Original Research
Nutritional and Plasmatic Antioxidant Vitamins Status of Ultra Endurance Athletes Guillaume Machefer, PhD, Carole Groussard, PhD, Hassane Zouhal, PhD, Sophie Vincent, PhD, Hala Youssef, Henri Faure, PhD, Ludivine Malarde´, Arlette Gratas-Delamarche, PhD Laboratoire de Physiologie et de Biome´canique de l’Exercice Musculaire, UFRAPS Rennes 2 (G.M., C.G., H.Z., S.V., H.Y., L.M., A.G.-D.), G.I.S. Sciences du Mouvement, Universite´ de Rennes 2 (G.M., C.G., H.Z., S.V., H.Y., A.G.-D.), Rennes Cedex, De´partement de Biologie Inte´gre´e, CHUG (H.F.), Grenoble, FRANCE Key words: antioxidant vitamins, diet nutrition, antioxidant status, vitamin E, endurance athletes. Objective: The “Marathon des Sables” (MDS) is a competition known to induce oxidative stress. Antioxidant vitamins prevent exercise-induced oxidative damages. The purpose of this study was to evaluate daily intake and plasma level of the main antioxidant vitamins (␣-tocopherol, vitamin C, -carotene and retinol) in 19 male athletes who participated in this competition. Methods: Data collected before the beginning of the competition included daily dietary intake using a 7-day food record and plasma biochemical measurements (␣-tocopherol, vitamin C, -carotene and retinol). Results: First, total energy intake was obviously lower than the energetic intake usually observed in well-trained endurance athletes. Second, antioxidant vitamins intake was also insufficient. Indeed, the intake was lower than the French Dietary Reference Intakes (DRI) for this population in 18 subjects for vitamin E and 6 subjects for vitamin C, -carotene and Retinol Equivalent. As a significant relationship was found between total energy intake and the intake of vitamin E (r ⫽ 0.73; p ⬍ 0.001) and vitamin C (r ⫽ 0.78; p ⬍ 0.001), the low total energy intake contributed partially to the insufficient antioxidant vitamins intake. The dietary questionnaire analysis also revealed a low intake of vegetable oils, fruits and vegetables. However, plasma concentrations of these antioxidant vitamins were similar to the literature data observed in athletes. Conclusion: This study evidenced obvious insufficient energy intake in ultra endurance athletes associated with a low antioxidant vitamin intake.
INTRODUCTION
competition. In this sense, the French DRI have been recently revised in sportsmen. In a previous study, we observed that an extreme competition, untitled the “Marathon des Sables” (MDS) induced an oxidative stress and altered plasma antioxidant vitamins concentrations [3]. This competition is a selfdietary race organized each year in April under high temperature conditions in the Moroccan desert. It consists of 6 exhaustive races (mean total distance ⬇ 230 km) during 7 days. Even if the beneficial effects of antioxidant vitamins supplementation on physical performances remain controversial [4 –5], insufficient intake and low plasma levels of these vitamins are recognized to increase oxidative damages and to limit physical performances in animal models [6 –7]. The objective of the study was to determine the dietary status and more specifically the antioxidant vitamin intake of athletes who voluntary participated to the MDS during the
Public health organizations have established the Dietary Reference Intakes (DRI) in order to inform the population of the physiologic requirements in micro- and macronutrients. Although DRI are considered to be enough to prevent deficiency signs in the non-trained population, some other concerns must be taken into account in well-trained athletes in order to cover the specific needs induced by training. For example, there is growing evidence that acute exhaustive exercise and training increase the need for antioxidant vitamins. Indeed, these vitamins (Vitamin E, vitamin C, -carotene and retinol) neutralize several ROS [1–2] and also play a major role in exercise-induced oxidative stress. Thus, the daily antioxidant vitamins intake of trained athletes must be sufficient to cover the additional needs induced by training or by a
Address reprint requests to: Guillaume Machefer, PhD, Laboratoire de Physiologie et de Biome´canique de l’Exercice Musculaire, UFRAPS Rennes 2, EA 1274, Avenue Charles Tillon, Campus la Harpe, CS 24414, 35044 Rennes Cedex, FRANCE. E-mail:
[email protected]
Journal of the American College of Nutrition, Vol. 26, No. 4, 311–316 (2007) Published by the American College of Nutrition 311
Antioxidant Vitamins in Sportsmen specific pre-competition period. The antioxidant vitamins intake was compared to the French DRI for sportsmen and linked with plasma antioxidant vitamins concentrations.
