Skip to main content
PLOS One logoLink to PLOS One
. 2024 May 31;19(5):e0304803. doi: 10.1371/journal.pone.0304803

Differences in urine creatinine and osmolality between black and white Americans after accounting for age, moisture intake, urine volume, and socioeconomic status

Patrick B Wilson 1,*, Ian P Winter 1, Josie Burdin 1
Editor: William M Adams2
PMCID: PMC11142698  PMID: 38820483

Abstract

Urine osmolality is used throughout research to determine hydration levels. Prior studies have found black individuals to have elevated urine creatinine and osmolality, but it remains unclear which factors explain these findings. This cross-sectional, observational study sought to understand the relationship of self-reported race to urine creatinine and urine osmolality after accounting for age, socioeconomic status, and fluid intake. Data from 1,386 participants of the 2009–2012 National Health and Nutrition Examination Survey were utilized. Age, poverty-to-income ratio (PIR), urine flow rate (UFR), fluid intake, estimated lean body mass (LBM), urine creatinine, and urine osmolality were measured. In a sex-specific manner, black and white participants were matched on age, dietary moisture, UFR, and PIR. Urine creatinine was greater in black men (171 mg/dL) than white men (150 mg/dL) and greater in black women (147 mg/dL) than white women (108 mg/dL) (p < .001). Similarly, urine osmolality was greater in black women than white women (723 vs. 656 mOsm/kg, p = .001), but no difference was observed between white and black men (737 vs. 731 mOsm/kg, p = .417). Estimated LBM was greater in black men (61.8 kg) and women (45.5 kg) than in white men (58.9 kg) and women (42.2 kg) (p≤.001). The strongest correlate of urine osmolality in all race-sex groups was urine creatinine (Spearman ρ = .68-.75). These results affirm that individuals identifying as black produce higher urine creatinine concentrations and, in women, higher urine osmolality after matching for age, fluid intake, and socioeconomic status. The findings suggest caution when comparing urine hydration markers between racial groups.

Introduction

Avoiding hypohydration is a significant element of maintaining good overall health and physical function. Studies, for example, have documented that hypohydration can impair cognitive function [1] and increase perception of effort during exercise [2]. Also, while still not yet confirmed as fully causal in nature, there is an observed relationship between levels of fluid-regulating hormones (i.e., arginine vasopressin) and metabolic function [3]. Given this evidence, the assessment of hydration status is broadly considered as important to researchers, clinicians, and practitioners.

A multitude of options exists for assessing hydration status, ranging from very practical/simple (urine color) to invasive (plasma osmolality). Experienced researchers in the field of hydration science routinely acknowledge that there is no single method that works best for every situation and that all assessment methodologies have advantages and disadvantages [4, 5]. However, given its relative ease of access and low cost, assessment of urine is frequently undertaken in both clinical and research settings. Of the three most common urine-based assessments available (color, specific gravity, and osmolality), urine osmolality and urine specific gravity (USG) are often viewed as the more valid options because evaluating urine color involves more sources of error (e.g., room lighting, evaluator experience, urine collection method) [6]. While there is variation in the literature, a threshold for urine osmolality that is frequently used to define hypohydration is ≥800 mOsm/kg [7, 8].

Based on a urine osmolality threshold of ≥800 mOsm/kg, it has been estimated that one-third of Americans are supposedly hypohydrated at any given time [9]. Furthermore, people identifying as black have been observed to have elevated urine osmolality, suggesting that the prevalence of hypohydration could be higher in this group [9, 10]. Studies showing lower water intakes among black individuals in the United States also support this hypothesis [10].

Recent research by Robinson et al. [11] suggests that these racial/ethnic differences in supposed hypohydration status are at least partly driven by differences in socioeconomic deprivation, which may ultimately impact the type and amount of fluid people consume. For example, black individuals may view tap water as less safe than whites do [12], a justifiable concern given the major water contamination crises that have occurred in black-majority localities like Flint, Michigan and Newark, New Jersey. However, another factor that may be playing a role in the higher rate of supposed hypohydration in blacks is fluid-independent variations in urine creatinine concentrations. Multiple large epidemiological studies have found that serum and urine creatinine are elevated in black individuals (e.g., [13, 14]), and while variances in fluid intake could provide an explanation for these findings, there is a strong basis for thinking that these differences may be partly independent of fluid intake. Specifically, serum creatinine increases in a linear fashion with African ancestry levels [15, 16]. In one study of UK Biobank participants, African ancestry levels explained over 70% of the variability in serum creatinine in men and women, and adjustment for socioeconomic deprivation did not attenuate the association [16].

With this literature in mind, this investigation’s goal was to examine if racial differences in urine creatinine and a hydration biomarker (urine osmolality) were apparent in a sample of American adults after accounting for the influences of age, fluid intake, urine flow rate, and socioeconomic status. We hypothesized that individuals identifying as black would have higher urine creatinine concentrations and urine osmolality than whites, even after accounting for age, socioeconomic status, fluid intake, and urine flow rate.

Methods

Design and participants

The present study involved a secondary analysis of publicly available cross-sectional data from the National Health and Nutrition Examination Survey (NHANES). Survey years 2009–2012 were utilized, as urine osmolality was available only for that timespan. Files containing de-identified individual-level data were downloaded from the NHANES website. The NHANES research protocols were reviewed and approved by the National Center for Health Statistics Ethics Review Board, and participants gave their written informed consent before participating. This study’s protocol was submitted to the Human Subjects Review Committee of the College of Health Sciences at Old Dominion University and was determined to have exempt status.

