This study was performed in a double-blind, placebo-controlled, randomized crossover design where only one investigator knew the identification of the supplementation; this investigator did not engage in the gathering or analysis of data. Participants were requested to attend the Exercise Physiology Lab on two separate occasions, with the second visit taking place within three to nine days after the first visit.

The initial visit comprised of obtaining the informed consent, HHQ, and anthropometric measures. Participant height cm and weight kg were gathered using an electronic physicians scale Tanita WB , Arlington Heights, IL.

Upon the completion of the ingestion period, a post-ingestion venipuncture was performed I2. Participants then performed a standardized warm-up prior to initiating the anaerobic exhaustive exercise protocol.

Immediately following the exercise protocol a post-exercise venipuncture was performed R1 and then the min recovery period was initiated. At the end of this recovery period the final venipuncture was taken R2. The study design can be seen in Fig. Participants were immediately walked to an electronically braked cycle ergometer Sport Excalibur, Lode BV, Groningen, The Netherlands , where the bike was adjusted to the appropriate settings in order to ensure the knee was at a slight bend at the bottom of the revolution.

Bike settings were repeated for both trials. Following the appropriate adjustments, participants feet were strapped into the pedals and the protocol was initiated.

Each Wingate test was s in duration and participants were encouraged to pedal at their maximal effort against a resistance of 0. There was a total of three Wingate tests performed with a two-minute active recovery period between each test.

At the completion of the last Wingate test, participants were walked to a separate room to undergo a post exercise venipuncture and to begin the recovery measures R1-R2. Pre-testing protocols on the electronically braked cycle ergometer followed manufacturer guidelines.

Blood draws were collected via the antecubital vein by a trained phlebotomist during four-time points throughout the study: I1, I2, R1, R2 Fig. In order to account for the plasma volume shifts following the exercise bout, all E and NE samples were normalized by using the established protocols of Dill and Costill [ 14 ].

Hematocrit Hct and hemoglobin Hb were collected via finger sticks at each venipuncture time point Alere Hemopoint 2. Blood glucose GLU was measured using a Medtronic Contour glucometer Bayer, Pittsburgh, PA via finger stick. The procedures and findings of plasma catecholamines were previously reported and permissions granted by the publishing Journal [ 15 ].

Citrus Aurantium and caffeine powder were purchased from Blackburn distributions Caffeine powder, Blackburn distributions limited, Nelson Lancashire, England; Citrus Aurantium powder, Blackburn distributions limited, Nelson Lancashire, England.

Each component was measured using an electronic supplement scale and encapsulated in green, non-translucent, size zero gelatin capsules. All data were analyzed using the statistical software package SPSS SPSS, Version 24 for Mac, Chicago, IL.

In order to determine the effect size, the recommend guidelines of Quintana were used. Of the fourteen participants who volunteered for the study, four were removed due to adverse reactions to the phlebotomy procedure i.

Therefore, a total of ten physically active males completed the study. Participant characteristics can be seen in Table 1. Plasma Insulin. Blood Glucose. Means ± SD can be seen in Fig. Plasma Triglycerides. Means ± SD can be seen in Figs. Plasma Epinephrine.

Plasma Norepinephrine. No significant trial differences occurred in insulin, lactate or triglycerides throughout the ingestion period.

Under normal fasted conditions it is not uncommon to observe a slight decrease in blood glucose with concurrent decreases in insulin concentration over a prolong period of rest [ 17 , 18 ]. Blood glucose concentration following the PLA trial is reflective of this response, with a significant drop occurring at I2.

No changes in glucose concentration occurred and was found to be significantly higher than that of the PLA trial at the I2 time point. The medium by which the supplements were delivered in the current study were capsules absent of carbohydrate and would rationalize the difference in observations between the two studies.

