Orange Juice, Hesperidin and Their Role in Vascular Health Benefit

Study Overview
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Brief Summary
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Although epidemiological studies have associated the consumption of sugary beverages with adverse health effects, experimental studies have demonstrated that the metabolic response of the human body to fruit juice as compared to artificial beverages is substantially different. Fruit juices do not just provide sugars and related calories, but they are rich sources of bioactive compounds especially of flavonoids. Flavanones constitute a class of flavonoids that are specifically and abundantly found in citrus fruits, with hesperidin being the major compound in orange. From prospective cohort studies, higher intakes of flavanones are associated with a lower incidence of mortality by cardiovascular disease (CVD). This relation is supported by results from a number of animal studies demonstrating a slowdown in atherosclerosis development and vascular protective effects in dietary interventions with flavanones. Randomized, controlled clinical trials to corroborate the suggested vasculo-protective effects of orange juice presumably mediated by the flavanones are scarce and available data do not allow to draw firm conclusions about their efficacy. To fill this gap, the HESPER-HEALTH study conducted in humans will assess the vascular protective effects of 100% orange juice consumption and evaluate the contribution of hesperidin in these effects.

Detailed Description
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This human dietary intervention study is a double blind, randomized, placebo controlled, cross over trial with 3 arms, carried out on subjects with predisposition to cardiovascular diseases (CVD) based on age and overweight. This study aims to demonstrate the vascular protective effects (with Flow Mediated Dilatation (FMD) as main criteria) of the consumption of a flavanone rich orange juice or of orange flavanones by comparison with a control sugary drink alone. The 42 recruited participants will receive the 3 drinks in a random order. For each subject, the study is divided into 3 identical experimental periods of 45 days (period 1,2,3): including 3 days prior to the beginning of the product intake, during which specific dietary guidelines, samplings and measures will be asked to be performed at home followed by a 6 weeks period of consumption of each of the 3 beverages). A period of 4 to 6 weeks of wash-out is planned between each experimental period. To summarize: Visit 1 (D-14) = inclusion, Visit 2 (D1: baseline) to 3 (D42) = period 1, Visit 3 (D42) to 4 (D70) = wash out 1, Visit 4 (D70) to 5 (D111) = period 2, Visit 5 (D111) to 6 (D139) = wash out 2, Visit 6 (D139) to 7 (D180) = period 3. The wash-out periods (minimum duration: 4 weeks) may be extended until 6 weeks for the convenience of participants. The protocol includes a total of 7 visits to PIC/CIC Inserm 1405 of the Clermont-Fd University Hospital. The total duration of the study will be between 28 and 34 weeks

Official Title
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Orange Juice, Hesperidin and Their Role in Vascular Health Benefit: a Human Double Blind, Randomized, Controlled, Cross Over Study

Conditions
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Metabolic Syndrome, Vascular Compliance, Predisposition to Cardiovascular Disease

Intervention / Treatment
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* Behavioral: Beverage consumption


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Man or post-menopausal woman ; 40-65 years old (inclusive) ; Body Mass Index (BMI)≤ 30 ; Waist circumference ≥80 cm for women, and ≥94 cm for men ; Weight > 46 kg Normal biological balance sheet or considered normal by the investigator No aversion or intolerance to citrus foods ; Accept to limit their total intake of flavonoid rich beverages (tea, coffee, cocoa, wine, fruit juice) to 250 mL/day ; Ability to give informed consent to participate in research ; Willingness to accept randomization and undergo the testing and intervention procedures and deliver stool, blood and urine samples for testing ; Affiliation to Social Security. Exclusion Criteria: Treated pre-diabetic or diabetic ; Treated for hypertension ; Use of statins or other medications for lowering cholesterol ; Treated with antibiotics, antifungals, probiotics or prebiotics in the 3 months before the enrolment ; Menopausal hormone replacement therapy ; Diagnosed gastrointestinal illness in the judgement of the investigator ; Any serious medical condition that precludes safe participation in the study, such as coronary artery disease, peripheral vascular disease, stroke, congestive heart failure, chronic obstructive pulmonary disease, insulin-dependent diabetes, psychiatric disease, renal disease, liver disease, active cancer and anemia ; History of eating disorders such as bulimia nervosa, anorexia nervosa and severe binge eating disorder in the last 5 years ; Digestive disorders with diarrhea during the 3 months preceding the beginning of the study ; Self-declared vegetarian, vegetalian, vegan ; History of substance abuse or alcohol abuse ; Involvement in a weight loss intervention program (including anti-obesity medication) within the past 3 months or who have had bariatric surgery ; Current smokers (within the last 30 days) ; Use of dietary supplements (vitamins, antioxidants) currently or in the past one month ; Strenuous exercise greater than 6 hours per week ; Anyone who in the opinion of the investigator is unlikely to be able to comply with the protocol ; Subjects involved in another clinical trial or being in the exclusion period of another study or having received a total compensation greater than 4,500 euros over the 12 months preceding the start of the trial ; Subject benefiting from a legal protection measure (curatorship, guardianship, safeguard of justice) ; Refusal to participate.

