Part A = Purpose of study Part A of the BOLT study will focus on characterising the total and essential amino acid concentrations for 4 hours following the consumption of a single serving of: 1) commercially available high-protein yoghurt drink, 2) a micellar casein drink, and 3) a flavoured water-based placebo drink. This is called a bioavailability study.

Part B = Purpose of study Part B of the BOLT study will examine the effect of a twice-daily ingestion of 1) high protein yoghurt drink (test product), or 2) flavoured water based drink (control drink) on biomarkers of 1) muscle damage/stress, 2) physical function, 3) self reported pain and muscle soreness, and 4) range of motion, sleep, and recovery assessments, at 24, 48, and 72 hours following acute muscle fatiguing exercise in 18-45-year-old male and female participants. Participants will be randomly allocated to the test product or control drink groups prior to beginning the first visit.

Part A =

This study has two parts: 1) an initial pre-screening assessment and 2) complete 3 separate trials where participants will be asked to ingest a nutritional test or control product drink. All lab-based sessions will take place at Compass House on East Road, Anglia Ruskin University, Cambridge. The study design is a three-way, single-blind, randomised, cross-over study design for bioavailability assessment of the amino acid profile of 3 nutritional drinks as outlined below, whereby participants will receive either 1 serving of the:

Test product: high protein yoghurt drink (1 serving = 25g protein, 300ml), Control product 1: micellar casein drink (1 serving = 25g protein, 300 ml), Control product 2: flavoured water-based placebo drink (1 serving = 0g protein, 300 ml), at each visit, in a randomised order.

Part 1 - Pre-screening assessment:

This will involve a full study briefing presentation, in which the investigators will go over the outline of the study, including expectations, lab visits and product information. Study staff will collect food and exercise diaries from participants (a 3-day food and exercise diaries, comprising of 1 weekend day, 1 rest day, and 1 exercise day within a week). If the inclusion criteria are met, participants will continue into the trial. Once participants have had a chance to ask any questions and should participants wish to volunteer, the investigators will ask participants to complete 2 consent forms (one for the main study and one for collection of samples as part of our internal processes).

Next, the investigators will check participants meet our study criteria via a standard health screen form as outlined above, along with assessment of participants body composition, body mass and height (for body mass index). The investigators will also collect data including, biological sex (male/female), age (years) and lifestyle parameters (alcohol consumption is classified as [on average during the last 6 months] none, ≤ 7 units per week, 8-14 units per week, 15-21 units per week, > 21 units per week, training history including amount of exercise undertaken per week/month [time and frequency] and general information from the health screen questionnaire, and then relevant medical history, prior and concomitant medication, and nutritional supplements. If exclusion criteria are met, participants will be withdrawn from the study. Assuming participants fully meet our criteria and are willing to take part, the investigators will then provide participants with key information and instructions for food diary assessment. Following this first assessment, the investigators will then provide participants with details to book in for each of the 3 trials.

Part 2 - Main study trials:

The main part of this study will involve 3 visits of >4 hours each, with each visit scheduled no more than 21 days apart.

Outline of each visit General - Participants will be required to refrain from physical activity and alcohol consumption 24 hours before each study visit. The evening before the lab visit, participants will be asked to eat a standardised meal to be consumed after 17:00 and arrive the labs after an overnight fast (at least 10 hours). Before arriving at the lab (~7-9am), participants can drink up to 500 mL of only water in the morning, but no sooner than 1 hour before each appointment. Participants will also be required to complete a habitual food diary which will be explained at the briefing session. At the end of each visit, the investigators will undertake product/protocol compliance audits to assess study product intake and adherence to assessment windows during the study visits.

