Sodium excretion in 24 h urine samples
The gold standard for population sodium intake assessment is the measurement of sodium excreted in a 24 h urine sample (5).
They are widely accepted as being the most reliable and practical method for assessing estimated population salt intakes.
The measurement of 24 h urine sodium excretion is a surrogate for 24 h sodium intake. The concentration of sodium in a 24 h collection, usually measured in mmol/L, can be multiplied by the volume in litres of urine collected to give mmol sodium excreted in 24 h (mmol/24 h) and subsequent conversion to salt equivalent.
Example protocols for 24 h urine collection:
UK NDNS surveys: Example method for study participants to take a 24 h urine sample using concomitant PABA
US CDC: Details of a 24 h urine collection protocol for study participants using a questionnaire to assess urine collection completeness
Completeness of 24 h urine samples should be assessed. Incomplete and/or under-collection of urine due to missed urine voids can result in falsely low 24-hour sodium and potassium excretion. Over-collection, beyond 24 hours, can skew results in the opposite direction (8).
Several methods are available to assess the completeness of 24 h urine collections:
- p-aminobenzoic acid (PABA) recovery (9);
- Urinary creatinine concentration correction (10, 11).
- Questionnaire.
PABA: PABA is a non-toxic B-complex vitamin that is thought to be fully absorbed and is readily analysed (8). PABA is an accepted measure to assess the completeness of 24 h urine collection (12). It involves the concomitant administration of 80 mg PABA tablets usually with main meals (12). The use of PABA in national surveys has been reported in the National Diet and Nutrition Survey: assessment of dietary sodium in adults 19-64 years in England (2014) (page 15, section 2.6) (13).
Though used as an objective measure of urine collection completeness, the use of PABA is not without issue due to potential variation in excretion rate with age, non-adherence to the dosage regimen and potential interaction with medication although this is less of a problem when PABA is measured by HPLC (12, 14).
UK NDNS surveys: Example method for study participants to take a 24 h urine sample using concomitant PABA
Method: SOP NBL PABA by HPLC
Kindly provided by the NIHR BRC Nutritional Biomarker Laboratory, University of Cambridge, Cambridge UK
Creatinine: Creatinine correction is an alternative method for 24 h urine collection completeness assessment. Creatinine values should be interpreted dependent on sex, protein intake, muscle mass, degree of malnutrition and ethnicity (10, 11). Since standard cut-offs do not widely exist, creatinine should be used with caution.
For guidance on creatinine measurement, see the OpeN-Global page on Common methods.
Questionnaire: Use of a questionnaire to assess 24 h urine collection completeness is reported in the US NHANES surveys (see link below). However, standardised procedures across study fieldworkers is paramount to reduce operator variation, and rigorous and intensive protocols are recommended to uphold data quality, including supervised urine collection (15).
Method: Details of a 24 h urine collection protocol for study participants using a questionnaire to assess urine collection completeness, from US CDC.
Sodium extretion in spot urine samples
Where collection of 24 h urine samples is impossible, spot urine samples may be a valuable alternative. Further, since spot samples are often routinely used in other health surveys, e.g. measurement of urinary iodine, additional assay can be easily integrated into survey or research protocols. Spot samples also remove the need for multiple visits, and therefore may be a more efficient use of resources.
Sample size estimates may need adjustment to provide the correct power and precision, and other conversion or correction factors (e.g. measurement of urinary creatinine) may be needed (14).
Measurements of creatinine concentration and sodium concentration in a spot urine sample combined with details of the individual’s sex, weight, height and age allow application of the Kawasaki formula which estimates 24 h urine excretion (16). It is recommended that this method, which is less accurate than using 24 h collections, can be used for population assessments provided that the sample size of the group is adequate (5).
Review the Kawasaki method
Methods
Note: urine is a potential biohazard, and safety precautions should be employed at all times whilst handling or manipulating urine samples.
Sample manipulation/processing and storage
Certain anticoagulants, preservatives, drugs and organophilic compounds may affect electrolyte determinations.
Visually turbid urine samples should be centrifuged prior to analysis.
Urine samples should be transferred to the required storage receptacle, e.g. Eppendorf tubes, which should be clean and free of contamination.
Samples should be frozen immediately if possible. If freezing is not possible, then refrigeration is preferred until the samples can be frozen.
Urine sample stability
Room temperature: Sodium is stable in urine for ≤ 45 days (17), though this is not recommended due to bacterial growth in the urine, and operator comfort during analysis.
Frozen: Sodium is stable indefinitely if stored frozen. Long-term storage for 20-25 years at -70 °C did not affect specimen desiccation (18).
Freeze-thaw cycles: Up to 6 freeze-thaw cycles did not affect sodium concentrations (18).
We advise OpeN-Global users to consult relative SOPs for other biomarker analyses to be conducted in the collected urine samples to ensure all stability restrictions are considered.
Analytical methods
The gold standard method is Inductively-Coupled Plasma Mass Spectrometry (ICP-MS), (See this method by NIST, p 7-1). This method can also be used on blood, serum and sweat.
The Ion-Selective Electrode method (ISE) method is also widely accepted and adopted for several electrolyte measurements, is accurate and precise and scalable for population studies e.g.US CDC NHANES surveys. See the CDC NHANES method.
The classical method for sodium assessment in a clinical laboratory is by Flame Atomic Emission Spectrometry (FAES) (also known as flame photometry), which is a selective method but not as easily automated nor as rapid.
Method: Sodium / potassium in urine by FAES
Kindly provided by the NIHR BRC Nutritional Biomarker Laboratory (NBL), University of Cambridge, Cambridge UK, as a legacy method; note: the NIHR BRC NBL no longer conducts this method in their laboratory.