Similar to glucose, insulin has been shown to be maintained or decrease during resting and fasted conditions [ 19 ]. This is in contrast to Graham et al. However, the differences in observations can likely be attributed to the dosage of caffeine Graham et al. The caffeine components role in sympathetic nervous system SNS mediated glucose release [ 22 ] may be another likely contributor to the observed glucose response.

Additionally, Stuart et al. The CA component of the complex is another mechanism by which the maintenance of blood glucose could have occurred. Specifically, the active ingredient p-synephrine acts on beta-3 receptors in order to increase lipolysis [ 1 ], thereby acting to spare blood glucose.

Future research should examine varying concentrations in order to determine a dose effect. The exhaustive exercise trial selected for this study was a repeated Wingate protocol designed to induce a high metabolic stress and fatigue. Following the completion of the trials, no differences in glucose, insulin, triglycerides, or catecholamines were observed.

However, insulin did not statistically elevate immediately post-exercise but demonstrated a non-statistical increase at the end of the recovery period. Previous research has demonstrated insulin spikes immediately following prolonged high-intensity protocols [ 25 ]; however, the duration of those protocols was ultimately longer than the one used previous studies and may have led to the different insulin response.

Though fat oxidation was not directly measured throughout this study, plasma triglycerides were obtained to determine changes in metabolic function. A primary function of the Citrus Aurantium is improved lipid peroxidation through p-synephrine and beta-3 activation, which may alter the release of triglycerides following exercise based on demand, and ultimately influence metabolic recovery.

Post-exercise plasma triglycerides have been shown to account for half of the delayed component of excess post exercise oxygen consumption EPOC [ 26 , 27 ], which is a beneficial response to high-intensity exercise. Interestingly, both trials showed spikes in plasma triglycerides at R1 when compared to I2, though no difference was observed between trials.

Furthermore, various dosages of this complex should be evaluated in order to better determine a dose-response effect. The markers used to examine metabolism were glucose, insulin, and triglycerides; future research should examine a more extensive metabolic profile including substrate utilization and free fatty acids.

Though a priori analysis based on a power of 0. However, this was not enough to elicit changes in resting insulin, or triglycerides. These findings suggest practical implications of hypoglycemic prevention during prolong i. Further research is needed to examine a dose and component response on these metabolic markers.

Stohs SJ, Preuss HG, Shara M. A review of the receptor-binding properties of p-synephrine as related to its pharmacological effects.

Oxid Med Cell Longev. Epub Aug 1. Ratamess NA, Bush JA, Kang J, Kraemer WJ, Stohs SJ, Nocera VG, Leise MD, Diamond KB, Campbell SC, Miller HB, et al.

The effects of supplementation with p-Synephrine alone and in combination with caffeine on metabolic, Lipolytic, and cardiovascular responses during resistance exercise.

J Am Coll Nutr. Article CAS Google Scholar. A review of the human clinical studies involving Citrus aurantium bitter orange extract and its primary protoalkaloid p-synephrine.

Int J Med Sci. The safety of Citrus aurantium bitter orange and its primary protoalkaloid p-synephrine. Phytother Res. Mohr M, Nielsen JJ, Bangsbo J.

Caffeine intake improves intense intermittent exercise performance and reduces muscle interstitial potassium accumulation. J Appl Physiol Goldstein ER, Ziegenfuss T, Kalman D, Kreider R, Campbell B, Wilborn C, Taylor L, Willoughby D, Stout J, Graves BS, et al.

International society of sports nutrition position stand: caffeine and performance. J Int Soc Sports Nutr. Article Google Scholar. Heckman MA, Weil J, Gonzalez de Mejia E. caffeine 1, 3, 7-trimethylxanthine in foods: a comprehensive review on consumption, functionality, safety, and regulatory matters.

J Food Sci. Evans SM, Griffiths RR. Caffeine tolerance and choice in humans. Robertson D, Wade D, Workman R, Woosley RL, Oates JA. Tolerance to the humoral and hemodynamic effects of caffeine in man. J Clin Invest. Zancheta R, Possi AP, Planeta CS, Marin MT. Repeated administration of caffeine induces either sensitization or tolerance of locomotor stimulation depending on the environmental context.