For the 2009–2012 survey cycles, 26,215 individuals were screened, and 20,015 completed interviews and examinations (76% response rate). Additional exclusions for this analysis were made for individuals who were pregnant, those less than 20 years of age, and those who had missing values for variables of interest, including urine osmolality, urine creatinine, urine flow rate, anthropometrics, dietary data, and income-to-poverty ratio (PIR). In addition, this analysis relied on a smaller subset of black and white adult participants who were matched on important characteristics (age, fluid intake, urine flow rate, socioeconomic status). The matching process is described in detail later. Ultimately, 305 black men, 305 white men, 388 black women, and 388 white women were included in the analysis.

Urine assessments

Spot urine samples were collected at mobile examination centers (MEC), with participants being asked to record the time of their last urination before arrival. Participants’ MEC visits were randomly assigned to occur in the morning, afternoon, or evening. Participants were sent a reminder letter before their scheduled visit with instructions to record the time of their last urine void on a card. At the MEC visit, participants provided a urine sample, with instructions to completely empty their bladder into a container. Urine volume and collection time were recorded by study staff. Based on this information, flow rate of urine was calculated as follows: volume of urine / time (min) since last urination. This was then recalculated to a daily value (mL/24 h).

Urine osmolality was quantified with an OSMETTE II Model 5005, Automatic Osmometer (Precision Systems, Inc), which uses a freezing point depression method.

Urinary creatinine concentration was assessed utilizing a Roche/Hitachi Modular P Chemistry Analyzer, which employs an enzymatic (creatinase) method.

Self-reported race

Background demographic information was collected in participants’ homes during interviews. Self-reported race/ethnicity was based on series of questions about the participant’s ethnic origins and racial identity. Responses to these questions were then used by NHANES personnel to group participants into one of the following five categories: Mexican American, other Hispanic, non-Hispanic white, non-Hispanic black, and other race or multi-racial. For the present analysis, only non-Hispanic black and white participants were included because differences in creatinine in previous studies have been most consistently observed in these two groups.

Other variables

Regarding sex, NHANES uses the term gender in their documentation, but this analysis uses the descriptor sex because there are only two possible responses for the variable in the NHANES data files (male or female). Socioeconomic status was quantified via the PIR, which is the total income of the household where the participant lives divided by the income associated with poverty guidelines relative to family size, as well as the appropriate year and state. Values of greater than 5.0 were recoded as 5.0 by NHANES staff because of potential disclosure concerns.

Because of their potential to impact urine osmolality [17, 18], intakes of fluid (moisture in g/day), sodium (mg/day), and protein (g/day) were assessed with a 24-hour recall method that is based on the United States Department of Agriculture Automated Multiple Pass Method. Dietary moisture reflects all water intake from foods and beverages. Of these three dietary variables, dietary moisture is likely the most important determinant of urine concentration and was selected as a variable for matching. The validity of the moisture intake estimates from this methodology is unclear [19]. Thus, we decided to not only match black and white participants on dietary moisture intake but also urine flow rate, because urine volume is a more objective measure and strongly associates with total fluid intake [20].

Anthropometric variables (body mass, height, waist circumference) were measured at in-person MEC visits. Because lean body mass (LBM) has been previously shown to associate positively with urine creatinine and markers of urine concentration [21, 22], validated equations were used to estimate LBM in kg [23]. The equations are as follows:

  • Men: 0.001 (age in y) + 0.064 (height in cm) + 0.756 (mass in kg)– 0.366 (waist circumference in cm) + 19.363 + (0 for white; 0.432 for black)

  • Women: -0.039 (age in y) + 0.186 (height in cm) + 0.383 (mass in kg)– 0.043 (waist circumference in cm)– 10.683 + (0 for white; 1.085 for black)

Case-control matching

Matching of black and white participants was carried out using the case-control matching function in SPSS (version 29, IBM Corp, Armonk, NY). Variables matched on were age, dietary moisture, urine flow rate, and PIR, with tolerance values of 3 years, 200 g, 200 mL/24 h, and 0.5, respectively. These tolerance values were selected in order to ensure sample sizes of at least 300 for each group. Matching was carried out for men and women separately.

Statistical analysis

Although NHANES data can be analyzed so that the estimates are nationally representative, the present analysis did not do that because of the case-control matching process, which resulted in a smaller subset of the original participants in the 2009–2012 NHANES. The distribution of variables was evaluated by inspecting histograms and Q-Q plots. Most of the variables, with the main exception of urine osmolality, showed a right-skewed distribution; consequently, descriptive statistics are reported using median (25th-75th percentiles). Potential differences between black and white participants on matched variables (age, PIR, moisture intake, urine flow rate) and non-matched variables (urine osmolality, urine creatinine, estimated LBM, dietary protein, dietary sodium) were evaluated using Mann-Whitney U tests. The Spearman’s rank-order correlation (ρ) was used to evaluate the strength of association between variables and urine osmolality. These correlations were carried out separately for black and white participants. Co-efficient ρ sizes of 0.0–0.19, 0.20–0.39, 0.40–0.59, 0.60–0.79, and 0.8–1.0 were used to determine correlation degrees of very weak, weak, moderate, strong, and very strong, respectively. All analyses were done separated by sex. A two-sided p < 0.05 was used as the threshold for statistical significance.

Results

Case-control matching resulted in samples of 305 black and 305 white men, as well as 388 black and 388 white women. Descriptive statistics for black and white participants by sex are reported in Table 1. As expected, there were no significant group differences for matched variables (age, PIR, moisture intake, urine flow rate), with p values all ≥0.87 for men and ≥0.89 for women. Further, dietary sodium and protein intakes did not differ significantly between black and white participants (all p values ≥.079). In contrast, the Mann-Whitney U tests identified statistically significant differences between black and white participants for estimated LBM (p≤.001) and urine creatinine (p < .001) in both men and women. Regarding urine osmolality, black women had higher values than white women (p = .001) but there was no racial difference in men (p = .417).

