where V̇O2(t) is oxygen uptake at time t, V̇O2(B) is baseline oxygen uptake (average V̇O2 during the minute before the onset of exercise), V̇O2 (EE) is end of exercise oxygen uptake (average V̇O2 during the last minute of exercise), G is the primary gain (i.e., the calculated change in oxygen uptake), TD is the on-kinetic time delay, and τ is the time constant (prime mark designates off-kinetics parameters). Mean response times for the on-kinetic responses (MRTon) and off-kinetic responses (MRToff) were calculated as TD + τ and TD′: + τ′:, respectively.
Blood sampling and analysis.
Venous blood was collected in a 5-mL container (Becton-Dickinson Ltd, UK) containing the anticoagulant ethylenediaminetetra-acid (EDTA). Several small aliquots were removed for the triplicate determination of blood lactate and glucose concentration (YSI 2300 Stat, Yellow Springs Instruments, U.S.), blood hemoglobin (Hb) concentration (Hemocue Ltd, UK), hematocrit (Hct) (Micro Hct MK IV, Hawksley, England) and changes in plasma volume as previously described (9). The remaining blood was centrifuged at 3000 × g for 15 min to obtain plasma, which was subsequently dispensed and frozen at −70°C. Two additional 7-mL blood samples were collected in serum separation tubes (Becton-Dickinson Ltd, UK), left to stand for 15 min, then centrifuged at 3000 × g for 15 min to obtain serum. The serum was transferred to appropriate containers and subsequently frozen at −70°C. Serum cortisol concentrations were determined using an automated time-resolved fluoroimmunoassay (AutoDELFIA™ Cortisol kit, Perkin Elmer, Life Sciences, UK).
Perceived feeling states.
Subjects were instructed to respond to the EFI as described in detail by Gauvin and Rejeski (11). Briefly, the subjects rated their feelings using the 12-item adjective scale on an analog scale from 0 (do not feel) to 5 (feel very strongly). The appropriate adjectives were averaged to obtain four perceived feeling states (positive engagement, revitalisation, tranquillity, and physical exhaustion) at each time point as previously described (11). The EFI was specifically developed to assess distinct feeling states that occur during stages of exercise and psychometric studies have indicated concurrent and discriminant validity (11). Moreover, the EFI has been demonstrated to be sensitive to interventions involving recreational active individuals (15).
Statistical analysis was carried out using SPSS software (version 12.0; SPSS Inc., IL). Group data were expressed as mean ± SEM and statistical significance was set at the P < 0.05 level. Subject characteristics were compared under supplementation groups using independent samples t-tests (Table 1). The exercise times to exhaustion during familiarisation, T1 and T2, were assessed using mixed-model repeated measures ANOVA (within-subject factors: trials; between-subject factor: supplementation groups) followed by simple main effect analysis. The remaining data, which contained multiple time points during each trial, were analyzed using mixed-model repeated measures ANOVA (within-subject factors: trial × time of sample; between-subject factor: supplementation groups). Mauchly's test was consulted and Greenhouse–Geisser correction was applied if the assumption of sphericity was violated. If a significant P value was identified for the three-way interaction effect (supplementation group × trial × time of sample), supplementation was deemed to have a significant effect. If a significant P value was identified for the main effect of time (time of sample), multiple pairwise comparisons were made using Bonferonni confidence interval adjustment, with statistical significance set at P < 0.01.
Exercise times to exhaustion at 85% V̇O2max were similar between groups during familiarization and T1 (PS, P: 8:02 ± 1:37, 7:46 ± 0:41; 7:51 ± 1:36, 8:09 ± 0:54 min:s). A significant interaction effect (supplementation group × trial, P = 0.007) indicated that supplementation had a significant effect on time to exhaustion; post hoc analysis revealed no differences between trials for P (P = 0.670) and significant differences between trials for PS (P = 0.001). The magnitude of change in exercise times to exhaustion (individual T2 values minus T1 values) in PS were 2:00 ± 0:28 min:s, whereas P remained similar (0:07 ± 0:13 min:s) (Fig. 2).
Supplementation did not significant effect last minute heart rates during the protocol (supplementation group × trial × time of sample, P = 0.058). Mean last-minute heart rates increased throughout the stages of the protocol (Table 2), being 119 ± 2, 137 ± 3, 156 ± 3, and 183 ± 2 beats·min−1, respectively at 45, 55, 65, and 85% V̇O2max.
