences based on the three groups: clinical safety markers, heart rate, blood pressure, questionnaires (VAS, Yale PAQ and POMS) and body composition. Subjects were instructed not to change their exercise habits or their standard dietary habits. The design allowed for examination of the effects of Teacrine® in a low and high dose with no diet or exercise regimen manipulations.

Subjects

Sixty men and women (mean ± SD age, height, weight, body fat percentage: 22.55 ± 4.6 years, 174.09 ± 12.4 cm, 77.47 ± 17.4 kg, 23.4 ± 9.9 %) participated in this study. Subjects were not allowed to participate in this study if they had any metabolic disorder including known; heart disease, arrhythmias, diabetes, cancer; if they were taking medications related to chronic disease; if they were taking or had taken dietary supplements (other than multi-vitamins and/or minerals) within 8 weeks prior to enrollment; if they had participated in another clinical trial within 8 weeks prior to enrollment; if they had any known allergies or sensitivity to any ingredient in the test product; and, if they were currently pregnant, nursing or became pregnant during the duration of the study. Subjects were asked to maintain their normal dietary intake and exercise habits for the duration of the study. Subjects meeting eligibility criteria were informed of the requirements for the study and signed approved informed consent statements in compliance with the guidelines of the Institutional Review Board at the University of Mary Hardin-Baylor (UMHB). All anthropometric and hemodynamic/ECG testing was conducted in the Human Performance Laboratory (HPL) and all blood processing was conducted in the Exercise Biochemistry Laboratory at UMHB.

Baseline (T1) testing

Prior to the baseline session, subjects were instructed to record all food intakes on dietary record forms for 3 days (3-d). At the end of the 3 days, diet logs were brought back to the lab to be entered into Esha Food Processor (ESHA Research, Salem, OR) for nutritional assessment. For all testing sessions, subjects were instructed to refrain from exercise for 48 h, abstain from smoking, caffeine, tobacco as well as fast for 12-h prior to baseline testing and refrain from alcohol consumption for 24 h. Subjects were encouraged to consume plenty of water the day prior to each testing session and to consume water upon waking prior to reporting to the lab.

The day of baseline testing (T1), subjects reported to the HPL during their scheduled time (between the hours of 0500–0800) to be weighed via TANITA electronic scale (Model TBF-310, TANITA, Arlington Heights, IL) and height measured using a SECA 242 instrument (242, SECA, Hanover, MD). Resting heart rate (RHR) and blood pressure (SBP & DBP) were obtained in a rested and seated position via OMRON Digital Blood Pressure Monitor (Model HEM-907XL, OMRON Healthcare, Inc. IL U.S.A.). HR, recorded as beats per minute, SBP and DBP, recorded as mmHg, were measured at the completion of the ECG. Body composition was determined using bioelectrical impedance analysis via InBody (Model 770, InBody Co., Ltd, Cerritos, CA). For InBody measurements, test-retest reliability in our lab are as follows: Fat Mass: ICC = 0.99; Lean Mass: ICC = 0.99; percent body fat: ICC = 0.998. All InBody tests were conducted by the same technician and strict manufacturer guidelines for calibration and testing procedures were followed.

Electocardiogram (ECG)

Once body composition was completed, subjects performed an electrocardiogram (ECG). Leads were placed in standard clinical fashion to produce a 12 lead ECG (I-III, V1-V6, aVR, aVL, aVF). Subjects remained in a supine position for 5 min. Cardiac rhythm was monitored through a Quinton Eclipse Premier Electrocardiograph (Cardiac Science Corporation, Bothell, WA). Data was printed from the 12-lead ECG machine and RR interval, RP interval, QRS duration, and QT interval were recorded.

