This was a randomized, double-blind, placebo- and diet-controlled, parallel groups with repeated measures study design. Both groups were assigned to a 12-week periodized RT protocol. Blinding occurred via an outside researcher who sent the supplement and placebo in identical opaque capsules. This researcher was not involved in direct data collection, and did not meet any of the subjects. For this reason neither the researchers conducting the study nor the subjects knew who was in each group. Moreover the code was not broken until after all of the data were entered into Microsoft® Excel, and sent to an outside researcher who was also blinded to the treatment groups. The protocol was divided into three phases. Phase one consisted of a three times per week non-linear periodized RT program for weeks 1–8, modified from Kraemer et al. . Phase two consisted of a two-week overreaching cycle during weeks 9 and 10. Finally, phase three consisted of participants tapering for weeks 11 and 12. Muscle mass and body composition were measured at baseline and at the end of weeks 4, 8, and 12. Muscle strength, vertical jump power, Wingate peak power (PP), creatine kinase (CK), C-reactive protein (CRP), free and total testosterone, and perceived recovery were measured at baseline and after weeks 4, 8, 9, 10 and 12. Additionally, CK, CRP, free and total testosterone, and perceived recovery were measured after week 1 of training. Protein breakdown was assessed using urinary 3-methylhistidine: Creatinine ratio (3-MH:Cr) at baseline and after weeks 1 and during the overreaching phase at weeks 8, 9, and 10. These variables were used to assess the effects of ATP on performance, hormone status, and indices of muscle damage and recovery during an overreaching cycle. The ClinicalTrials.gov registration ID was NCT01508338.
Twenty-four resistance-trained males were selected for the study. However, due to injury three subjects dropped out of the study leaving 21 subjects (11 ATP supplemented and 10 placebo supplemented) aged 23.4 ± 0.7 years, with an average one-repetition maximum (1RM) squat, bench press, and deadlift of 1.71 ± 0.04, 1.34 ± 0.03 and 2.05 ± 0.04 times their bodyweight were recruited for the study. Participants could not participate if they were taking an anti-inflammatory agent, a performance-enhancing supplement, if they smoked, or if they had consumed nutritional supplements during the three months prior to data collection. Each participant signed an informed consent approved by the University of Tampa Institutional Review Board before participating in the study.
Muscle strength, power, body composition and skeletal muscle hypertrophy testing
After familiarization procedures, muscle strength was assessed via 1RM testing of the back squat, bench press, and deadlift. Each lift was performed as described by the International Powerlifting Federation rules . We used an intraclass correlation coefficient (ICC) (2,k) formula  to determine the reliability of repeated measures within testers. The ICC for strength measures ranged from 0.96 to 0.98. Body composition (lean body mass, fat mass, and total mass) was determined by dual-energy x-ray absorptiometry (DXA; Lunar Prodigy enCORE 2008, Madison, Wisconsin, U.S.A.). Skeletal muscle hypertrophy was determined via the combined changes in ultrasonography-determined muscle thickness of the vastus lateralis (VL) and vastus intermedius (VI) muscles. The mean of three measurements by the same blinded investigator were taken at 50% of femur length over the mid-belly of the muscle with the subjects lying in a supine position. The precision for the test-retest of muscle thickness measurements was 0.975.
Muscle power was assessed during maximal cycling and jumping movements. During the cycling test, volunteers were instructed to cycle against a predetermined resistance (7.5% of body weight) as fast as possible for 10 seconds . The saddle height was adjusted to the individual’s height to produce a 5–10° knee flexion while the foot was in the low position of the central void. A standardized verbal stimulus was provided to each participant. Power output was recorded in real time during the 10-second sprint test, by a computer connected to the standard cycle ergometer (Monark model 894e, Vansbro, Sweden). Peak power (PP) was recorded using Monark Anaerobic Wingate Software, Version 1.0 (Monark, Vansbro, Sweden). The ICC of muscle peak power was 0.96.