Visit 1 - Upon arrival the investigators will assess participants morning body mass. A member of the research team will then fit an intravenous cannula via a short scratch, using a butterfly needle (which is a short, flexible tube that is placed into a vein on participants arm or back of participants hand) and withdraw 10 mL of blood as a baseline sample. Participants will then be asked to consume of 1 of 3 drinks (as outlined above), where participants will have up to 10 mins to drink the assigned drink. Blood samples will then be drawn at time intervals including 15, 30, 45, 60, 90, 120, 150, 180, 210, and 240 min after the study product intake. This will equate to 11 samples of 10 mL blood collection (within 2 x 5 mL tubes per timepoint), which equals 110 mL blood collection in total per visit (or 330 mL in total over the study). Per trial (110 mL of blood collection) and also over the course of the study (330 mL of blood collection), the amount of blood drawn is less than the maximum permitted extra corporeal volume for male (552-971 mL) and female (486-908 mL) donors of varying body masses and heights (50-100kg, 150-183 cm; JPAC, 2013).

This constitutes the 'bioavailability' test for this study. The investigators will ask participants to rest comfortably (please bring something to read or a laptop). Throughout the study visits, participants will not be able to eat, but will be permitted to drink water ad libitum after the first 2 hours (with a maximum of 250 mL) - the volume consumed at visit trial 1 will be replicated on subsequent visits. In addition, participants will be asked to abstain from any physical exercise other than incidental walking during the study visits.

Visits 2 and 3 - will be at least 5 days apart but no more than 21 days between a visit; participants will undergo the same protocol method as outlined above, with a different study product.

About the nutrition products the investigators will assessing:

This study is a project being undertaken between Anglia Ruskin University and Danone Global Research & Innovation Center, Netherlands. All products will be supplied independently by Danone Global Research & Innovation Center as a nutrition company and have been verified for human consumption and safety. All ingredients are commonly found in food, with the main test product available in supermarkets. For the purposes of this study, the investigators will be using 3 different products in a random order:

Test product: high protein yoghurt drink (1 serving = 25g protein, 300ml) that is found in supermarkets throughout the UK, Control product 1: micellar casein drink (1 serving = 25g protein, 300 ml), which is a protein supplement that is the predominant protein in cow's milk.

Control product 2: flavoured water-based placebo drink (1 serving = 0g protein, 300 ml),

Part B = This study has two parts: 1) an initial pre-screening assessment and 2) complete 4 separate trials where participants will be asked to ingest a nutritional test or control product drink. All lab-based sessions will take place at Compass House on East Road, Anglia Ruskin University, Cambridge.

The study is an exploratory interventional study, utilising double blind, randomised, parallel group design for the assessment of a nutritional drink as outlined below on the recovery from acute muscle fatiguing exercise protocol. Participants will receive 2 servings of a Test product: high protein yoghurt drink (1 serving = 25g protein, 300 ml) or a Control product: flavoured water-based placebo drink (1 serving = 0g protein, 300 ml), per day for 3 days over the course of the study.

Participants will receive a copy of this information on the study, to read over and decide upon their voluntary participation. If interest, participants will be requested to complete 3-day food and exercise diaries (as outlined/provided below), comprising of 1 weekend day, 1 rest day, and 1 exercise day within a week before attending the pre-screening assessments where they will be asked to provide their consent for participation.

Visit 1:

Part 1 - Pre-screening assessment:

This will involve a full study briefing presentation, in which the investigators will go over the outline of the study, including expectations, lab visits and product information. Participants will refrain from physical activity and alcohol consumption 24 hours before the familiarisation visit.

Study staff will collect food and exercise diaries from participants. If the inclusion criteria are met, participants will continue into the trial. Once participants have had a chance to ask any questions and should participants wish to volunteer, the investigators will ask participants to complete 2 consent forms (one for the main study and one for collection of samples as part of our internal processes).

