Do Triathletes Periodize Their Diet and Do Their Mineral Content, Body Composition and Aerobic Capacity Change during Training and Competition Periods?

athlon is a demanding endurance multisport, which may strongly affect the nutri- tional status of athletes. The aim of this study was to nd whether there are any differences in energy value and nutrient intake, body mass and body composition, aerobic performance and hair mineral status between training and competition periods and to assess whether there is a link between hair mineral content and physical capacity and nutrition. This observational study covered 20 triathletes aged 32 7 years. The results of our study indicated performance improvement during the competi- tion period (longer time to exhaustion (p= 0.025) and lower maximal oxygen uptake at the ventilatory threshold (%VO 2max_VT;p= 0.047)). However, no differences were recorded in nutrition and body composition between two training vs. competition periods. There was a signi cant depletion in hair iron content during the competition period (p= 0.010). Furthermore, there

indicated performance improvement during the competi- tion period (longer time to exhaustion (p= 0.025) and lower maximal oxygen uptake at the ventilatory threshold (%VO 2max_VT;p= 0.047)). However, no differences were recorded in nutrition and body composition between two training vs. competition periods. There was a signi cant depletion in hair iron content during the competition period (p= 0.010). Furthermore, there were signi cant relationships between hair calcium content and absolute maximal oxygen uptake and %VO 2max_VT during the training period. It is necessary to introduce nutritional education in the group of triathletes focused on exercise-oriented nutritional periodization following the requirements of the training and competition periods, thus preventing the risk of nutrient de ciencies. Keywords:triathlon; nutrition assessment; aerobic capacity; elemental hair analysis 1. Introduction The triathlon is a demanding endurance multisport, which is a combination of three consecutive disciplines, i.e., swimming, cycling and running, completed over a variety of distances. During a competition, there is only a brief transition period between these disciplines within a speci c zone [1]. The overall duration of the event may last from 20 min up to even 8–9 h, depending on the many variations in distance, which demands the contribution and interaction of different energy systems and necessitates athletes having well-developed anaerobic qualities to meet speci c requirements. In turn, during training, athletes focus on developing mainly aerobic capabilities, but also anaerobic power and capacity by high-intensity and speed work programmes [2]. The athletic success achieved in competitive sports is in uenced by many factors, both exogenous and endogenous. Some of them, i.e., the training regime, nutrition, physical and physiological indicators, experience and technique, mental state and motivation, are shaped in the preparatory process by athletes [1,3–5]. Combining three different disciplines into one competition formula requires athletes to take up greater training volume and loads than training individual disciplines separately. Endurance training, which leads to changes in metabolism, respiratory, circulatory and muscular systems, is one of the most Nutrients2023,15, 6.

formula requires athletes to take up greater training volume and loads than training individual disciplines separately. Endurance training, which leads to changes in metabolism, respiratory, circulatory and muscular systems, is one of the most Nutrients2023,15, 6.

Nutrients2023,15, 6 2 of 16 important parts of the training process for a triathlon. Adaptations of the organism to undertake long-term exercise allow for delaying the onset of fatigue while undertaking physical activity [1,3]. The ability to perform long-term efforts is determined to the greatest extent by the high aerobic capacity potential, the availability of energy substrates, enzyme activity, and the ef ciency of thermoregulation and the cardiorespiratory system. This predictor is the basis for assessing an athlete's tness level and is shaped by genetic factors, training regime, nutrition and body composition, among other factors [1,6]. Furthermore, it should be emphasised that intensive training, nutritional discrepancy and mental stress, cause muscle fatigue and damage, as well as oxidative stress, which shape the speci c nutritional needs of physically active people [1,6]. For competitive athletes undergoing strenuous training sessions and genetically pre- disposed to succeed in their sport, proper nutrition determines the ability to participate in high-impact sports training and may make the difference between victory or failure [7]. Proper diet and supplementation signi cantly support training-induced stimulation and exercise adaptation, as they support the process of generating energy during physical exer- tion and post-recovery ef ciency (including, i.e., energy substrates' resynthesis after the regeneration of damaged tissues) [7]. Although the aforementioned role of sports nutrition is widely known, the current state of knowledge about the nutrition of various groups of athletes shows the prevalence of qualitative nutritional mistakes and the limited scale of implementation of the recommendations of the Swiss nutrition pyramid. Results from the study conducted on a representative group of Polish athletes revealed the main mistakes concerning insuf cient frequency of consuming different groups of food products, a high prevalence of irregular meals and non-limiting intake of energy drinks and sweetened beverages [8]. Furthermore, another important aspect from the point of view of sports training is to achieve and maintain an adequate nutritional status. In this respect, the basic factor providing information about the nutritional status is body composition, and the analysis of the concentration of selected clinically relevant biochemical markers (e.g., in blood, saliva, urine and hair)

