Subjects

Nine high-level male athletes (age 21 ± 5.5), elite artistic gymnasts, were recruited for this study. Subjects competed in the Italian premier league for the CorpoLibero Gymnastics Team ASD, Padova, Italy and include two athletes belonging to the Italian national team. The mean volume of weekly training was about 30 hours. During the VLCKD period (30 days) the athletes were asked to keep to their normal training schedule. During a preliminary meeting it was explained that during the first three weeks it was necessary to almost totally exclude carbohydrates and a detailed menu containing permitted and non-permitted foods was provided to each participant, along with the components of the ketogenic diet with phytoextracts diet described below. All gymnasts read and signed an informed consent with the testing procedures approved by the council of the Human Anatomy and Physiology Department, University of Padova.

Experimental design

Subject measurements were taken, according to the methodology described below, before starting the VLCKD and repeated after thirty days of VLCKD. Since we chose a within subject design to strengthen the study (Subjects served as their own control), the athletes were re-tested during a second training period comparable in terms of intensity and volume of training to the first one.. The work load between athletes was similar because the team training regimes are strictly controlled, and recorded, due to the elite nature of their competition. The protocol took place three months later to ensure a comparable training load and achieve this goal the intensity and volume of training during the two periods (hours of training, kind of exercises, etc.) was carefully measured. During the second experimental session the subjects followed their normal diet (WD) instead of the VLCKD. The test procedure before and after WD was the same as the first testing session (Figure 1). As in the experimental period also during the control period the athletes were invited to continue their normal training. One subject withdrew from the study due to injury.

Figure 1 Scheme of the experimental protocol. Full size image

Diet

Before the start of the study, each athlete was given a detailed list containing the foods permitted and prohibited in a ketogenic diet. The diet consumed was primarily made of beef and veal, poultry, fish, raw and cooked green vegetables without restriction, cold cuts (dried beef, carpaccio and cured ham), eggs and seasoned cheese (e.g. parmesan). The drinks allowed were infusion tea, moka coffee and the herbal extracts. The foods and drinks that athletes avoided included alcohol, bread, pasta, rice, milk, yogurt, soluble tea and barley coffee. In addition to facilitate the adhesion to the nutritional regime, each athlete was given a variety of speciality meals constituted principally of protein and fiber These meals (TISANOREICA® by Gianluca Mech SpA, Asigliano Veneto, Vicenza, Italy) which are composed of high quality protein (equivalent to 18 grams/portion) and virtually zero carbohydrate (but that mimic their taste) were included in the standard ration [16, 24].

Both the foods mentioned in the list and the standard ration could be consumed during the same meal and VLCKD was taken by athletes ad libitum. During the VLCK diet, the athletes also consumed some specific herbal extracts: 20 ml of extract A, 20 ml of extract B and 50 ml of extract C as described in Tables 1 and 2. Moreover, during ketogenic diet periods, athletes assumed 1 caplet in of a multivitamin-mineral supplement each morning ([19, 25, 26]. The composition of the caplets was: Magnesium19 mg, Calcium 16 mg, Phosphorus 8 mg, Zinc 4.5 mg, Iron 4.62 mg, Manganese 1 mg, Potassium 0.5 mg, Copper 0.4 mg, Chromium 28.55 μg, Selenium 4 μg, Niacin 10 mg, Beta carotene 1.8 mg, Folic Acid 66 μg, Biotin 30 μg, Vitamin C 19.8 mg, Vitamin E 3.3 mg, Pantothenic Acid 1.98 mg, Vitamin B6 0.66 mg, Vitamin B2 0.53 mg, Vitamin B1 0.426 mg, Vitamin D3 1.65 μg, Vitamin B12 0.33 μg (Multivitaminico Balestra e Mech, Gianluca Mech SpA, Asigliano Veneto VI).

Table 1 Plant extracts used during the VLCKD Full size table

Table 2 Main actives ingredients of phytoextracts and their reported beneficial effects Full size table

During the second period of the study, the athletes themselves assumed the function of a control group and consumed their usual diet that is very close to the present US diet western diet (WD) [38] with the exception of fat origin (monounsaturated fats, i.e. olive oil in our subjects vs. saturated fats in the typical US diet). The diet consumed was composed mainly of potatoes, whole grains (bread, pasta, whole wheat, rice, and potatoes), meat, fish, eggs, poultry, vegetables, legumes, fruits, condiments (mainly olive oil), whole milk and wine. As with the ketogenic diet, the WD was taken ad libitum according to the nutritional habits of the athletes.