Ages Eligible for Study
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Minimum Age: 40 Years
Maximum Age: 65 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Basic Science
Allocation: Randomized
Intervention Model: Crossover Assignment
Masking: Quadruple


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: Orange juice (A)<br>42 subjects between 40 and 65 years old with predisposition to cardiovascular disease will consume daily 330 ml of orange juice naturally rich in hesperidin (drink A) during 6 weeks | Behavioral: Beverage consumption<br>* Volunteers will consume, in random order, daily 330 ml of 1 experimental beverage per period (Orange Juice, Control Beverage, Control Beverage supplemented with hesperidin) for 6 weeks in each period. At the beginning and the end of each period, exploration will be conducted at fasted state and at post-prandial state after the administration of a high-fat high-sugar meal.<br>|
| Placebo Comparator: Control beverage (B)<br>42 subjects between 40 and 65 years old with predisposition to cardiovascular disease will consume daily 330 ml of control beverage (drink B)- a soft drink with sugar concentration identical to drink A - during 6 weeks | Behavioral: Beverage consumption<br>* Volunteers will consume, in random order, daily 330 ml of 1 experimental beverage per period (Orange Juice, Control Beverage, Control Beverage supplemented with hesperidin) for 6 weeks in each period. At the beginning and the end of each period, exploration will be conducted at fasted state and at post-prandial state after the administration of a high-fat high-sugar meal.<br>|
| Experimental: Control beverage supplemented with hesperidin formulation (B+HESP)<br>42 subjects between 40 and 65 years old with predisposition to cardiovascular disease will consume daily 330 ml of control beverage (identical to drink B but supplemented with hesperidin to reach level of drink A) during 6 weeks | Behavioral: Beverage consumption<br>* Volunteers will consume, in random order, daily 330 ml of 1 experimental beverage per period (Orange Juice, Control Beverage, Control Beverage supplemented with hesperidin) for 6 weeks in each period. At the beginning and the end of each period, exploration will be conducted at fasted state and at post-prandial state after the administration of a high-fat high-sugar meal.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 1 | 
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 42 | 
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 70 | 
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 111 | 
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 139 | 
| Brachial artery Flow Mediated Dilation (FMD) | The endothelial function will be assessed using the non-invasive ultrasound technique of flow mediated dilatation of the brachial artery. FMD measure is the percentage of dilation of brachial artery in response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery in fasted state | Day 180 | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Sex | Participant sex (man/women) will be requested. | Day -14 | 
| Age | Participant age (years) will be requested. | Day 34 | 
| Basal Systolic Blood Pressure | Blood pressure measure (mm Hg) with monitor. | Day -14 | 
| Basal Heart Rate | Heart rate measure (beat/min) with monitor. | Day -14 | 
| D-3 visit systolic Blood Pressure (BP) | Self-monitored blood pressure (mmHg) at home with tensiometer, at home the morning, 3 days before the visit, in fasted state. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| D-3 visit diastolic BP | Self-monitored blood pressure (mmHg) at home with tensiometer, at home the morning, 3 days before the visit, in fasted state. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| D-3 visit Heart Rate | Self-monitored heart rate (beat/min) at home with tensiometer,at home the morning, 3 days before the visit, in fasted state. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| FMD post prandial endothelial response 3h after a challenge meal | Assessment of the postprandial endothelial response to a challenge meal (900kcal, fresh cream, sucrose and milk proteins) by measuring FMD (percent) using ultrasound technique, 3h after intake of the full daily dose of study products concomitantly with challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| FMD post prandial endothelial response 6h after a challenge meal | Assessment of the postprandial endothelial response to a challenge meal by measuring FMD (percent) using ultrasound technique, 6h after intake of the full daily dose of study products concomitantly with challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Arterial compliance assessment | Noninvasive measure with Sphygmocor (AtCor Medical Pty. Ltd) of pulse transit time between carotid artery and femoral artery. The carotid-femoral Pulse Wave Velocity (PWV) (m/sec) is an established index of arterial stiffness,in fasted state. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Rest flow by Flowmetry Laser Doppler (FLD) in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the rest flow using laser-Doppler system at the level of the skin of the hand, in fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Occlusion area by FLD in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the occlusion area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement) in fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area by FLD in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement) in fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area / occlusion area ratio by FLD in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the ratio hyperaemia area / occlusion area determined by FLD. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Maximal flow by FLD in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the maximal flow using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), in fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia half time by FLD in fasted state | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia half time using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), in fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Rest flow by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the rest flow using laser-Doppler system at the level of the skin of the hand,3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Occlusion area by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the occlusion area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area / occlusion area ratio by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the ratio hyperaemia area/ occlusion area determined by FLD, 3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Maximal flow by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the maximal flow using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia half time by FLD 3h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia half time using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 3h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Rest flow by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the rest flow using laser-Doppler system at the level of the skin of the hand, 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Occlusion area by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the occlusion area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia area using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia area / occlusion area ratio by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the ratio hyperaemia area / occlusion area ratio determined by FLD, 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Maximal flow by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the maximal flow using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hyperaemia half time by FLD 6h after a challenge meal | Vascular endothelial function in the micro-vascular compartment will be assessed using the measurement of the hyperaemia half time using laser-Doppler system at the level of the skin of the hand by following the response to a reactive hyperaemia induced by the release of a transient occlusion of the brachial artery (same stimulus as for FMD measurement), 6h after intake of the full daily dose of study products concomitantly with challenge meal | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin concentration in 24h urine | Concentration of hesperetin in urine (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin concentration in plasma on fasted state | Concentration of hesperetin in plasma (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin catabolites concentration in plasma on fasted state | Concentration of hesperetin microbial catabolites in plasma (nM) will be measured, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin concentration in plasma on 3h post prandial test | Concentration of hesperetin in plasma (mM) will be measured, in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin catabolites concentration in plasma on 3h post prandial test | Concentration of hesperetin microbial catabolites in plasma (mM) will be measured, in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin concentration in plasma on 6h post prandial test | Concentration of hesperetin in plasma (mM) will be measured, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Hesperetin catabolites concentration in plasma on 6h post prandial test | Concentration of hesperetin microbial catabolites in plasma (mM) will be measured, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin concentration in 24h urine | Concentration of naringenin in urine (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin catabolites concentration in 24h urine | Concentration of naringenin microbial catabolites in urine (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin concentration in plasma on fasted state | Concentration of naringenin in plasma (nM) will be measured, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin catabolites concentration in plasma on fasted state | Concentration of naringenin microbial catabolites in plasma (nM) will be measured, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin concentration in plasma on 3h post prandial test | Concentration of naringenin in plasma (mM) will be measured, in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin catabolites concentration in plasma on 3h post prandial test | Concentration of naringenin microbial catabolites in plasma (mM) will be measured, in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin concentration in plasma on 6h post prandial test | Concentration of naringenin in plasma (mM) will be measure, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Naringenin catabolites concentration in plasma on 6h post prandial test | Concentration of naringenin microbial catabolites in plasma (mM) will be measured, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone concentration in 24h urine | Concentration of flavanone in urine (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone catabolites concentration in 24h urine | Concentration of flavanone microbial catabolites in urine (nM) will be measured. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone concentration in plasma on fasted state | Concentration of flavanone in plasma (nM) will be measured, on fasted state. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone catabolites concentration in plasma on fasted state | Concentration of flavanone microbial catabolites in plasma (nM) will be measured, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone concentration in plasma on 3h post prandial test | Concentration of flavanone in plasma (mM) will be measured, in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone catabolites concentration in plasma on 3h post prandial test | Concentration of flavanone microbial catabolites in plasma (mM) will be measured,in post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone concentration in plasma on 6h post prandial test | Concentration of flavanone in urine (nM) will be measured, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Flavanone catabolites concentration in plasma on 6h post prandial test | Concentration of flavanone microbial catabolites in plasma (mM) will be measured, in post prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma nitrites dosage | Determination of nitrite plasma concentration (nM) (a biomarker of endothelial activation), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma nitroso-thiols dosage | Determination of nitroso-thiols plasma concentration (nM) (a biomarker of endothelial activation), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma Inter-Cellular Adhesion Molecules (ICAM) dosage on fasted state | Determination of ICAM plasma concentration (ng/ml) (a biomarker of endothelial activation), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma ICAM dosage on 6h post prandial state | Determination of ICAM plasma concentration (ng/ml) (a biomarker of endothelial activation), on 6h post prandial state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma Vascular-CAM (VCAM) dosage on fasted state | Determination of VCAM plasma concentration (ng/ml) (a biomarker of endothelial activation), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma VCAM dosage on 6h post prandial state | Determination of VCAM plasma concentration (ng/ml) (a biomarker of endothelial activation), on 6h post prandial state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma e-selectin dosage on fasted state | Determination of e-selectin plasma concentration (ng/ml) (a biomarker of endothelial activation), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma e-selectin dosage on 6h post prandial state | Determination of e-selectin plasma concentration (ng/ml) (a biomarker of endothelial activation), on 6h post prandial state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma Extracellular Vesicles analyses (EVs) on fasted state | EVs (a biomarker of endothelial activation) will be isolated from platelet-poor plasma samples and analyzed, on fasted state | Day 42, Day 111, Day 180 | 
| EVs analyses on 3h post prandial state | EVs (a biomarker of endothelial activation) will be isolated from platelet-poor plasma samples and analyzed, on post prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 42, Day 111, Day 180 | 
| Plasma oxylipins identification on fasted state | Determination of oxylipins present in plasma (a biomarker of inflammation and oxidative stress) using a method of profiling, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma oxylipin concentration on fasted state | Determination of oxylipin plasma concentration (a biomarker of inflammation and oxidative stress), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma oxylipins identification on 6h post prandial state | Determination of oxylipins present in plasma (a biomarker of inflammation and oxidative stress) using a method of profiling, on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 42, Day 111, Day 180 | 
| Plasma oxylipin concentration on 6h post prandial state | Determination of oxylipin plasma concentration (a biomarker of inflammation and oxidative stress), on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 42, Day 111, Day 180 | 
| Plasma Interleukin 6 (IL-6) dosage | Determination of IL-6 plasma concentration (pg/ml) (a biomarker of inflammation and oxidative stress), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma Tumor Necrosis Factor α (TNFα) dosage | Determination of TNFα plasma concentration (pg/ml) (a biomarker of inflammation and oxidative stress), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma high-sensitivity C-reactive protein (hs-CRP) dosage | Determination of hs-CRP plasma concentration (mg/L) (a biomarker of inflammation and oxidative stress), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma glucose dosage on fasted state | Determination of glucose plasma concentration (mM) (a metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma glucose dosage on 3h post prandial state | Determination of glucose plasma concentration (mM) (a metabolic parameter), on post-prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma glucose dosage on 6h post prandial state | Determination of glucose plasma concentration (mM) (a metabolic parameter), on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| -Plasma Triacylglycerol (TAG) dosage on fasted state | Determination of TAG plasma concentration (g/L) (a metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma TAG dosage on 3h post prandial state | Determination of TAG plasma concentration (g/L) (a metabolic parameter), on post-prandial state 3h after the concomitant consumption of the study drink and challenge meal, on post-prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma TAG dosage on 6h post prandial state | Determination of TAG plasma concentration (g/L) (a metabolic parameter), on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma uric acid dosage on fasted state | Determination of uric acid plasma concentration (mM) (a metabolic parameter) method, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma uric acid on 3h post prandial state | Determination of uric acid plasma concentration (mM) (metabolic parameter), on post-prandial state 