Last-minute oxygen uptake data are presented in Table 2. These data increased progressively with exercise intensity (47 ± 1, 58 ± 1, 70 ± 1, and 97 ± 2% V̇O2max; time of sample effect, P < 0.001) and the supplement × trial × time of sample interaction was not significant (P = 0.160). Steady state was confirmed in all stages at exercise intensities up to and including 65% V̇O2max. Mean carbohydrate oxidation, fat oxidation, and total energy expenditure were 1.40 ± 0.06, 1.95 ± 0.11, and 2.49 ± 0.14 g·min−1; 0.28 ± 0.02, 0.27 ± 0.02, and 0.27 ± 0.02 g·min−1; and 34.0 ± 0.8, 42.3 ± 1.0, and 51.1 ± 1.1 kJ·min−1, respectively at each exercise stage (Fig. 3). Supplementation had no effect on carbohydrate oxidation (supplementation group × trial × time of sample, P = 0.596), fat oxidation (supplementation group × trial × time of sample, P = 0.187), or calculated energy expenditure (supplementation group × trial × time of sample, P = 0.595).
The mean response times for the on-kinetic response (MRTon) were significantly higher at 85% V̇O2max than at 45–65% V̇O2max (time of sample effect, P = 0.019); however, the three-way interaction was not significant (P = 0.069) (Fig. 4). Supplementation had no effect on MRToff (supplementation group × trial × time of sample, P = 0.449), and exercise intensity had no significant effect (time of sample effect, P = 0.055) (Fig. 4).
Estimated plasma volume fell by 7–11% during the first two exercise stages and remained below preexercise values throughout exercise (Table 3); supplementation had no effect (supplementation group × trial × time of sample, P = 0.392), and no statistical differences were identified between T1 and T2 (trial effect: P = 0.491).
Blood lactate concentrations were significantly elevated from preexercise values from Post-65% (Table 4) and peaked at Post-85% (mean value for all trials was 7.70 ± 0.33 mmol·L−1). Blood glucose concentrations did not differ significantly from preexercise values at any time point (Table 4), and no differences were identified between groups or as a result of supplementation.
Serum cortisol concentrations were significantly elevated by exercise (time of sample effect, P < 0.001) from preexercise values of 378 ± 21 nmol·L−1 to postexercise values of 554 ± 32 nmol·L−1 (Fig. 5). Serum cortisol concentrations were not significantly affected by supplementation (supplementation group × trial × time of sample, P = 0.118).
Significant temporal changes were observed in all of the EFI subscales; subjects reported decreases in revitalisation, positive engagement and tranquility, and increases in physical exhaustion through exercise (Table 5). The three-way interactions were not significant in any of the subscales (Table 5).
The primary finding of this investigation was that oral supplementation with 750 mg·d−1 S-PtdSer for 10 d significantly affected exercise capacity in a group of recreationally active subjects during a staged intermittent cycling protocol. Furthermore, the enhancements in exercise capacity in PS ranged from 0:15 to 3:47 min:s, whereas the exercise times to exhaustion remained unchanged in P.
It was originally hypothesized that PtdSer would influence the primary oxygen uptake kinetic response and thereby increase exercise capacity. However, supplementation did not significantly affect MRTon. Furthermore, no differences were observed between trial or supplementation group in MRToff; therefore, the current data presents insufficient evidence to support any change in the primary oxygen uptake kinetic response following supplementation.
The causes of fatigue during cycling at 85% V̇O2max (within an intensity domain that has been previously classified as very heavy exercise (19)) have not been fully elucidated and may include central and peripheral components (14). Exercise during the final bout was associated with near maximal oxygen uptakes and heart rates in addition to relatively high blood lactate concentrations; therefore, it can be assumed that heavy demands were placed on both oxidative and nonoxidative phosphorylation. Metabolic acidosis has been implicated as a mechanism of peripheral fatigue either through direct effects on the contractile proteins or through inhibition of key regulatory enzymes such as phosphofructokinase (7); however, it may be more likely that other ionic imbalances contribute to fatigue in this exercise model (1). In vitro studies have demonstrated that low concentrations of PtdSer are effective in activating (Na+-K+)–dependent ATPase in mammalian kidney (25) and brain (28) preparations. Similarly, Ca2+-ATPase, an enzyme primarily responsible for Ca2+ re-uptake from the muscle cyctosol into the sarcoplasmic reticulum, is known to require PtdSer (18,24). Therefore, it is plausible that exogenous S-PtdSer delayed the onset of fatigue by maintaining ionic homeostasis for longer during exercise.