Pharmacol Rep. Sokmen B, Armstrong LE, Kraemer WJ, Casa DJ, Dias JC, Judelson DA, Maresh CM. Caffeine use in sports: considerations for the athlete. J Strength Cond Res. Medicine ACoS. ACSM's guidelines for exercise testing and prescription. Google Scholar. MacIntosh BR, Rishaug P, Svedahl K.

Assessment of peak power and short-term work capacity. Eur J Appl Physiol. Dill DB, Costill DL. Calculation of percentage changes in volumes of blood, plasma, and red cells in dehydration. J Appl Physiol. Kliszczewicz B, Bechke E, Williamson C, Bailey P, Hoffstetter W, McLester J, McLester C.

The influence of citrus aurantium and caffeine complex versus placebo on the cardiac autonomic response: a double blind crossover design. Quintana DS. Statistical considerations for reporting and planning heart rate variability case-control studies.

Garg S, Jovanovic L. Relationship of fasting and hourly blood glucose levels to HbA1c values: safety, accuracy, and improvements in glucose profiles obtained using a 7-day continuous glucose sensor.

Diabetes Care. Legro RS, Finegood D, Dunaif A. A fasting glucose to insulin ratio is a useful measure of insulin sensitivity in women with polycystic ovary syndrome.

J Clin Endocrinol Metab. CAS PubMed Google Scholar. Dekker MJ, Gusba JE, Robinson LE, Graham TE. Glucose homeostasis remains altered by acute caffeine ingestion following 2 weeks of daily caffeine consumption in previously non-caffeine-consuming males.

Br J Nutr. Graham TE, Sathasivam P, Rowland M, Marko N, Greer F, Battram D. Caffeine ingestion elevates plasma insulin response in humans during an oral glucose tolerance test.

Can J Physiol Pharmacol. Shi X, Xue W, Liang S, Zhao J, Zhang X. Acute caffeine ingestion reduces insulin sensitivity in healthy subjects: a systematic review and meta-analysis.

aurantium is easily applied in weight loss strategies 2 and, thus, contributes to the restoration of hunger and satiety balance regulation of blood glucose, insulin, and triglycerides 3.

P-synephrine has an affinity with β3-adrenergic receptors, seems capable of stimulating lipolysis without compromising cardiovascular activity at rest, unlike other substances e.

Recently, Guitiérrez-Hellín et al. aurantium supplementation could elevate fat consumption rates in submaximal aerobic exercise and, therefore, this has made C. aurantium a widely used substance to cut the levels of body fat. Nevertheless, there are no assessments of the cardiovascular safety of C.

aurantium in combination with aerobic submaximal exercise, and evidence regarding its use is rare but needed to guide clinical prescriptions that have C. aurantium as a therapeutic option. In this way, the analysis of cardiorespiratory parameters in combination with autonomic control of heart rate HR after physical exercise has been widely enforced to assess cardiovascular risk.

Through the scrutiny between heartbeats RR intervals or HR variability HRV , it is possible to study the efferent flow of sympathetic and parasympathetic autonomic to the heart. During exercise, there is a vagal or parasympathetic withdrawal and, then, there is an upsurge in sympathetic modulation to the heart, revealing a surge in HR and cardiac contractility.

Upon cessation of exercise, it is expected that there will be a fast reactivation of vagal modulation, which will provide an abrupt reduction and recovery in HR 6.

Recent studies have fixated on examining whether nutritional interventions e. Based on these aforementioned considerations, it was probed as to whether the supplementation of C. aurantium prior to aerobic physical exercise could impact the autonomic control of HR and interfere with cardiovascular recovery following exercise.

