A total of twenty proteinogenic amino acids are present in the human body1. Due to their
chemical structure, three of the amino acids - isoleucine, leucine and valine, are termed
branched chain amino acids (BCAAs). Humans consume BCAAs through protein containing foods
such as meat, eggs and milk2,3. However, recent studies, have revealed links between elevated
plasma levels of BCAAs and type 2 diabetes (T2D), insulin resistance (IR) and obesity4-6.
Such is the strength of these links that elevated blood concentration of BCAAs are considered
biomarkers and prognostic factors for obesity, insulin resistance and type 2 diabetes7-9.
Collectively, these data suggest that targeting dysregulation of BCAAs by reducing BCAA
levels could offer important therapeutic value. Various methods can be implemented in order
to reduce circulating fasting BCAA concentrations one method that our department has
successfully implemented includes the restriction of BCAAs in 6 healthy volunteer
participants through diet control. By restricting BCAA in the diets of healthy individuals,
we successfully reduced fasting circulating BCAAs by 50% in just seven days. However, the
construction of the diet was very time consuming, tedious, and longer studies would be
plagued with compliance issues. This has led us to try an alternative method, which is to
lower BCAA levels using Sodium Phenylbutyrate (NaPB). NaPB is a commonly used medication to
treat patients with urea cycle disorders (UCD). However, it seems a secondary effect of the
drug is to reduce circulating BCAA levels in volunteers. Lindsey Burrage et al 2014, reported
significantly lower circulating BCAAs in volunteers receiving NaPB when compared to subjects
not receiving NaPB10. Although the mechanisms are not fully understood it has been suggested
that NaPB causes depletion of glutamine in the blood which may activate BCAA catabolism and
thus cause a reduction in circulating plasma BCAAs11. It is our aim to use NaPB to lower
BCAAs in prediabetic and obese participants.
Screening Visit Volunteers will sign a consent form, A health assessment, including an ECG to
monitor your heart, a blood pressure check, height and weight measurement and a medical
history check A fasted blood sample to measure health parameters, including glucose, HbA1c,
liver and kidney function, lipid profiles, thyroid function and full blood count.
Once we received the details and results (within 1 week) from your screening session, we will
contact you regarding your suitability. If you are suitable, we (you and the research team
together) will decide on the dates to start the programme.
Instruct you in the completion of a food diary. STUDY DAY 0 For this day, we will ask
volunteers to attend after an overnight fast (except water). We will take small samples of
your blood (fasting sample).
After the blood samples participants will receive a 'heavy water' drink called Deuterium
Oxide (D2O). This is almost the same as normal water, but a small and harmless chemical
change allows us to calculate how much new muscle your body is making. The amount of D2O you
are given will be calculated based on your body weight. Participants will provide saliva
samples before and after receiving D2O. Before they go home they will receive small daily
top-ups of D2O and asked to collect your saliva 3 hours after taking your daily top-ups for
the first week, and twice weekly thereafter and keep the saliva samples in a fridge/freezer.
STUDY DAY 7 On return for the second study day participants will also bring with them saliva
samples from the last 7 days. For this day, again volunteers are to attend after an overnight
fast (except water). We will perform a muscle biopsy (fasting sample) to use as our
baseline/pre-intervention measures. A fully qualified medical practitioner will take muscle
biopsies. Biopsies are sterile procedures, with local anaesthetic.
Muscle biopsies Muscle biopsies will be taken from the thigh. Before the biopsy, the area
will be cleaned and anaesthetised with local anaesthetic, after which a small (approx. 1cm
wide, 2-3cm deep) incision will be made through the skin. The muscle biopsy (smaller than the
size of a pea) will then be removed from the thigh using conchotome forceps (essentially a
sharp pair of tweezers), with 3-5 small pieces taken to ensure adequate sampling. The
incision will then be stitched shut and covered with a waterproof adhesive dressing. Muscle
biopsies are usually well tolerated, largely painless (though the insertion is felt) and the
complication rate is very low.
Biopsy Aftercare At the end of the biopsy, you will receive advice and information about how
to take care of the wound and given an information leaflet. Five to seven days after each
muscle biopsy, the stitches will be removed and the wounds will be checked.
In addition to the above, volunteers will also have a series of minimally and non-invasive
tests to assess different aspects of your health and metabolic status:
We will measure how well the body is able to process sugar by performing the oral
glucose tolerance test (OGTT). This is usually used to diagnose diabetes, or stratify
the future risk of diabetes. For this test, you will be given 75g of a sugary drink.
Small blood samples will be taken prior to the drink and every 15 minutes (for 2 hours)
from a cannula (thin plastic line) sited in a vein in your hand, which will be placed in
a warm box.
A light lunch will be provided at midday. After lunch the first oral dose of phenyl
butyrate will be given.
Volunteers will receive the first dose of Phenylbutyrate after lunch. Volunteers will
receive 5 grams of sodium Phenylbutyrate every day. 5 g/m2/day of Sodium Phenylbutyrate
will be split into three and taken with each meal.
Participants will receive Sodium Phenylbutyrate to last one week. STUDY DAY 10 For this
day, we will ask you to attend after an overnight fast (except water). We will take
small samples of your blood (fasting sample). In addition, blood pressure, weight and
height will be recorded. Volunteers will also have an ECG. It is anticipated study day
11 will take up to 1 hour to complete.
STUDY DAY 17 For this day, we will ask you to attend after an overnight fast (except water).
We will take small samples of your blood (fasting sample) and a saliva sample to use as our
midpoint intervention measures. In addition, blood pressure, weight and height will be
recorded. Volunteers will then be provided with another medication organiser box with enough
Sodium Phenylbutyrate to last them until the end of the study. It is anticipated study day 17
will take up to 1 hour to complete. We will also request a test for Leukopenia.
STUDY DAY 28 This is the final day of the study, again volunteers are to attend after an
overnight fast (except water). A fully qualified medical practitioner will take muscle
biopsies. Biopsies are sterile procedures, with local anaesthetic. Following this, we will
perform the oral glucose tolerance test (OGTT), volunteers will be given 75g of a sugary
drink. Small blood samples will be taken prior to the drink and every 15 minutes (for 2
hours) from a cannula which will be placed in a warm box In addition, blood pressure, weight
and height will also be recorded. In addition a Fructosamine test will also be requested pre
and post study.