Conjugated linoleic acids (CLAs) are naturally occurring positional and geometric isomers of linoleic acid, also known as octadecadienoic acid, found primarily in ruminant meats (i.e., lamb and beef) and dairy products (i.e., milk and cheese) (46). Conjugated linoleic acid contains 2 double bonds separated by a single bond in a cis, trans configuration. These bonds commonly occur between the 8th and 13th carbon positions, and the two most common isomers of CLA are trans-10, cis-12 and cis-9, trans-11 (17). Conjugated linoleic acid may have several health benefits, including improved insulin sensitivity (40,45,48), decreased insulin and glucose concentrations (11,32), and enhanced energy expenditure (27,32,59). Although a pseudo meta-analysis by Terpstra (55) concluded that CLA has no positive effects on insulin sensitivity, insulin, or glucose concentrations. Thus, conflicting evidence exists regarding the potential health-related benefits of CLA supplementation.
The effects of CLA on fat oxidation in humans have been even less clear (5,55). Several studies have demonstrated that CLA supplementation may lead to decreases in body weight or body fat (5,10,11,56), whereas others have reported no effects of CLA on body composition (26,32). It has been hypothesized that CLA may enhance β-oxidation, carnitine palmitol transferase activity (10,27,28,41,59), expression of uncoupling protein 2 (UCP-2), or expression of peroxisome proliferator-activated receptor δ (PPARδ) (28,29). Theoretically, because of these cellular effects in skeletal muscle, CLA may indirectly enhance endurance performance (11,28,29) through a glycogen sparing mechanism. For example, studies (28,29,37) have demonstrated that CLA supplementation increased fat utilization, reduced the consumption of liver glycogen and production of lactate, and improved the time to exhaustion during aerobic exercise in mice. However, the effects of CLA on endurance exercise performance in humans have not been thoroughly investigated (34).
We are aware of only 6 studies that have examined the effects of CLA on exercise performance in humans (11,18,31,32,42,59). Zambell et al. (59) and Lambert et al. (32) demonstrated no effects of CLA on substrate utilization during steady-state walking and submaximal cycling, respectively. Krieder et al. (29) reported that CLA did not augment muscle strength during 4 weeks of resistance training, whereas Pinkoski et al. (42) showed greater increases in bench press strength with CLA supplementation after 7 weeks of resistance training. Colakoglu et al. (11) reported that CLA provided no additional benefit to aerobic exercise alone on running velocity and 30 minutes running performance, whereas Ha and Jeong (18) indicated that CLA improved the number of sit-ups in 1 minute, standing long jump, and shuttle run performance in obese adolescent boys. Thus, again, conflicting evidence exists regarding the efficacy of CLA for enhancing exercise performance. Given the inconclusive evidence (11,31,32,42,59) and the proposed glycogen sparing mechanism (28,29) for CLA to impact endurance performance, there is a need to investigate the potential ergogenic effects of CLA using a reliable and sensitive exercise test, such as the physical working capacity at the fatigue threshold (PWCFT), that has been successfully used to test the efficacy of other dietary supplements (9,50–53). Therefore, the purpose of this study was to examine the effects of CLA supplementation in conjunction with 6 weeks of aerobic exercise training on the PWCFT, timed sit-ups, and the standing long jump.
Previous studies have demonstrated the ability of dietary supplements to improve exercise performance in humans by reducing glycogen utilization (i.e., the glycogen sparing effect), enhancing fat oxidation, and reducing lactate accumulation (25,49,58). Based on the aforementioned studies and the existing evidence on CLA (11,18,28,29,31,37,42), we hypothesized that CLA combined with aerobic exercise training would improve the PWCFT to a greater extent than aerobic exercise alone. We also hypothesized that CLA supplementation would improve the maximum number of sit-ups and the standing long jump based specifically on the results of Ha and Jeong (18).
