Impacts of Intermittent Fasting on Energy Balance and Associated Health Outcomes

Obesity Clinical Trial

Official title:

Impacts of Intermittent Fasting in Adults on Energy Balance, Body Composition, Postprandial Hormone Profiles and Gene Expression in Adipose Tissue

Clinical Trial Details — Status: Completed

Administrative data

• NCT number: NCT02498002
• Other study ID #: IMF-02
• Status: Completed
• Phase: N/A
• Start date: May 2015
• Est. completion date: September 28, 2018

Study information

• Verified date: April 2023
• Source: University of Bath
• Contact: n/a
• Is FDA regulated: No
• Study type: Interventional

Clinical Trial Summary

Obesity is a global concern with links to conditions such as diabetes. Historically, these conditions have been managed by reducing energy intake on a daily basis, which is often hampered by low adherence rates and compensatory reductions in physical activity. Intermittent fasting (IMF) has become popular as an alternative method of weight loss, where periods of normal eating are punctuated by days of fasting. However, despite its popularity, IMF is scarcely examined within the literature.

This study will examine the impact of IMF on energy balance and the associated health outcomes relating to conditions such as diabetes. To achieve this, 36 lean adults and 36 overweight/obese adults will be recruited, excluding those with conditions which may be adversely affected by fasting. Once enrolled, participants will complete a one month control phase, incorporating 12 days of diet and activity monitoring, to ensure their weight is stable before moving in to the intervention phase.

The intervention phase will begin with a lab protocol designed to examine body composition, resting metabolic rate and metabolic responses to two successive meals. Once completed, participants will be randomised to one of three diets which are listed below. Comparing the changes seen with diets 1 and 2 will provide the first direct comparison of IMF against more conventional strategies, while diet 3 will allow us to ascertain the role of weight loss in mediating any effects of IMF.

1. Daily calorie restriction - Reduce energy intake by 25% on a daily basis

2. IMF with weight loss - Alternate between 24 hour cycles of fasting and feeding with 150% of normal intake when feeding

3. IMF without weight loss - Alternate between 24 hour cycles of fasting and feeding with 200% of normal intake when feeding

All three diets will last for 20 consecutive days, with transitions between each 24 hour diet cycle occurring at 15:00 each day to allow at least one main meal per day. When fasting, participants will only be permitted water and black tea/coffee to eliminate energy intake. Once again physical activity and diet will be monitored for 12 days during the intervention to examine changes in energy expenditure and ensure compliance.

Having completed all 20 diet cycles participants will return to the laboratory to repeat the protocol described earlier. Comparing these results against the values seen before the intervention will then allow the effects of IMF to be identified and compared.

Description:

Recruitment

Local advertising in the form of posters will distributed throughout the research institution, nearby organisations and the city of Bath. The posters will encourage interested persons to contact the chief investigator to receive more information on what the study will involve. The research team will be looking to recruit 72 (36 lean, 36 overweight/obese) individuals who satisfy the entry criteria (discussed later).

Enrolment

Potential volunteers will use the contact details provided in advertising materials to express their interest to the research team who will then provide them with a copy of the participant information sheet to read through at their convenience. Those interested in participating will then be invited to a pre-enrolment consultation where they will be given the opportunity to ask any questions they may have about the study before being screened for the inclusion and exclusion criteria. This will involve measurements of weight and height, as well as the completion of an entry criteria questionnaire, the Eating Disorder Examination Questionnaire and the Physical Activity Readiness Questionnaire. Providing they satisfy the relevant criteria they will be talked through the study and before being asked to complete an informed consent form. As part of this, the procedure for an adipose tissue biopsy will be discussed with the participant and they will be asked to decide whether or not they would like to include this as it is an optional test.