3h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma uric acid on 6h post prandial state | Determination of uric acid plasma concentration (mM) (metabolic parameter) will be determined, on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma total cholesterol dosage | Determination of total cholesterol plasma concentration (mM) (a metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma High Density Lipoprotein cholesterol (HDL-chol) dosage | Determination of HDL-chol plasma concentration (mM) (a metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma High Density Lipoprotein cholesterol (LDL-chol) calculation | Determination of LDL-chol plasma concentration (mM) (metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma Total Fatty Acids (FA) dosage | Determination of total FA plasma concentration (mM) (a metabolic parameter), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Serum Insulin dosage | Determination of insulin serum concentration (mU/L or pM) (a metabolic parameter), on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Weight measure | the body weight (kg) will be recorded with a bathroom scale, on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Height measure | the body height (cm) will be measured with a stadiometer. | Day -14 | 
| Body Mass Index (BMI) calculation | the BMI (kg/m²) will be calculated, on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Waist circumference measure | the waist circumference (cm) will be recorded with a measuring tape, on fasted state | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Fat mass ratio determination | The percentage of fat mass (percent) (body composition) will be determined on each participant using a multi-frequency bioelectrical Impedance Analyzer, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Lean mass ratio determination | The percentage of lean mass (percent) (body composition) will be determined on each participant using a multi-frequency bioelectrical Impedance Analyzer, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Water mass ratio determination | The percentage of water (percent) (body composition) will be determined using a multi-frequency bioelectrical Impedance Analyzer, on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| RNA profiling on fasted state | Nutrigenomic analysis will be performed from total Ribo Nucleic Acid (RNA) isolated blood collected in PAXgene Blood RNA Tube. The isolated RNA will be used to perform microarray analyses that allow identification of expression of all genes of the genome, on fasted state | Day 42, Day 111, Day 180 | 
| RNA profiling on 6h post prandial state | Analysis will be performed from total RNA isolated blood collected in PAXgene Blood RNA Tube. The isolated RNA will be used to perform microarray analyses that allow identification of expression of all genes of the genome, on post-prandial state 6h after the concomitant consumption of the study drink and challenge meal. | Day 42, Day 111, Day 180 | 
| Gut microbiota profiling | Identification of the microbiota composition of feces samples (collected by subjects) by performing a genetic sequencing analysis of bacterial DNA. | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma carotenoids dosage | Carotenoids will be quantitated from plasma (nM), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Plasma vitamine C dosage | Vitamin C status will be quantified in deproteinized plasma (mg/L), on fasted state | Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 
| Treatment compliance | Treatment compliance (percent) will be determined using counting of empty orange drinks brought back by volunteers after the consumption periods | Day 42, Day 111, Day 180 | 
| Polyphenol intake | Polyphenol intake (ml/day) in basal and during treatment consumption periods will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Low-polyphenol diet compliance | Respect (y/n) of a low-polyphenol diet in basal (between Visit 1 - Visit 2) and during treatment consumption periods (between Visit 2 - Visit 3, Visit 4 - Visit 5, Visit 6 - Visit 7) will be determined by a dietician using 3 days food reports completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Calorie intake | Calorie (kcal/day) intake in basal and during treatment consumption periods will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Protein intake | Protein (g/day) intake in basal and during treatment consumption periods will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Lipid intake | Lipid intake (g/day) in basal and during treatment consumption periods will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Carbohydrate intake | Carbohydrate intake (g/day) in basal and during treatment consumption periods will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Diet division of protein/lipid/carbohydrate intakes | The overall distribution of the protein/lipid/carbohydrate food intakes (%) will be determined by a dietician using food report completed by volunteers. | Day 1, Day 42, Day 111, Day 180 | 
| Diet stability | Diet stability (y/n) in terms of polyphenols, calories, proteins, carbohydrates and lipids intakes during the whole study will be determined by a dietician using the 4 food reports completed by volunteers. | Day 180 | 
| Biobank for food metabolome | Urine samples will be stored for further assessment of food metabolome to potentially identify new bioactive compounds present in the juice that could contribute to the biological response after orange juice/HESP intake. | Day -14, Day 1, Day 42, Day 70, Day 111, Day 139, Day 180 | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Cardiovascular disease prevention, Endothelial dysfunction, Orange juice, Flavanone, Hesperidin, Nutrition Assessment, Nutritional Physiological Phenomena