Experimental Approach to the Problem
This study used a randomized, double-blind, placebo-controlled, parallel design. During visit 1, the subjects were familiarized with the testing procedures. One to 3 days after visit 1, the subjects returned for visit 2 to begin the pretraining assessments and performed a standing long jump and maximal sit-up test. Two to 4 days after the completion of visit 2, the subjects returned for visit 3 to perform an incremental test to exhaustion on a calibrated electronically braked cycle ergometer (Lode Corival, Groningen, the Netherlands) to determine the PWCFT. Two to 4 days after visit 3, the subjects were randomly assigned to either the supplement (CLA; n = 17) or placebo (PLA; n = 16) group and began 6 weeks of cycle ergometer training at a workload associated with 50% of V[Combining Dot Above]O2peak for 30 minutes, 2 times per week. Two to 4 days after the last exercise training session, subjects returned to the laboratory for visit 4 to begin posttraining assessments. At visit 4, the subjects completed an incremental test to exhaustion to determine the PWCFT (replication of visit 3). The subjects were instructed to consume the same diet leading up to visit 4 as was consumed before visit 3 and were asked to refrain from consuming caffeine on the days of PWCFT testing. Two to 4 days after visit 4, the subjects performed the standing long jump and maximal sit-up tests (replication of visit 2). The subjects continued to take their supplement (CLA or PLA) up to and including visit 5. Figure 1 shows the timeline for this study design.
Forty men volunteered to participate in this study, however, only the data from thirty-three men (mean ± SD; age = 21.6 ± 2.8 years; height = 179.5 ± 6.0 cm; mass = 77.5 ± 9.6 kg) were analyzed for this study. Five subjects did not complete the study for the following reasons: an adverse event unrelated to the study or study product (n = 1), enrollment in another clinical trial after completing the consent process in this study (n = 2), and unspecified reasons unrelated to the study or study product (n = 2). In addition, the investigators were unable to determine PWCFT values before or after the intervention for 2 subjects, so their data were excluded from analyses. All subjects were considered untrained to moderately trained and reported no more than 4 hours of aerobic exercise per week. All of the subjects had a body mass index below 30 kg·m−2 were free from musculoskeletal injuries or neuromuscular diseases and did not report any medical disorders or medicinal or supplement usage that could have effected the outcome of this study. Subjects were paid $200 to complete the study. This study was approved by the university's Institutional Review Board for the protection of human subjects, and all subjects completed a health history questionnaire and informed consent document before any testing.
Each subject consumed eight 1-ml capsules per day split into 2 doses: (a) 4 capsules with a morning meal and (b) 4 capsules with an evening meal. Supplementation began 2–4 days after visit 3 on the first day of training and continued throughout the end of the study. Each CLA capsule contained 1 ml of CLA oil (75% active CLA isomers). Each PLA capsule contained 1 ml of high oleic sunflower oil. The manufacturing and blinding of the CLA and PLA supplements were provided to the study site by the funding company, Stepan Specialty Products, LLC (Koog aan de Zaan, the Netherlands). Testing of the CLA capsules before study initiation confirmed that the capsules contained 1 ml of oil, which consisted of 78% CLA (Clarinol A-80): 74% cis-9, trans-11 and trans-10, cis-12 CLA isomers. Compliance was assessed by weighing each subject's container containing the CLA or PLA capsules before and after the supplementation period and expressing the difference as a percentage of the capsule weight that was intended to be consumed as follows:
The average supplementation compliance was (mean ± SD) 95.2 ± 9.0%. None of the subjects were below 80% compliance.
Determination of V[Combining Dot Above]O2peak
Each subject performed an incremental test to exhaustion on a calibrated electronically braked cycle ergometer at a pedal cadence of 70 revolutions per minute. Seat height was adjusted so that the subject's legs were at near-full extension during each pedal revolution. In addition, toe clips were used to ensure that each subject maintained pedal contact throughout the ride. All subjects wore a nose clip and breathed through a 2-way valve (2700; Hans Rudolph, Kansas City, MO, USA). Expired gas samples were collected and analyzed using a TrueMax 2400 metabolic cart that was calibrated before every test (Parvo Medics, Sandy, UT, USA). The test began at 50 W and the power output was increased by 30 W every 2 minutes until the subject reached voluntary exhaustion or could no longer maintain a pedal cadence of 70 revolutions per minute despite strong verbal encouragement. The V[Combining Dot Above]O2peak was defined as the highest relative V[Combining Dot Above]O2 value recorded during the test (ml·kg−1·min−1). The intraclass correlation coefficient for V[Combining Dot Above]O2peak testing from our laboratory is R = 0.95, with no significant mean difference between the test and retest values (p > 0.05).
Subjects performed 6 weeks of aerobic training on a mechanically braked bicycle ergometer (Monark, model 818) at the workload associated with 50% of the V[Combining Dot Above]O2peak workload for 30 minutes, twice per week separated by at least 48 hours (total training sessions = 12). The cadence was set at 70 revolutions per minute, and the seat height was adjusted and remained constant throughout the training period so that the subject's legs were at near-full extension during each pedal revolution. Subjects were considered compliant if they completed 10 or more of the intended training sessions. All subjects completed at least 10 training sessions, and the average session completed was (mean ± SD) 11.5 ± 0.7.
