However, recent investigations by Church et al. A decrease in muscle histidine levels may compromise physiological function by reducing protein synthesis and lowering hematocrit and hemoglobin levels Kriengsinyos et al.

An interesting difference between these studies is that Church et al. It is possible that the different pharmacokinetic properties of sustained-release and rapid-release β-alanine may affect concentrations of intramuscular histidine content in a different manner.

In addition, the effect of these different β-alanine formulations on changes in muscle performance was also examined. Each participant reported to the Human Performance Laboratory at the University of Central Florida on two occasions separated by a period of four weeks 28 days.

Informed consent was obtained from all individual participants included in the study. On the first day of testing, each participant was randomly assigned into one of three groups: rapid-release β-alanine RR , sustained-release β-alanine SR , or placebo PLA.

This was a double-blinded experimental design. Prior to each testing session, participants were instructed to fast for a minimum of 2 h and avoid lower body physical activity for 48 h prior to testing. During each visit, body composition measures and a muscle biopsy were obtained from participants prior to completing a fatiguing knee extensor protocol.

Thirty-seven physically active men and women were recruited for this study. Participants were stratified into one of the three groups SR, RR, or PLA in a counterbalanced fashion based on the peak torque PT values obtained during unilateral maximal voluntary isometric contraction MVIC performed on the first testing day.

Participants were instructed to maintain normal food and exercise habits throughout the duration of the study. This investigation was approved by the New England Institutional Review Board for human subjects, and all procedures were in accordance with the ethical standards of the Helsinki Declaration and its later amendments.

Following an explanation of all procedures, risks, and benefits, each participant provided their written informed consent to participate in the study. All participants were required to be free of any physical limitations as determined by medical history questionnaire and PAR-Q and not to have consumed β-alanine for at least 9 weeks prior to enrollment in the study.

Although being a vegetarian was not part of the exclusion criteria, none of the participants recruited reported to be a vegetarian. The demographics of each group can be seen in Table 1. Participants were instructed to consume their supplements with regular meals breakfast, lunch, dinner.

Both RR and SR supplements were provided in tablet form 1 g and were identical in appearance. Participants in the placebo group were provided with the same number of tablets that were also identical in appearance to the β-alanine tablets.

Active and placebo tablets were provided by Natural Alternatives International Carlsbad, CA, USA. Supplementation compliance was tracked by self-reported supplement logs and by inspection of the number of tablets remaining in each bottle upon return for follow-up testing.

Anthropometric measurements were assessed for each participant during both visits to the laboratory. Upon arrival to the laboratory, participants were instructed to void their bladder in order to properly assess body composition.

Body composition was assessed via air displacement plethysmography BodPod ® , COSMED, Chicago, IL, USA. Anthropometric measures of each group are depicted in Table 1.

The muscle biopsy procedures used in this investigation have been previously reported Varanoske et al. Prior to testing, all participants were instructed to wear shorts on testing day to expose the upper portion of their thigh. A B-mode, linear probe ultrasound General Electric LOGIQe, Wauwatosa, WI, USA , coated with transmission gel Aquasonic ® , Parker Laboratories, Inc.

The biopsy area was washed with antiseptic soap and cleaned with rubbing alcohol. A small area of the clean skin approximately 2 cm in diameter was then anesthetized with a 2. The biopsy site was further cleansed by swabbing the area with betadine.

Once anesthetized, a spring loaded reusable microbiopsy instrument with a disposable gauge needle Argon Medical Devices Inc. Approximately, 5—6 muscle samples were extracted from each participant on each occasion, with the goal of obtaining about 15—20 mg of total wet tissue weight.

After removal, each muscle sample was transferred to a petri dish placed on ice to trim adipose tissue from the muscle specimens. The isometric testing and the muscle-fatiguing protocol used in this investigation have been previously reported Varanoske et al.

Due to the participants not being specifically trained in resistance exercise, an isokinetic dynamometer was used to elicit muscle fatigue and evaluate muscle strength while preventing injury. Briefly, as participants were reporting to the lab on a 2-h minimum fast, 8 oz of a carbohydrate-containing beverage 60 calories, 16 g carbohydrates, 0 g fat, and 0 g protein was provided to the participants following the muscle biopsy, minutes prior to the isokinetic muscle-fatiguing protocol.