Next, the investigators will check participants meet our study criteria via a standard health screen form as outlined above, along with assessment of participants body composition, body mass and height (for body mass index). The investigators will also collect data including, biological sex (male/female), age (years) and lifestyle parameters (alcohol consumption is classified as [on average during the last 6 months] none, ≤ 7 units per week, 8-14 units per week, 15-21 units per week, > 21 units per week, training history including amount of exercise undertaken per week/month [time and frequency] and general information from the health screen questionnaire, and then relevant medical history, prior and concomitant medication, and nutritional supplements. If exclusion criteria are met, participants will be withdrawn from the study. Assuming participants fully meet our criteria and are willing to take part, the investigators will then provide participants with key information and instructions for food diary assessment.

Participants will then undergo familiarisation of the primary dependent measures and exercises (as outlined below) that will be completed during the study. Study staff will measure participants' body mass and hydration status, and maximal voluntary contraction (MVC) of knee extension/flexion. Study staff will familiarise participants with the perceived pain threshold test, range of motion (ROM) test, 75% of MVC, drop jumps, 30 second counter movement jump (CMJ30), and 6 second peak power test (PPT6).

Participants will be informed of the set standardised meals for breakfast, lunch and dinner across visit 2-5 and will be asked to keep their habitual physical activity pattern of no more than 4 exercise sessions per week, of which no more than 2 sessions are light resistance exercise sessions. Between visit 2-5 participants will be asked not engage in exercise, except for the exercise that is part of this study. Participants will record any deviations from the standardised meal plan in a food compliance diary throughout.

Participants will be randomly allocated to receive either the test product or the control product. Following this first assessment, the investigators will then provide participants with details to book in for each of the 4 trials.

Part 2 - Main study trials:

The main part of this study will involve 4 visits of ~2 hours each, across 4 consecutive days, occurring at the same time of day.

Outline of each visit General - Participants will be required to refrain from physical activity and alcohol consumption 24 hours before each study visit. Participants will be asked to adhere to the recommended standardised meals for breakfast, lunch and dinner across visit 2-5. Between visit 2-5, participants will be asked not engage in exercise, except for the exercise that is part of this study. Participants will record any deviations from the standardised meal plan in a food compliance diary throughout.

After an overnight fast (at least 10 hours) for each visit, participants will come to the study site to participate in the study visit. Before arriving to our lab, participants can drink up to 500 mL of only water in the morning. Participants will also be required to complete a habitual food diary which will be explained at the briefing session. At the end of each visit, the investigators will undertake product/protocol compliance audits to assess study product intake at the study visits and adherence to assessment windows.

Visit 2 - Upon arrival to the laboratories, participants will be asked to provide their completed food compliance diary. Study staff will then measure participants body mass, hydration status and collect one 10 mL whole blood sample via venepuncture by a trained researcher or phlebotomist (i.e. blood collector). Participants will complete the sleep quality questionnaire, recovery questionnaires, perceived pain threshold test, record heart rate variability (HRV) and range of motion (ROM) test. Next, participants will complete maximal voluntary contraction (MVC), 75% of MVC, 30 second countermovement jump (CMJ30) test and a 6 second peak power output test (PPT6). Participants will then complete the muscle fatiguing exercise protocol (6 sets, whilst wearing a weighted vest) of 25 drop jumps, with passive rest between sets. Study staff will measure the participants' rating of perceived exertion (RPE) and heart rate before, during and after the protocol, as well as a capillary blood lactate concentration from the participants' fingertip immediately before and after the protocol. Participants will then repeat the baseline protocol (e.g. blood sample, perceived pain threshold, ROM, MVC, CMJ30 and PPT6 tests). Within 30-min after the acute muscle fatiguing exercise protocol, participants will consume the first serving of the study product at the site.

Before leaving the study site, participants will receive recommendations for standardised meals for the next 3 days, the second serving of the study product, and study product compliance diaries. In the afternoon, participants will consume standardised lunch, test or control product and dinner (described below).

