A
B
C
D
~30
30
60
4
Time line [min]
A
=
Stabilization period plus attachment of equiment (electrodes and blood pressure cuffs)
B
=
Hemodynamic baseline measurement
C
=
Treatment ingestion (one of four treatments; randomized fashion)
D
=
Post treatment observational time
Treatment 1 (200 mL still mineral water at room temperature)
SCHEMATIC STUDY PROTOCOL – DOSE-RESPONSE WATER STUDY
Treatment 2 (400 mL still mineral water at room temperature)
Treatment 3 (600 mL still mineral water at room temperature)
Treatment 4 (800 mL still mineral water at room temperature)
Data supplement Figure 2 Blood pressure responses
1
Figure 2 – Supplement
Figure 2 – Supplemental data. Left panels A and B: Time course for changes in systolic-
and diastolic blood pressure after ingestion of water at different volumes (200 mL, 400 mL, 600 mL, and 800 mL) investigated in five young, non-obese subjects. Each mean value (i.e. 10 min blocks) corresponds to the average over the respective interval that was subtracted by
its baseline values. Right panels A and B: Floating bars (min to max) represent individual
mean responses (open circles: ○) averaged over min 0 to 60 post-drink relative to baseline values and presented as a delta (Δ) for each drink volume, respectively. Treatment versus time interaction effects were analysed by two-way repeated measures ANOVA with time and treatment (200 mL, 400 mL, 600 mL, and 800 mL) as interaction factors (no interaction effect
Data supplement Figure 2 Blood pressure responses
2 was observed). BL indicates baseline. Data are presented as means ± standard deviation (Panel A).
Systolic and diastolic blood pressure were measured continuously and beat-to-beat by using a Task Force Monitor (CNSystems, Graz, Austria) for a 30 min baseline and for a 60 min post-drink period. Detailed information concerning this technique can be accessed from the references below:
Monnard CR, Grasser EK. Cardiovascular responses to a glucose drink in young male individuals with overweight/obesity and mild alterations in glucose metabolism, but without impaired glucose tolerance. Eur J Nutr 2019; doi: 10.1007/s00394-019-02120-3.
Grasser EK, Yepuri G, Dulloo AG, Montani JP. Cardio- and cerebrovascular responses to the energy drink Red Bull in young adults: a randomized cross-over study. Eur J Nutr
2014;53(7):1561-71.
Moreover, a critical methodological discussion for continuous compared with discontinuous blood pressure measurements can be accessed from the reference below:
Erik Konrad Grasser, Jennifer Lynn Miles-Chan, Nathalie Charrière, Cathríona R Loonam, Abdul G Dulloo, and Jean-Pierre Montani. Energy Drinks and Their Impact on the
Data supplement Table 1 Subject characteristics
Table 1 – Supplement: Anthropometric- and averaged beat-to-beat cardiovascular baseline data
over the last 10 min prior to ingesting each of the drinks (data are presented as means ± standard deviation)
n: number of subjects; VLF_RRI_LN: Very low frequency component of RR-interval (0.003 to 0.04 Hz) after natural logarithm transformation; LF_RRI_LN: Low frequency component of RR-interval (0.04 to 0.15 Hz) after natural logarithm transformation; HF_RRI_LN: High frequency component of RR-interval (0.15 to 0.4 Hz) after natural logarithm transformation; PSD_RRI_LN: Power spectral density of RR-interval (0.003 to 0.4 Hz) after natural
logarithm transformation; HF_RRI_%: Normalized high frequency power component in percentage. Statistical analysis was performed using one-way repeated measures ANOVA with Holm-Sidak’s multiple comparison test comparing each condition among themselves and
p < 0.05 was considered as significant difference between test drinks.