Baseline Lab Protocol (LAB 1)

Having formally enrolled in the study, participants will be invited to begin their involvement in the study with a 60 minute visit to the University of Bath following a 12 hour fast. During this visit, the researcher will obtain a urine sample to check hydration levels using an Osmometer, before taking measurements of height, weight and waist circumference. A fasting blood sample will also be obtained using venepuncture to measure resting levels of key hormones and metabolites. To conclude this visit participants will be asked to perform a submaximal exercise test to calibrate the 'Actiheart' monitor being used throughout the study to measure physical activity energy expenditure. For this, participants will be asked to remain on a treadmill as the intensity increases over the following four stages:

• Stage 1 - level walking at 3.2 km/h

• Stage 2 - brisk walking at 5.2 km/h

• Stage 3 - brisk walking at 5.8 km/h with a steep gradient of +10.2%

• Stage 4 - level running at 12.5 km/h

Stage 4 will only be included for those who are formal exercisers, as non-exercisers are unlikely to be doing this kind of activity in their normal routine rendering a calibration at this intensity unnecessary. Formal exercisers will be identified using the entry criteria questionnaire. In addition, the treadmill protocol will be stopped earlier if the participant asks to stop or if their heart rate exceeds 85% of their age-predicted maximum.

Monitoring Phase

Before leaving the lab, participants will be provided with an Actiheart monitor, a small device applied to the chest which measures physical activity by recording heart rate and movement every minute. They will also be provided with a set of kitchen scales and a log book to create a weighed record of their food and fluid intake and a set of body weight scales to track their weight and help keep it stable.

During the four week monitoring phase, participants will be asked to complete four 3-day monitoring periods, each comprising two weekdays and one weekend day whilst they continue in their normal dietary and exercise habits. The researchers will arrange the collection and delivery of monitoring equipment where necessary. The purpose of this is firstly to ensure weight stability to minimise any confounding influences, secondly to provide accurate baseline values for key variables such as physical activity, and finally to ensure the dietary records provided are a valid reflection of habitual intake.

Provided the data obtained in the control phase satisfies the relevant reliability criteria, participants will be invited to return to the physiology lab at least four weeks after their first visit. In preparation for this, participants will be asked to avoid strenuous exercise, alcohol and caffeine, before fasting overnight until they arrive at the lab the following morning.

Pre-Intervention Lab Protocol (LAB 2)

Before beginning their intervention participants will be asked to attend a pre-intervention lab protocol, as outlined below. Female participants will be asked to schedule this so that it coincides with the follicular phase of their menstrual cycle (3-10 days after the onset of menses) as this is when their hormone profiles are at their most stable.

On the day of their visit participants will be asked to consume one pint of water at 06:30 before preparing themselves to arrive at the lab by 07:30, wearing light clothes (i.e. shorts, t-shirt) with no metal components or jewellery. They will then need to provide a small urine sample to check hydration levels before measurements of height, weight and waist circumference are taken (NOTE: If weight varies by more than 1.0 kilograms compared to their baseline measurement they may be asked to repeat the monitoring phase).

At 07:45 participants will be asked to lie in a semi-recumbent position and rest for 30 minutes. This is in preparation for the measurement of resting metabolic rate and substrate oxidation which will be completed at 08:15 over three 5-minute periods using indirect calorimetry while the participant remains rested. Following this an intravenous cannula will be fitted to the antecubital vein at the elbow for blood sampling throughout the visit. At this point a 12 ml fasting blood sample will also be drawn to measure resting levels of key metabolites (e.g. glucose, insulin, cholesterol, leptin, ghrelin, etc.).

At 08:45, participants who have elected to include the adipose tissue biopsy will undergo this procedure. If included, an area on the side of the abdomen just above the waistband will be sterilised and injected with an anaesthetic (Lidocaine Hydrochloride) to numb the area. Using a larger needle a 0.5 gram sample of adipose tissue will then be taken from just below the skin. There will some bruising, but if left to heal there should be no scarring. Those who choose not to include this biopsy will simply continue resting.

At 09:00 participants will be asked to complete visual analogue scales (VAS) which will provide subjective measures of appetite, fullness and desire to eat. Immediately after this, participants will be provided with a test breakfast of instant porridge made with whole milk and sugar which they will need to consume within 10 minutes. Key measures will then be taken at the following intervals:

• 12 ml blood sample - 15, 30, 45, 60, 90, 120, 180

• Visual analogue scales - 30, 60, 90, 120, 150, 180

• Metabolic rate (indirect calorimetry) - 60. 120, 180

(Note: total urine output will also be quantified throughout the postprandial period to adjust substrate oxidation values for protein oxidation)

At 12:05 participants will be asked to place the hand of their cannulated arm in a padded Perspex box which will be heated to 55 degrees. This is to allow the venous blood to be arterialised which will provide a better indication of whole body glucose metabolism in the test which follows. At 12:15 a 7 ml blood sample will be drawn after which the participant will be asked to consume a milkshake which provides 33% of their energy requirements. Blood samples (7 ml) will then be drawn every 15 minutes for the ensuing two hours to test for the presence of a second meal effect. At 14:15 the final blood sample for this visit will be drawn and the cannula will be removed.