We assume that C. aurantium would not affect the recovery of cardiovascular and autonomic parameters after exercise. Given these declarations, we intended to assess the effect of C. aurantium supplement on cardiovascular recovery and autonomic constraints after submaximal aerobic exercise.

This is a randomized study, double-blind, placebo-controlled crossover clinical trial. According to the Declaration of Helsinki, the intervention protocols were approved by the Research Ethics Committee Institutional—Brazil Process: The register of study details on Clinicaltrials.

We recruited 17 male subjects via social media e. to participate in the study. Participants were between 18 and 30 years of age, had a body mass index BMI between Through screening, we studied the presence of some conditions that would make them ineligible to participate in the study, for instance, smoking, present and past anabolic steroid usage, cardiorespiratory, neurological or musculoskeletal disorders, use of pharmacotherapies that affect the autonomic nervous system.

At the end of the study, the sample consisted of 12 subjects Figure 1. Additionally, baseline values of heart rate beats per minute , systolic blood pressure SBP , and diastolic blood pressure DBP mmHg were logged Table 1. Table 1. Mean values followed by their respective standard deviations minimum and maximum of age, mass, height, BMI, heart rate, SBP and DBP.

The intervention protocols were split into three phases, with an interval of 48—72 h between each protocol to provide time for the participants' physical recovery.

On the first day, an initial interview was completed with the participating candidates in the study. After screening, eligible applicants were provided with a list of guidelines to abstain from certain citrus fruits mandarin, sweet, and bitter orange , alcoholic and caffeinated beverages, or nutriments coffee, sports drinks, chewing gum, chocolate , and exercise 24 h prior to the ensuing study sessions.

Participants were told to have a light meal e. Through a random sequence, in the second step, participants were randomized to consume a capsule containing mg C.

This amount was selected as it is regularly applied in clinical practice In the final stage, the participants received the protocol contrary to the previous one to safeguard the study's crossover. An independent researcher who did not participate in the data logging was responsible for randomizing the interventions, choosing the capsules, and assigning them to the investigator.

The capsules were opaque and visibly identical; neither the participant nor the investigator could recognize the capsules' contents. The capsules were attained in commercial form from a reliable provider Florien Fitoativos ® Ltd. Naringin and hesperidin concentrations were not analyzed. The participants persisted for 20 min walking at this speed, and at the end of the activity, the participants were once more seated and monitored for an additional 60 min 8.

SBP and BPD measurements were completed indirectly using a stethoscope Littman Classic II, Saint Paul, USA and aneroid sphygmomanometer Welch Allyn Tycos, New York, USA on the participants' left arm For HR and HRV indices scrutiny, cardiac activity was logged beat by beat throughout the data logging technique with a sampling rate of 1 kHz using a Polar ® heart rate monitor model RSCX.

The HR and HRV recordings were logged at the following epochs: Rest R1: 90th to 95th min of resting after capsule ingestion , and all through exercise recovery: 0 to 5th min; 5th to 10th min; 15th to 20th min; 25th to 30th min; 35th to 40th min; 45th to 50th min, and; 55th to 60th min Figure 2.

Series with regular heartbeats R-R intervals were required to make the HRV indices, as recommended by the Task Force of the European Society of Cardiology and the North American Society of Pacing and Electrophysiology In these series, digital and manual filters were executed to remove artifacts.

After collection, the RR intervals were exported to the software program Kubios ® HRV Analysis to produce the linear indices of the frequency domain and time domain Frequency domain analysis was accomplished via spectral analysis by means of the Fast Fourier Transform FFT to cause the high frequency HF index with a sampling rate of 0.

The non-linear HRV analysis was achieved using the PyBios ® software Biomedical Signal Analysis in Python Version 1. We dispersed the number of RR intervals through six levels 0—5 , transforming them into a spatial methodology; a sequence of three symbols. All patterns were independently assembled into two clusters, according to the number and type of variation between symbols:.