Unilateral MVICs and an isokinetic muscle-fatiguing protocol were performed on an isokinetic dynamometer System 4, Biodex Medical System, Inc. To avoid any residual effects of the muscle biopsy, the right leg of each participant was tested.

The lower portion of the leg was secured to the dynamometer arm just above the medial and lateral malleoli. Participants were seated in the dynamometer with a hip angle of ° and strapped to the chair at the waist, shoulders, and across the left thigh.

Chair and dynamometer settings were adjusted for each participant but kept consistent between visits to properly align the axis of rotation of the knee with the lateral condyle of the femur.

Range of motion was assessed for each participant. All participants were able to achieve a range of motion of 90—° without discomfort. The gravity effect of moment was measured at ° of knee flexion ° representing full extension and subsequently corrected during testing Beyer et al.

Each contraction was initiated from a position of 90° knee flexion and was continued to the point of full knee extension. Each bout of 50 contractions was separated by a s recovery period. Participants were encouraged during the first three contractions to make sure that they were contracting maximally from the start of each bout.

Two MVICs were performed prior to the isokinetic muscle-fatiguing protocol, separated by a period of 3 min. Of the two MVICs performed prior to the isokinetic muscle-fatiguing protocol, the one that produced the greatest PT was saved and used for later analysis MVIC1.

Additionally, one MVIC was performed s after the final set of the muscle-fatiguing protocol was completed MVIC2. During these tests, the knee angle was fixed at °, and all MVICs were held for a total of 6 s. PT was recorded during each MVIC.

Torque signals were sampled at 1 kHz with a Biopac data acquisition system MP Biopac Systems, Inc. PT was identified as the greatest torque achieved on the torque-time curve.

Participants were instructed to maintain their normal dietary intake habits throughout the investigation. Total energy, macronutrient, and histidine intakes were monitored using recorded food logs during the h period prior to each visit.

The FoodWorks Dietary Analysis Software, Version 13 The Nutrition Company, Long Valley, NJ, USA was used to analyze dietary recalls.

Participants were required to record side effects that were associated with consuming the supplement on a daily calendar. Side effects were recorded subjectively, through explanation of the symptom and by noting the time during which the symptom occurred.

Side effects were analyzed by quantifying the total amount of days during which paresthesia was encountered over the day supplementation period for each subject. Muscle biopsy samples were homogenized with three volumes of 0.

Louis, MO, USA and subsequently centrifuged at 4 °C for 20 min at 10, rpm. Muscle homogenates were deproteinized with three volumes of acetonitrile BDH VWR Analytical, Radnor, PA, USA , and left to stand at 4 °C for 20 min.

Then, the sample was centrifuged at 4 °C for 10 min at 10, rpm. The supernatant was collected and subsequently analyzed. The experimental methods were performed as described by Mora et al.

Calibration standards were prepared in the range of 0. Chromatography was performed on an Agilent Infinity HPLC Agilent Technologies, Santa Clara, CA, USA and separation was carried out using an Atlantis hydrophilic interaction chromatography HILIC silica column 4.

Mobile phase consisted of solvent A, containing 0. Louis, MO, USA , pH 5. Solvents were filtered through a 0. The column was equilibrated for 10 min under initial conditions before each injection.

The separation was monitored using a diode array detector at a wavelength of nm for carnosine and histidine. Peak areas were correlated to compound concentration by interpolation in the corresponding calibration curve. Duplication of retention times for a known standard was used to verify column equilibrium prior to analysis.

Each sample was run in duplicate; the average intra-assay CV of carnosine was 1. Calibration standards were prepared in the range of 1—0. Louis, MO, USA. Norvaline Nva; Ark Pharm, Arlington Heights, IL, USA was used as an internal standard at a concentration of 0.

Afterwards, the supernatant was transferred to vials and automatic pre-column derivatization with ortho-phthalaldehyde OPA; Agilent Technologies, Santa Clara, CA, USA was performed at room temperature.

