Non-targeted metabolomics analyses by mass spectrometry to explore metabolic stress after six training weeks in high level swimmers.

The objective was to compare the metabolic responses of high-level national swimmers to threshold or polarised training. 22 swimmers (n = 12 males and 10 females) participated in a 28-week cross-over intervention study consisting of 2 × 6 period weeks of training. Swimmers were assigned randomly to either training group for the first period: polarised (POL) (81% in energetic zone 1: blood lactate [La]b ≤ 2 mmol.L-1; 4% in zone 2: 2 mmol.L-1 <[La]b ≤ 4 mmol.L-1; 15% in zone 3: [La]b > 4 mmol.L-1) or threshold (THR) (65%/25%/10%). Before and after each training period, urine samples were collected for non-targeted metabolomics analysis. Mixed model analysis was performed on metabolomics data including fatigue class factors and/or training and/or interaction. Ion intensities of 6-keto-decanoylcarnitine (+31%), pregnanediol-3-glucuronide (+81%), P-cresol sulphate (+18%) were higher in the threshold group (P < 0.05) indicating higher glycogenic depletion and inflammation without alteration of the neuroendocrine stress axis. 4-phenylbutanic acid sulphate was 200% higher in less fatigued swimmers (P < 0.01) linking the anti-inflammatory activity at the cell membrane level to the subjective perception of fatigue. This research suggests the importance of replenishing glycogen stores and reducing inflammation during high thresholds training loads.

A laboratory approach for characterizing chronic fatigue: what does metabolomics tell us?

INTRODUCTION: Manifestations of fatigue range from chronic fatigue up to a severe syndrome and myalgic encephalomyelitis. Fatigue grossly affects the functional status and quality of life of affected individuals, prompting the World Health Organization to recognize it as a chronic non-communicable condition. OBJECTIVES: Here, we explore the potential of urinary metabolite information to complement clinical criteria of fatigue, providing an avenue towards an objective measure of fatigue in patients presenting with the full spectrum of fatigue levels. METHODS: The experimental group consisted of 578 chronic fatigue female patients. The measurement design was composed of (1) existing clinical fatigue scales, (2) a hepatic detoxification challenge test, and (3) untargeted proton nuclear magnetic resonance (1H-NMR) procedure to generate metabolomics data. Data analysed via an in-house Matlab script that combines functions from a Statistics and a PLS Toolbox. RESULTS: Multivariate analysis of the original 459 profiled 1H-NMR bins for the low (control) and high (patient) fatigue groups indicated complete separation following the detoxification experimental challenge. Important bins identified from the 1H-NMR spectra provided quantitative metabolite information on the detoxification challenge for the fatigue groups. CONCLUSIONS: Untargeted 1H-NMR metabolomics proved its applicability as a global profiling tool to reveal the impact of toxicological interventions in chronic fatigue patients. No clear potential biomarker emerged from this study, but the quantitative profile of the phase II biotransformation products provide a practical visible effect directing to up-regulation of crucial phase II enzyme systems in the high fatigue group in response to a high xenobiotic-load.

Influence of a physical exercise until exhaustion in normothermic and hyperthermic conditions on serum, erythrocyte and urinary concentrations of magnesium and phosphorus.

AIM: The aim of this study was to evaluate the effect of the performance of a maximal exercise test until exhaustion in normothermic and hyperthermic conditions on body concentrations of magnesium (Mg) and phosphorus (P). METHODS: 19 adult males (age: 22.58 ±â€¯1.05 years) performed two maximum incremental exercise tests on a cycloergometer separated by 48 h. The first was performed in normothermia (22 ±â€¯2 °C) and the second in hyperthermic conditions induced with a sauna (42 ±â€¯2 °C). Blood and urine samples were taken before and after each test. RESULTS: The tests in hyperthermia did not produce ergospirometric alterations or a noticeable cardiovascular drift. Serum Mg concentrations underwent a reduction after the stress test in hyperthermia (p > 0.05) but not in normothermia. Nevertheless, urinary and erythrocyte concentrations of Mg, and urinary, erythrocyte and serum concentrations of P did not undergo alterations in either conditions. CONCLUSIONS: It seems that exercise in hyperthermic conditions induces a tissue redistribution of Mg in the body, a fact which was not observed in normothermic conditions.