MATERIALS AND METHODS Subjects Data were collected on nineteen healthy male long-distance runners. These athletes voluntarily participated in the 2001 and 2002 edition of the competition named “Marathon des Sables” (MDS). All subjects were non-smokers and did not take any supplements including vitamins and medications. They gave their informed consent prior to the beginning of the study. The protocol of the study was reviewed and approved by the local ethic committee for human subjects.
Protocol and Blood Samples Collection Blood samples were collected at rest, 3 weeks prior to the competition, between 0900 h and 1000 h from an antecubital vein. An heparinized vacutainer was used to determine the plasma concentration of ␣-tocopherol, Vitamin C, -carotene and retinol. The blood samples were centrifuged (1500 ⫻ g, 5 min, 4°C) and 1 mL for plasma ␣-tocopherol, -carotene and retinol measurement was stored at ⫺80°C. For vitamin C determination, an aliquot (500 L) of plasma was added to 4.5 mL of MPA (5% w/v) prior to storage at ⫺80°C. The anthropometric characteristics of the athletes were recorded. Body fat percentage was estimated from 4 skinfold thicknesses [8]. Maximal oxygen uptake (VO2max) was measured using a graded treadmill test performed until exhaustion [9]. An electrocardiogram was recorded throughout the test and VO2 was measured using a breath-by-breath automated exercise metabolic system (CPX, Medical Graphics, St. Paul, Minn., USA).
Biochemical Analysis Alpha-tocopherol, retinol and -carotene concentrations in plasma were measured using a reversed high performance liquid chromatography (HPLC) procedure [10]. Plasma vitamin C concentration was determined by HPLC [11].
utilized and that 1 mg of food -carotene is equivalent to 0.167 mg of retinol. The adequacy of nutrient intake of energy and each vitamin was assessed by comparing it with the French 2001 DRI for sportsmen [13]. This approach takes into account that vitamin needs for athletes are higher than for non-trained subjects. According to the French DRI, non-trained subjects have to consume 12 mg 䡠 day⫺1 in ␣-tocopherol, 110 mg 䡠 day⫺1 in vitamin C, 2.4 mg 䡠 day⫺1 in -carotene and 800 g 䡠 day⫺1 in RE. These intakes have to be increased in athletes respectively of 12 mg 䡠 day⫺1 for ␣-tocopherol, 100 mg 䡠 day⫺1 for vitamin C, 1 mg 䡠 day⫺1 for -carotene and 200 g 䡠 day⫺1 for RE per 4180 kJ (1000 kcal) of total energy intake above 9196 kJ (2200 kcal) [13].
Statistics The results are presented in mean ⫾ SEM. A student t test was used to compare the dietary intake of the athletes with the French DRI for sportsmen [13]. A Pearson test was used to measure the relationship between two parameters. The limit of significance was set at p ⬍ 0.05.
RESULTS Subject Characteristics Anthropometric characteristics and training status of all the athletes are summarized in Table 1. As indicated by their training habits and by their laboratory performances (VO2max ⫽ 59.8 ⫾ 1.4 mL.kg⫺1.min⫺1) recorded before the beginning of the MDS, all the participants were well-trained endurance athletes. The subjects carried out the distance of the competition at mean speed 7.9 ⫾ 0.4 km.h⫺1 i.e 44.3 ⫾ 1.6% of VMA.