Separation was carried out using a Poroshell HPH-C18 column 3. Mobile phase consisted of solvent A which contained 10 mM sodium phosphate dibasic Sigma-Aldrich, St. Louis, MO, USA , 10 mM sodium tetraborate decahydrate Alfa Aesar, Tewksbury, MA, USA , and 5 mM sodium azide BDH VWR Analytical, Radnor, PA, USA , pH 8.

Primary amino acids derivatized with OPA were detected at nm. Each sample was run in duplicate; the average intra-assay CV for β-alanine was 1. Prior to statistical procedures, all data were assessed for normality, homogeneity of variance, and sphericity. If the assumption of sphericity was violated, a Greenhouse—Geisser correction was applied.

PRE- and POST-values were used as the covariate and dependent variable, respectively. To analyze differences in supplement compliance, side effects, and participant characteristics between groups, a one-way ANOVA was performed.

In the event of a significant F ratio for any of these analyses, LSD post hoc comparisons were performed. Outliers were identified when values exceeded 1. Recruitment of participants, screening, and progression through the study are presented in Fig.

A total of 10 participants withdrew from the study prior to group assignment due to reasons unrelated to the investigation. One participant was removed from the final data analysis due to the lack of compliance to the supplementation protocol.

Four participants were removed from the final data analysis because they were deemed to be outliers. Two additional participants from PLA were excluded from the final analysis due to inabilities to fulfill the time commitments of the study.

Additionally, 3 subjects in RR were removed from the final analysis due to errors in data collection. The demographics of participants included in the final analysis are reported in Table 1. Participant recruitment, sampling, and progression through the study.

Dietary analysis revealed that at PRE, the adjusted average nutrient intake for participants during the 72 h prior to the testing session was: No significant difference in supplement compliance was noted between the groups.

Participants consuming RR formulation reported paresthesia on significantly more days Changes in skeletal muscle carnosine are depicted in Fig. At PRE, the average-adjusted muscle carnosine content was 7. Muscle carnosine content in participants consuming SR The unadjusted change in muscle carnosine values from PRE to POST for participants consuming the SR formulation was 3.

These changes reflected a Unadjusted values for PRE- and POST-supplementation skeletal muscle carnosine content. PRE before 28 days of supplementation; POST after 28 days of supplementation, SR sustained-release formulation of β-alanine, RR rapid-release formulation of β-alanine, PLA placebo.

Changes in skeletal muscle histidine and β-alanine are depicted in Figs. At PRE, the average-adjusted muscle histidine content was 0. PRE before 28 days of supplementation, POST after 28 days of supplementation, SR sustained-release formulation of β-alanine, RR rapid-release formulation of β-alanine, PLA placebo.

The average-adjusted muscle β-alanine content at PRE was 0. Unadjusted values for PRE- and POST-supplementation decline in peak torque.

Initial studies reporting significant increases in skeletal muscle carnosine from β-alanine supplementation used an RR formulation Harris et al. Symptoms of paresthesia were a common side effect associated with β-alanine ingestion, with greater symptoms associated with larger daily doses Decombaz et al.

More recently, an SR form of β-alanine has become available, which delays the release of β-alanine and prevents or attenuates symptoms of paresthesia Decombaz et al. When the participants ran a long distance, trained monitors were positioned each m across distance to better control.

A 2 × 2 group × moment repeated measures analysis of variance RMANOVA with the Bonferroni adjustment for multiple comparisons was used to compare lactate concentration and performance.

The partial eta-squared η 2 was calculated for moment. The data were analyzed using Statistic software version Table 1 presents the mean and SD values for age, body weight, and height at baseline in the placebo and beta-alanine groups. There were no statistically significant differences between groups at baseline for any variable investigated.

Figure 2 shows the differences in performance and delta for time between the placebo and β-alanine groups. FIGURE 2. Comparison between placebo and beta-alanine group according to km running performance.

Effect sizes were moderate for β-alanine group 0. Figure 3 presents the differences in the lactate concentration between the placebo and β-alanine groups.

FIGURE 3. Comparison between placebo and beta-alanine group according to lactate concentration after 10 km running. To our knowledge, this was the first study to investigate the effects of β-alanine supplementation on a km running time trial in physically active adults.