Visit 3, 4 and 5 - On visit 3-5, participants will attend the laboratory once again and return completed food compliance diaries and study product intake compliance diaries from the previous day and receive the same empty diaries for the next day. On visit 3-5 study staff will again measure the participants' body mass, hydration status and draw a 10 mL venous blood sample. Participants will then complete the sleep quality questionnaire, recovery questionnaire, perceived pain threshold test, record HRV and range of motion test. Next, participants will complete MVC, 75% of MVC, CMJ30, and PPT6 tests. Within 30-min testing, participants will consume the study product at the site. Participants will consume a standardised lunch, the second serving of the study product, and dinner at home. On visit 5, participants will end the study after the performance and fatigue measures followed by lunch.

About the nutrition products the investigators will assessing:

This study is a major project being undertaken between Anglia Ruskin University and Danone Global Research & Innovation Center, in the Netherlands. All products will be supplied independently by Danone Global Research & Innovation Center as a world-leading nutrition company and have been verified for human consumption and safety. For the purposes of this study, there will be 2 parallel groups undertaking 1 of 2 different products:

Test product: high protein yoghurt drink (1 serving = 25g protein, 300 ml), Control product: flavoured water-based placebo drink (1 serving = 0g protein, 300 ml)

Inclusion Criteria:

• Age ≥ 18 and ≤ 45 years.
• Body Mass Index (BMI) ≥ 18.5 and ≤ 29.9 kg/m2 (measured at the first assessment),
• Written informed consent (undertaken at the first assessment).
• Willingness and ability to comply with the protocol in the opinion of the investigator.
• Judged by the investigator to be in good health as assessed by a health screening questionnaire.
• Currently identifies as the same biological sex as at birth.

Exclusion Criteria:

• Any known ongoing medical condition that might interfere with absorption and digestion and/or gastrointestinal (GI) function (e.g. inflammatory bowel disease, gastroesophageal reflux disease, celiac disease, diaphragmatic hernia or diaphragmatic surgery, gastric ulcer, gastritis, gall bladder problems, pancreatitis, GI cancer, oesophageal and/or gastric surgery).), in opinion of the investigator,
• Known musculoskeletal or soft tissue injury.
• Known cardiovascular disease, disease related to the immune system and/or the respiratory system.
• Known renal or hepatic failure or known thyroid dysfunction.
• Known Diabetes Mellitus type I or type II, insulin resistance or metabolic syndrome.
• Any ongoing cancer (except for basal cell carcinoma) and/ or cancer treatment,
• Known anaemia or low haemoglobin or low iron status.
• Any known bleeding disorder or reaction to withdrawal of blood samples.
• Use of oral and systemic use of prokinetics, laxatives, antidiarrheals, anticoagulants within 2 weeks prior to screening.
• Use of systemic antibiotics within 4 weeks prior to screening.
• Any known dietary allergies.
• Any known intolerances to ingredients of the study product, i.e. dairy, cow's milk allergies, lactose intolerance.
• Self-reported adherence to a strict dietary regime (e.g. vegetarian/ vegan/ paleo/ketogenic/intermittent fasting/ high protein diet (>1.6 g/kg body weight/day), or protein supplementation or a weight loss program, as confirmed via the screening process.
• Use of any nutritional supplements or additional protein supplements or nutritional support within 4 weeks prior to screening.
• Engage (on average) in more than 2 resistance exercise sessions and/or more than 4 exercise sessions in total weekly within 2 months before screening,
• Considered as endurance trained, competitive athletes or trained weightlifters, powerlifters or bodybuilders, in the opinion of the investigator.
• Known pregnancy and/or lactating.
• Current smoking or stopped smoking for <1 month prior to screening (except for incidental smoking of ≤3 cigarettes/cigars/pipes per week on average in the last month).
• Average alcohol use of >21 glasses* per week for men or >14 glasses per week for women (on average during the last 6 months) or drug/ medicine abuse in opinion of the investigator.
• Participation in any other clinical study with investigational or marketed products concomitantly or within 4 weeks before screening.
• Major medical or surgical event requiring hospitalization within the preceding 3 months and/or scheduled in the period of study participation relevant in the opinion of the investigator.