Serum, erythrocyte and urinary concentrations of iron, copper, selenium and zinc do not change during an incremental test to exhaustion in either normothermic or hyperthermic conditions.

AIM: The aim of this study was to evaluate the effect of the performance of an incremental exercise test until exhaustion in normothermic and hyperthermic conditions on serum, erythrocyte and urine concentrations of Iron (Fe), Copper (Cu), Selenium (Se) and Zinc (Zn). METHODS: Nineteen adult males (age: 22.58 ±â€¯1.06 years) performed two maximum incremental exercise tests on a cycloergometer in normothermia (22 ±â€¯2 °C) and hyperthermia (42±2 °C) separated by 48 h. Urine, serum and erythrocyte samples were collected before and after each test. RESULTS: Serum Se (p < 0.01) and Cu (p < 0.05) levels were altered after each test, but the significance disappeared with the correction for haematocrit. The rest of the values did not undergo alterations in either condition. CONCLUSIONS: It seems that a higher stimulus is necessary to obtain changes in these minerals. The study reveals the need to correct serum concentrations concerning possible changes in these volumes after an acute effort.

Changes in urinary amino acids excretion in relationship with muscle activity markers over a professional cycling stage race: in search of fatigue markers.

The aim of this study was to identify the relationship between metabolic effort, muscular damage/activity indices, and urinary amino acids profile over the course of a strenuous prolonged endurance activity, as a cycling stage race is, in order to identify possible fatigue markers. Nine professional cyclists belonging to a single team, competing in the Giro d'Italia cycling stage race, were anthropometrically characterized and sampled for blood and urine the day before the race started, and on days 12 and 23 of the race. Diet was kept the same over the race, and power output and energy expenditure were recorded. Sera were assayed for muscle markers (lactate dehydrogenase, aspartate aminotransferase, and creatine kinase activities, and blood urea nitrogen), and creatinine, all corrected for plasma volume changes. Urines were profiled for amino acid concentrations, normalized on creatinine excretion. Renal function, in terms of glomerular filtration rate, was monitored by MDRD equation corrected on body surface area. Creatine kinase activity and blood urea were increased during the race as did serum creatinine while kidney function remained stable. Among the amino acids, taurine, glycine, cysteine, leucine, carnosine, 1-methyl histidine, and 3-methyl histidine showed a net decreased, while homocysteine was increased. Taurine and the dipeptide carnosine (ß-alanyl-L-histidine) were significantly correlated with the muscle activity markers and the indices of effort. In conclusion, the metabolic profile is modified strikingly due to the effort. Urinary taurine and carnosine seem useful tools to evaluate the muscle damage and possibly the fatigue status on a long-term basis.

Cancer-related fatigue and chemotherapy-associated adverse effects: correlation with TNF-α, IL-1 and 17-hydroxycorticosteroids.

AIM: We sought to determine the relationship between cancer-related fatigue, chemotherapy-associated adverse effects in patients with advanced stages of cancer, and the levels of TNF-α, IL-1 and 17-hydroxycorticosteroids (17-HCS). PATIENTS & METHODS: Two hundred cancer patients were recruited. They were given a Cancer Fatigue Scale survey to assess their general state of health before and after chemotherapy. Their plasma levels of TNF-α and IL-1 and urine levels of 17-HCS were also measured. RESULTS: Increased levels of TNF-α and IL-1 are common in cancer patients. Thirty-five (17.5%) patients suffered from chemotherapy-associated adverse effects, but their plasma levels of TNF-α and IL-1 were not significantly elevated after chemotherapy. However, the urinary levels of 17-HCS levels were significantly elevated in 23 patients after chemotherapy. CONCLUSION: Patients who had elevated urinary levels of 17-HCS before chemotherapy are accompanied by chemotherapy-associated adverse effects. Thus, elevated 17-HCS in urine could be a possible predictor for chemotherapy-associated adverse effects.

The effect of sleep restriction and psychological load on the diurnal metabolic changes in tryptamine-related compounds in human urine.