Energy Intake and Macronutrient Distribution Total energy intake and the contribution of proteins, lipids and carbohydrates are presented in Table 2. There are no Table 1. Physiologic Characteristics, Training Status and Performances of the Athletes Athletes (n ⫽ 19)
Dietary Intake A quantitative assessment of dietary intake was provided by means of a 7-day food record. The food records were analyzed using a computer dietary analysis (Profil V.6, C.I.A.M., SaintChouldard, France) employing the Ciqual table of food composition. This analysis takes into account the loss of the vitamins induced by the cooking preparation but not those induced by the transport, the storage and the reheating of the food. The total retinol value of the diet was evaluated by the Retinol Equivalent (RE) measurement that was calculated according to Biery and Kanna [12] by assuming that dietary retinol is 100%
312
Age (years) Height (cm) Weight (kg) Fat (%) VO2max (mL.min⫺1.kg⫺1) Training history (years) Weekly exercise sessions Weekly distance of training (km) Best marathon time (min) Performances during the MDS Mean speed (km.h⫺1) Mean speed (% VMA)
41.4 ⫾ 1.8 173.6 ⫾ 1.5 67.3 ⫾ 1.9 17.2 ⫾ 0.8 59.8 ⫾ 1.4 11.5 ⫾ 1.5 5.6 ⫾ 0.4 87.0 ⫾ 6.7 180.7 ⫾ 5.4 7.9 ⫾ 0.4 44.3 ⫾ 1.6
VOL. 26, NO. 4
Antioxidant Vitamins in Sportsmen Table 2. Daily Energy and Macro-Nutrients Intakes
Total energy intakes: MJ.day⫺1 kJ.kg⫺1.day⫺1 kcal.kg⫺1.day⫺1 Carbohydrates (%) Lipids (%) Proteins (%) 1
Subjects (n ⫽ 19)
DRI1
9561.4 ⫾ 578.2 145.4 ⫾ 10.6 34.8 ⫾ 2.5 53.7 ⫾ 1.3 27.9 ⫾ 1.4 18.5 ⫾ 0.6
55–70 20–30 12–16
Dietary Reference Intakes for endurance athletes [13].
specific DRI for mean energy intake in trained athletes. However, daily energy intake was significantly lower than the DRI for the non-trained French population. Expressed as a relative value of energy intake (after exclusion of alcoholic calories), and compared to the usual recommendations for endurance athletes, the contribution of carbohydrates to energy was too low while the contribution of protein was slightly too high. The contribution of lipids was within the range of the recommendations [13].
Antioxidant Vitamins Intake Figs. 1 and 2 present the distribution of the athletes in comparison to the calculated DRI. The results indicated that
Fig. 2. Daily -carotene and Retinol Equivalent intakes. The circles indicate distribution of the athletes in comparison to their respective DRI.
⬇32% of marathon runners had lower daily vitamin C, -carotene and RE intakes than the recommendations. Concerning ␣-tocopherol, ⬇95% of the athletes questioned consumed less than 100% of the daily requirement.
Plasma Antioxidant Vitamin Concentrations Biochemical measurements of plasma ␣-tocopherol, retinol, -carotene and vitamin C are indicated in Table 3. There is nowadays no real norm in France for the mean plasma antioxidant vitamin concentrations in sportsmen. Table 3 gives the limit threshold [13] below which there is a real risk of vitamin deficiency and disease and some mean values found in the literature on sportsmen. The plasma antioxidant vitamin concentrations were quit similar to those previously published.
Relationships
Fig. 1. Daily vitamin E and vitamin C intakes. The circles indicate distribution of the athletes in comparison to their respective DRI.