To conclude the pre-intervention protocol participants will be taken to a separate lab for a dual-energy x-ray absorptiometry (DEXA) scan. This is a specialized form of whole-body scan which measures how a person's weight is distributed between fat and muscle tissue. It takes approximately eight minutes and requires the participant to lie still throughout. It does administer a small dose of ionizing radiation, but this is equivalent to 7 microSieverts which is accompanied by a risk of detrimental radiation effects in the region of 1 in 10 million. The application of this method has been approved in this study by an independent medical physics expert and a clinical radiation expert.

Intervention Phase

Before exiting the lab participants will be randomised to one of three treatment conditions outlined below. All three diet groups will last for 20 days in total, with transitions between each 24 hour cycle occurring at 15:00 each day, the idea being that it will allow them to consume at least one main meal per day in all cases. For the intermittent fasting groups (IMF), please note that during fasted cycles participants will be asked to only consume water and black tea/coffee with no milk or sugar. During feeding cycles, and throughout the daily calorie restriction diet, participants will be asked to recreate their control phase diet record with appropriately modified quantities to give 75%, 150% or 200% of their normal calorie intake in accordance with the group they are assigned to. Where recreation is not possible, participants will be asked to note the item that has been removed and/or the item that was added in a new diet record that will be provided before they leave the lab. Physical activity will also be monitored for the first and last week of the intervention using an Actiheart monitor to examine behavioural adaptations to the diets.

1. Daily calorie restriction - Reduce energy intake by 25% on a daily basis

2. IMF with weight loss - Alternate between 24 hour cycles of fasting and feeding with 150% of normal intake when feeding

3. IMF without weight loss - Alternate between 24 hour cycles of fasting and feeding with 200% of normal intake when feeding

At 15:00 on the day of their initial lab visit, male participants will begin their assigned diet, although this can be postponed by up to one week if necessary. Female participants will be asked to delay this by six days in all cases in order to ensure their post-intervention lab visit falls within the same phase of their menstrual cycle to minimise the impact of monthly hormonal fluctuations.

Post-Intervention Lab Protocol (LAB 3)

Following the completion of 20 consecutive 24 hour dietary cycles, participants will return to their normal diet for one day (prescribed) whilst avoiding strenuous exercise, smoking, alcohol and caffeine, before fasting until they arrive at the lab the following morning for their post-intervention lab visit. Having consumed a pint of water at 06:30 and arrived at 07:30 in light clothing, participants will undergo the same lab protocol described earlier for their pre-intervention visit. However, instead of being randomised to a diet group they will be asked to complete a feedback questionnaire on their experience of the study.

Follow-up

Four weeks after their post-intervention lab visit participants will be invited to a feedback session to discuss all available results. The results from the meal test and the study as a whole will follow at a later date due to logistics and cost-effectiveness. Where convenient participants will also be asked to include a weight measurement to provide an indication of weight loss sustainability.

Aim

Once datasets are obtained for all 72 participants, the daily calorie restriction group will be compared against the IMF with weight loss group. This will permit the comparison of the effects of an IMF diet against the effects of a more conventional approach when matched for energy deficit. The final group, IMF without weight loss, will then highlight the relative contributions of weight loss and fasting-dependent mechanisms in producing any observed effects.