drinks and sweetened beverages [8]. Furthermore, another important aspect from the point of view of sports training is to achieve and maintain an adequate nutritional status. In this respect, the basic factor providing information about the nutritional status is body composition, and the analysis of the concentration of selected clinically relevant biochemical markers (e.g., in blood, saliva, urine and hair) should be indicated at the forefront. Monitoring the body composition indicators of athletes allows, inter alia, to assess the effectiveness of dietary and training interventions. On the other hand, manipulation of the body composition allows for support of physical performance, adaptation to the requirements of a given sport discipline and taking on high-intensity exercise loads [9–12]. Similarly, an important element is also the nutritional status assessment involving the analysis of selected nutrients' balance—especially over a longer perspective and time (which is possible, for example, by elemental hair analysis). Finally, it should be pointed out that the above-mentioned elements are interrelated and may determine the proper body homeostasis and training process of an athlete, al- though their joint analysis is rarely carried out. Therefore, the objective of this study was to assess differences in nutrition and nutritional status related to body composition and hair mineral content, as well as physical capacity between two periods of macrocycles (training and competition), which differ between each other based on focusing on devel- oping different qualities and capabilities in triathletes. To specify, body mass and body composition, nutritional value of a habitual diet, indices of aerobic capacity, as well as hair content of selected minerals were assed in both training and competition periods, to assess whether triathletes modify their diet in different athletic macrocycles and if this aspect may in uence the changes in nutrition status and aerobic capacity and if changes in nutritional status may indicate possible variations in nutritional requirements during different macrocycle periods. Subsequently, the regression analysis between hair mineral content vs. energy value and nutrient intake and vs. indices of aerobic capacity was per- formed within both evaluated periods to verify the impact of nutrition on nutritional status and the link

status and aerobic capacity and if changes in nutritional status may indicate possible variations in nutritional requirements during different macrocycle periods. Subsequently, the regression analysis between hair mineral content vs. energy value and nutrient intake and vs. indices of aerobic capacity was per- formed within both evaluated periods to verify the impact of nutrition on nutritional status and the link between nutritional status and aerobic capacity. It was hypothesized that there would be differences between two macrocycles and that a correlation between hair mineral content of athletes and nutrition, aerobic capacity and body composition will be vital.

Nutrients2023,15, 6 3 of 16 2. Materials and Methods 2.1. Study Group Initially, the study group consisted of 50 triathletes (Figure). Due to injuries or other health problems disallowing the athlete to train or to take part in both periods of the study phases, refusal to continue participation in the research, or the suspicion of the authors of this paper of the athlete's non-compliance with the recommendations that formed the basis for maintaining the appropriate conditions of the research conducted, the number of athletes participating in the research decreased. Finally, the study group consisted of 20 athletes (2 females and 18 males) practicing for triathlons at the moderate (but competitive) and professional levels. The average training experience of athletes was 8.5 4 years, and the average age was 32 7 years (Table). All athletes declared good health and voluntary willingness to participate in the research. The study protocol was reviewed and approved by the Bioethics Committee of the Poznan University of Medical Sciences, reference numbers 681/16 and 683/16 (10 November 2016). All study participants gave written informed consent. All procedures were carried out in accordance with the ethical standards of the Helsinki Declaration of 1975. Figure 1.Flow chart of the study design. Table 1.Characteristics of the tested group after enrolment in the studies. Variable Mean SD Min–Max Age [years] 32 7 20–40 Body mass [kg] 80.5 14.4 55.8–123.2 Body height [cm] 179 9 157–190 Training experience [years] 8.5 4.0 2–17 Weekly training length [h] 11.0 5.5 1–20