In addition, Tibbits et al. (27) reported that extended exercise training increased the levels of phospholipids, especially PtdSer content, in rat cardiac sarcolemma. This adaptation to training might suggest that additional PtdSer within the heart muscle has functional benefits during exercise. An increase in membrane bound PtdSer may have the potential to enhance myocardial excitation–contraction coupling, potentially through the activation of different protein kinase C isoforms (26) and/or enhanced calcium uptake (20). Thus, it is plausible that these mechanisms also may have contributed to delaying fatigue in the present study. However, without corroborating data from further studies that investigate the in vivo pharmacological actions of S-PtdSer, the proposed mechanisms remain speculative.
The significant rise in serum cortisol concentration that followed the final bout of exercise suggested that the protocol activated the HPA axis (16). However, supplementation with PtdSer did not significantly influence serum cortisol concentrations (Fig. 5). This finding does not concur with the results of Fahey and Pearl (10), who found that S-PtdSer, using a similar supplementation regime, attenuated serum cortisol concentrations following resistance training. Furthermore, Monteleone et al. (17) reported that 800 mg·d−1 BC-PtdSer resulted in significant reductions in plasma cortisol and adrenocorticotrophic hormone (ACTH) concentrations during submaximal cycle exercise in untrained subjects.
The elevation in blood cortisol is a generic response to stress from both psychological and physical origin; consequently, there is considerable interindividual variability in response to exercise. Although the choice of experimental design in the current study investigated individual changes in response (pre- to postsupplementation) and, therefore, reduced the possible effect of subject selection–related bias, the possibility exists that the current dose may have been insufficient to attenuate the cortisol response in these active individuals. Alternatively, the current exercise protocol required that all participants continued the final exercise bout until exhaustion in both trials; therefore, it remains plausible that any effects of PtdSer supplementation on cortisol concentrations were masked as the PS group completed significantly more work in T2 when compared with T1.
Blood glucose concentrations remained unchanged throughout all trials. This finding was in agreement with previous studies using similar exercise protocols (16,17). The concomitant effects of reduced insulin and elevations in ACTH, cortisol, and epinephrine are responsible for controlling blood glucose during exercise (3). Therefore, any effects that S-PtdSer supplementation may have had on blood ACTH and cortisol did not appear to have overchallenged blood glucose homeostasis during exercise. Furthermore, the calculated rates of carbohydrate, fat, and combined fuel oxidation during the steady-state stages of exercise were similar in all trials, suggesting that any change in the HPA axis induced by S-PtdSer supplementation did not affect substrate oxidation during moderate exercise stages.
All subscales of the EFI were sensitive to change during the exercise. However, the three-way interaction did not reach significance in any subscale; therefore, there was no evidence to suggest that feeling states differed following supplementation in either supplementation group. The participants in the current study provided baseline responses that were similar to those of other recreationally active populations before exercise training (15), indicating that the testing procedures did not induce large changes in feeling states before exercise in these subjects. Benton et al. (4) reported improvements in mood after mental stress within a subgroup of young healthy adults following chronic S-PtdSer supplementation. Nevertheless, these improvements were only identifiable in a subgroup of subjects who scored higher than the median for neuroticism; people who score highly on this dimension are known to display strong emotional reactions to stress (4). Consequently, the baseline emotional state of an individual might influence the efficacy of S-PtdSer in altering feeling states during exercise.
To our knowledge, this is the first study to identify the ergogenic properties of phosphatidylserine; therefore, we suggest that further studies are required to substantiate these findings and to investigate the potential uses of S-PtdSer in exercise and physical activity. In addition, future studies are warranted to investigate the mechanism by which S-PtdSer may act physiologically.
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Keywords:©2006The American College of Sports Medicine
PHOSPHOLIPIDS; SUPPLEMENTATION; ERGOGENIC AID; ENDURANCE; HUMANS