Ergogenic effect of pre-exercise chicken broth ingestion on a high-intensity cycling time-trial.

BACKGROUND: chicken meat extract is a popular functional food in Asia. It is rich in the bioactive compounds carnosine and anserine, two histidine-containing dipeptides (HCD). Studies suggest that acute pre-exercise ingestion of chicken extracts has important applications towards exercise performance and fatigue control, but the evidence is equivocal. This study aimed to evaluate the ergogenic potential of the pre-exercise ingestion of a homemade chicken broth (CB) vs a placebo soup on a short-lasting, high-intensity cycling exercise. METHODS: fourteen men participated in this double-blind, placebo-controlled, crossover intervention study. Subjects ingested either CB, thereby receiving 46.4 mg/kg body weight of HCD, or a placebo soup (similar in taste without HCD) 40 min before an 8 min cycling time trial (TT) was performed. Venous blood samples were collected at arrival (fasted), before exercise and at 5 min recovery. Plasma HCD were measured with UPLC-MS/MS and glutathione (in red blood cells) was measured through HPLC. Capillary blood samples were collected at different timepoints before and after exercise. RESULTS: a significant improvement (p = 0.033; 5.2%) of the 8 min TT mean power was observed after CB supplementation compared to placebo. Post-exercise plasma carnosine (p <  0.05) and anserine (p <  0.001) was significantly increased after CB supplementation and not following placebo. No significant effect of CB supplementation was observed either on blood glutathione levels, nor on capillary blood analysis. CONCLUSIONS: oral CB supplementation improved the 8 min TT performance albeit it did not affect the acid-base balance or oxidative status parameters. Further research should unravel the potential role and mechanisms of HCD, present in CB, in this ergogenic approach.

Specificity, Dose Dependency, and Kinetics of Markers of Chicken and Beef Intake Using Targeted Quantitative LC-MS/MS: A Human Intervention Trial.

SCOPE: Common methods for food intake assessment are error-prone. Estimating food intake via metabolite biomarkers in blood/urine is challenged by inter-individual variation. Here, meat intake markers based on criteria defined within the FoodBAll consortium, including dose dependency, specificity, kinetics, and their ability to predict meat dose, are evaluated. METHODS AND RESULTS: In two randomized human interventions, meat at different doses are consumed. Plasma concentrations of 100 analytes, including previously proposed meat intake markers, are determined at different time points up to 24 h after meat ingestion using targeted liquid chromatography-tandem mass spectrometry. Plasma concentrations of π-methylhistidine (π-M-His) correlated best with the chicken meat amount consumed even after 24 h (R2 = 0.96). Both, anserine and π-M-His show first-order elimination kinetics, irrespective of meat dose (t1/2 is 1.4 and 5.9 h, respectively). Surprisingly, π-M-His best predicted the amount of beef consumed, albeit at lower concentrations. Furthermore, trimethylamine-N-oxide (TMAO) increases only after beef, while dimethylglycine only after chicken consumption. The lack of baseline concentrations for π-M-His and anserine is likely the strength of these compounds to predict meat dose. CONCLUSION: Quantitative assessment of meat intake within 24 h is most accurate with π-M-His, whereas TMAO and dimethylglycine best discriminate between chicken and beef.

Plasma concentrations of anserine, carnosine and pi-methylhistidine as biomarkers of habitual meat consumption.

BACKGROUND/OBJECTIVES: Dietary intake of red and processed meat has been associated with disease risk. Since dietary intake assessment methods are prone to measurement errors, identifying biomarkers of meat intake in bio-samples could provide more valid intake estimates. We examined associations of habitual red and processed meat, poultry, fish, and dairy products consumption with plasma concentrations of anserine, carnosine, pi-methylhistidine (Π-MH), tau-methylhistidine (T-MH), and the ratio of T-MH to Π-MH in a cross-sectional study. SUBJECTS/METHODS: Plasma anserine, carnosine, Π-MH, and T-MH concentrations were measured using ion-pair LC-MS/MS in 294 participants in the second Bavarian Food Consumption Survey (BVS II). Habitual food consumption was assessed using three 24-h dietary recalls. Associations between plasma metabolites concentrations and meat, fish, eggs, and dairy products consumption were assessed by fitting generalized linear model, adjusted for age, sex, and BMI. RESULTS: Total meat intake was associated with plasma concentrations of anserine, carnosine, Π-MH and, the ratio of T-MH to Π-MH. Red meat intake was related to carnosine (p-trend = 0.0028) and Π-MH plasma levels (p-trend = 0.0493). Poultry (p-trend = 0.0006) and chicken (p-trend = 0.0003) intake were associated with Π-MH. The highest anserine concentrations were observed in individuals consuming processed meat or turkey. For T-MH we did not observe any association with meat intake. CONCLUSIONS: Our results indicate an association between habitual meat consumption and plasma concentrations of anserine, carnosine, Π-MH and the ratio of T-MH to Π-MH. Intervention studies should clarify whether the analyzed plasma metabolites are indicative for a specific type of meat before proposing them as biomarkers of habitual meat intake in epidemiologic studies.