200 mL 400 mL 600 mL 800 mL p values n
Age, years 23.9 ± 2.9 --- --- --- --- 8
Height, m 1.81 ± 0.07 --- --- --- --- 8
Weight, kg 77.9 ± 9.5 --- --- --- --- 8
Body mass index, kg · m-2 23.7 ± 2.2 --- --- --- --- 8
Heart rate, bpm 65.6 ± 6.2 66.1 ± 5.9 68.1 ± 8.7 71.4 ± 10.0 > 0.10 8 VLF_RRI_LN 6.1 ± 0.8 5.9 ± 0.7 5.3 ± 0.8 5.4 ± 1.1 > 0.10 8 LF_RRI_LN 6.4 ± 0.6 6.6 ± 0.5 6.4 ± 0.6 6.5 ± 0.6 > 0.74 8 HF_RRI_LN 5.9 ± 0.9 6.2 ± 0.9 5.9 ± 1.1 5.7 ± 1.1 > 0.45 8 PSD_RRI_LN 7.2 ± 0.7 7.5 ± 0.5 7.2 ± 0.7 7.2 ±0.7 > 0.58 8 HF_RRI_% 43.3 ± 11.7 44.5 ± 13.8 40.2 ± 14.5 39.5 ± 18.1 > 0.89 8 Systolic blood pressure, mmHg 120.2 ± 7.1 121.2 ± 10.3 117.6 ± 7.4 116.6 ± 4.3 > 0.36 5 Diastolic blood pressure, mmHg 78.0 ± 5.7 76.5 ± 6.3 76.6 ± 6.5 75.8 ± 5.3 > 0.87 5
Data supplement Table 1 Subject characteristics
Subjects
Eight healthy, young male subjects were recruited and paid (Table 1). All of the test subjects were non-smokers and classified as non-obese with normal resting blood pressure and normal resting heart rate and were free of any diseases nor were taking any medication that would affect cardiovascular and/or autonomic regulation. All participants fasted (no food or water) for ≥ 12 h and abstained from alcohol, smoking and caffeine, as well as from vigorous
exercise for 24 h prior to each test and were advised not to change their dietary habits between the tests.
Data supplement Table 2 Heart rate variability parameters
1 1
Table 2 - Supplement: Heart rate variability parameters from spectral analysis in response to
water at different drink volumes and presented as an averaged delta response relative to baseline values.
Holm-Sidak test 2F-ANOVA Variable Post-drink time periods (averaged delta responses to baseline
values) Overall effect Interaction ΔVLF_RRI_LN 10 min 20 min 30 min 40 min 50 min 60 min p - value None 200 mL -0.0 ± 0.9 -0.2 ± 0.6 -0.3 ± 1.0 -0.4 ± 0.6 -0.2 ± 0.9 -0.1 ± 0.9 0.43
400 mL 0.1 ± 0.4 0.3 ± 0.6 0.0 ± 0.4 0.2 ± 0.4 0.1 ± 0.5 0.1 ± 0.3 0.53 600 mL 0.9 ± 0.7* 1.0 ± 0.6* 0.4 ± 0.5 0.9 ± 0.9 0.6 ± 0.5* 0.6 ± 0.4* 0.02 800 mL 0.8 ± 0.8 0.7 ± 1.0 0.4 ± 1.2 0.4 ± 1.0 0.3 ± 1.1 0.3 ± 1.1 0.15
ΔLF_RRI_LN 10 min 20 min 30 min 40 min 50 min 60 min p - value None 200 mL 0.6 ± 0.4 0.1 ± 0.4 0.2 ± 0.5 0.3 ± 0.4 0.3 ± 0.4 0.5 ± 0.5 0.03
400 mL 0.4 ± 0.4 0.1 ± 0.6 0.1 ± 0.5 0.2 ± 0.6 0.1 ± 0.5 0.0 ± 0.5 0.19 600 mL 0.8 ± 0.6* 0.2 ± 0.6 0.4 ± 0.4 0.0 ± 0.6 0.2 ± 0.4 0.1 ± 0.5 < 0.005 800 mL 0.5 ± 0.6 0.2 ± 0.6 -0.1 ± 0.7 0.0 ± 0.7 0.0 ± 0.4 0.1 ± 0.6 0.10