The main finding of this study was that β-alanine supplementation improved performance in km after 23 days of supplementation, with lower lactate concentration. A meta-analysis conducted by Hobson et al. They found that β-alanine supplementation was most effective in high-intensity exercise with a duration between 1 and 4 min, showing no effect of β-alanine supplementation in exercises shorter than 60 s.

Another meta-analysis found similar results, in which β-alanine supplementation had greater impact in exercises with a duration between 0. Furthermore, the majority of investigations of β-alanine in the literature used a cycle ergometer, but few studies have analyzed the influence of β-alanine supplementation on running performance.

Smith et al. On the other hand, Ducker et al. In accordance with Ducker et al. The ergogenic effect of β-alanine supplementation is widely due to the increase in intramuscular carnosine content, which improves skeletal muscle buffering capacity Culbertson et al.

Although long-distance running relies mainly on aerobic energy metabolism, some studies have demonstrated that mean running speed in prolonged running is dependent on lactate concentration, showing an association between lower lactate accumulations and higher running speed and anaerobic threshold Sjodin and Jacobs, ; Fohrenbach et al.

Our findings showed that β-alanine supplementation decreased lactate concentration after a km running trial, suggesting that the improvement in performance was due in part to lower blood lactate accumulation.

Previous studies have investigated the influence of β-alanine on lactate accumulation during exercise. These findings corroborate with others Jordan et al.

We hypothesize that the increase in km running performance after β-alanine supplementation observed in the present study may be in part due to the increased muscular buffering capacity, mainly through lower demand on anaerobic glycolysis, generating lower lactate accumulation.

Furthermore, long running duration induced physiological and neuromuscular alterations that impair running speed Davies and Thompson, ; Giandolini et al. Lower muscular excitability induced by prolonged running may be associated with the reduction in muscle glycogen and higher production of reactive oxygen species ROS Duhamel et al.

In addition, carnosine has also been reported to decrease ROS production, with an anti-oxidant activity Kohen et al. We hypothesize that the improvement in km running performance induced by β-alanine supplementation in this study could also be explained by the effect of carnosine on intramuscular calcium influx and anti-oxidant activity, delaying neuromuscular fatigue.

However, more studies are needed to better understand this mechanism. Despite the importance of this study, some limitations need to be mentioned, such a lack of intramuscular analysis, muscle carnosine concentration, and muscle buffering capacity.

Therefore, we suggest further research to analyze the effects of β-alanine supplementation on running time trials over different distances and investigate muscular adaptations in different populations, such as athletes. In summary, β-alanine supplementation improved a km running time trial and decreased blood lactate concentrations in physically active adults.

These results suggest that β-alanine supplementation has positive effects on prolonged running. The present study suggests that β-alanine supplementation can be used as a nutritional strategy to improve performance in km running by lowering blood lactate accumulation.

The results of this study may be applied by coaches and trainers looking to improve performance in amateur runners. EC devised the study design, participated in the interpretation of data, and drafted the manuscript. JS and DdS carried out the data collection, participated in the interpretation of data, and assisted in the writing of the manuscript.

MdF, FL, and JR-N participated in the interpretation of data and drafted the manuscript. FR performed all statistical analysis, participated in the interpretation of data, and assisted in the writing of the manuscript.

All authors read and approved the final manuscript. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Bex, T. Muscle carnosine loading by beta-alanine supplementation is more pronounced in trained vs. untrained muscles.

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Sport Nutr. Duhamel, T. Manipulation of dietary carbohydrates after prolonged effort modifies muscle sarcoplasmic reticulum responses in exercising males. Dutka, T. Effect of carnosine on excitation-contraction coupling in mechanically-skinned rat skeletal muscle.

Muscle Res. Cell Motil. Fohrenbach, R. Determination of endurance capacity and prediction of exercise intensities for training and competition in marathon runners. Sports Med. Privacy Copyright. Skip to main content Home About FAQ My Account.

Authors B McGonigle J Arnold M Lockard. Abstract B. McGonigle, J. Arnold, M. Lockard Willamette University, Salem, OR Increasing total power and decreasing overall fatigue in anaerobic performance are sought-after physical abilities among various athletic and fitness disciplines.

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