AWM: Eating Frequency and Patterns (2013)

Nonino-Borges CB, Martins Borges R, Bavaresco M, Suen VM, Moreira AC, Marchini JS. Influence of meal time on salivary circadian cortisol rhythms and weight loss in obese women. Nutrition, 2007; 23 (5): 385-391.
PubMed ID: 17483007
Study Design:
Randomized Crossover Trial
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Quality Rating:
Neutral NEUTRAL: See Quality Criteria Checklist below.
Research Purpose:

The objective of this study was to determine whether ingestion of a low-calorie diet by obese women at different times during the day would alter circadian cortisol rhythms and to determine the effects on loss of body weight, body composition, RMR and nitrogen balance values.

Inclusion Criteria:
  • Obese women with a body mass index (BMI) of more than 40kg/m2
  • No other endocrine, renal, hepatic or severe psychiatric disorders
  • Not using prescribed drugs or other pharmacologic treatment
  • Provided a signed term of informed consent to participate in the study.
Exclusion Criteria:
  • History or presence of endocrine, renal, hepatic or severe psychiatric disorders
  • Using prescribed drugs or other pharmacologic treatment.
Description of Study Protocol:
  • Recruitment: Selected from among outpatients of the Nutriology Clinic of the Department of Internal Medicine of the University Hospital of Ribeirao Preto, University of Sao Paulo
  • Design: Randomized crossover trial
  • Blinding used: Implied with measurements.


  • Stage One (control stage): The diet was portioned into five meals per day: Breakfast, lunch, afternoon snack, dinner and supper, given at 9:00 a.m., 11:00 a.m., 4:00 p.m. and 8:00 p.m.
  • Stage Two: The subjects received the same diet, but only during the period from 9:00 a.m. to 11:00 a.m.
  • Stage Three: They received the same diet only during the period from 6:00 p.m. to 8:00 p.m.
  • Subjects selected were admitted to a metabolic unit for a period of 64 days that was divided into three hospitalizations of 18 days each, separated by a five-day washout period, which the subjects consumed their usual diets.

Statistical Analysis

  • Two-way analysis of variance was used for the statistical analysis of repeated measurements of changes in salivary cortisol, food intake, RMR, body composition and anthropometric variables across the three stages of treatment
  • Paired T-tests were used to determine the significance of differences between variables measured at the beginning and at the end of each stage
  • The significance level was set at P<0.05.
Data Collection Summary:

Timing of Measurements

Clinical and dietary histories were obtained and anthropometric, physical and laboratory examinations were performed before admission for each subject.

Dependent Variables

  • Salivary circadian cortisol rhythms: Saliva was collected at 8:00 a.m., 12 noon, 5:00 p.m., 7:00 p.m. and 9:00 p.m. on Days One and 18 after admission. Saliva was also collected at 8:00 a.m. on the subsequent day for cortisol measurement. Saliva samples were then transferred to the laboratory and centrifuged at 2,000rpm and stored until assayed.
  • Weight loss: Subjects were weighed on a digital platform-type balance that had a 300-kg capacity and 0.2-kg precision. Height was measured with a vertical rod graduated in 0.5cm. Waist circumference was measured above the umbilical scar. Hip circumference was measured by placing the metric tape around the largest circumference between the waist and knee.
  • Body composition: Evaluated by bioimpedance with a Quantum BIA instrument. Fat-free mass and body fat were estimated by mathematical equations developed for obese women.
  • Resting metabolic rate: Measured at room temperature by indirect calorimetry at the alert subject's bedside after the subject had fasted for 12 hours and slept for eight hours
  • Urine samples: Collected and used for nitrogen determination by the Kjeldahl method. Nitrogen balance was calculated by subtracting urinary urea nitrogen (24-hour urine) plus imperceptible losses from ingested nutrition nitrogen.