Table 1. Descriptive statistics.

Men Women
White (n = 305) Black (n = 305) p White (n = 388) Black (n = 388) p
Age (years) 53.0 (34.0–64.5) 53.0 (33.5–63.5) .912 50.0 (36.0–63.0) 50.0 (36.0–62.8) .903
PIR 2.1 (1.2–4.5) 1.9 (1.3–4.5) .892 1.6 (1.0–3.6) 1.7 (1.0–3.8) .999
UFR (mL/24 h) 893 (616–1,320) 890 (625–1,300) .873 716 (477–1,037) 719 (473–1,032) .891
Moisture (g) 2,650 (2,136–3,227) 2,671 (2,117–3,222) .915 2,174 (1,674–2,776) 2,150 (1,647–2,754) .916
LBM (kg) 58.9 (53.2–65.9) 61.8 (53.8–70.5) .001 42.2 (37.2–47.5) 45.5 (41.2–51.0) < .001
Dietary sodium (mg) 3,728 (2,716–4,867) 3,859 (2,768–5,082) .616 2,591 (1,897–3,554) 2,790 (1,967–3,772) .079
Dietary protein (g) 84 (64–111) 90 (64–122) .149 60 (46–80) 64 (45–86) .184
Urine creatinine (mg/dL) 150 (99–200) 171 (127–228) < .001 108 (68–160) 147 (94–211) < .001
Urine osmolality (mOsm/kg) 737 (559–877) 731 (582–888) .417 656 (436–817) 723 (516–878) .001

LBM, lean body mass; PIR, poverty-to-income ratio; UFR, urine flow rate. Values are shown as median (25th-75th percentile).

Among black men, there were significant correlations between urine osmolality and age (ρ = -.39, p < .001), urine creatinine (ρ = .68, p < .001), LBM (ρ = .29, p < .001), and urine flow rate (ρ = -.25, p < .001). The effect sizes of these relationships are weak for age, LBM, and urine flow rate, while the effect size is strong for urine creatinine. Among white men, there were significant correlations between urine osmolality and age (ρ = -.23, p < .001), urine creatinine (ρ = .70, p < .001), LBM (ρ = .20, p < .001), and urine flow rate (ρ = -.32, p < .001). As with black men, the effect sizes of these relationships for white men are weak for age, LBM, and urine flow rate, while the effect size is strong for urine creatinine. Moisture intake was not significantly associated with urine osmolality in men. Since urine creatinine was most strongly associated with urine osmolality, Fig 1 shows the association between these two variables for black and white men separately, along with median values presented using horizontal and vertical lines.

Fig 1. Relationship between urine creatinine and urine osmolality in white (triangles) and black (circles) men.

Fig 1

Median values are represented by solid (black men) and dashed (white men) lines.

Among black women, there were significant correlations between urine osmolality and age (ρ = -.39, p < .001), urine creatinine (ρ = .75, p < .001), LBM (ρ = .17, p < .001), moisture intake (ρ = -.11, p = .037), and urine flow rate (ρ = -.31, p < .001). Effect sizes of these relationships are very weak for LBM and moisture intake, weak for age and urine flow rate, and strong for urine creatinine. Among white women, there were significant correlations between urine osmolality and age (ρ = -.31, p < .001), urine creatinine (ρ = .72, p < .001), LBM (ρ = .17, p < .001), moisture intake (ρ = -.17, p < .001), and urine flow rate (ρ = -.27, p < .001). Effect sizes of these relationship for white women are very weak for LBM and moisture intake, weak for age and urine flow rate, and strong for urine creatinine. Since urine creatinine was most strongly associated with urine osmolality in women, Fig 2 shows the association between these two variables for blacks and whites separately, along with median values presented using horizontal and vertical lines.

Fig 2. Relationship between urine creatinine and urine osmolality in white (triangles) and black (circles) women.

Fig 2

Median values are represented by solid (black women) and dashed (white women) lines.

Discussion

The main hypothesis of this study was that black individuals, as compared to their white counterparts, would produce higher urine creatinine concentrations and urine osmolality after accounting for age, socioeconomic status, fluid intake, and urine flow rate. Indeed, black women were found to have significantly higher values of both urine osmolality and urine creatinine than white women. Black men were found to have significantly higher values of urine creatinine than white men, but there were no differences found in urine osmolality between black men and white men. The consistent finding of elevated urine creatinine in black women and men is most likely explained by the fact that increasing African ancestry levels are strongly and positively associated (R2 = 0.7) with serum creatinine levels [16]. While self-reported race is not equivalent to measuring genetic ancestry, it is generally effective for classifying people into ancestral clusters [24]. In one study of women from New York City, for example, genetic ancestry levels were 77.6% African and 75.1% European for those who self-identified as black and white, respectively [25].

Black women had significantly higher urine osmolality compared to white women, while on the other hand, there was no significant difference in urine osmolality between black men and white men. This could be due to the relative sex differences in urine creatinine between black and white participants. Specifically, the median urine creatinine for black women was 36% higher than for white women, while this relative racial difference was only 14% among men. A potential explanation for the larger relative urine creatinine difference in women is that median estimated LBM was 7.8% higher in black women relative to white women, while for black and white men this relative difference in LBM was apparently smaller (4.9%). Creatinine is a byproduct of muscle metabolism and generally associates strongly with total amounts of skeletal muscle mass [26], a major component of LBM. Thus, a larger relative difference in LBM between black and white women could translate to a larger relative difference in urine creatinine as compared to men.