ΔHF_RRI_LN 10 min 20 min 30 min 40 min 50 min 60 min p - value None 200 mL 0.5 ± 0.8 0.3 ± 0.9 0.2 ± 0.7 0.2 ± 0.7 0.2 ± 0.8 0.2 ± 0.7 0.23
400 mL 0.5 ± 0.4 0.3 ± 0.3 0.3 ± 0.5 0.3 ± 0.5 0.1 ± 0.5 -0.1 ± 0.3 0.06 600 mL 0.7 ± 0.5* 0.7 ± 0.5* 0.5 ± 0.6 0.3 ± 0.6 0.4 ± 0.7 0.1 ± 0.5 < 0.01 800 mL 0.8 ± 0.6* 0.7 ± 0.2*** 0.5 ± 0.3** 0.4 ± 0.2*** 0.2 ± 0.1** 0.3 ± 0.3* < 0.005
ΔPSD_RRI_LN 10 min 20 min 30 min 40 min 50 min 60 min p - value None 200 mL 0.5 ± 0.4* 0.2 ± 0.2 0.2 ± 0.2 0.2 ± 0.1* 0.3 ± 0.2* 0.4 ± 0.3* 0.01
400 mL 0.4 ± 0.3 0.2 ± 0.5 0.2 ± 0.5 0.2 ± 0.5 0.1 ± 0.5 0.0 ± 0.4 0.11 600 mL 0.8 ± 0.4*** 0.5 ± 0.4* 0.5 ± 0.3* 0.3 ± 0.4 0.3 ± 0.4 0.2 ± 0.4 < 0.005 800 mL 0.6 ± 0.4 0.4 ± 0.5 0.1 ± 0.5 0.2 ± 0.4 0.1 ± 0.3 0.1 ± 0.5 0.03
ΔHF_RRI_% 10 min 20 min 30 min 40 min 50 min 60 min p - value None 200 mL -2.5 ± 9.1 0.5 ± 11.3 -3.8 ± 9.2 -4.0 ± 12.6 -5.0 ± 13.6 -6.8 ± 12.0 0.28
400 mL 0.4 ± 14.6 4.1 ± 11.8 4.8 ± 11.6 0.4 ± 11.1 -0.2 ± 11.3 -2.9 ± 9.6 0.30 600 mL -1.9 ± 15.3 8.5 ± 16.2 5.8 ± 11.5 7.0 ± 14.4 3.0 ± 14.8 0.4 ± 16.2 0.13 800 mL 2.1 ± 13.3 7.0 ± 10.2 11.2 ± 10.5 7.4 ± 11.3 4.5 ± 8.1 1.5 ± 7.3 0.07
VLF_RRI_LN: Very low frequency component of RR-interval (0.003 to 0.04 Hz) after natural logarithm transformation; LF_RRI_LN: Low frequency component of RR-interval (0.04 to 0.15 Hz) after natural logarithm transformation; HF_RRI_LN: High frequency component of RR-interval (0.15 to 0.4 Hz) after natural logarithm transformation; PSD_RRI_LN: Power spectral density of RR-interval (0.003 to 0.4 Hz) after natural logarithm transformation; HF_RRI_%: Normalized high frequency power component in
Data supplement Table 2 Heart rate variability parameters
2 2 percentage. Δ refers to averaged post-drink responses, which were subtracted from respective baseline values for that variable and presented as deltas: *p < 0.05, **p < 0.01, ***p < 0.005, statistical significant difference between a post-drink period and its respective baseline value by using one-way repeated measures ANOVA with the Holm-Sidak’s multiple comparison test. Interaction refers to treatment versus time effects by two-way repeated measures ANOVA analysis with time and treatment (i.e. 200 mL, 400 mL, 600 mL, and 800 mL) as interaction factors. Values are presented as means ± standard deviation.