OBJECTIVE: The effect of a severely stressful situation (sleep restriction and psychological load) on the diurnal changes in novel tryptamine-related compounds (hydroxydiacetyltryptamine, sulphatoxymelatonin, and dihydromelatonin) was evaluated in human subjects for 16 days. METHODS: The subjects were allowed to sleep for 5 h on days three through 12 and for 8 h on the other days. On days three through 12, the subjects were asked to perform a psychological task. The first two and the last 4 days were viewed as control days. A performance test was administered to evaluate the extent of the subjects' fatigue. Total urine was sampled by collecting it into bottles three times a day [(1) during the sleeping period, (2) in the morning, and (3) in the afternoon]. Seven tryptamine-related compounds in urine were assayed using HPLC-fluorometry. RESULTS: The urine melatonin level was high at night and low during the day. In contrast, urinary levels of hydroxydiacetyltryptamine and sulphatoxydiacetyltryptamine were low at night and high during the day. Dihydromelatonin was undetectable in urine during the sleeping period. Sleep restriction and psychological load did not affect diurnal changes in urinary melatonin, hydroxydiacetyltryptamine, sulphatoxydiacetyltryptamine, or N-acetylserotonin levels. The concentrations of hydroxymelatonin and sulphatoxymelatonin in urine did not show diurnal changes and decreased gradually during the experimental days. A principal component analysis confirmed the diurnal changes and suggested two novel metabolic pathways: (1) N-acetylserotonin to sulphtoxydiacetyltryptamine via hydroxydiacetyltryptamine, and (2) melatonin to dihydromelatonin. CONCLUSION: Severely stressful situations did not affect diurnal changes in melatonin, hydroxydiacetyltryptamine, sulphatoxydiacetyltryptamine, or N-acetylserotonin levels in urine.

Changes of Differential Urinary Metabolites after High-Intensive Training in Teenage Football Players.

OBJECTIVE: The mechanism underlying the fatigue of football players is closely related to the energy depletion and accumulation of metabolites; the present study tries to explore the metabolic mechanism in teenage football players during exercise-induced fatigue. METHODS: 12 teenage football players were subjected to three groups of combined training by using a cycle ergometer, with the subjective Rating of Perceived Exertion (RPE) as a fatigue criterion. The following indicators were measured in each group after training: maximum oxygen uptake (VO2max), anaerobic power, and average anaerobic power. Urine samples were collected before and after the training. Gas chromatography-mass spectrometry (GC-MS) was performed for the metabonomics analysis of the samples. The metabolism data was analyzed by using principal component analysis (PCA) and orthogonal partial least squares analysis (OPLS-DA), through the Kyoto Encyclopedia of Genes and Genomes (KEGG) database to confirm the potential differences between metabolites, and the MetPA database was used to analyze the related metabolic pathways. RESULTS: There was no significant difference between the maximal oxygen uptakes among the three groups. Compared with group 1, the maximum and average anaerobic power in group 3 significantly decreased (p < 0.05) at the end of training. GC-MS detected 635 metabolites in the urine samples. Through PCA, OPLS-DA analysis, and KEGG matching, 25 different metabolites (3↑22↓) that met the conditions were finally selected. These different metabolites belonged to 5 metabolic pathways: glycine-serine-threonine metabolism, citrate cycle, tyrosine metabolism, nitrogen metabolism, and glycerophospholipid metabolism. CONCLUSIONS: During the combined exercise of aerobic and anaerobic metabolism, teenage football players show a significant decrease in anaerobic capacity after fatigue. The metabolic mechanism of exercise fatigue was related to disorders in amino acid and energy metabolism.

Mental and physical fatigue-related biochemical alterations.