Although urine osmolality did not vary significantly between black and white men (even with a difference in urine creatinine), it is possible that other urine-based measures of hydration status may differ by self-reported race. USG, for instance, is a common measure of hydration status that tends to be more frequently used in field settings than urine osmolality due to the availability of portable, relatively inexpensive urine refractometers. Notably, as compared to urine osmolality, USG is more substantially impacted by variations in urine creatinine [27]. This is because USG is influenced by both molecule number and size, while urine osmolality depends only on molecule number [27]. The relatively large molecular weight of creatinine means that any increases in creatinine will raise USG more than urine osmolality. In support of this contention, a prior study of NHANES showed that black individuals were more likely than whites to have an elevated USG (>1.02) [28]. Unfortunately, USG is not available for the 2009–2012 years of NHANES, meaning we were unable to address this possibility directly.

Despite significant correlations between several variables of interest, only urine creatinine had a large effect size with urine osmolality. Other factors with a weaker relationship to urine osmolality included age, LBM, and urine flow rate for both sexes and races, and moisture intake for females of both races. These findings emphasize the strong relationship between urine creatinine and urine osmolality. Additionally, it further supports the idea that any factor that is associated with higher creatinine—including self-reported black race or higher African ancestry—is a potentially important determinant of urine osmolality.

Despite having a weaker effect size, estimated LBM was statistically significant in its correlation with urine osmolality. The relatively weak association (ρ = 0.17–0.29) is likely due to using an estimated value of LBM derived from predictive equations [23]. It is possible that LBM and osmolality would have had a stronger association if more direct measures of LBM were taken. Additionally, muscle mass is just one of the components of LBM, with additional components such as water weight, organ tissue, and bone mass. This could have also contributed to the weaker association between LBM and osmolality as compared to what would be expected between muscle mass and osmolality.

Another finding of this study is that black individuals had higher amounts of LBM than whites, which could at least partially explain the racial differences in urine creatinine. Indeed, some limited research supports the notion that having a high African ancestry admixture is associated with lower fat mass and higher LBM [29, 30]. Thus, the previously noted positive association between African ancestry and concentrations of serum creatinine [16] may be partly driven by racial differences in LBM.

Another possibility to consider is that the higher LBM observed among black participants may reflect a truly greater daily water requirement. In adults, fat-free mass is typically comprised of 70–75% water [31], meaning that those with greater amounts of LBM could theoretically require greater water intakes to maintain their body water stores. Since we did not match on LBM, it is therefore possible that the higher LBM and urine creatinine observed in black participants reflects a truly greater water need. However, other research has shown that the variance in daily water turnover is not explained well by anthropometric variables like weight, height, or body mass index [32], and the necessity of taking body size into consideration when making fluid intake recommendations for adults is uncertain [31].

The results of this research have allowed further insight into the differences in urine creatinine and osmolality between black and white Americans. However, there are a few limitations of the research to take into consideration. While urine osmolality and USG are both commonly used to assess hydration status, this specific study only considered urine osmolality due to a lack of USG data for years 2009–2012. Implementing both USG and urine osmolality assessments while evaluating urine creatinine would allow for a more detailed assessment of how urine creatinine impacts both of these urine markers in different racial groups. Another limitation of the study is the assessment of urine flow rate. Participants were reminded to record the time of their last urine void before their scheduled visit. This time was used in the urine flow rate calculation, which is easily subject to error. For example, if participants estimated the time this occurred instead of recording it in real time, the urine flow rate calculation may have been altered significantly, negatively affecting the present results. In addition, the use of spot urine sampling, which is the method employed by NHANES, is not as valid as 24-hour urine collections, especially when it comes to being used as an indicator of daily fluid intake [20].

Conclusion

Prior research has found elevated urine osmolality in individuals self-identifying as black [9, 10], but the reasons for this difference remain under-investigated. Although studies support the idea that fluid intakes are lower among individuals self-identifying as black in the United States, and that socioeconomic deprivation likely plays an important role in this disparity, previous research has also found that serum creatinine levels increase in those with higher levels of African ancestry, even when adjusted for socioeconomic deprivation [16]. The present analysis extends these prior results to a large sample of American adults, including black and white women and men matched on age, fluid intake measures (dietary moisture and urine flow rate), and socioeconomic status. Overall, urine creatinine was elevated among black men and women compared to whites after accounting for the previously listed influencing factors, although the effect was larger in women and a difference in urine osmolality was not observed in men. Further research should be conducted to examine whether the greater LBM and urine creatinine observed in black participants reflect an actual increase in water requirements. For the time being, our results suggest that researchers and practitioners should use caution when directly comparing urine hydration markers between racial groups.

Supporting information

S1 File. Dataset.

(XLS)

pone.0304803.s001.xls (1.1MB, xls)

Data Availability

All relevant data are within the manuscript and its Supporting Information files.

Funding Statement

The author(s) received no specific funding for this work.