To record electromyographic (EMG) signals during the incremental tests, pregelled bipolar surface electrodes (AccuSensor Ag/AgCl Electrodes; Lynn Medical, Wixom, MI, USA) were placed on the vastus lateralis muscle of the right thigh with an interelectrode distance of 30 mm. The center of the bipolar electrode pair was placed at 66% of the distance between the anterior superior iliac spine and the lateral superior border of the patella (20). The electrodes were placed 5 cm lateral to the reference line so that they laid over the vastus lateralis muscle (35). A goniometer (Smith & Nephew Rolyan, Inc., Menomonee Falls, WI, USA) was used to orient the electrodes so that the longitudinal axis of the bipolar electrode pair was parallel to the angle of pennation of the vastus lateralis fibers (approximately 20°) (8,16). The reference electrode was placed over the right iliac crest. To increase the signal-to-noise ratio (2), local areas of the skin were shaved, abraded, and cleaned with isopropyl alcohol before the placement of the electrodes. The EMG signal was amplified (gain: ×1,000) using differential amplifiers (EMG100; Biopac Systems, Inc., Santa Barbara, CA, USA; bandwidth = 10–500 Hz), digitally bandpass filtered (zero-phase shift, fourth-order Butterworth) at 10–499 Hz, and the amplitude (microvolts root mean square, μVrms) values were calculated using custom written software (LabVIEW Version 12.0; National Instruments, Austin, TX, USA).
Determination of PWCFT
The PWCFT was determined using the EMG amplitude values (µVrms) recorded from the vastus lateralis muscle using the procedures adapted from deVries et al. (14). Specifically, during each 2 minutes stage of the incremental cycle ergometer test, six 10-second EMG samples were recorded, and the amplitude values (µVrms) from each 10-second epoch were plotted across time for each power output of the test. The PWCFT was defined as the average of the highest power output that resulted in a nonsignificant (p > 0.05; single-tailed t-test) slope coefficient for the EMG amplitude vs. time relationship, and the lowest power output that resulted in a significant (p ≤ 0.05) positive slope coefficient. For 6 subjects, a PWCFT could not be identified from the first pre- or postsupplementation incremental test. These subjects were retested 48–72 hours later. A PWCFT was determined on the second attempt for 4 subjects; however, for 2 subjects, a PWCFT was not identified. It has previously been reported (24) that a PWCFT cannot be determined in about 10% of subjects. The intraclass correlation coefficient for the PWCFT from our laboratory is R = 0.95, with no significant mean difference between the test and retest values (p > 0.05).
Maximum Number of Sit-ups
For local muscular endurance, the subjects performed a maximal sit-up test (23). Each subject lay on his back with his knees bent, feet flat on the floor, heels about 18 inches from the buttocks, and the fingers next to the ears. An investigator held each subject's feet firmly on the floor for stability. The subjects then touched each elbow to the opposite knee, alternately, and performed as many sit-ups as possible in one minute. The number of correctly performed sit-ups in 1 minute was recorded and used for analysis. Brotons-Gil et al. (6) demonstrated intraclass correlation coefficients for a flexion-rotation sit-up test of 0.87–0.93 with no significant difference between test and retest scores (p > 0.05) in 35 recreationally active, college-age men.
Standing Long Jump Performance
Standing long jumps were performed as a measure of muscular power (23). Subjects began by facing the starting line with their feet parallel and shoulder width apart. Each subject then dropped into a squat position with an approximate knee joint angle of 90° while thrusting their arms backwards before rapidly reversing direction, transitioning into a horizontal jump. The standing long jump score was then recorded as the distance (23) from the starting line to the most rearward heal after landing. A minimum of 3 and maximum of 5 attempts were performed with the best jump used for analysis. Markovic et al. (36) reported an intraclass correlation coefficient for the standing long jump of 0.95 with no significant difference between the test and retest scores (p > 0.05) in a sample of 93 healthy, college-age men.
Each subject completed a 3-day food log during the 3 days leading up to the pre- and posttraining PWCFT incremental tests to ensure that there were no differences between the CLA or PLA groups.
