The doubly labeled water (DLW) technique is a noninvasive method for measuring total energy expenditure in humans, which utilizes isotopes of hydrogen and oxygen to calculate carbon dioxide production and energy metabolism. The protocol involves administering a dose of labeled water, followed by urine and saliva sampling to measure isotopic enrichment over a defined period. Although recognized as the gold standard, the method faces limitations such as high costs and the inability to provide specific data on physical activity.

1. DOUBLY LABELLED WATER TECHNIQUE
By
E.SRILALITHA
MSC SPORTS NUTRITION
ROLL NO-10

2. INTRODUCTION
• (DLW) technique was developed as a method for measurement of
free-living energy expenditure in animals (Lifson and McClintock,
1966). The method has since been validated in many animal species
(for summary see Roberts, 1989; Schoeller, 1988).
• 2H2
18O,technique to calculate total production of carbondioxide(CO₂)
&from this measurement of mean respiratory quotient and total
energy expenditure was originally developed in small animals

3. THEORY
• The DLW method is based on the premise that after a dose of doubly
labeled water, 2H2
18O, the two isotopes equilibrate with total body
water (TBW) and then are eliminated differentially from the body .
• Deuterium (2H) leaves the body as water, while 18O leaves as water
(H2O) and carbon dioxide (CO2).
• Therefore, CO2 production can be calculated by subtracting 2H
elimination from 18O elimination.

6. Doubly Labeled Water Protocol
• The study begins with the collection of baseline urine and/or saliva
samples, followed by oral administration of the 2H2
18O dose.
• Saliva samples are obtained 2 to 4 hours after the dose for calculation
of dilution spaces (Schoeller et al., 1980).
• Urine samples then are collected at the end of the period for
measurement of final isotopic enrichment.
• The length of time a DLW study can be carried out depends on the
turnover of the two isotopes, which is dependent on H2O and CO2
output.
• For studies in typical adults, the optimal metabolic period is 4 to 21
days (Schoeller, 1988).

8. CALCULATION
• The isotope elimination rates (kH and kO) can be calculated by two
point method using the:
initial (i) isotopic enrichment
the final (f) enrichments
k = (ln enrichmentf-ln enrichmenti)/Δ t
where ln is the natural log,
enrichment is the enrichment above baseline,
Δt is the number of days between the initial and final sample

9. CALCULATION
• CO2 production has been calculated according to Schoeller (1988)
N is the TBW calculated from the 18O enrichment in the 4-h saliva
rH2Of is the rate of fractionated evaporative water loss,which is
estimated to be 1.05N(1.01kO -1.04kH).

10. CALCULATION BY WEIR'S FORMULA
• The rate of CO2 production (rCO2) is calculated as the difference
between the elimination rates of 2H and 18O, using the following
formula [31]:
rCO2 (mol/day) = 0.4554 × TBW (mol) × (1.007 ko − 1.041 kh)
• where ko and kh (day-1) are the elimination rates of 2H and 18O,
respectively.
• The TEE is calculated by using the modified version of Weir's formula
based on rCO2 and food quotient (FQ) [31]:
TEE (kcal/day) = 22.4 × (3.9 × [rCO2/FQ] + 1.1 × rCO2)

11. RQ for carbohydrates is 1.0
RQ for fats is 0.69-0.73
RQ for proteins is 0.82

12. DOSAGE OF DLW
• The dose of DLW is based on the body size of subject in order to
match the body water enrichments to the isotope-ratio mass
spectrometry precision.
• In most DLW studies, investigators have assumed TBW as 60% of
body weight. The 99 atom % deuterium 2H2 and 10 atom % 18O, are
the most commonly used for enrichments of the labeled water
available on the market.
• The International Atomic Energy Agency (IAEA) recommends doses of
0.12 g·kg−1 body water of 99 atom % deuterium labeled water and
1.80 g·kg−1 body water of 10 atom % 18O, When the more highly
enriched 18O, water is used, the dose should be reduced [25].

13. ADVANTAGES
• The DLW method uses stable isotopes of oxygen (18O) and hydrogen
(2H) for the measurement of TEE.
• The DLW method is widely recognized as the gold standard for the
measurement of TEE.
• DLW method presents the advantage of its noninvasive nature and
possibility for the subjects to continue their normal activities during
the measurement period.

14. LIMITATIONS
• The method is its high cost due to the high price of DLW, the
expensive equipment and expertise required for analysis.
• DLW technique is that it provides the overall measure of averaged
daily TEE over the measurement period, but it does not provide any
specific details on physical activity.

15. REFERENCES:
• www.ncbi.nlm.nih.gov/books/NBK233774/
• https://doi.org/10.7762/cnr.2017.6.2.68
• Scholler et al 1983