Konstantinos Havenetidis1, Ourania Matsouka2 , Carlton Brian Cooke1 and Apostolos Theodorou1

¹ Leeds Metropolitan University, School of Leisure and Sports Studies, Leeds, UK
² Democritus University of Thrace, Department of Sports Science and P.E., Komotini, GREECE

Currently, an increasing number of investigators (Balsom et al., 1993; Kamber et al., 1999; Rockwell et al., 2001) have investigated the potential benefit of creatine as an ergogenic aid for improving exercise performance. In the majority of the creatine supplementation studies the dosages used during an Acute Creatine Loading (ACRL) were 20g-25g day^-1 for a period of 4-5 days (Greenhaff et al., 1994; Birch et al., 1994; Rockwell et al., 2001) totalling 100g of creatine. This supplementation regimen was based on the results of preliminary studies (Harris et al., 1992) which showed that dosages of 20g. day^-1 and 30g. day^-1 for a period of 3.5 days (n=1) and 4 days (n=3),  7 days (n=1) in group of  five sedentary subjects led to an increase of 20% in total creatine pool. However, Harris et al. (1992) did not proceed to any comparisons in exercise performance data for each creatine dosage separately, in order to establish the most beneficial creatine loading regimen, possibly due to the small size of the sample group. Using a different experimental design Vandenberghe et al. (1999) reported that 25g day^-1 for the first two days resulted an increase of 11% in muscle CP and 5-13% in muscle torque, whilst continuation of supplementation for another three days did not add to these effects. That report reverses the ‘‘traditional’’ 100g ACRL and there emerges the need for further research regarding creatine dosage optimisation and exercise potentiation. Such research will ensure the effectiveness of an ACRL, help avoid any overdose side effects, and present some reference creatine values for minimising urine creatine excretion alongside with the maximum beneficial effect on performance. Therefore, the aim of the present study was to establish an ACRL, which would cause the greatest benefit on repetitive all-out cycle performance on active individuals.

Since each subgroup was following a different ACRL (40g, 100g, 135g) they performed the exercise testing on separate weeks in order to make management of the experiment feasible. One reason was the required commitment and consistency of the subjects in consuming the beverage at a specific time each day was essential for the fulfilment of the purposes of the study. Therefore, subgroup 1, 2, and 3 performed the exercise testing in weeks 4-6, 7-9 and 10-12 respectively. For the subgroups 1, 2 and 3 the first, second and third week represented the baseline, placebo and creatine condition respectively. The control group performed the exercise testing in weeks 1-3 where the first week represented the baseline condition and the second and third the placebo condition. Since the kinetics of creatine washout has not been established yet, the investigator provided placebo and creatine supplements in the 1^st and 2^nd supplementation period respectively in order to avoid the carry-over effect. Even through placebo and creatine conditions were always administrated in this order they were conducted as a double blind design for the subjects and the test administrator.

All supplements were prepared by the investigators at University Campus, in powder form with similar texture, taste and appearance and were independently packaged in generic foil packets for double-blind administration. The supplement administrator who was not aware of the content of the packets dissolved placebo or creatine in 300ml of hot-warm water and hand over the solution to the subjects with morning, mid-day and evening meals. Two, five and seven solutions were prepared per day for the 40g, 100g and 135g ACRL respectively. Each solution was prepared by the supplement administrator immediately prior to the ingestion, and in the absence of the subjects.

The 4^th day and one-hour after the last placebo dose all subjects performed three Anaerobic Wingate Tests (AWT) interspersed with 6 minutes active recovery (60rpm). A standardised warm-up was performed which involved 5 min of cycling at 60 Watts immediately followed by two 2 s sprints. The warm up was followed by a 5 min rest period where the subjects remained seated before the start of the AWT. The ergometer used during the AWT testing was a MONARK 814E (Sweden) brated pre, and post each test was bolted to the floor in order to provide greater stability during maximal cycling. Before starting the test a belt attached to the wall at one end, was passed around the subject’s waist in order to prevent the subject rising out of the saddle. Saddle height adjustments for each subject were made prior testing as well a series of  6 s cycle sprints as part of a habituation process with AWT (Havenetidis et al., 2003). The experimental conditions and procedures for performing the AWT were those described by Bar-Or (1987). The exercise protocol was repeated the following week with the use of creatine supplements under the same conditions and by following the same experimental procedures. The present exercise protocol is considered an ideal model for assessing the ergogenic properties of creatine as reported by other investigators (Dawson et al., 1995; Prevost et al., 1997). Repeated sprints are associated with cumulative fatigue, a condition, which is preferable in studies investigating the ergogenic effect of various supplements. Using a single exercise bout a less marked creatine phosphate depletion (a metabolic state which is linked with fatigue) would have been produced and consequently a failure to quantify the relationship between the elevated muscle creatine phosphate stores (following creatine supplementation) and the prevention of performance reduction. In contrast, the present exercise protocol has shown to maximally stress the creatine phosphate pathway as creatine phosphate concentration at the end of the 3^rd AWT represented only 14% of the values measured at rest (Havenetidis et al., 2002).