OBJECTIVE: To confirm fatigue-related biochemical alterations, we measured various parameters just before and after relaxation and fatigue-inducing mental or physical sessions. METHODS: Fifty-four healthy volunteers were randomized to perform relaxation and fatigue-inducing mental and physical sessions for 4 h in a double-blind, three-crossover design. Before and after each session, subjects were asked to rate their subjective sensations of fatigue, and blood, saliva, and urine samples were taken. RESULTS: After the fatigue-inducing mental and physical sessions, subjective scores of fatigue were increased. After the fatigue-inducing mental session, the vanillylmandelic acid level in urine was higher and plasma valine level was lower than after the relaxation session. In contrast, after the fatigue-inducing physical session, serum citric acid, triacylglycerol, free fatty acid, ketone bodies, total carnitine, acylcarnitine, uric acid, creatine kinase, aspartate aminotransferase, lactate dehydrogenase, cortisol, dehydroepiandrosterone, dehydroepiandrosterone sulfate, plasma branched-chain amino acids, transforming growth factor-beta1 and -beta2, white blood cell and neutrophil counts, saliva cortisol and amylase, and urine vanillylmandelic acid levels were higher and serum free carnitine and plasma total amino acids and alanine levels were lower than those after the relaxation session. CONCLUSION: Some mental or physical fatigue-related biochemical changes were determined. Various biochemical alterations reflecting homeostatic perturbation and its responses might be shown. We believe that our results contribute to clarifying the mechanism of fatigue, developing evaluation methods, and establishing a basis for treatment.

Characterisation of Patients with Systemic Lupus Erythematosus in Malta: A Population Based Cohort Cross-Sectional Study.

Systemic Lupus Erythematosus (SLE) is a multisystemic autoimmune disorder. The aim of this study was to characterise the SLE patients living in Malta in order to estimate the prevalence and incidence of SLE and characterise the clinical presentation as well as identify any unmet needs. 107 SLE patients who fulfilled SLICC classification criteria were identified. These were invited to participate in the study by means of an interview, blood and urine tests, and filling of the following questionnaires: Fatigue Severity Scale (FSS), visual analogue scale (VAS) for fatigue, Hospital Anxiety and Depression Scale (HADS), VAS for pain, Pittsburgh Sleep Quality Index (PSQI), and modified Health Assessment Questionnaire (mHAQ). The estimated prevalence of SLE in Malta is 29.3 patients per 100,000 and the estimated incidence is 1.48 per 100,000 per year. 93.5% of SLE patients were female, and the mean age at diagnosis was 33.1 years. 60.8% were overweight or obese and body mass index (BMI) had a significant positive correlation with daily dose of prednisolone (R=0.177, p=0.046). 20.7% and 3.3% had a moderate and high disease activity, respectively, as measured by SLEDAI-2K. Disease activity had a significant positive correlation with functional disability measured by mHAQ (R=0.417, p<0.001). 56.5% had an abnormal level of fatigue (FSS >3.7) and 57.6% had a high level of anxiety (HADS ≥8). This study has identified a number of unmet needs of SLE patients, including obesity, uncontrolled disease activity, fatigue, and anxiety.

L-Carnitine treatment reduces severity of physical and mental fatigue and increases cognitive functions in centenarians: a randomized and controlled clinical trial.

BACKGROUND: Centenarians are characterized by weakness, decreasing mental health, impaired mobility, and poor endurance. L-Carnitine is an important contributor to cellular energy metabolism. OBJECTIVE: This study evaluated the efficacy of L-carnitine on physical and mental fatigue and on cognitive functions of centenarians. DESIGN: This was a placebo-controlled, randomized, double-blind, 2-phase study. Sixty-six centenarians with onset of fatigue after even slight physical activity were recruited to the study. The 2 groups received either 2 g levocarnitine once daily (n = 32) or placebo (n = 34). Efficacy measures included changes in total fat mass, total muscle mass, serum triacylglycerol, total cholesterol, HDL cholesterol, LDL cholesterol, Mini-Mental State Examination (MMSE), Activities of Daily Living, and a 6-min walking corridor test. RESULTS: At the end of the study period, the levocarnitine-treated centenarians, compared with the placebo group, showed significant improvements in the following markers: total fat mass (-1.80 compared with 0.6 kg; P < 0.01), total muscle mass (3.80 compared with 0.8 kg; P < 0.01), plasma concentrations of total carnitine (12.60 compared with -1.70 mumol; P < 0.05), plasma long-chain acylcarnitine (1.50 compared with -0.1 micromol; P < 0.001), and plasma short-chain acylcarnitine (6.0 compared with -1.50 micromol; P < 0.001). Significant differences were also found in physical fatigue (-4.10 compared with -1.10; P < 0.01), mental fatigue (-2.70 compared with 0.30; P < 0.001), fatigue severity (-23.60 compared with 1.90; P < 0.001), and MMSE (4.1 compared with 0.6; P < 0.001). CONCLUSIONS: Our study indicates that oral administration of levocarnitine produces a reduction of total fat mass, increases total muscular mass, and facilitates an increased capacity for physical and cognitive activity by reducing fatigue and improving cognitive functions.