Recruitment information / eligibility

• Status: Completed
• Enrollment: 54
• Est. completion date: September 28, 2018
• Est. primary completion date: September 4, 2018
• Accepts healthy volunteers: Accepts Healthy Volunteers
• Gender: All
• Age group: 18 Years to 65 Years

Eligibility

Inclusion Criteria:

• Body Mass Index of 20.5 kg/m2 or greater

• Stable body weight for preceding three months (<3 kg increase or decrease)

• Able and willing to safely comply with study procedures

• Be able to attend the laboratory and willing to participate in necessary protocols

• Be willing to undertake the durations of fasting required by the study

• Have the capacity and willingness to provide informed consent (oral and written)

Exclusion Criteria:

• Have a body weight greater than 120 kg

• Will be undertaking any other fasting practices during their participation in the study (can enrol once stable weight returns after fasting is completed)

• Currently engaged in or planning to engage in another weight management programme (e.g. weight watchers) or exercise programme (e.g. hypertrophy training or marathon training) during the study

• Have previously suffered or are suffering from an eating disorder as assessed using the Eating Disorder Examination Questionnaire, Version 6.0 (Fairburn & Beglin, 2008)

• Have been diagnosed with either type 1 or type 2 diabetes

• Undertaking any form of medical treatment which may interfere with study variables (e.g. chemotherapy, liposuction, taking metformin, statins/other lipid-lowering medications)

• Are peri-menopausal or menopausal (absence of menses for 3-24 months)

• Pregnant, recently pregnant (within last 6 months), planning to become pregnant (within next six months), or currently breastfeeding.

• Have donated more than 500 ml of blood in the last three months before the initial laboratory visit

• Lack of metal capacity or language skills to independently understand/follow the study protocol

• Physical disability which impacts study variables or ability to independently follow study protocol (e.g. paralysis)

• Cannot consume test meals due to intolerances (i.e. lactose)

• Report medical contraindications on the 'Physical Activity Readiness Questionnaire' and are unable to obtain medical approval for the treadmill test.

Related Conditions & MeSH terms

• Obesity
• Type 2 Diabetes

Intervention

Device:
• Physical Activity Monitoring (Actiheart)
Monitor physical activity using an Actiheart monitor during the required periods.

Other:
• Energy Intake Monitoring
Monitor energy intake by weighing and recording all foods/drinks consumed during the required periods.
• Body Weight Monitoring
Monitor body weight during monitoring phase to ensure stability despite other monitoring activities.

Procedure:
• Intravenous Cannulation
Fitting of an intravenous cannula to a vein on the forearm for blood sampling during laboratory protocols.
• Adipose Tissue Biopsy
This is an optional feature which if included involves the removal of a 0.5 gram sample of adipose tissue from just below the skin using a needle.

Drug:
• Lidocaine Hydrochloride
The anaesthetic that will be used to numb the site of the adipose tissue biopsy in the event it is included.

Other:
• Post-Prandial Meal Tests
Consume two successive meals with regular blood samples throughout the ensuing hours (2-3 hours each).

Radiation:
• DEXA Scan
Undergo a dual energy x-ray absorptiometry scan to examine body composition.

Other:
• Actiheart Calibration
Submaximal treadmill test to calibrate monitoring equipment.
• Daily Calorie Restriction
Reduce energy intake by 25% on a daily basis.
• Intermittent Fasting with Weight Loss
Alternate between 24 hour cycles of fasting and feeding with 150% of normal intake when feeding.
• Intermittent Fasting without Weight Loss
Alternate between 24 hour cycles of fasting and feeding with 200% of normal intake when feeding.

Locations

Country	Name	City	State
United Kingdom	University of Bath	Bath	Somerset

Sponsors (1)