JOURNAL OF THE AMERICAN COLLEGE OF NUTRITION
The daily ␣-tocopherol and vitamin C intakes are linked with the total energy intake (Fig. 3). Moreover, the intake of vitamin E and vitamin C are linked with total lipid intake and total carbohydrate intake respectively (Fig. 3). No relationships between vitamin E and C intakes and their respective plasma concentrations are observed. The daily intake of -carotene was significantly related to its plasma concentration (Fig. 4). Plasma retinol concentrations are significantly linked with the
313
Antioxidant Vitamins in Sportsmen Table 3. Plasma Antioxidant Vitamins Concentrations
␣-tocopherol (mol.L⫺1) Retinol (mol.L⫺1) -carotene (mol.L⫺1) Vitamin C (mol.L⫺1) 1 2
Athletes (n ⫽ 19)
Martin [13] (minimum value)1
Rokitzki [4]2
Liu [20]2
30.57 ⫾ 1.67 2.18 ⫾ 0.12 0.91 ⫾ 0.14 65.27 ⫾ 3.84
14.0 0.7 -
23.1 ⫾ 6.3 1.76 ⫾ 0.31 0.80 ⫾ 0.33 42.6 ⫾ 9.6
17.5 ⫾ 1.9 2.13 ⫾ 0.16 0.53 ⫾ 0.04 86 ⫾ 10
This value is the limit threshold below which there is a real risk of vitamin deficiency and disease. Example of values published in the literature on marathon runners.
Fig. 3. Relationships between total energy intake, total lipid and carbohydrate intake and vitamin E and C intakes.
RE intake (Fig. 4). There are no statically significant relations between performances (expressed as mean speed or % of VMA) during the MDS competition and intakes or plasma concentrations of antioxidant vitamins.
DISCUSSION This study compared the nutritional intakes and plasma concentrations of antioxidant vitamins (vitamin E, vitamin C, retinol and -carotene) in nineteen male marathon runners prior to the 2001 and 2002 edition of the MDS. Almost all of the marathon runners had low intake of vitamin E. Thirty two percent of the marathon runners consumed less than the recommended amounts of vitamin C, retinol and -carotene. However, no significant plasma deficiencies were observed for all these antioxidant vitamins. Finally, the total energy intake of this population was obviously insufficient.
314
Antioxidant Vitamin Intake A few studies [14 –19] have investigated the antioxidant vitamin intake in sportsmen. These studies concerned different populations of young athletes. In these studies, the mean vitamin C and RE intakes were between 95 to 203 mg 䡠 day⫺1 and 1.0 to 1.7 mg 䡠 day⫺1 respectively. The daily vitamin E intake ranged between 3.6 to 11 mg 䡠 day⫺1, a value lower than DRI for non-trained subjects (12 mg 䡠 day⫺1). Thus, the results of the current study well agreed with previous data indicating that moderately or well-trained subjects consumed insufficient antioxidant vitamin intake. In this study, the analysis is based on individual values, considering that the theoretical French DRI correspond to the minimal quantity necessary to obtain a sufficient protection for each subject. By this approach, it appears that 32% of the marathon runners had insufficient vitamin C as well as -carotene and RE intakes and 63 % are really at risk of vitamin E deficiency since consuming less than 50 % of the French DRI. The mean
VOL. 26, NO. 4
Antioxidant Vitamins in Sportsmen mean total energy intake. As a consequence, probably more than 32 % of the athletes had insufficient consumption of vitamin C, -carotene and RE, and we could hypothesize that the deficiency in vitamin E intake was also much more important. Unsurprisingly, the deficiency in vitamin E was linked to the concomitant insufficient fat intake as shown in Fig. 3. We noticed that the consumption of fats, rich in vitamin E, was very low: butter, vegetable oil and oleaginous plants. The relationship between vitamin C intake and total carbohydrate intake could be attributed to fruit consumption. Fruits are rich in fructose and many of them contain vitamin C. However, as vitamin C, -carotene and retinol are essentially present in various fruits but also in vegetables that bring few calories, the deficiency in these vitamins was less pronounced. In fact, even if fresh fruits and vegetables are less available at the end of the winter, we noticed that the athletes obviously made a real effort to diversify their diet as advised by the nutritional rules.
