Once participants have consented to be screened, they will come to the lab for a medical screening. If the medical screening is satisfactory, all participants will be asked to undergo a one-week pre-study standardization period. Participants will be asked to maintain their normal dietary and physical activity patterns throughout this period. Participants will be asked to complete a 3-day food diary, which will be analyzed to evaluate their habitual food intake and temporal distribution of energy intake throughout the day. In addition, participants will be provided with and instructed on how to use, an Actiheart (a device that combines accelerometry with heart rate monitoring). This device will be placed on the participants' chest using two standard electrocardiogram (ECG) electrodes to record HR, heart rate variability and activity level (to allow estimation of total energy expenditure) for one week. Both items will be returned before the next visit, which will be the pre-intervention measurement. After the pre-intervention visit, participants will be then randomly assigned to two weeks of either (1) the CER of 20% restriction below their estimated requirement (\~1600 kcal) 7 days/week or (2) the 5:2 IER of 70% restriction (\~600 kcal) delivered for two non-consecutive days/week and no restriction (so sufficient energy to meet their requirement) on the other 5 days/week. Seven days after the pre-intervention visit, participants will then start their 2-week intervention. A 7-day gap is designed in a bid to ensure both pre- and post-intervention measurements will be conducted during the second week of the follicular phase for females. The foods, drinks and overall energy restriction will be the same on each arm. Using estimated BMR based on the Henry equation and the individual daily physical activity level (PAL), estimated from the Actiheart during a one-week standardization period and the physical activity questionnaire, allowed the estimation of daily energy requirements for each subject using basal metabolic rate x PAL. Thus energy intake will be tailored to each participant's specific energy requirements. According to the suggestion from The Association of UK Dietitians, 2 non-consecutive ER days can be achieved on Day 1 and Day 4 of an IER week, thus the investigators will fix and prescribed the two ER days in a week with 3 days apart.
Food on intervention days will be prepared based on a 3-day sequence of menus to avoid boredom but be practical regarding the provision of foods. Each menu provides three meals/day to avoid confounding effects causing by altering meal frequency between the interventions. Three meals/day (and no snacks) is selected to make each eating incident acceptable on the most restricted day. Three meals with a macronutrient distribution of \~50% energy from carbohydrates, \~31% from fat and \~19% from protein will be prescribed. The distribution of energy across the day will be 25% of daily energy requirements from breakfast, 37.5% from lunch and 37.5% from the evening meal. To avoid confounding by altering the mealtime, and the period between which meals are consumed, breakfast will be instructed to be consumed between 8 am-8:30 am; lunch between 1 pm-1:30 pm and evening meal between 6 pm-6.30 pm. Menu designed so that it could be proportionally increased and decreased depending on the intervention requirement and informed by cooking facilities available and optimizing food safety. Participants will be instructed to adhere to the prescribed meal plan and record their food intake by taking photos.
On Day 7 of each experimental trial, participants will report to the unit in the morning after an overnight fast (\~12h) for the assessment of body weight. The investigators will also use this visit to attach a subcutaneous continuous glucose monitoring (CGM) device, which will be placed on the abdomen to allow for the estimation of continuous blood glucose over the last 7 days of each trial. Also, the investigators will download the data from the pedometer and use it to assess compliance with the requirements of the study during the first week. CGM and pedometer will be removed upon arrival on the post-intervention visit.
On the day after one week pre-study standardization period and the day after the 2-week intervention, the participants will be asked to attend the laboratory after an overnight fast to undergo a series of appetite and metabolic measurements in response to a standardised breakfast in the form of a milk-based test drink (in a volume of 10 kcal/kg body weight and energy distribution from the macronutrients will be 47.9 % carbohydrate, 35.1% fat, and 17% protein) and a subsequent ad libitum pasta lunch containing cheese, olive oil, and tomato sauce, which will provide 148.24 kcal/100g with 49.8%, 15.4% and 34.8% energy provided by carbohydrates, protein and fat, respectively.