References

  • 1.Wittbrodt MT, Millard-Stafford M. Dehydration impairs cognitive performance: A meta-analysis. Med Sci Sports Exerc. 2018;50(11):2360–2368. doi: 10.1249/MSS.0000000000001682 [DOI] [PubMed] [Google Scholar]
  • 2.Deshayes TA, Pancrate T, Goulet ED. Impact of dehydration on perceived exertion during endurance exercise: A systematic review with meta-analysis. J Exerc Sci Fit. 2022;20(3):224–235. doi: 10.1016/j.jesf.2022.03.006 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Vanhaecke T, Perrier ET, Melander O. A journey through the early evidence linking hydration to metabolic health. Ann Nutr Metab. 2021;76(Suppl. 1):4–9. [DOI] [PubMed] [Google Scholar]
  • 4.Armstrong LE. Assessing hydration status: the elusive gold standard. J Am Coll Nutr. 2007;26(sup 5):575S–584S. doi: 10.1080/07315724.2007.10719661 [DOI] [PubMed] [Google Scholar]
  • 5.Barley OR, Chapman DW, Abbiss CR. Reviewing the current methods of assessing hydration in athletes. J Int Soc Sports Nutr. 2020;17(1): 52. doi: 10.1186/s12970-020-00381-6 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Wardenaar F, Armistead S, Boeckman K, Butterick B, Youssefi D, Thompsett D, et al. Validity of urine color scoring using different light conditions and scoring techniques to assess urine concentration. J Athl Train. 2022;57(2):191–198. doi: 10.4085/1062-6050-0389.21 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Kenefick RW, Cheuvront SN. Hydration for recreational sport and physical activity. Nutr Rev. 2012;70(suppl 2):S137–S142. doi: 10.1111/j.1753-4887.2012.00523.x [DOI] [PubMed] [Google Scholar]
  • 8.Armstrong LE, Johnson EC, McKenzie AL, Munoz CX. An empirical method to determine inadequacy of dietary water. Nutr. 2016;32(1):79–82. doi: 10.1016/j.nut.2015.07.013 [DOI] [PubMed] [Google Scholar]
  • 9.Chang T, Ravi N, Plegue MA, Sonneville KR, Davis MM. Inadequate hydration, BMI, and obesity among US adults: NHANES 2009–2012. Ann Fam Med. 2016;14(4):320–324. doi: 10.1370/afm.1951 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Brooks CJ, Gortmaker SL, Long MW, Cradock AL, Kenney EL. Racial/ethnic and socioeconomic disparities in hydration status among US adults and the role of tap water and other beverage intake. Am J Public Health. 2017;107(9):1387–1394. doi: 10.2105/AJPH.2017.303923 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Robinson AT, Linder BA, Barnett AM, Jeong S, Sanchez SO, Nichols OI, et al. Cross-sectional analysis of racial differences in hydration and neighborhood deprivation in young adults. Am J Clin Nutr. 2023;118(4):822–833. doi: 10.1016/j.ajcnut.2023.08.005 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Onufrak SJ, Park S, Sharkey JR, Merlo C, Dean WR, Sherry B. Perceptions of tap water and school water fountains and association with intake of plain water and sugar‐sweetened beverages. J School Health. 2014;84(3):195–204. doi: 10.1111/josh.12138 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Jones CA, McQuillan GM, Kusek JW, Eberhardt MS, Herman WH, Coresh J, et al. Serum creatinine levels in the US population: third National Health and Nutrition Examination Survey. Am J Kidney Dis. 1998;32(6):992–999. doi: 10.1016/s0272-6386(98)70074-5 [DOI] [PubMed] [Google Scholar]
  • 14.Jain RB. Trends in the levels of urine and serum creatinine: data from NHANES 2001–2014. Environ Sci Pollut Res. 2017;24(11):10197–10204. doi: 10.1007/s11356-017-8709-y [DOI] [PubMed] [Google Scholar]
  • 15.Udler MS, Nadkarni GN, Belbin G, Lotay V, Wyatt C, Gottesman O, et al. Effect of genetic African ancestry on eGFR and kidney disease. J Am Soc Nephrol. 2015;26(7):1682–1692. doi: 10.1681/ASN.2014050474 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Mariño-Ramírez L, Sharma S, Rishishwar L, Conley AB, Nagar SD, Jordan IK. Effects of genetic ancestry and socioeconomic deprivation on ethnic differences in serum creatinine. Gene. 2022;837:146709. doi: 10.1016/j.gene.2022.146709 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Miller M, Price JW, Longley LP. Effect of varying intake of protein and salts on the composition and specific gravity of urine. J Clin Investig. 1941;20(1):31–38. doi: 10.1172/JCI101192 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 18.Heer M, Frings-Meuthen P, Titze J, Boschmann M, Frisch S, Baecker N, et al. Increasing sodium intake from a previous low or high intake affects water, electrolyte and acid–base balance differently. Br J Nutr. 2009;101(9):1286–1294. doi: 10.1017/S0007114508088041 [DOI] [PubMed] [Google Scholar]
  • 19.Gandy J. Water intake: validity of population assessment and recommendations. Eur J Nutr. 2015;54(Suppl 2):11–16. doi: 10.1007/s00394-015-0944-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Perrier E, Rondeau P, Poupin M, Le Bellego L, Armstrong LE, Lang F, et al. Relation between urinary hydration biomarkers and total fluid intake in healthy adults. Eur J Clin Nutr. 2013;67(9):939–943. doi: 10.1038/ejcn.2013.93 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Ehlert AM, Wilson PB. The associations between body mass index, estimated lean body mass, and urinary hydration markers at the population level. Meas Phys Educ Exerc Sci. 2021;25(2):163–170. [Google Scholar]
  • 22.Wilson PB. Associations of urine specific gravity with body mass index and lean body mass at the population level: Implications for hydration monitoring. Int J Sport Nutr Exerc Metab. 2021;31(6):475–481. doi: 10.1123/ijsnem.2021-0140 [DOI] [PubMed] [Google Scholar]
  • 23.Lee DH, Keum N, Hu FB, Orav EJ, Rimm EB, Sun Q, et al. Development and validation of anthropometric prediction equations for lean body mass, fat mass and percent fat in adults using the National Health and Nutrition Examination Survey (NHANES) 1999–2006. Br J Nutr. 2017;118(10):858–866. doi: 10.1017/S0007114517002665 [DOI] [PubMed] [Google Scholar]
  • 24.Yaeger R, Avila-Bront A, Abdul K, Nolan PC, Grann VR, Birchette MG, et al. Comparing genetic ancestry and self-described race in African Americans born in the United States and in Africa. Cancer Epidemiol Biomarkers Prev. 2008;17(6):1329–1338. doi: 10.1158/1055-9965.EPI-07-2505 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Lee YL, Teitelbaum S, Wolff MS, Wetmur JG, Chen J. Comparing genetic ancestry and self-reported race/ethnicity in a multiethnic population in New York City. J Genet. 2010;89:417–423. doi: 10.1007/s12041-010-0060-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Wang Z, Gallagher D, Nelson ME, Matthews DE, Heymsfield SB. Total-body skeletal muscle mass: evaluation of 24-h urinary creatinine excretion by computerized axial tomography. Am J Clin Nutr. 1996;63(6):863–869. doi: 10.1093/ajcn/63.6.863 [DOI] [PubMed] [Google Scholar]
  • 27.Voinescu GC, Shoemaker M, Moore H, Khanna R, Nolph KD. The relationship between urine osmolality and specific gravity. Am J Med Sci. 2002;323(1):39–42. doi: 10.1097/00000441-200201000-00007 [DOI] [PubMed] [Google Scholar]
  • 28.Mao W, Zhang H, Xu Z, Geng J, Zhang Z, Wu J, et al. Relationship between urine specific gravity and the prevalence rate of kidney stone. Transl Androl Urol. 2021;10(1):184–194. doi: 10.21037/tau-20-929 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Cardel M, Higgins PB, Willig AL, Keita AD, Casazza K, Gower BA, et al. African genetic admixture is associated with body composition and fat distribution in a cross-sectional study of children. Int J Obes. 2011;35(1):60–65. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Shaffer JR, Kammerer CM, Reich D, McDonald G, Patterson N, Goodpaster B, et al. (2007). Genetic markers for ancestry are correlated with body composition traits in older African Americans. Osteoporos Int. 2007;18:733–741. [DOI] [PubMed] [Google Scholar]
  • 31.EFSA Panel on Dietetic Products, Nutrition, and Allergies (NDA). Scientific opinion on dietary reference values for water. EFSA J. 2010;8(3):1459. [Google Scholar]
  • 32.Raman A, Schoeller DA, Subar AF, Troiano RP, Schatzkin A, Harris T, et al. Water turnover in 458 American adults 40–79 yr of age. Am J Physiol Renal Physiol. 2004;286(2):F394–F401. doi: 10.1152/ajprenal.00295.2003 [DOI] [PubMed] [Google Scholar]