Appearance of Di- and Tripeptides in Human Plasma after a Protein Meal Does Not Correlate with PEPT1 Substrate Selectivity.

SCOPE: Peptide transporter 1 (PEPT1) function is well understood, yet little is known about its contribution toward the absorption of dietary amino acids in the form of di- and tripeptides. In the present human study, changes in plasma concentrations of a representative oligopeptide panel are investigated after meat intake. METHODS AND RESULTS: Based on a method for quantitative analysis of a panel of selected di- and tripeptides in biological samples, the kinetics of plasma changes of peptides derived from a widely accessible dietary protein source are described. The findings demonstrate postprandial changes of a whole spectrum of dipeptides of different size, charge, and polarity in peripheral blood in a dose-dependent manner after consumption of chicken breast in healthy human volunteers. Although the substrate specificity of PEPT1 is well known, the spectrum of peptides appearing in blood cannot be matched to the affinity to PEPT1. Stability against hydrolysis by exo- and endopeptidases appears to be another factor influencing their presence in blood. In addition, the study shows that dipeptides, including gamma-glutamyl-peptides, as well as tripeptides are common components present in human plasma. CONCLUSION: Besides amino acids, human peripheral blood contains numerous di- and tripeptides. The dietary source determines their abundance and composition.

Development and validation of a sensitive LC-MS/MS assay for the quantification of anserine in human plasma and urine and its application to pharmacokinetic study.

Carnosine (beta-alanyl-L-histidine) and its methylated analogue anserine are present in relevant concentrations in the omnivore human diet. Several studies reported promising therapeutic potential for carnosine in various rodent models of oxidative stress and inflammation-related chronic diseases. Nevertheless, the poor serum stability of carnosine in humans makes the translation of rodent models hard. Even though anserine and carnosine have similar biochemical properties, anserine has better serum stability. Despite this interesting profile, the research on anserine is scarce. The aim of this study was to explore the bioavailability and stability of synthesized anserine by (1) performing in vitro stability experiments in human plasma and molecular modelling studies and by (2) evaluating the plasma and urinary pharmacokinetic profile in healthy volunteers following different doses of anserine (4-10-20 mg/kg body weight). A bio-analytical method for measuring anserine levels was developed and validated using liquid chromatography-electrospray mass spectrometry. Both plasma (CMAX: 0.54-1.10-3.12 µM) and urinary (CMAX: 0.09-0.41-0.72 mg/mg creatinine) anserine increased dose-dependently following ingestion of 4-10-20 anserine mg/kg BW, respectively. The inter-individual variation in plasma anserine was mainly explained by the activity (R2 = 0.75) and content (R2 = 0.77) of the enzyme serum carnosinase-1. Compared to carnosine, a lower interaction energy of anserine with carnosinase-1 was suggested by molecular modelling studies. Conversely, the two dipeptides seems to have similar interaction with the PEPT1 transporter. It can be concluded that nutritionally relevant doses of synthesized anserine are well-absorbed and that its degradation by serum carnosinase-1 is less pronounced compared to carnosine. This makes anserine a good candidate as a more stable carnosine-analogue to attenuate chronic diseases in humans.

Antioxidant status of turkey breast meat and blood after feeding a diet enriched with histidine.

The objective of this study was to investigate the effects of 1) spray dried blood cells rich in histidine and 2) pure histidine added to feed on the antioxidant status and concentration of carnosine related components in the blood and breast meat of female turkeys. The experiment was performed on 168 Big7 turkey females randomly assigned to 3 dietary treatments: control; control with the addition of 0.18% L-histidine (His); and control with the addition of spray dried blood cells (SDBC). Birds were raised for 103 d on a floor with sawdust litter, with drinking water and feed ad libitum. The antioxidant status of blood plasma and breast muscle was analyzed by ferric reducing ability (FRAP) and by 2,2-Azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radicals scavenging ability. The activity of antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) was analyzed in the blood and breast meat, with the content of carnosine and anserine quantified by HPLC. Proximate analysis as well as amino acid profiling were carried out for the feed and breast muscles. Growth performance parameters also were calculated. Histidine supplementation of the turkey diet resulted in increased DPPH radical scavenging capacity in the breast muscles and blood, but did not result in higher histidine dipeptide concentrations. The enzymatic antioxidant system of turkey blood was affected by the diet with SDBC. In the plasma, the SDBC addition increased both SOD and GPx activity, and decreased GPx activity in the erythrocytes. Feeding turkeys with an SDBC containing diet increased BW and the content of isoleucine and valine in breast muscles.