The diets were explained to all subjects by a qualified dietician during an individual visit. Dietary intake was measured by validated 3- day food diary [39] that has been used in the past in studies with athletes [40] and analysed by Dietnext® (Caldogno, Vicenza, Italy) software. During the ketogenic period the prescribed daily intake of carbohydrate was 22 g. The percentage distribution of total daily energy macronutrients was 54.8% fat, 40.7% protein and 4.5% carbohydrates. The total amount of daily kilojoules was 8254.5 ± 1136. During the WD period the macronutrients were distributed in the following order: 46.8% carbohydrate, 38.5% lipids, 14.7% protein. The Western diet provided a total daily kJ 9520.7 ± 1080.71 (Table 3).

Table 3 Average macronutrient and total energy intake during the ketogenic and the free diet period Full size table

Measurements

Before and after the diet period, muscle and fat amounts and percentages, were assessed by skinfold measurement which are highly related to percent body fat in fit and healthy young men [41–43]. We used software (Fitnext®, Caldogno, Vicenza, Italy) that includes 9 skinfolds (triceps, biceps, pectoral, subarmpit, subscapular, iliac crest, mid-abdominal, anterior thigh, medial calf), 6 bone circumferences (arm, forearm, waist, hip, thigh, calf), 4 bone diameters (elbow, wrist, knee, ankle), waistline and hip circumference measurement [24, 44]). Anthropometric measurements were performed according to the Anthropometric Standardization Reference Manual [45]. Weight was measured to the nearest 0.1 kg using an electronic scale (Tanita BWB-800 Medical Scales, USA), and height to the nearest 1 cm using a Harpenden portable stadiometer (Holtain Ltd, UK). Skinfolds were measured to the nearest 1 mm using a Holtain caliper (Holtain Ltd, UK), and circumferences to the nearest 0.001 m using an anthropometric tape. All measurements were taken by the same operator (LC) before and during the study according to standard procedures [45, 46].

Following the anthropometric assessment a standardized warm-up lasting 15 minutes consisting of callisthenic exercise was carried out. After 5–8 minutes all the athletes underwent the following strength tests: squat jump (SJ), counter movement jump (CMJ), 15 seconds of consecutive CMJs, push-ups test, reverse grip chins test, legs closed barrier maximum test, parallel bar dips test. Jump tests were performed on a contact mat (Ergojump—Bosco system, srl, S. Rufina di Cittaducale, Rieti, Italia), that allowed the measurement of height of jump, time of flight and time of contact. The height of jumps was calculated according to the Asmussen and Bonde-Petersen formula [47]. All jump test techniques assume that the athlete’s position on the mat is the same both at take-off and landing. During jumps athlete’s hands were kept on hips to minimize upper limbs contribution and trunk was maintained erect. The SJ test was performed from the seated position maintained at least for 1 second (knee secured at 90° of knee flexion) then athletes were asked to jump. The CMJ starting from a standing position, then subjects were instructed to perform a rapid downward movement to about 90° of knee flexion immediately followed by an upward movement. The CMJs were consecutively repeated during 15 seconds without recovery between jumps. For CMJs mean jump height and mechanical power per kilogram of body weight were computed [48]. For all three test types the subjects were requested to jump as high as possible. SJ and CMJ were performed three times with two minutes rest between each trial. The best performance was retained and included in the test [49]. The exercises for the upper part of the body were carried out by each athlete until exhaustion. In the push-up test the subjects were positioned with the palms of the hands in support on the floor at shoulder width; at the start of the exercise, the subjects folded their arms while contemporaneously lowering the trunk to the floor.

In the reverse grip chins test the athletes grabbed the bar (as used in artistic gymnastics) at shoulder width; the subjects first brought the chest to the bar height. In the legs closed barrier maximum test, the subjects grab the bar and without oscillating the pelvis elevated the lower limbs to bring the back of both feet in contact with the bar. During parallel bar dips test the subjects lowered themselves to the limit allowed by the shoulder joint.

Test-retest reliability for all exercises obtained in our setting was consistent with previous findings: ICCr: SJ O.97, CMJ 0.99, push-up 0.98, reverse grip chins 0.96, leg closed barrier 0.90, parallel dips 0.95 [50–55].

Statical analysis

A one-way Anova for repeated measurements was used with significance placed at p < 0.05. When appropriate a Bonferroni post hoc test was used to compare selected data.