A pilot study conducted with six participants performed the sample size calculation. We applied the root mean square of successive differences between RR intervals in the online software at www. br , which provided the magnitude of the difference.

We measured a standard deviation of The Shapiro-Wilk statistical test was enforced to assess data normality. For the cardiovascular recovery and autonomic reactivity analysis during the experimental protocols Rest vs.

recovery , One-way analysis of variance ANOVA1 for repeated measures and the Bonferroni post-test was enforced when the assumption of data normality was attained. Friedman's test followed by Dunn's post-test was required for data that did not acquire a normal distribution. Cohen's d calculated effect sizes to measure the magnitude of changes for significant differences.

Assessments were achieved using Statistical Package for the Social Sciences SPSS IBM ® SPSS Statistics v. The descriptive data of twelve healthy males that met the study criteria are included in Table 1. These datasets strengthen the homogeneity of our sample.

The HR recovery analysis revealed no significant differences between the protocols. In the placebo protocol, the comparison of resting and after exercise established an increase in HR from 0 to 5th min of recovery Rest vs.

In the C. aurantium protocol, the same results were attained, and HR values remained significantly enlarged from 0 to 5th min of recovery Rest vs. Table 2. No significant changes were identified in the C. aurantium intervention during the recovery analysis rest vs.

recovery for DBP, MAP, and PP. Only SBP demonstrated significant changes in 1 min following exercise Rest vs.

aurantium protocol. During the placebo protocol, SBP remained significantly higher during 3 min of recovery compared to rest Rest vs. Table 3. Time and frequency domain indices in addition to non-linear analyzes revealed that autonomic heart rate recovery occurred more quickly in the C.

aurantium protocol compared to the placebo protocol. In the placebo protocol, the investigation of recovery rest vs. recovery of the HF index revealed that its values remain depressed throughout 10 min of recording after exercise Rest: Figure 3.

In the placebo protocol, pNN50 index values continued to be significantly decreased throughout 20 min of recovery related to resting values Rest: Our findings demonstrate that the ingestion of C.

aurantium p-synephrine mg prior to exercise fast-tracks the fall in SBP after physical exertion. Earlier studies propose that one of the benefits of using C. aurantium equated to other adrenergic substances e.

Activation of β-3 adrenergic receptors triggers reverse inotropic effects, antagonizing the activation of further classes of adrenoreceptors β-1 and β-2 in cardiac tissue and, thus, decreasing sympathetic modulation to the heart.

This clarifies why overall, the binding of p-synephrine with β-3 adrenergic receptors does not increase BP or HR, displaying cardioprotective effects In this study, in the placebo intervention, for the spectral analysis, the HF index, representative of parasympathetic modulation, needed 10 min after termination of exercise to recover.

aurantium protocol, we did not find substantial changes in the HF index in exercise recovery vs. Analogous deviations occurred in the pNN50 index and were reduced 20 min after cessation of exercise in the placebo protocol.

While in the protocol with C. aurantium , this index continued to be reduced for only 10 min after exercise. aurantium protocol, transformations were only following 5 min of recovery. However, in the C.

aurantium protocol, the values were only meaningfully reduced for 5 min after the cessation of exercise.

These observations make C. aurantium a safe nutritional compound to be applied during exercise, which supports the recovery of autonomic parameters following exercise. Since a slow post-exercise autonomic recovery is linked with an increased cardiovascular risk 25 , the results of our study indicate that C.

aurantium compounds have a potential preventive role on the onset of cardiovascular complications in physical exercise.

As caffeine and C. aurantium are frequently sold as complementary formulas for use in humans, preceding studies have assessed the effects of using these substances alone and in combination.

Through a randomized clinical trial, Guitiérrez-Hellín et al. aurantium alone or in combination with caffeine would have different results for fat utilization during aerobic physical exercise. No superiority was found between C.

aurantium alone and combined with caffeine on the total values of fat consumption during the physical exercise session, while both interventions were superior to the placebo treatment. This supports the isolated use of C. aurantium an alternate way to be applied as an adjunct in cutting body fat without inducing cardiac risk.