Decision Letter 0

William M Adams

23 Apr 2024

PONE-D-24-10364Differences in urine creatinine and osmolality between black and white Americans after accounting for age, moisture intake, urine volume, and socioeconomic statusPLOS ONE

Dear Dr. wilson,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Jun 07 2024 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

William M. Adams

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at 

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and 

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information. 

3. We are unable to open your Supporting Information file "Final dataset_matched_blackwhite_bothsexes.sav". Please kindly revise as necessary and re-upload.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: I read with great interest this manuscript and I would like to congratulate with the authors for this well-done work. The manuscript is well-written, the topic is of interest and the discussion provides severaly hypotheses explaining the reported findings.

I have only a question: did the authors consider salt intake as a variable that might affect hydration parameters and possible differences among diet between the two selected populations? Despite matching for social-economic status could have reduced differences between black and white people, do you think that black individuals might be characterized by a higher salt intake (Yoon et al., 2024)? Maybe another point of discussion might be related to renal handling and reabsorption of salt and water, if differences are present based on racial/ethnic differences.

Reviewer #2: In this manuscript, the authors use NHANES data combined from 2009-2012 to examine differences in hydration status (urine osmolality) and urine creatinine. Based on higher urine osmolality in black women compared to white women and correlations between urine osmolality and urine creatinine among racial groups, the authors recommend caution when comparing urine hydration markers between racial groups.

Major Comments:

I am not sure that the case matching makes sense in this case. Can the authors better describe why each group was matched for both moisture intake and UFR simultaneously? I would expect these variables to be correlated with one another, in which case wouldn’t it make sense to only match by UFR if the authors believe that is more representative of the actual fluid intake of participants? https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2827892/

Can the authors also further describe how the case-control matching was performed (i.e., how was this specific subset generated)? Particularly because the sample size has reduced considerably to achieve this matching.

Dietary salt intake may also influence creatinine clearance and appears to be something that should be accounted for: https://pubmed.ncbi.nlm.nih.gov/35157527/

Is LBM not also associated with urinary creatinine? It may be that the LBM differences (and perhaps physical activity as well) are driving these differences in creatinine. You acknowledge this in your female group as this possibly accounting for the differences. The study you cite regarding the genetic ancestry differences contributing to creatinine differences included BMI but not lean body mass in the model. So could it actually be the LBM driving these differences?

The lean body mass differences between each group would likely necessitate greater fluid intake between groups. Thus, because fluid intake was equated, it seems natural that there is higher osmolality in the group with higher fluid intake requirements, at least in the case of the females where this difference was more apparent.

I am not sure how the conclusion has been reached that caution should be used when directly comparing urine hydration markers and rates of hypohydration between racial groups. Based on the results presented, it would seem more accurate to conclude when accounting for age, PIR, UFR, and moisture intake that there are no significant differences in urine osmolality between men of different races. However, when accounting for these factors in females, urine osmolality remained higher in black females, perhaps related to the greater magnitude of LBM and perhaps fluid requirements of those specific females. Also, the “rates” of hypohydration were not examined in this study (for example, by using the cutoff of >800mOsm/kg), so I think it may be a stretch to include this as part of the conclusion.