Plasma Antioxidant Vitamin Status
Fig. 4. Relationships between intake and plasma concentrations of antioxidant vitamins.
vitamin E intake was lower than the recommended intakes: only one of the athletes consumed enough vitamin E. The present study also indicated that the limited vitamin E and C intakes were linked with the limited mean total energy intake. Here, total energy intake was even lower than in previous studies [16 –17, 19]. Even if the 7-day food record may underestimate the real total energy intake by voluntary or involuntary omission of some nutrients by the athletes or by underestimation of the quantity of food ingested, the very low values observed in the present study could not be only explained by this parameter. Indeed, the runners were very compliant and particularly motivated by this investigation. In fact, similar results have been already found in other sports activities (skating, ski jumping) where sportsmen feared any body weight increase at risk of compromising their performances [14 –15]. The inverse relation between % of fat and the performances during the MDS competition expressed as mean speed (r ⫽ ⫺0,70, p ⬍ 0.05) observed in our study seemed to support this hypothesis. So, the athletes involved in the MDS competition probably restricted their food intake to prevent any body weight increase. Consequently, not only carbohydrates intake was insufficient, but also that of fats, even if their relative contribution to energy was correct. Thus, the vitamin DRI used for this specific population were obviously underestimated since the DRI for each antioxidant vitamin are calculated taking into account the
JOURNAL OF THE AMERICAN COLLEGE OF NUTRITION
Despite the obviously insufficient intake in antioxidant vitamins, corresponding plasma concentrations of vitamin E and -carotene were near the data previously published in marathon runners [4,20]. Plasma concentrations of vitamin C were also quite similar than values published by [4] and [20]. These findings strongly suggest that the runners had no plasma deficiency in the current antioxidant vitamins. The insufficient intake of antioxidant vitamins had no significant effect on plasma antioxidant vitamin concentrations. The possibility for dietary intakes to be linked with plasma concentrations is still controversial [16,21–22]. In the present study, only two low relations were observed and concerned -carotene and RE intakes and their respective plasma concentrations (Fig. 4). In fact, plasma vitamin concentrations reflect the balance between many factors such as intake, transport and storage and did not really reflect vitamin intake. So, our data suggested that more specific markers were needed to determine status intakes, e.g. isolated lipoproteins, white cells, specific binding proteins. Greater specificity would enhance sensitivity, and then allow determination of changes that might occur following the competition.
CONCLUSION The present study evidenced significant nutritional mistakes in runners who restricted their total energy intake as to limit any body weight increase. Indeed, 32 % of the 2001–2002 MDS runners had lower vitamin C, -carotene and RE intakes than their theoretical respective DRI before the beginning of the competition. Except one, all the athletes also consumed insufficiently vitamin E due to a too low fat intake. However, that does not seem to affect the plasmatic concentrations nor the performances during the MDS competition.
315
Antioxidant Vitamins in Sportsmen
ACKNOWLEDGMENTS The authors thank Dominique Paul, Marie-The´re`se Gougeon, Martine Godard and Michelle Jester for their technical assistance.
