2.3. Urine Collection (24 h) and Measurements of the Urine Volume, Sodium, Potassium and Creatinine Excretions

To determine daily sodium and potassium excretion, one 24 h urine collection per participant was used. The urine volume and creatinine excretion were determined. All procedures were conducted in accordance with the existing guidelines [44].

The urine collection was performed by collecting participant’s urine in a special container over a 24 h period. On the day of the test, the participants were asked to void the bladder in the morning, discard the urine and note the time as the beginning of the 24 h urine collection period. All urine voided in the next 24 h was collected in the same container. The collected urine samples were transported to the Department of Laboratory Diagnostics, General Hospital Nova Gorica, Slovenia, the same morning as the end of the 24 h urine collection period.

Creatinine excretion in urine occurs at a fairly constant rate over 24 h. A typical 70-kg adult man produces about 2 g of creatinine per day and there is a continual production and excretion of creatinine in the urine [45]. The 24 h creatinine excretion in urine was therefore used as a measure for excluding any urine collection samples judged to be incomplete according to creatinine values [46,47,48,49,50]. Creatinine was measured using the Jaffe method [51]. Thus, a 24 h urine collection was accepted for further analysis, if the creatinine excretion was 10.0–17.0 mmol/day for men and 7.5–12.0 mmol/day for women aged 45–60 years, or 8.5–14.25 mmol/day for men and 5.75–10.5 mmol/day for women aged 60–75 years, or 4.5–11.0 mmol/day for men and 3.5–8.5 mmol/day for women aged over 75 years [52]. Furthermore, urine collections were not considered appropriate and were thus not analysed if the 24-h urinary volumes were smaller than 250 mL [53].

The sodium and potassium 24 h excretions were determined by indirect potentiometry [54], while considering the total volume of urine collected (litres; L) and the precise time of urine collection. The sodium and potassium excretions were calculated individually as the product of the urine sodium or potassium concentration (mmol/L) and 24 h urinary volume (L/day), and were expressed in mmol/day. The daily Na and K excretion estimates (g/day) were calculated by multiplying the urine sodium or potassium concentration (mmol/day) with the respective atomic weights (Na: 23 mg/mmol and K: 39.1 mg/mmol) [55].

Considering that about 90% of the ingested sodium [37,56,57] and potassium [23,24] is excreted in urine and the remaining 10% is excreted in faeces and sweat, the total daily intakes of sodium and potassium were callculated by dividing the daily Na and K excretion estimates (g/day) by 0.9.

For the conversion from sodium (Na) to sodium chloride (NaCl, i.e., salt), a factor of 2.54 was used (NaCl (g) = Na (g) × 2.54) [58].