Blood collection protocol and questionnaires

Blood was collected from an antecubital vein with the subject in a supine position. To negate diurnal variation in hormonal status sampling was taken at the same time for each of the testing days. Upon extracting the blood into two 7.5 ml tiger top Vacutainer®, the sample was inverted gently (5–6 times) and was sent to the blood analysis room to allow time to clot and was centrifuged for 15 min within 30–60 min at room temperature to yield serum samples for complete metabolic and lipid panel measures. A 4 mL purple top Vacutainer® was drawn from the forearm vein and was mixed gently via inversion and then refrigerated for CBC measurements. All samples were then refrigerated and sent to Quest (Quest Diagnostics Inc., Irvine, TX) for analysis. All blood collection and processing protocols were in line with the recommendations from Quest diagnostics to ensure optimal samples for analysis.

After blood collection subjects were given the supplement based on their randomly assigned group and told to ingest the supplement with 8 fluid ounces of water and remained in the HPL for 30 min to ensure no allergic reaction occurred. Immediately after ingestion of the supplement, subjects were given a symptoms survey, Profile of Mood States (POMS), the Visual Analog Scale (VAS) (including: energy, focus, concentration, sleep, vigor, and motivation to exercise), and YALE Physical Activity Survey (YALE) to establish baseline measures of these dependent variables. The VAS was structured as a 15-cm scale that was labeled as “lowest possible” and “highest possible” for each VAS variable as used in previous research [10, 11]. The YALE survey has been used in various settings as a measure of physical activity [12, 13].

Supplementation protocol

Subjects were matched according to T1 body mass and gender, then in a randomized, double-blind manner were assigned to consume a low dose of 200 mg of TeaCrine® (TC-LD) (n = 19, 23.5 ± 6 years, 175.6 ± 13.3 cm, 78 ± 18.4 kg, 23.4 ± 10 %BF), a higher dose of 300 mg of TeaCrine® (TC-HD) (n = 21, 21.38 ± 2.3 years, 172.7 ± 12 cm, 77 ± 17.3 kg, 24 ± 10.2 %BF) or 300 mg of maltodextrin (PLA) (n = 20, 22.8 ± 4.7 years, 174 ± 12.1 cm, 77 ± 17.2 kg, 22.3 ± 9.9 %BF) once per day, after breakfast but before lunch, with 8 oz of water for the 8-week study protocol. Dosing was selected based on previous research that has been presented at the ISSN Annual Meeting [2]. The terms high and low dose are relative to what was utilized in this study and future research is needed to truly establish what determines a true “high” and “low” dose of TeaCrine®. Compliance of pill ingestion was monitored by research assistants every 2 weeks in the HPL when subjects would return to get a new 2 week supply of pills for their respective treatment group.

Follow-up testing and dietary intake

Throughout the 8-week protocol, subjects returned to the HPL for follow-up testing at week 4 (T2) and week 8 (T3) and repeated identical testing procedures as discussed during T1 testing. Note that prior to T1 testing subjects were instructed to record all food intakes on 3-d dietary record forms. Subsequently, subjects were told to maintain their diets as closely as possible to the previously 3-d recorded intakes (on T1). Dietary intakes for calories and macronutrient profiles are displayed in Table 1.

Table 1 Macronutrient intakes Full size table

Statistics

Statistical analyses was performed using SPSS v22.0 (Chicago, IL, USA). An alpha level (α) of p ≤ 0.05 was used to determine significance within or between groups.

Mixed-factorial ANOVA’s with repeated measures [group (PLA × TC-LD × TC-HD) × gender (M × F) × time (T1, T2, T3)] were used for all dependent variables. If a significant [(time), (time*group), (time*group*gender)] interaction was observed, additional post hoc analyses were performed as follows: 1) for significant time effects, within-group repeated measures ANOVAs with Bonferroni’s corrections were performed, 2) for significant time*group interactions, within-group repeated measures ANOVAs with Bonferroni’s corrections, and one-way ANOVAs at each time point with Tukey post hoc analyses were performed, and 3) for significant time*group*gender interactions, within-group repeated measures ANOVAs with Bonferroni’s corrections were performed for each gender, and one-way ANOVAs with Tukey post hoc analyses were performed for each gender. Unless otherwise noted, data are presented as mean ± SD.