Performance Measurements
The indices measured during the AWT were: Mean power (MP) and Minimum Power (MIP). MP was calculated as the average mechanical power output during each 10 s interval (MP 0-10s, 10-20s, 20-30s), whilst MIP as the lowest power output during any 1s period during the whole 30s AWT. All AWT indices were measured using a computer package (Lakomy, 1986). Physiological and performance measurements were not made in the control group because the experimental group served as its own control throughout the first series of measurements which were carried out without any supplement administration (control week).

No significant differences in protein and carbohydrate intake were observed among the control, placebo and creatine weeks across all groups (Table 1).

In the current study body mass (Mean ± SD) increased (p<0.01) from 82.5 ± 11.7 kg, to 82.9 ± 11.7 kg, following creatine supplementation across the three supplementation groups (t-value –4.084). Body mass values at placebo vs creatine supplementation for the 40g, 100g and 135g ACRL were 84.8 ± 15.7 kg vs 85.0 ± 15.5 kg, 76.9 ± 6.4 kg vs 77.2 ± 6.5 kg and 85.6 ± 10.8 kg vs 86.3 ± 10.8 kg respectively. Details on experimental group anthropometry are shown in Table 2.

Blood and urinary creatine were also significantly (p<0.01) increased and that increase was proportional to the creatine dosage ingested (Table  3).

Performance Data
On all occasions the highest power output was achieved during the first seconds of each AWT and then was followed by a steady decline. For the control group the statistics showed no significant differences in performance across the three weeks of testing, which indicates that repeated 30 s cycle sprints are not characterised by a significant learning effect (Figure 1). Average difference of performance between each AWT was ± 2.5% a value that was not statistically significant.

In contrast, among the three experimental groups significant differences were observed on condition (placebo vs creatine) and group by condition interaction (40g ACRL creatine condition versus 100g ACRL creatine condition, p<0.01 and 40g ACRL creatine condition versus 135g ACRL creatine condition, p<0.01). A summary of AWT data for all groups during baseline, placebo and creatine condition is presented in Table 3.

The present data showed that performance potentiation was influenced by the creatine dosage used. Figure 2 shows the percentage difference between creatine and placebo condition across all AWT indices for each supplementation group.

The present data also indicates that performance potentiation was not consistent for all supplementation groups and throughout the 30s period. This is more profound when power output expressed  per second (Figures 3 - 5).

As shown by Figures 3 to 5 there is an increase in power output throughout the 30 s period for the 100g and 135g ACRL in creatine to the placebo condition. In contrast, for the 40g ACRL the power slopes, after the 4^th second, for the placebo and creatine conditions were almost identical.

The present study showed that creatine supplementation per se does not necessarily improve exercise performance, but it is influenced by the amount of creatine ingested during a four-day ACRL. The amount of creatine ingested in the present ACRL was 40g, 100g and 135g and led to an average (across all AWT indices) improvement of 0.7%, 11.8% and 11.1% respectively. This difference in performance potentiation could be related to differences in blood creatine concentration (post supplementation) which influences creatine muscle uptake (Harris et al., 1992). Fitch and Shields (1966) who developed a model for creatine entry into muscle based on results with the use of guinea pigs showed that the facilitation of creatine entry is much greater when blood creatine concentration exceeds 1 mmol·l^-1. Based on the short time period that plasma creatine remains elevated (half-life 1-1.5 hours) it is possible that the effect of creatine ingestion on muscle entry did not last long, especially if the plasma levels (1.1 ± 0.1 mmol·l^-1) for the 40g ACRL were just above the threshold. The latter, together with an initial high muscle total creatine concentration could have resulted a decreased creatine uptake by the muscle. Alternatively, for the 100g and 135g ACRL significantly higher blood creatine concentrations were presented, alongside with a greater potentiation of performance compared to the 40g ACRL, which seems that is not sufficient to achieve the highest values of muscle creatine uptake. In contrast, creatine dosages totaling 100g or greater (135g) produce a blood creatine concentration far beyond the threshold of 1 mmol·l^-1, therefore facilitating creatine entry into the muscle. It is noteworthy that blood creatine values at placebo were similar among groups and within the normal values reported by other investigators (Tortora and Anagnostakos, 1990; Engelhardt et al., 1998) therefore, no initial differences existed between groups prior supplementation.

The use of a 100g compared to a 40g acute creatine loading produced a greater and constant potentiation of sprint cycle performance, whilst no significantly greater benefit occurred with the use of a greater dosage (135g). These findings support previous reports that the use of the commonly accepted creatine-loading regimen of 100g may provide ergogenic benefit. Performance potentiation is greater in the first seconds of repeated sprint cycling (even for the low creatine dosage) and progressively diminishes towards the end of a 30s period. This ergogenic pattern could be attributed primary to an elevated pre exercise creatine phosphate concentration and to a lesser extent to a more efficient buffering capacity. Additional research however, should evaluate the use of varying creatine dosages, in relation to body mass, as studies show that sample group specificity seems to affect the magnitude of performance potentiation.

The authors would like to thank the State Scholarship Foundation of Greece for the financial support of the present study. Our gratitude is extended to R.F.G.J. King, Research Fellow in Leeds General Infirmary, U.K. and R. Butterly, Senior Lecturer in Leeds Metropolitan University, for their advice and guidance throughout the period of this study. Many thanks also to all the staff in the Department of Chemical Pathology (Leeds General Infirmary) for their precious help in the blood and urine sample collection.