Anserine inhibits carnosine degradation but in human serum carnosinase (CN1) is not correlated with histidine dipeptide concentration.

BACKGROUND: We reported an association of a particular allele of the carnosinase (CNDP1 Mannheim) gene with reduced serum carnosinase (CN1) activity and absence of nephropathy in diabetic patients. Carnosine protects against the adverse effects of high glucose levels but serum carnosine concentration was generally low. METHODS: We measured the concentration of two further histidine dipeptides, anserine and homocarnosine, via HPLC. CN1 activity was measured fluorometically and for concentration we developed a capture ELISA. RESULTS: We found an association between the CNDP1 Mannheim allele and reduced serum CN1 activity for all three dipeptides but no correlation to serum concentrations although anserine and homocarnosine inhibited carnosinase activity. Patients with liver cirrhosis have low CN1 activity (0.24 ± 0.17 µmol/ml/h, n=7 males; normal range: 3.2 ± 1.1, n=104; p<0.05) and CN1 concentrations (2.3 ± 1.5 µg/ml; normal range: 24.9 ± 8.9, p<0.05) but surprisingly, histidine dipeptide concentrations in serum are not increased compared to controls. CONCLUSIONS: Serum histidine dipeptide concentrations are not correlated to CN1 activity. The protective effect of low CN1 activity might be related either to turnover of CN1 substrates or a protective function of dipeptides might be localized in other tissues.

Profiling histidine dipeptides in plasma and urine after ingesting beef, chicken or chicken broth in humans.

The in vitro metabolic stability of histidine-dipeptides (HD), carnosine (CAR) and anserine (ANS), in human serum, and their absorption kinetics after ingesting pure carnosine or HD rich foods in humans have been investigated. Healthy women (n = 4) went through four phases of taking one dose of either 450 mg of pure carnosine, 150 g beef (B), 150 g chicken (C), or chicken broth (CB) from 150 g chicken with a >2-week washout period between each phase. Blood samples were collected at 0, 30, 60, 100, 180, 240, and 300 min, and urine samples before and after (up to 7 h) ingesting pure carnosine or food. Both plasma and urine samples were analyzed for HD concentrations using a sensitive and selective LC-ESI-MS/MS method. CAR was undetectable in plasma after ingesting pure carnosine, B, C or CB. By contrast, plasma ANS concentration was significantly increased (P < 0.05) after ingesting C or CB, respectively. Urinary concentrations of both CAR and ANS were 13- to 14-fold increased after ingesting B, and 14.8- and 243-fold after CB ingestion, respectively. Thus, dietary HD, which are rapidly hydrolyzed by carnosinase in plasma, and excreted in urine, may act as reactive carbonyl species sequestering agents.

Intestinal absorption and blood clearance of L-histidine-related compounds after ingestion of anserine in humans and comparison to anserine-containing diets.

Anserine is a bioactive dipeptide found in muscles and brains of vertebrates, but little is known about the kinetics of its absorption into blood and the clearance after the ingestion of anserine or anserine-containing diets. This study investigated time-dependent changes in the concentrations of l-histidine-related compounds from deproteinized blood. The concentration of anserine peaked and then decreased to zero, whereas the concentration of pi-methylhistidine gradually increased, at which point anserine was not detected. Thus, ingested anserine is absorbed intact in human blood and is hydrolyzed to pi-methylhistidine and beta-alanine by serum and tissue carnosinases. Moreover, the crossover study suggests that there was no significant difference in absorption under curves of anserine between anserine alone and anserine-containing diet, whereas there was significant difference in the peak concentration of anserine. This is the first study to demonstrate intestinal absorption and blood clearance of anserine.

Carnosine and anserine ingestion enhances contribution of nonbicarbonate buffering.