Minor comments:

Lines 60-68 – I do not think urine specific gravity needs to be described here since it is not mentioned elsewhere in the results. At least not in terms of its commonly used cutoff point.

Can the authors also briefly comment somewhere in the limitations on the drawbacks of spot urine samples vs 24hr urine samples?

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

PLoS One. 2024 May 31;19(5):e0304803. doi: 10.1371/journal.pone.0304803.r002

Author response to Decision Letter 0


8 May 2024

We appreciate the reviewers’ time and effort providing feedback on our manuscript. We have used their helpful comments and suggestions to improve the overall quality of the manuscript. Each of the reviewer’s comments is listed below, followed by our responses.

Reviewer #1

I read with great interest this manuscript and I would like to congratulate with the authors for this well-done work. The manuscript is well-written, the topic is of interest and the discussion provides severaly hypotheses explaining the reported findings.

• Thank you for taking the time to review our manuscript. We appreciate the positive comments regarding our work.

I have only a question: did the authors consider salt intake as a variable that might affect hydration parameters and possible differences among diet between the two selected populations? Despite matching for social-economic status could have reduced differences between black and white people, do you think that black individuals might be characterized by a higher salt intake (Yoon et al., 2024)? Maybe another point of discussion might be related to renal handling and reabsorption of salt and water, if differences are present based on racial/ethnic differences.

• This is an interesting question. We did consider including sodium in the analysis during the initial planning stages of the project. However, based on some previous work from our lab that showed sodium was not a major determinant of urine specific gravity in American adults (https://pubmed.ncbi.nlm.nih.gov/34470907/), we ultimately decided to omit it from the original analysis. That being said, we agree with the reviewer that it may be valuable for readers to see the data on dietary sodium. Thus, we have added the median (25-75th percentile) values along with Mann Whitney U test results to Table 1. In both men and women, there were no statistically significant differences in sodium intakes between black and white individuals. We also added the statistics for dietary protein, since it could potentially impact urine osmolality via the production of urea. Similar to sodium, there were no statistically significant differences in protein intakes between black and white individuals.

Reviewer #2:

In this manuscript, the authors use NHANES data combined from 2009-2012 to examine differences in hydration status (urine osmolality) and urine creatinine. Based on higher urine osmolality in black women compared to white women and correlations between urine osmolality and urine creatinine among racial groups, the authors recommend caution when comparing urine hydration markers between racial groups.

Major Comments:

I am not sure that the case matching makes sense in this case. Can the authors better describe why each group was matched for both moisture intake and UFR simultaneously? I would expect these variables to be correlated with one another, in which case wouldn’t it make sense to only match by UFR if the authors believe that is more representative of the actual fluid intake of participants? https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2827892/

• Thank you for the insightful comment/question. In general, we decided to use the case-control matching design for a couple of reasons. The primary reason is that we find that, in comparison to some other statistical techniques, the results from case-control matching are fairly simple for most readers (including non-scientist practitioners) to understand and interpret. For example, multivariate regression may have some advantages over case-control matching, particularly from a statistical power perspective, but the output is more difficult for many practitioners to comprehend. In contrast, with case-control matching, the reader can easily understand that we made the two groups roughly equivalent on the matched variables.

• As to why we decided to match on both moisture intake and urine volume, we feel that it does provide a modestly higher degree of rigor than matching on one or the other. Yes, we agree that urine volume is likely to correlate relatively well with total fluid intake, but the correlations can range from 0.3 to 0.8 depending on whether 24-h vs. spot urine collections are used (https://www.nature.com/articles/ejcn201393; https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8411265/). In addition, it is certainly possible that a person could have a high fluid intake but still have a low-to-moderate urine output due to high sweat losses. While we acknowledge that that scenario likely represents a minority of people, we still feel that matching on both dietary moisture intake and urine volume is the most robust approach.

Can the authors also further describe how the case-control matching was performed (i.e., how was this specific subset generated)? Particularly because the sample size has reduced considerably to achieve this matching.

• Case-control matching was carried out with SPSS software. An example of how the process works can be viewed in the following video: https://www.youtube.com/watch?v=E6nzTTGIWss. In our study, we selected 4 variables to match white and black participants on (age, PIR, moisture intake, and urine flow rate). These four variables were selected because they are potential confounders of the relationship between race and our outcome urine measures (creatinine and osmolality). Tolerance/fuzz values are set for each variable, which means that each member of a matched pair needs to have values for a matched variable within the range of tolerance set.

• As pointed out by the reviewer, we acknowledge that the case-control matching process reduced our sample size. However, all our sex-race subgroups still had a sample size of >300, which is large enough to detect relatively small effect sizes. A bigger sample size could potentially yield more statistically significant effects, but the practical relevance of those very small effects would be questionable.

Dietary salt intake may also influence creatinine clearance and appears to be something that should be accounted for: https://pubmed.ncbi.nlm.nih.gov/35157527/

• Thanks for the suggestion. Reviewer 1 also requested that we take into consideration dietary sodium intake. We have added median values of sodium intake to Table 1. In both sexes, there were no statistically significant differences in sodium intake between black and white participants.

Is LBM not also associated with urinary creatinine? It may be that the LBM differences (and perhaps physical activity as well) are driving these differences in creatinine. You acknowledge this in your female group as this possibly accounting for the differences. The study you cite regarding the genetic ancestry differences contributing to creatinine differences included BMI but not lean body mass in the model. So could it actually be the LBM driving these differences?