Three separate analyses of covariance were used to compare groups (CLA vs. PLA) for the PWCFT, standing long jump, and maximal sit-up data using the respective pretraining scores as the covariate and the adjusted posttest scores as the dependent variable. Multiple a priori planned comparisons were used to analyze the differences within conditions (CLA and PLA) from pre- to posttraining for the PWCFT, standing long jump, and maximal sit-up data using dependent samples t-tests. Independent samples t-tests were used to analyze total caloric (kilocalories) and macronutrient (grams of protein, carbohydrate, and fat) intake at pre- and posttraining. All statistical analyses were performed by a third party investigator who was not present for data collection (J.P.W.) using custom written software (LabVIEW Version 12.0; National Instruments, Austin, TX, USA). A type I error rate of 5% was considered statistically significant for all comparisons.
There were no significant (all p ≥ 0.23) differences between the CLA and PLA groups for the adjusted mean values for PWCFT, sit-ups, or standing long jump scores at posttraining (Figure 2). The planned comparisons indicated that PWCFT increased from pre- to posttraining in the CLA (p = 0.003) and PLA (p = 0.003) groups. Ten subjects increased by 12–39% from pre- to posttraining in the CLA group, whereas 7 subjects increased by 16–39% from pre- to posttraining in the PLA group. There were no differences (p > 0.05) from pre- to posttraining for sit-ups and standing long jump scores in either the CLA or PLA groups (Figure 2). There were no differences (p > 0.05) between groups for kilocalories, protein, carbohydrate, or fat (Figure 3).
This study was the first to examine the effects of CLA vs. PLA on the PWCFT. The PWCFT represents the highest power output during cycle ergometry that can be maintained for an extended period of time without neuromuscular evidence of fatigue (15) and has been sensitive to dietary supplementation (9,50–53). For example, increases in the PWCFT have been observed after creatine (50,52), beta-alanine (51), and arginine (9) supplementation, which have collectively suggested that these dietary supplements may beneficially prolong the onset of neuromuscular fatigue. The mechanisms of action for the supplement-induced increases in the PWCFT were increased muscle phosphocreatine stores (43,57), improved H+ buffering capacity from intramuscular carnosine (53), and increased coronary and peripheral blood flow (30,44), respectively. However, in this investigation, there was no effect of CLA on the training-induced increases in the PWCFT, nor were there any effects of CLA or aerobic training on the maximum number of sit-ups or standing long jump. Consequently, these findings suggested that 8 ml of CLA supplementation per day for 6 weeks does not have a beneficial effect on the improvements in neuromuscular fatigue after 6 weeks of aerobic training or on local muscle endurance or power.
Although many studies have examined the effects of CLA on body composition (4,5,27,31,56,59), energy expenditure (27,32,59), plasma lipoprotein concentrations (3,4,32,40,45,48), and insulin concentrations and sensitivity (11,32,40,45,48), little is known about the effects of CLA in conjunction with aerobic exercise training on performance (11,32,56,59). It has been hypothesized that CLA may enhance endurance performance by facilitating intramuscular fatty acid oxidation through increases in carnitine palmitol transferase turnover and β-oxidation (27,41,59), as well as the upregulation of UCP-2 and PPARδ (28,29). In turn, the CLA-induced increases in intramuscular fatty acid oxidation may reduce the reliance on anaerobic metabolism, limit the production of metabolic byproducts (i.e., lactate, H+, and ammonia), and limit decreases in intramuscular pH, which may initiate or contribute to the onset of fatigue (1,12,19,21,22,33,38,39,47). Consequently, through these mechanisms, CLA supplementation might be expected to delay the onset of neuromuscular fatigue (PWCFT) and improve endurance performance. However, Lambert et al. (32) and Zambell et al. (59) failed to show an effect of CLA and aerobic training on substrate utilization (i.e., respiratory exchange ratio) during submaximal exercise. Colakoglu et al. (11) demonstrated that 6 weeks of CLA supplementation and aerobic exercise training increased the running velocities and 30 minutes running performance in young healthy females to the same extent as aerobic training alone. In this investigation, the CLA and PLA groups demonstrated average increases of 16.4 and 16.1% in the PWCFT, respectively, with no difference between groups. The daily dosage of CLA used in this study was also greater (8 ml·d−1) than the dosages of CLA used previously by Lambert et al. (3.9 g·d−1) (31), Zambell et al. (3.0 g·d−1) (59), and Colakoglu et al. (3.6 g) (11). Furthermore, Bulut et al. (7) previously demonstrated that CLA supplementation and exercise may reduce low-density lipoproteins, very low-density lipoproteins, and triglycerides in 30 days, which was similar to the 4-week period used in this study. Therefore, the results of this study indicated that CLA supplementation had no beneficial effects on the aerobic training–induced increases in the PWCFT, which extended the findings of Colakoglu et al. (11), Lambert et al. (32), and Zambell et al. (59) to include the onset of neuromuscular fatigue, despite using a higher daily dosage of CLA than used previously (11,32,59).