REFERENCES 1. Powers SK, DeRuisseau KC, Quindry J, Hamilton: Dietary antioxidants and exercise. J Sports Sci 22:81–94, 2004. 2. Yu BP: Cellular defences against damage from reactive oxygen species. Physiol Rev 74:139–162, 1994. 3. Machefer G, Groussard C, Rannou-Bekono F, Zouhal H, Faure H, Vincent S, Cillard J, Gratas-Delamarche A: Extreme running competition decreases blood antioxidant defense capacity. J Am Coll Nutr 23:358–364, 2004. 4. Rokitzki L, Logemann E, Sagredos AN, Murphy M, Wetzel-Roth W, Keul. J: Lipid peroxidation and antioxidant vitamins under extreme endurance stress. Acta Physiol Scand 151:149–158, 1994. 5. Sacheck JM, Roudenoff R: Nutrition in the exercising elderly. Clin Sports Med. 18:656– 583, 1999. 6. Davies KJA, Quintanilha AT, Brooks GA, Packer L: Free radicals and tissue damage produced by exercise. Biochem Biophys Res Comm 107:1198–1205, 1982. 7. Gohil K, Packer L, De Mumen B, Brooks GA, Terblanche SE: Vitamin E deficiency and vitamin C supplements: exercise and mitochondrial oxidation. J Appl Physiol 60:1986–1991, 1986. 8. Durnin J, Rahaman M: The assessment of the amount of fat in human body from measurements of skinfold thickness. Brit J Nutr 21:681–689, 1967. 9. Flandrois R, Grandmontagne M, Mayet MH, Favier R, Frutoso G: La consommation maximale d’oxyge`ne chez le jeune franc¸ais, sa variation avec l’aˆge, le sexe et l’entraıˆnement. J. Physiol (Paris) 78:186–194, 1982. 10. Steghens J-P, Van Kappel AL, Riboli E, Collombel C: Simultaneous measurement of seven carotenoids, retinal and ␣-tocopherol in serum by high-performance liquid chromatography. J Chromatrogr 694:71–81, 1997. 11. Speek SJ, Schriver J, Schreurs WHP: Fluorometric determination of total vitamin C in whole blood by high-performance liquid
316
12.
13. 14.
15.
16.
17.
18.
19.
20.
21.
22.
chromatography with precolumn derivatization. J Chromatrogr 13: 53–60, 1984. Biery JG, Kanna MC: Expressing dietary values for fat-soluble vitamins : changes in concept and terminology. Am J Clin Nutr 34:289–295, 1981. Martin A: Apports nutritionnels conseille´s pour la population franc¸aise. (3e`me Ed). Paris, Ed Tec & Doc 608p. 2001. Rankinen T, Lyytikainen S, Vanninen E, Penttila I, Rauramaa R, Uusitupa M: Nutritional status of the Finnish elite ski jumpers. Med Sci Sports Exerc 30:1592–1597, 1998. Ziegler PJ, Nelson JA, Jonnalagadda SS: Nutritional and physiological status of U.S. national figure skaters. Int J Sport Nutr 9:345–60, 1999. Guilland JC, Penaranda T, Gallet C, Boggio V, Fuchs F, Klepping J: Vitamin status of young athletes including the effects of supplementation. Med Sci Sports Exerc 21:441–9, 1989. Margaritis I, Palazzetti S, Rousseau A-S, Richard M-J, Favier A: Antioxidant supplementation and tapering exercise improve exercise-induced antioxidant response. J Am Coll Nutr 22:147–156, 2003. Palazzeti S, Rousseau A-S, Richard M-J, Favier A, Margaritis I: Antioxidant supplementnation preserves antioxidant response in physical training and low antioxidant intake. Brit. J. Nutr 91:91– 100, 2004. Groussard C, Machefer G, Rannou-Bekono F, Gratas-Delamarche A: Evaluation de l’apport en vitamines antioxydantes chez des sportifs. Sci Sport 18:108–110, 2003. Liu ML, Bergholm R, Ma¨kimattila S, Lahdenpera¨ S, Valkonene M, Hilden H, Yki-Ja¨rvinen H, Taskinen M-R: A marathon run increases the susceptibility of LDL to oxidation in vitro and modifies plasma antioxidants. Am J Physiol 276:E1083–E1091, 1999. Hercberg S, Preziosi P, Galan P, Devanlay M, Keller H, Bourgeois C, Potier de Courcy G, Cherouvrier F: Vitamin status of a healthy French population: dietary intakes and biochemical markers. Int J Vit Nutr Res 64:220–232, 1994. Stryker WS, Kaplan LA, Stein EA, Stampfer MJ, Sober A, Willett WC: The relation of diet, cigarette smoking, and alcohol consumption to plasma beta-carotene and alpha-tocopherol levels. Am J Epidemiol. 127:283–96, 1988.
Received October 14, 2004; Accepted July 21, 2006
VOL. 26, NO. 4