• Yes, we do think that it is likely that some of the observed racial differences in urine creatinine are being driven by LBM differences, or more precisely, differences in skeletal muscle mass (which is not available in the NHANES dataset). We briefly mentioned this in our Discussion section (“Another finding of this study is that black individuals had higher amounts of LBM than whites, which could partly explain the racial differences in urine creatinine”), but we have added some additional text to the Discussion about this possibility. Specifically, some limited research supports the notion that having a high African ancestry admixture is associated with lower fat mass and higher lean mass. Thus, the association between African ancestry and higher concentrations of serum creatinine may be at least partly driven by racial differences in lean mass.

The lean body mass differences between each group would likely necessitate greater fluid intake between groups. Thus, because fluid intake was equated, it seems natural that there is higher osmolality in the group with higher fluid intake requirements, at least in the case of the females where this difference was more apparent.

• We see where the reviewer is coming from, but we do not necessarily agree that having a higher lean body mass necessitates a higher fluid intake. In multiple population studies, there is little-to-no relationship between BMI (or body size) and urine volume, meaning that larger individuals generally probably aren’t naturally choosing to consume larger amounts of fluid on a daily basis than smaller individuals (see example studies listed below). We realize that BMI is not the same as LBM, but the two measures tend to correlate moderately with one another, at least when quantified as absolute amounts (total kg). Furthermore, a study by Raman et al. (2004) reported that anthropometric variables explained little of the variance in daily water turnover in adults (https://journals.physiology.org/doi/full/10.1152/ajprenal.00295.2003). In total, there is not clear evidence that fluid intake requirements or water turnover consistently depend on body size. Although we agree that there could in theory be a higher daily fluid intake requirement for larger individuals with more LBM, that idea is still up for debate. Nonetheless, we have added the following text to the Discussion section to address the issue raised by the reviewer. “Another possibility to consider is that the higher LBM observed among black participants may reflect a truly greater daily water requirement. In adults, fat-free mass is typically comprised of 70-75% water [31], meaning that those with greater amounts of LBM could theoretically require greater water intakes to maintain their body water stores. Since we did not match on LBM, it is therefore possible that the higher LBM and urine creatinine observed in black participants reflects a truly greater water need. However, other research has shown that the variance in daily water turnover is not explained well by anthropometric variables like weight, height, or body mass index [32], and the necessity of taking body size into consideration when making fluid intake recommendations for adults is uncertain [31].”

https://journals.lww.com/CJASN/fulltext/2011/11000/Urine_Volume_and_Change_in_Estimated_GFR_in_a.14.aspx

https://link.springer.com/article/10.1186/1471-2369-14-246

https://ehp.niehs.nih.gov/doi/full/10.1289/ehp.1408944

I am not sure how the conclusion has been reached that caution should be used when directly comparing urine hydration markers and rates of hypohydration between racial groups. Based on the results presented, it would seem more accurate to conclude when accounting for age, PIR, UFR, and moisture intake that there are no significant differences in urine osmolality between men of different races. However, when accounting for these factors in females, urine osmolality remained higher in black females, perhaps related to the greater magnitude of LBM and perhaps fluid requirements of those specific females. Also, the “rates” of hypohydration were not examined in this study (for example, by using the cutoff of >800mOsm/kg), so I think it may be a stretch to include this as part of the conclusion.

• We appreciate the reviewer’s perspectives on our conclusions. Taking their comments into consideration, we have removed reference to “rates of hypohydration.” In addition, we have added a sentence highlighting the need for additional research on whether the greater amounts of LBM and urine creatinine observed in black participants reflect an actual increase in water requirements.

Minor comments:

Lines 60-68 – I do not think urine specific gravity needs to be described here since it is not mentioned elsewhere in the results. At least not in terms of its commonly used cutoff point.

• We have edited this section as recommended.

Can the authors also briefly comment somewhere in the limitations on the drawbacks of spot urine samples vs 24hr urine samples?

• We have added some text to the limitations section regarding the use of spot urine samples.

Decision Letter 1

William M Adams

20 May 2024

Differences in urine creatinine and osmolality between black and white Americans after accounting for age, moisture intake, urine volume, and socioeconomic status

PONE-D-24-10364R1

Dear Dr. wilson,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. If you have any questions relating to publication charges, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

William M. Adams

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: I would like to thank the authors for addressing my comment and I am totally satisfied with their responses.

Thank you for adding this information on salt and protein intake.

Reviewer #2: All comments have been addressed well by the authors. This manuscript appears suitable for publication in its present form. Well done!

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

**********

Acceptance letter

William M Adams

22 May 2024

PONE-D-24-10364R1

PLOS ONE

Dear Dr. wilson,

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team.

At this stage, our production department will prepare your paper for publication. This includes ensuring the following:

* All references, tables, and figures are properly cited

* All relevant supporting information is included in the manuscript submission,

* There are no issues that prevent the paper from being properly typeset

If revisions are needed, the production department will contact you directly to resolve them. If no revisions are needed, you will receive an email when the publication date has been set. At this time, we do not offer pre-publication proofs to authors during production of the accepted work. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few weeks to review your paper and let you know the next and final steps.

Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

If we can help with anything else, please email us at customercare@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. William M. Adams

Academic Editor

PLOS ONE

Associated Data

    This section collects any data citations, data availability statements, or supplementary materials included in this article.

    Supplementary Materials

    S1 File. Dataset.

    (XLS)

    pone.0304803.s001.xls (1.1MB, xls)

    Data Availability Statement

    All relevant data are within the manuscript and its Supporting Information files.


    Articles from PLOS ONE are provided here courtesy of PLOS

    RESOURCES