Measurements of PP were also taken during a vertical jump (VJ) test performed on a multicomponent AMTI force platform (Advanced Mechanical Technology, Inc., Watertown, MA), interfaced with a personal computer at a sampling rate of 1000 Hz . Data acquisition software (LabVIEW, version 7.1; National Instruments Corporation, Austin, TX) was used to calculate PP. Peak power was calculated as the peak combination of ground reaction force and peak velocity during the accelerated launch on the platform. The ICC of VJ power was 0.97.
Supplementation and diet control
Prior to the study, participants were randomly assigned to receive either 400 mg per day of ATP disodium or maltodextrin (placebo), consumed orally via a two-piece gelatin capsule 30 minutes prior to RT sessions. On non-training days, participants were instructed to consume one dose on an empty stomach prior to breakfast. The placebo and ATP capsules (400 mg) were obtained directly from the commercial manufacturer (TSI USA Inc., Missoula, MT) and were produced in compliance with U.S. cGMPs for dietary supplements. The Certificate of Analysis for both the placebo and ATP capsules were provided and verified the supplement contents. Moreover, we verified the capsule’s purity by HPLC. Two weeks prior to and throughout the study, participants were placed on a diet consisting of 25% protein, 50% carbohydrates, and 25% fat by a registered dietician who specialized in sport nutrition. Participants met as a group with the dietitian, and they were given individual meal plans two weeks prior to the onset of the study. Diet counseling was continued on an individual basis throughout the study.
Resting blood draws
All blood draws were obtained after an overnight 12-hour fast via venipuncture by a trained phlebotomist, and were scheduled at the same time of day to avoid influences of hormonal variations. Whole blood was collected and transferred into appropriate tubes and centrifuged at 1500 x g for 15 min at 4°C. Resulting serum and plasma were then aliquoted and stored at −80°C until subsequent analyses. A portion of the blood samples taken at weeks 0, 4, 8, and 12 were used for measurements (ANY LAB TEST NOW®, Tampa, Fl) of glucose, blood urea nitrogen, creatinine, eGFR, Na, K, Cl, CO2, Ca, protein, albumin, globulin, albumin:globulin ratio, total bilirubin, alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase. A complete blood count was also performed on each blood sample. The commercial laboratory performing the safety analysis was instructed to inform investigators of any abnormal values in any parameters measured during the study time points measured. In the case of an abnormal value, the subject would be told and not be allowed to continue the study. Because no abnormal values were detected during the study in these healthy and highly fit subjects, the total aggregate numbers were not presented to the investigators until the end of the study.
Samples were thawed once and analyzed in duplicate for each analyte. Serum free and total testosterone, cortisol, and C-reactive protein (CRP) were assayed via ELISA kits obtained from Diagnostic Systems Laboratories (Webster, TX). All hormones were measured within the same assay and on the same day to avoid compounded inter-assay variance. Intra-assay variance was less than 3% for all analytes. Serum creatine kinase (CK) was measured using colorimetric procedures at 340 nm (Diagnostics Chemicals, Oxford, CT).
Perceived recovery status scale
Perceived Recovery Status (PRS) Scale was measured at all measurement times and in particular at weeks 8, 9, and 10 to assess participant recovery during the overreaching phase. The PRS Scale consists of values between 0–10, with 0–2 being very poorly recovered and characterized by anticipated declines in performance; 4–6 is defined as low-to-moderately recovered and characterized by no expected change in performance; and, 8–10 represents high perceived recovery and correlates strongly with increases in performance .
A one-way ANOVA model was used to analyze the baseline characteristic data using the Proc GLM procedure in SAS® (SAS Institute, Cary, NC). The main effect of treatment (Trt) was included in the model. Actual values for muscle strength and power, body composition, muscle damage, hormonal status, and PRS changes over the 12-week study were analyzed by using a repeated measures ANOVA using the Proc Mixed procedure in SAS®. The initial baseline value (e.g. week 0) was used as a covariate with the main effects of Time, Trt, and the interaction Trt*Time in the model. The overreaching cycle of the study was further assessed by using repeated measures ANOVAs with the Proc Mixed procedure in SAS®. Values measured at the week-8 time point were used as a covariate with the main effects of Time, Trt, and Trt*Time for the overreaching phase (phase two). The Least Squares Means procedure was used to compare Trt means at each time point. Statistical significance was determined at p ≤ 0.05.