In the study by Guitiérrez-Hellín et al. aurantium isolated supplement. In contrast, the HR and SBP were significantly higher when caffeine was included in the formulation. Our study achieved no changes for HR, and SBP was lessened more quickly following exercise. The identification of β-3 adrenoreceptors in cardiovascular tissues posed challenges to the paradigm of sympathetic regulation by β-1 and β-2 adrenoceptors.

The binding response of p-synephrine to the β-3 receptor may elucidate why no increase in HR or BP is detected when C. aurantium is enforced alone. In contrast, when C.

aurantium is combined with caffeine in dietary supplements, it is capable of affecting these parameters, particularly in caffeine-sensitive individuals It has been revealed that the combination of these substances promotes a significant increase in the concentration of plasma catecholamines e.

The study by Kliszczewicz et al. aurantium upsurges sympathetic modulation to the heart throughout rest and corroborates the increases in HR and SBP achieved in the study by Guitiérrez-Hellín et al.

It is assumed that caffeine alone can increase HR during physical exercise Despite that, a recent meta-analysis demonstrated that caffeine could not delay vagal return to the heart after exercise, evaluated by the HF and root mean square of successive differences between RR intervals RMSSD indices Equally, Kliszczewicz et al.

aurantium combined. Caffeine and C. aurantium combination have no extra effects on exercise fat utilization 5. These substances appear to exhibit the opposite cardiovascular effects and, thus, caffeine seems to overlap the beneficial effects of the isolated use of C.

aurantium on cardiovascular health. In this study, C. aurantium supplementation alone optimized the recovery of SBP and HRV indices after exercise. The nutritional characteristics demonstrated in the flavonoids e. aurantium perform antioxidant and anti-inflammatory activities, which are partly answerable for accelerating the return of parasympathetic control of heart rate seen by vagal indices of HRV.

Such properties can hasten the removal of metabolites produced by physical exercise, restoring baroreflex sensitivity and decreasing metaboreflex activation more quickly at the end of physical exercise While C. aurantium exhibited cardioprotective effects, it is essential to be careful with its usage.

Bui et al. Yet, in other studies that enforced doses beneath mg in an acute 5 , 30 , 31 and chronic for 15 days 32 form, no changes were achieved for the HR, SBP, and DBP values, nor electrocardiographic disturbances.

Likewise, our results do not support the findings of Bui et al. The results from the study of Ratamess et al. In your results, the p-synephrine supplementation mg did not evoke changes in HR before, during, and following resistance exercise unless mg of caffeine was added to the formulation.

The same occur in the rest situation, in another study by Ratamess et al. The study of Bui et al. Although it is a randomized and crossover study, there is a lack of information about allocation order in the study. aurantium, and provoked adjustments in blood pressure, because of higher sweet and fat content e.

Furthermore, the authors did not report guarantees that snack was equal on the others evaluation days. Bitter orange caused cardiovascular effect was only observed based on statistical adjustments.

A difference was seen compared to placebo but not when compared to baseline. All these factors raise questions about the validity of their conclusions. The results recognized in our analyses will advance health professionals' conduct who work with the prescription of nutritional supplements.

Consequently, it may be an alternative way to replace other compounds that demonstrate similar contributions regarding fat utilization during exercise but that promote unwanted cardiovascular effects e. Our study highlights important points about the study population, given that it is restricted to healthy and physically active males.

Notwithstanding the number of participants having exceeded the sample size calculation, the final sample is considered small.

With the desire to improve body composition. In spite of this, these facts do not allow these results to be extrapolated to other populations and, therefore, further research with obese individuals is needed to confirm the safety of using C. aurantium in combination with exercise.

For the time being, we prefer to use a healthy population free from metabolic disorders to prevent possible adverse events from C.

aurantium supplementation.