It has been demonstrated that aerobic training improves the PWCFT. For example, deVries et al. (13) reported 29.8 and 38.4% improvements in the PWCFT in elderly men and women after 10 weeks of low- (30 min·d−1, 3 d·wk−1, 70% PWCFT) or high-intensity (30 min·d−1, 3 d·wk−1, 85% PWCFT) aerobic training, respectively. The authors (13) concluded that the PWCFT was a useful measure for evaluating fitness in the elderly. This study was the first, to the best of our knowledge, to report improvements in the PWCFT after only 12 aerobic training sessions in young healthy men. The 16% increase in the PWCFT in the current study was smaller than the 30–38% increase observed by deVries et al. (13), however, the subjects completed 40% of the training volume reported previously (13). In addition, the magnitude of increase in the PWCFT (16%; approximately 23 W) observed in this study was 3 times greater than the SEs of the PWCFT (±3.7 to 4.1%; ±7.1 to 7.9 W) reported in a similar sample (9). Therefore, our findings suggested that the PWCFT was sensitive to moderate-intensity (50% V[Combining Dot Above]O2peak), short-term (6 weeks) aerobic training performed twice per week. Future studies are needed to establish the minimum amount of aerobic exercise necessary to elicit neuromuscular adaptations manifested through the PWCFT.
In addition to reductions in body weight and body fat in obese adolescent boys, Ha and Jeong (18) showed that 12 weeks of CLA supplementation also increased sit-up and standing long jump performance. The authors (18) concluded that the beneficial effects of CLA supplementation on muscular endurance and power were a result of decreased body weight. However, caution should be used when interpreting the findings of Ha and Jeong (18) because the volunteers were 14–16 years old and experienced maturation during the study. For example, mean height increased from 168 to 173 cm in the CLA group during the supplementation period. It is possible that other aspects of growth and development, such as muscle mass and strength, in boys of this age may have influenced their findings (18). In contrast, the results of this study indicated that neither the CLA supplementation nor the aerobic training had any effect on sit-up or standing long jump performance. The disparity between the results of this study and those of Ha and Jeong (18) may have been due to the differences in subject demographics.
There was no effect of CLA on the training-induced increases in PWCFT, nor were there any effects of CLA or aerobic training on the maximum number of sit-ups or standing long jump. Consequently, CLA did not exhibit ergogenic benefits on this model of aerobic training–induced improvements in neuromuscular fatigue, or on field tests of muscle endurance and power. Future studies should examine the effects of CLA supplementation in conjunction with aerobic training on other performance-related variables, such as maximal aerobic capacity and the ventilatory threshold. Future studies may also wish to use the PWCFT as a method to evaluate neuromuscular adaptations to short-term or moderate-intensity aerobic exercise. Finally, moderate-intensity (50% V[Combining Dot Above]O2peak), short-term (6 weeks) aerobic training performed twice per week may be sufficient to improve aerobic fitness by delaying the onset of neuromuscular fatigue in untrained individuals.
Conjugated linoleic acid is an increasingly popular fat supplement often marketed as an ergogenic aid. Recent animal studies (28,29) have suggested that CLA enhances endurance capacity in animal models. In fact, based on their data demonstrating that CLA can enhance running capacity through enhanced fat oxidation and reductions in the consumption of stored glycogen during exercise, Kim et al. (28) suggested that CLA may have potential as a “novel ergogenic aid for future human applications.” In addition, it has been suggested that aerobic exercise training may augment the effects of CLA supplementation (32). However, no studies have examined the effects of concomitant CLA supplementation and exercise on aerobic exercise performance. Our data suggested that 8 ml of daily CLA supplementation did not exhibit ergogenic effects on the aerobic training–induced improvements in neuromuscular fatigue or field tests of muscle endurance and power. Therefore, although CLA may be marketed as a performance-enhancing supplement, allied health professionals, athletes, or recreationally active individuals may wish to avoid recommending or consuming CLA as an ergogenic aid for improving endurance performance in light of the current data. Future studies, however, should examine the effects of CLA supplementation in conjunction with aerobic training on other performance-related variables.
This study was funded by a research grant from Stepan Specialty Products, LLC. Stepan Specialty Products, LLC had no involvement in the data collection, analysis and interpretation of the data, writing of the article, or in the decision to submit the article for publication. The results of the present study do not constitute endorsement of the product by the authors or the National Strength and Conditioning Association.
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