PURPOSE: The purpose of the present study was to investigate the effect of supplementation with chicken breast extract (CBEX), which was a rich source of carnosine and anserine, on acid-base balance and performance during intense intermittent exercise. METHODS: Eight male subjects performed intense intermittent exercise that consisted of 10 x 5-s maximal cycle ergometer sprints with a 25-s recovery period between each sprint. The subjects ingested 190 g of the test soup containing either CBEX or a placebo 30 min before the commencement of exercise. Arterial blood samples were collected at rest and during exercise to estimate the carnosine and anserine concentrations, pH, and bicarbonate concentration ([HCO3-]). RESULTS: Concentrations of anserine and its related amino acid significantly increased 30 min after CBEX supplementation, as compared with their values at rest. However, carnosine did not increase significantly. Following CBEX supplementation, the pH was significantly higher (P < 0.05) at the end of exercise, and [HCO3-] was also significantly higher (P < 0.05) during the latter half of exercise and after exercise. There were no significant differences in the total power and mean power of each set between the CBEX and placebo supplemented groups. CONCLUSION: Although oral supplementation with CBEX (which is a rich source of carnosine and anserine) increased the contribution of the nonbicarbonate buffering action and decreased bicarbonate buffering action in blood, intense intermittent exercise performance did not improve significantly.

Peculiarities of carnosine metabolism in a patient with pronounced homocarnosinemia.

The article describes a case of homocarnosinemia with increased liquor and plasma content of homocarnosine, increased urinary excretion of homocarnosine, and low activity of serum carnosinase. These metabolic disturbances were accompanied by moderate neurological disorders. Changes in carnosine metabolism in family members were less pronounced and not accompanied by neuropathological symptoms.

Effect of dietary carnosine on plasma and tissue antioxidant concentrations and on lipid oxidation in rat skeletal muscle.

The effect of dietary carnosine supplementation on plasma and tissue carnosine and alpha-tocopherol concentrations and on the formation of thiobarbituric acid reactive substances (TBARS) in rat skeletal muscle homogenates was evaluated. Plasma, heart, liver and hind leg muscle was obtained from rats fed basal semipurified diets or basal diets containing carnosine (0.0875%), alpha-tocopheryl acetate (50 ppm), or carnosine (0.0875%) plus alpha-tocopheryl acetate (50 ppm). Dietary carnosine supplementation did not increase carnosine concentrations in heart, liver and skeletal muscle. Dietary supplementation with both carnosine and alpha-tocopherol increased carnosine concentrations in liver 1.56, 1.51- and 1.51-fold as compared with diets lacking carnosine, alpha-tocopherol or both carnosine and alpha-tocopherol, respectively. Dietary supplementation with both carnosine and alpha-tocopherol also increased alpha-tocopherol concentrations in heart and liver 1-38-fold and 1.68-fold, respectively, as compared to supplementation with alpha-tocopherol alone. Dietary supplementation with carnosine, alpha-tocopherol or both carnosine and alpha-tocopherol was effective in decreasing the formation of TBARS in rat skeletal muscle homogenate, with dietary alpha-tocopherol and alpha-tocopherol plus carnosine being more effective than dietary carnosine alone. The data suggest that dietary supplementation with carnosine and alpha-tocopherol modulates some tissue carnosine and alpha-tocopherol concentrations and the formation of TBARS in rat skeletal muscle homogenates.

Determination of carnosine and other biogenic imidazoles in equine plasma by isocratic reversed-phase ion-pair high-performance liquid chromatography.

The isocratic reversed-phase ion-pair high-performance liquid chromatographic technique presented provides a sensitive, rapid and reproducible analytical method for the selective determination of carnosine and other biogenic imidazoles in equine plasma. Plasma was deproteinized with 5-sulphosalicylic acid and the compounds of interest were isolated by sorbent extraction on Bond Elut PRS cartridges. Recoveries were 97-105% and the lowest limits of detection were 58.3-80.1 nM. All compounds of interest were well resolved within a maximum retention time of 9.2 min. The mean equine plasma carnosine level determined by this method was 11.31 microM. Comparative determinations were made in canine and human plasma. Carnosine was not detected in human plasma. Concentrations of imidazole in canine plasma are reported here for the first time.

A sensitive and specific high performance liquid chromatographic assay for imidazole dipeptides and 3-methylhistidine in human muscle biopsies, serum and urine.

A sensitive and specific high performance liquid chromatographic method is described for measuring imidazole dipeptides and 3-methylhistidine in human muscle biopsies, serum and urine. Muscle extract, serum or urine was reacted with o-phthaldialdehyde and the derivatives were separated by reversed phase chromatography with column switching and fluorescence detection.