[No relationship between so-called 'haemopyrrollactamuria' and fatigue in secondary-school students]. / Geen verband tussen zogeheten 'hemopyrrollactamurie' en vermoeidheid bij gymnasiasten.

OBJECTIVE: To determine the relationship between urinary haemopyrrollactam (HPL) and subjective fatigue in secondary-school students. DESIGN: Cross-sectional and descriptive. METHOD: Urine samples were collected from 75 secondary-school students (43 boys and 32 girls, aged 16-18 years) in Nijmegen, the Netherlands. The samples were tested for HPL complex at the Centre for Environmental Medicine (Klinisch Ecologisch Allergie Centrum; KEAC) in Weert. Students also completed the 'Abbreviated fatigue questionnaire'. A score of 21 points was considered the threshold for fatigue. RESULTS: Of the 75 participants, 22 (29%) had increased HPL complex excretion, including 15 boys (35%) and 7 girls (22%). Overall, 19 (25%) had a fatigue score > 21: 16 boys (37%) and 3 girls (9%). There was no statistical relationship between increased HPL complex excretion and fatigue score. CONCLUSION: These findings do not support a relationship between fatigue and increased urinary HPL complex excretion.

Relationship between Carnitine Pharmacokinetics and Fatigue in Patients Treated with Cisplatin-Containing Chemotherapy.

BACKGROUND: Approximately 70% of the patients who receive chemotherapy suffer from fatigue, which lowers their quality of life and also has a negative influence on therapeutic efficacy. Previous studies have suggested a relationship between blood carnitine levels and fatigue. We conducted a prospective observational study to examine the relationship between carnitine pharmacokinetics and chemotherapy-induced fatigue in patients receiving cancer chemotherapy regimens that include cisplatin. PATIENTS AND METHODS: 11 patients receiving chemotherapy including cisplatin (60-80 mg/m2) were included in the study. We performed 24-h urine collections and took blood samples on day 1 (before the initiation of chemotherapy) and days 2, 3, 4, and 8 in order to measure the carnitine concentrations in the serum and urine. These were compared with measures of self-reported fatigue. The primary endpoint was the change in self-reported fatigue subscales from baseline to day 8. RESULTS: Urinary carnitine concentrations differed significantly on days 2 and 3 (p = 0.003). The Functional Assessment of Chronic Illness Therapy-Fatigue scale version 4A score on day 8 indicated significantly higher levels of fatigue as compared to day 1 (p = 0.013). CONCLUSION: This study suggests that there is an association between urinary carnitine levels and self-reported fatigue.

Separation of Key Biogenic Amines by Capillary Electrophoresis and Determination of Possible Indicators of Sport Fatigue in Athlete's Urine.

This article aims to build up a simple, rapid and accurate capillary zone electrophoresis (CZE) method for the separation of biogenic amines (BAs). Here, 10 key BAs (phenethylamine, histamine, tryptamine, tyramine, 5-hydroxytryptamine, octopamine, dopamine, norepinephrine, epinephrine and carnosine) owning significant functions were chosen for method development. The baseline separation and identification of 10 standards of the mixture by CZE were eventually achieved in 150.0 mmol/L phosphate buffer (Na2HPO4-NaH2PO4) containing 1.0 mmol/L borax at a pH of 6.1. The addition of borax was found effective for improving the isomeric separation of octopamine and dopamine. The proposed method allowed the limits of detections of BAs to be in the range of 0.2-1.2 µmol/L at UV detection (200 nm); the relative standard deviation of the migration time and the peak area were in the ranges 0.08-0.12 and 2.74-4.63% (n = 5), respectively. Formaldehyde (a possible antiseptic in urine) and five main matrices in urine were studied for the identification of BAs. Finally, profiling of BAs in actual urine from athletes was carried out. Currently, only phenethylamine, norepinephrine and carnosine were designated by spiking the standards. In addition, their variation in athletes' urine has been checked at different states of sport fatigue with the goal of obtaining possible indicators of sport fatigue.

