NSA: Serum Proteins (2009)

Study Design:
- Click here for explanation of classification scheme.
Quality Rating:
Research Purpose:

To assess the changes in traditional serum biochemical indicators of nutritional status in a one-year follow-up of patients with restricting type anorexia nervosa.

Inclusion Criteria:

Patients were selected from the Hospital de Mostoles in Madrid, Spain who were:

  • Female
  • Met DSM IV criteria for diagnosis of anorexia nervosa (AN)
  • Adolescent age
  • First in-hospital stay
  • Restrictive type AN.
Exclusion Criteria:
  • Patients who did not meet the inclusion criteria were not considered
  • Of the 26 original participants, data is provided on 14, as they were the ones that completed all of the timed checkpoints over one year.
Description of Study Protocol:


Patients were admitted for in-hospital treatment of anorexia nervosa.


Anthropometric measures and fasting serum samples were collected from participants at:

  • Initial hospitalization (within 10 days of admission, allowing rehydration and stabilization)
  • One month later
  • Six months after admission
  • 12 months after admission.

Subjects were compared with a control group of healthy, non-AN free-living females of similar age (data for initial only).

Statistical Analysis  

  • Mean ± Standard Deviation (SD)
  • Bonferroni methods for pairwise comparison between time points (P<0.01)
  • Student's T-test (P<0.05)
  • Pearson's correlation test
  • SPSS Windows statistical software package version 11.0.1 (SPSS Inc., 1989-2001).
Data Collection Summary:

Timing of Measurements

Anthropometric and biochemical measures were taken at initial (admission up to day 10 of hospitalization), one month, six months and twelve months.

Dependent Variables

  • Albumin
  • Prealbumin
  • Retinol binding protein
  • Transferrin
  • Complement factors C3 and C4
  • Zinc
  • Iron.

Independent Variables


Control Variables

Subjects changes over time were compared to their own data as well as to a control group of healthy adolescent girls (one-time assessment of nutritional status).

Description of Actual Data Sample:
  • Initial N: 26 female patients, 15 female controls
  • Attrition (final N): 12 subjects did not complete the follow-up review points; therefore, final N=14
  • Age: Adolescent.


Some characteristics of the study population are provided below. (Note: AN=patients with anorexia nervosa.)

  Controls AN Initial AN One Month AN Six Months AN 12 Months
Age (year) 15.10±1.40 15.10±2.56 15.19±2.56 15.62±2.57 16.13±2.59
BMI (kg/m2) 20.24±1.25 15.24±1.71 16.59±1.12 17.29±1.11 17.34±1.97
Weight gain (kg) N/A N/A 3.6±2.1 5.4±3.7 5.8±4.5


Madrid, Spain.


Summary of Results:

 Key Findings

  • Weight and weight-related parameters increased significantly after one month in patient treatment and were maintained thereafter
  • Time effect was significant for serum total protein, transferrin, complement factors C3 and C4, but NOT albumin, prealbumin and retinol binding protein (RBP)
  • Transferrin was lower in patients that did not gain weight after the initial month and transferrin increased in patients that gained weight
  • Ferritin values decreased significantly and progressively over time and were higher than controls at all times
  • Phosphorous, calcium, zinc and iron did not change significantly over time for patients with anorexia nervosa. Calcium was significantly lower in patients at all time points than in controls.
  Controls AN Initial AN One Month AN Six Months AN 12 Months ANOVA P-value
Total Protein (g per dL) 7.47±0.27 6.98±0.67 6.96±0.91 7.70±0.52 7.47±0.30 0.007
Albumin (g per dL) 4.41±0.30 4.51±0.69 4.92±0.61 4.67±0.51 4.77±0.45 0.700
Prealbumin (mg per dL) 27.8±5.0 26.3±5.6 28.5±5.2 29.6±5.9 25.8±6.8 0.085
RBP (mg per dL) 4.13±0.87 3.67±0.80 3.89±0.98 4.01±1.02 3.66±1.08 0.493
Transferrin (mg per dL) 285.9±21.0 250.9±54.9 290.2±60.4 320.3±39.5 279.3±66.5 <0.001
C3 (mg per dL) 81.2±11.7 91.0±13.4 94.2±21.9 91.6±18.7 74.9±18.4 0.015
C4 (mg per dL) 24.3±4.3 35.8±11.3 39.4±14.0 39.3±25.9 25.5±12.4 <0.001
Phosphorous (mg per dL) 4.64±0.48 4.67±0.73 4.50±0.52 4.40±0.69 4.91±0.58 0.195
Calcium (mg per dL) 9.56±0.25 8.85±0.37 9.08±0.35 9.08±0.40 9.12±0.24 0.241
Zinc (mcg per dL) 105.0±15.1 101.4±26.9 120.3±35.3 88.9±9.9 91.4±17.7 0.105
Iron (mcg per dL) 78.3±25.5 81.9±22.9 79.9±25.9 81.4±19.0 71.5±25.9 0.366
Ferritin (mcg per dL) 16.4±9.6 114.8±89.8 74.3±60.6 43.0±25.2 37.0±23.9 0.022
Transferrin (mg per dL) 285.9±21.0 250.9±54.9 290.2±60.4 320.3±39.5 279.3±66.5 <0.001
TIBC (mcg per dL) 411.1±26.2 367.4±68.6 416.5±75.4 454.1±49.4 402.8±83.1 <0.001
Transferrin saturation (percentage) 19.13±6.37 20.16±6.68 21.76±7.00 15.91±5.98 20.80±5.60 0.103





Author Conclusion:

There is a variable response to treatment among patients with anorexia nervosa patients during the outpatient period. The serum concentrations of most of the liver-synthesized proteins confirm that the visceral protein compartment is usually well preserved in patients with anorexia nervosa. However, the changes in transferrin, C3 and C4 during outpatient treatment reveal an increased risk of relapse after one year. Transferrin, ferritin and zinc appear sensitive to physical recovery with transferrin increasing with weight gain and recovery and ferritin and zinc decreasing with weight gain and recovery.

Funding Source:
University/Hospital: Department of Metabolism and Nutrition, Institute of Frio, CSIS and Pediatric Hospital of Mostoles, Madrid, Spain
Other: Funding not clearly stated
Reviewer Comments:
  • Strengths: Detailed data collection and follow-up
  • Weaknesses: Small sample size.
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? No
  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? No
  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) Yes
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? ???
  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? Yes
  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")? Yes
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%.) No
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? Yes
  4.4. Were reasons for withdrawals similar across groups? Yes
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? Yes
5. Was blinding used to prevent introduction of bias? No
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? No
  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? No
  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? No
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? No
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? N/A
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? No
  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? Yes
  6.6. Were extra or unplanned treatments described? Yes
  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? Yes
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? Yes
  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? Yes
  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)? No
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? No
  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? ???
  10.1. Were sources of funding and investigators' affiliations described? No
  10.2. Was the study free from apparent conflict of interest? Yes