Nutrients2023,15, 6 4 of 16 2.2. Study Design This observational study was conducted during two triathlon-speci c training macro- cycles, i.e., the preparatory training period of February–March and the competition period of June–August. 2.3. Anthropometric and Body Composition Measurements The study group was informed that at least 24 h before the body composition analysis, they should refrain from strenuous exercise and alcohol consumption and limit their coffee intake. The participants were also asked to pay special attention to proper hydration. They were aware that inadequate hydration due to excessive uid loss or improper uid intake can result in unreliable results from body composition analysis by bioelectrical impedance analysis (BIA). Males were also asked to shave their facial hair due to the effect of body hair on the results of body composition analysis by plethysmography. Prior to body composition analysis, height and body mass were measured in duplicate each time the participants visited the laboratory using a calibrated scale with a stadiometer (WPT 60/150 OW, Radwag ® , Radom, Poland) in a fasted state to the nearest 0.1 kg and 0.1 cm, respectively. The total body water and hydration level were assessed by bioelectric impedance with Bodystat 1500 (Bodystat Inc., Douglas, UK) and via urine specific gravity measurement with URYXXON ® Relax (Macherey-Nagel, Düren, Germany); values < 1.020 indicated proper hydration. Only properly hydrated participants were approved for testing. During the bioimpedance analyses, the recommended measurement conditions were strictly followed [13]. Fat-free mass and fat mass were assessed by air displacement plethysmography (Bod Pod ® , Cosmed, Rome, Italy) as described previously [14,15]. The excellent repeatability and reliability of the applied parameters in the current study methods for body composition analysis were previously insightfully evaluated and are published elsewhere [ 2.4. Nutritional Assessment The assessment of the diet was made on the basis of the open-ended dietary recording method from the period of three consecutive days before each test round. Participants were trained in the dietary recording method by a dietitian. They were also provided with special food diary forms and a photo album of food products and dishes prepared by

Nutritional Assessment The assessment of the diet was made on the basis of the open-ended dietary recording method from the period of three consecutive days before each test round. Participants were trained in the dietary recording method by a dietitian. They were also provided with special food diary forms and a photo album of food products and dishes prepared by the National Food and Nutrition Institute in Warsaw [16]. At each testing visit, a face-to-face dietary interview with each participant was undertaken, to discuss any doubts regarding dietary recording diaries. The quantitative analysis of the composition of daily food rations was carried out using the Dietetyk 2011 software package (Jumar, Poland), which uses a database developed by the National Food and Nutrition Institute in Warsaw [17]. 2.5. Aerobic Capacity Assessment An incremental cycling test (ICT) was conducted to evaluate the aerobic tness and capacity of the study group during two periods of the training macrocycles. Before ICT, athletes were advised to consume a standard pre-workout meal at 2 h before performing the exercise test. Prior to the test, athletes were asked to apply adequate recovery and to resign from any training the day before the test. ICT was carried out on the Kettler X1 cycloergometer (Kettler, Ense-Parsit, Germany) using a Quark CPET ergospirometer (Cosmed, Rome, Italy) to assess aerobic fitness and capacity according to the procedure described previously [15]. Before the test, a mask was applied on the athlete's face and a belt, with electrodes for heart rate (HR) measurement was placed on the chest. The athlete's task was to maintain a constant cadence of~70 5 RPM during the exercise test. Participants started with a low load, i.e., 100 W for males and 75 W for females, which increased by 25 W every 1.5 min of exercise. The test continued until the subjective feeling of exhaustion of the athlete, i.e., refusal to undertake further physical exertion. During the test, selected respiratory and cardiovascular indices were analysed, i.e., minute oxygen uptake (VO2), minute ventilation (VE) and HR. Aerobic fitness and capacity was expressed by the maximal oxygen uptake (VO2max) and ventilatory

25 W every 1.5 min of exercise. The test continued until the subjective feeling of exhaustion of the athlete, i.e., refusal to undertake further physical exertion. During the test, selected respiratory and cardiovascular indices were analysed, i.e., minute oxygen uptake (VO2), minute ventilation (VE) and HR. Aerobic fitness and capacity was expressed by the maximal oxygen uptake (VO2max) and ventilatory threshold (VT) markers.