An UPLC-MS-based metabolomics investigation on the anti-fatigue effect of salidroside in mice.

An ultra-performance liquid chromatography-quadrupole time-of-flight-based metabolomic approach was developed to study influence of salidroside, an anti-fatigue ingredient from Rhoiola rosea, on urinary metabolic profiling of rats to a single dose of 180 mg/kg per day. Unsupervised principal component analysis (PCA) and supervised orthogonal pre-projection to latent structures discriminate analysis (OPLS-DA) on metabolite profiling revealed obvious differentiation between the salidroside treated groups and controls in both positive and negative ion modes. Eleven urinary metabolites contributing to the differentiation were identified as anti-fatigue biomarkers: N-acetylserotonin, 2-Methoxyestrone 3-glucuronide, Taurine, Melatonin, Sorbitol, Geranyl diphosphate, Z-nucleotide, Cortisone, Dihydrocortisol, Sebacic acid, Pregnenolone sulfate. The physiological significance of these biomarkers is discussed. The work showed that metabolomics is a powerful tool in studying the anti-fatigue effects of natural compound salidroside on multiple targets in vivo.

Association of habitual dietary resveratrol exposure with the development of frailty in older age: the Invecchiare in Chianti study.

BACKGROUND: Resveratrol may play a protective role against the frailty syndrome (FS) because of its antioxidant and anti-inflammatory properties. OBJECTIVE: We prospectively evaluated the association between habitual dietary resveratrol exposure and the development of FS after 3-, 6-, and 9-y follow-up periods in a community-dwelling older population. DESIGN: We conducted a longitudinal analysis with the use of data from 769 participants aged ≥65 y from the Invecchiare in Chianti (Aging in Chianti) study. Total dietary resveratrol (TDR) intake was estimated at baseline with the use of a validated food-frequency questionnaire, which was developed to assess participants' usual food intakes over the previous year, and an ad hoc resveratrol database. Total urinary resveratrol (TUR) was analyzed with the use of liquid chromatography-tandem mass spectrometry with a previous solid-phase extraction at baseline. The combination of both measures [total dietary resveratrol plus total urinary resveratrol (TDR+TUR)] was computed with the use of the Howe's method. FS was assessed at baseline and at 3-, 6-, and 9-y of follow-up and was defined as the presence of ≥3 of the following 5 criteria: shrinking, exhaustion, sedentariness, slowness, and weakness. RESULTS: TDR+TUR concentrations were inversely associated with FS risk over 3-y of follow-up (OR for comparison of extreme tertiles: 0.11; 95% CI: 0.03, 0.45; P-trend = 0.002) but not after 6- and 9-y of follow-up in multinomial logistic regression models adjusted for baseline frailty status and potential confounders. These results did not differ when analyses were further adjusted for inflammatory markers. CONCLUSION: Higher habitual dietary resveratrol exposure was associated with lower risk of older community dwellers developing FS during the first 3 y of follow-up but not after longer follow-up periods.

The circadian covariation of fatigue and urinary melatonin.

Twelve healthy male subjects were kept under constant conditions (no sleep, isolation from time cues, controlled activity, etc.) for 64 hr. Urinary melatonin values, self-rated sleepiness, and vigilance performance scores were obtained every 3 hr. All variables showed a pronounced circadian rhythmicity. Vigilance performance and self-rated sleepiness showed, in addition, a gradual decrease and increase, respectively, with increasing sleep deprivation. The correlation of melatonin with performance and ratings was highly significant, high melatonin levels being associated with reduced performance and increased sleepiness. Aligning self-ratings and behavioral data with respect to the melatonin troughs and peaks showed that the former coincided with performance and alertness peaks and the latter with the troughs. It was concluded that under these conditions there is a strong circadian covariation between melatonin and indices of fatigue/sleepiness.