Independent Variables

  • Assignment to Stage One, Stage Two or Stage Three
  • During the periods of hospitalization, all food was weighed or measured before and after each meal to calculate amounts ingested.
Description of Actual Data Sample:
  • Initial N: 12 women with a body mass index over 40kg/m2
  • Attrition: 0% dropout rate, all 12 women completed the trial
  • Age: 27±8 years
  • Ethnicity: Not described
  • Other relevant demographics: Outpatients of the nutriology clinic of the Department of Internal Medicine of the University Hospital of Ribeirao Preto, University Sao Paulo
  • Anthropometrics: All subjects had a history of long-term weight gain lasting for more than 10 years and 83% had a family history of obesity
  • Location: University of Sao Paulo, Brazil.
Summary of Results:

Key Findings

  • No significant changes in circadian rhythm for salivary cortisol were noted when the meal times were changed
  • There were no significant differences in mean daily cortisol concentrations across the first, second and third stages on Day One (593±262ng, 655±260ng and 720±391ng per dL) or Day 18 (802±473ng, 670±319ng, and 782±362ng per dL)
  • There were no significant differences across stages in the changes in total energy or nutrient composition that occurred between Days Four and 18 and the first three days of admission
  • Significant reductions in all variables except waist-to-hip ratio were noted in all three stages (P<0.05).

Other Findings

  • Correlations between caloric ingestion and nitrogen balance (R=0.84, P<0.05) and between the amount of protein ingested and nitrogen balance (R=0.93, P<0.05) were noted, as was a correlation between total urinary nitrogen and urinary urea nitrogen (R=0.90, P<0.05)
  • Urinary urea nitrogen averaged 78.7% of total urinary nitrogen.
Author Conclusion:
  • The administration of a hypocaloric diet led to changes in weight, body composition, resting metabolic rate and nitrogen balance, but did not significantly alter salivary circadian cortisol rhythms
  • Further studies of cortisol rhythm and food intake in this field will be required.
Funding Source:
University/Hospital: University of Sao Paulo, Brazil
Reviewer Comments:

Small sample size, power calculation not reported.

Quality Criteria Checklist: Primary Research
Relevance Questions
  1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies) Yes
  2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? Yes
  3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice? Yes
  4. Is the intervention or procedure feasible? (NA for some epidemiological studies) Yes
Validity Questions
1. Was the research question clearly stated? Yes
  1.1. Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified? Yes
  1.2. Was (were) the outcome(s) [dependent variable(s)] clearly indicated? Yes
  1.3. Were the target population and setting specified? Yes
2. Was the selection of study subjects/patients free from bias? ???
  2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Yes
  2.2. Were criteria applied equally to all study groups? Yes
  2.3. Were health, demographics, and other characteristics of subjects described? Yes
  2.4. Were the subjects/patients a representative sample of the relevant population? ???
3. Were study groups comparable? Yes
  3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) No
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Yes
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) Yes
  3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? N/A
  3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.) N/A
  3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")? N/A
4. Was method of handling withdrawals described? Yes
  4.1. Were follow-up methods described and the same for all groups? Yes
  4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Yes
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? Yes
  4.4. Were reasons for withdrawals similar across groups? N/A
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? N/A
5. Was blinding used to prevent introduction of bias? Yes
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? N/A
  5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) Yes
  5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? N/A
  5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? N/A
  5.5. In diagnostic study, were test results blinded to patient history and other test results? N/A
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? Yes
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? Yes
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? N/A
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Yes
  6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? Yes
  6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? N/A
  6.6. Were extra or unplanned treatments described? N/A
  6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? Yes
  6.8. In diagnostic study, were details of test administration and replication sufficient? N/A
7. Were outcomes clearly defined and the measurements valid and reliable? Yes
  7.1. Were primary and secondary endpoints described and relevant to the question? Yes
  7.2. Were nutrition measures appropriate to question and outcomes of concern? Yes
  7.3. Was the period of follow-up long enough for important outcome(s) to occur? Yes
  7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Yes
  7.5. Was the measurement of effect at an appropriate level of precision? Yes
  7.6. Were other factors accounted for (measured) that could affect outcomes? Yes
  7.7. Were the measurements conducted consistently across groups? Yes
8. Was the statistical analysis appropriate for the study design and type of outcome indicators? ???
  8.1. Were statistical analyses adequately described and the results reported appropriately? Yes
  8.2. Were correct statistical tests used and assumptions of test not violated? ???
  8.3. Were statistics reported with levels of significance and/or confidence intervals? Yes
  8.4. Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? N/A
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? N/A
  8.6. Was clinical significance as well as statistical significance reported? Yes
  8.7. If negative findings, was a power calculation reported to address type 2 error? No
9. Are conclusions supported by results with biases and limitations taken into consideration? Yes
  9.1. Is there a discussion of findings? Yes
  9.2. Are biases and study limitations identified and discussed? Yes
10. Is bias due to study's funding or sponsorship unlikely? Yes
  10.1. Were sources of funding and investigators' affiliations described? Yes
  10.2. Was the study free from apparent conflict of interest? Yes