Nutrients2023,15, 6 5 of 16 From the collected data, time to exhaustion (T exh), VO2max, and VO2at VT (VO2VT), percentage of VO2maxat VT (%VO2max_VT), HR maximum (HRmax) and at VT (HRVT), as well as workload expressed in watts at VT (WVT) were determined. The excellent repeatability and reliability of the aerobic capacity methodology were previously evaluated, and the results are published elsewhere [15]. 2.6. Hair Sampling, Preparation and Analysis of the Mineral Content Hair samples were collected during each test round. Proximal parts (1–2 cm in length, total weight of 0.5 g) of occipital scalp hair strands were collected after cutting hair using ethanol-precleaned stainless steel scissors. Hair samples were stored in a laboratory at room temperature until analysis. The collected hair samples were washed with acetone and deionized water with subsequent drying on air to a stable weight. Hair samples were mineralized using a microwave digestion system (Speedwave Xpert, Berghof, Eningen, Germany) by digesting in 65% (w/w) spectra pure HNO3(Merck, Kenilworth, NJ, USA). The analysis of copper (Cu), iron (Fe), zinc (Zn), calcium (Ca) and magnesium (Mg) levels in the samples was performed using an AAS-3 spectrophotometer (Carl Zeiss, Ger- many). The accuracy of the assay was 94%, 94%, 102%, 95% and 91% for magnesium, iron, copper, zinc and calcium, respectively, as veri ed by certi ed reference materials (Human Hair NCS DC73347a, LGC). The results of hair analysis were expressed as g/g for all elements. 2.7. Statistical Analysis All variables were checked for normal distribution using the Shapiro–Wilk test. Results are presented as mean 1 standard deviation (SD) and 95% con dence interval (95% CI). Comparisons between two training periods were performed usingT-test for dependent samples (normal distribution of the data) or Wilcoxon signed-rank test (non-normal distri- bution of the data). The relationships between nutrition or physical capacity indices and the content of minerals in collected hair samples were analysed using Spearman's rank correlation (due to non-normal distribution of at least of one variable in all the analysis). Statistical signi cance was set atp< 0.05. Data were analysed using STATISTICA 13.3 (StatSoft Inc., Tulsa, OK, USA) software. 3.

of the data). The relationships between nutrition or physical capacity indices and the content of minerals in collected hair samples were analysed using Spearman's rank correlation (due to non-normal distribution of at least of one variable in all the analysis). Statistical signi cance was set atp< 0.05. Data were analysed using STATISTICA 13.3 (StatSoft Inc., Tulsa, OK, USA) software. 3. Results 3.1. Anthropometric and Body Composition Results Body mass slightly decreased from 80.5 14.4 kg in the training period to78.6 11.2 kg in the competition period, especially due to the insigni cant reduction of body fat (Table). Furthermore, other body composition indices were not different between training and competition periods. However, it should be mentioned that the average % of fat mass was above the reference values for triathletes (5–12%) [18]. The average water content in the athletes' bodies was within the generally accepted reference limits (45–75%) [19], both in the training and competition period, and amounted to 58.7 5.1 and 59.9 4.1%, respectively (Table). Table 2. Assessment of the anthropometric and body composition indices of athletes during the training and competition period. Training Period Competition Period Variable Mean SD 95% CI Mean SD 95% CI p Body mass [kg] 80.5 14.4 73.7–87.2 78.6 11.2 73.3–83.8 0.078 † Fat mass [kg] 15.1 9.0 10.9–19.4 12.9 5.4 10.2–15.6 0.157 § [%] 18.1 8.2 14.3–22.0 16.4 6.3 13.3–19.6 0.244 †

This study assesses differences in nutrition and performance between training and competition periods in triathletes.