Lead Sponsor	Collaborator
University of Bath

Country where clinical trial is conducted

United Kingdom, 

Outcome

Type	Measure	Description	Time frame	Safety issue
Primary	Body Fat Percentage	Comparison of body fat percentage (measured by dual-energy x-ray absorptiometry) from pre-intervention time-point to post-intervention time-point.	Pre - Post (3 weeks)
Primary	Lean Body Mass	Comparison of lean body mass (measured by dual-energy x-ray absorptiometry) from pre-intervention time-point to post-intervention time-point.	Pre - Post (3 weeks)
Primary	Fasting Plasma Glucose Concentration	Comparison of glucose concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Plasma Insulin Concentration	Comparison of insulin concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Plasma Triglyceride Concentration	Comparison of triglyceride concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Total Cholesterol Concentration	Comparison of total cholesterol concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting LDL Cholesterol Concentration	Comparison of LDL cholesterol concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting HDL Cholesterol Concentration	Comparison of HDL cholesterol concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Leptin Concentration	Comparison of leptin concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Adiponectin Concentration	Comparison of adiponectin concentration in the blood plasma at rest across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Fasting Resting Metabolic Rate	Comparison of resting metabolic rate (measured by indirect calorimetry) across all three measurement points to ensure stability in the monitoring phase and examine change during the intervention phase.	Baseline - Pre - Post (7 weeks)
Primary	Post-Prandial Glucose AUC	Comparison of the area under the plasma glucose curve seen during the 180 minute post-prandial period following a test meal conducted before and after the intervention phase.	Pre - Post (3 weeks)
Primary	Post-Prandial Insulin AUC	Comparison of the area under the plasma insulin curve seen during the 180 minute post-prandial period following a test meal conducted before and after the intervention phase.	Pre - Post (3 weeks)
Primary	Post-Prandial Ghrelin AUC	Comparison of the area under the plasma ghrelin curve seen during the 180 minute post-prandial period following a test meal conducted before and after the intervention phase.	Pre - Post (3 weeks)
Primary	Post-Prandial Peptide-YY AUC	Comparison of the area under the plasma peptide-YY curve seen during the 180 minute post-prandial period following a test meal conducted before and after the intervention phase.	Pre - Post (3 weeks)
Primary	Physical Activity Energy Expenditure	Comparison of the mean physical activity energy expenditure values recorded using the Actiheart monitor during the monitoring and intervention phase to see the impact of the three treatments on physically active behaviours.	Monitoring - Intervention (7 weeks)
Primary	Physical Activity Intensity	Comparison of the mean physical activity intensity values recorded using the Actiheart monitor during the monitoring and intervention phase to see the impact of the three treatments on physically active behaviours.	Monitoring - Intervention (7 weeks)
Primary	CLOCK Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	NPAS2 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PER1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	CRY1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	LEP Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ADIPO-Q Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	IL-6 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	TNF Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	LPL Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	FABP4 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PPARG Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PPARGC1A Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	SREBP1C Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	FSP27 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PRKAA1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ANGPTL4 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	HSL Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ATGL Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PNPLA3 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	CIDE-A Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	GLUT-4 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	AKT2 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PDK4 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	SIRT1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	SIRT3 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	UCP2 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	IGF1R Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	IRS1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	IRS2 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	PIK3R1 Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ChREBP Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ACACA Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	FASN Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Primary	ACADM Gene Expression in Subcutaneous Adipose Tissue		Pre - Post (3 weeks)
Secondary	Fasting Carbohydrate Oxidation	Comparison of carbohydrate oxidation rate measured by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre - Post (3 weeks)
Secondary	Fasting Lipid Oxidation	Comparison of lipid oxidation rate measured by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre - Post (3 weeks)
Secondary	Fasting Protein Oxidation	Comparison of protein oxidation rate measured by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre - Post (3 weeks)
Secondary	Post-Prandial Resting Metabolic Rate	Comparison of post-prandial metabolic rate measured 60, 120 and 180 minutes after a test meal by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre-intervention 60, 120 and 180 minutes after meal - post-intervention 60, 120 and 180 minutes after meal (3 weeks)
Secondary	Post-Prandial Carbohydrate Oxidation	Comparison of post-prandial carbohydrate oxidation rate measured 60, 120 and 180 minutes after a test meal by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre-intervention 60, 120 and 180 minutes after meal - post-intervention 60, 120 and 180 minutes after meal (3 weeks)
Secondary	Post-Prandial Lipid Oxidation	Comparison of post-prandial lipid oxidation rate measured 60, 120 and 180 minutes after a test meal by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre-intervention 60, 120 and 180 minutes after meal - post-intervention 60, 120 and 180 minutes after meal (3 weeks)
Secondary	Post-Prandial Protein Oxidation	Comparison of post-prandial protein oxidation rate measured 60, 120 and 180 minutes after a test meal by indirect calorimetry before and after the intervention phase to examine the effect of the three treatment arms.	Pre-intervention 60, 120 and 180 minutes after meal - post-intervention 60, 120 and 180 minutes after meal (3 weeks)