FNOA: Antioxidants (2011-2012)

Citation:
 
Study Design:
Class:
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Quality Rating:
Research Purpose:

To assess the zinc status of healthy late-middle-aged and older Europeans and to investigate the effects of zinc supplementation on their cognitive functioning.

Inclusion Criteria:

Inclusion criteria were:

  • Men and women aged between 55 and 87 years
  • Body mass index (BMI) between 20 and 30kg/m2
  • Good health
  • Mini Mental State Examination (MMSE) score greater than 23
  • Geriatric depression scale score less than six
  • Negative serology for the HIV and hepatitis C viruses.
Exclusion Criteria:

Participants were excluded based on the following criteria:

  • Tobacco consumption of more than 10g per day
  • Alcohol consumption of more than 30g per day (men) or 20g per day (women) 
  • Unconventional dietary habits (i.e., vegetarians, vegans)
  • Use of a mineral supplement during the preceding three months
  • Use of more than three (55 to 70 years) or four (70 to 87 years) prescription drugs per day
  • Use of antidepressant, laxatives or hormone replacement therapy
  • Pathological diseases including cancer and diabetes
  • After a full blood profile and tests of kidney and liver function, insufficient renal and hepatic performance, malabsorption or inflammatory chronic pathologies.
Description of Study Protocol:

Recruitment  

Participants were recruited through posters, leaflets, local television and radio and community groups and organizations serving older individuals living independently. 

Design

Randomized double-blind placebo-controlled trial with zinc supplementation of zero, 15 or 30mg per day administered as zinc gluconate, participants taking two tablets at the same time each day for six months. Cognitive function was assessed in the laboratory at baseline and after three and six months of supplementation using different parallel versions of the Cambridge Automated Neuropsychological Test Battery (CANTAB) tests on each occasion.

Blinding Used

Double blinding with tablets identified by code so that neither the experimenter nor the participant knew the dose.

Intervention

Placebo, 15mg zinc per day or 30mg zinc per day for six months. 

Statistical Analysis

  • Baseline measures were entered into two-way ANOVA with age group (younger vs. older) and subsequent treatment (zero, 15 and 30mg per day) as between subject factors
  • The main analyses were three-way repeated measures, ANOVA with age group (younger vs. older) and treatment (zero,15 and 30mg per day) as between subjects factors and time (zero, three and six months) as a within subjects factor
  • Initial analyses included: Center (four levels: Coleraine; Clermont-Ferrand; Rome; Grenoble) as a between-subject factor. These revealed few center effects or interactions involving center that could not be attributed instead to differences in age between participants tested in Coleraine and Clermont-Ferrand (younger) and Rome and Grenoble (older). Therefore, data from centers testing participants of the same age were combined  in all the reported analyses.
  • Where there was evidence of departure from the sphericity assumption, Greenhouse-Geisser corrections are reported
  • In addition, correlations were calculated between zinc measures (intake and blood  and urine levels) and cognitive measures (MMSE and CANTAB scores) at baseline with age partialled out, separately for males and females.

     

 

Data Collection Summary:

Timing of Measurements

Baseline (zero months), three months and six months.

Dependent Variables

CABTAB tests including:

  • Pattern recognition memory (PRM) accuracy (percentage correct)
  • PRM mean correct latency (ms)
  • Spatial working memory (SWM) total errors
  • SWM strategy
  • Spatial span (SSP) span length
  • Matching to sample visual search (MTS) mean corrected latency (ms)
  • Five-choice reaction time mean correct latency (ms)
  • Five-choice movement time mean correct latency (ms).

Independent Variables

Placebo, 15mg zinc per day or 30mg zinc per day for six months.  

Control Variables

  • Age
  • Education
  • Sex
  • Retirement status
  • Baseline (CABTAB) Mini Mental State Examination (MMSE) and Geriatric Depression Scale (GDS) scores
  • Baseline zinc levels (blood and urine levels)
  • Zinc intake.
Description of Actual Data Sample:
  • Initial N: 433
  • Attrition (final N): 387 (196 men and 191 women)
  • Age: 55 to 87 years
  • Ethnicity: European
  • Other relevant demographics: 188 younger and 199 older
  • Anthropometrics:  Participants in both age groups were reasonably well matched across the three treatments on measures at baseline. Younger adults differed from older adults only in terms of erythrocyte (higher) and urinary (lower) zinc levels.
  • Location: 
    • Coleraine, UK
    • Clermont-Ferrand, France
    • Rome, Italy
    • Grenoble, France.

 

Summary of Results:

Key Findings

Zinc Measures

  • For both serum and urinary zinc, there were highly significant two-way interactions between treatment and time [F(0.758) 16.95, P<0.001; F(3.6) 12.43, P<0.001, respectively]
  • Changes between baseline, three months and six months for serum zinc (Zn) were as follows:
  Serum Zn Urinary Zn Erythrocyte Zn
Placebo group -0.25μmol per L -10μmol per L -3.15μmol per L
15mg per day group +1.02μmol per L +0.13μmol per L +7.14μmol per L
30mg per day group +2.12μmol per L +0.44μmol per L +8.16μmol per L
  • Although the trends for erythrocyte zinc were similar to the other trends, the treatment Χ time interaction did not reach significance in this case [F(4.738) 1.79; P>0.1]
  • These data demonstrate significant biological consequences of zinc supplementation and provide confirmation of the participants' compliance with the study's requirements
  • For all three measures, the three-way interaction between age, treatment and time was not significant (all P>0.1) indicating that younger and older adults reacted similarly in biological terms to the zinc supplementation.

Cognitive Measures

Out of 72 zinc-cognition correlations at baseline, only four reached significance:

Serum zinc and PRM latency for males P<0.05 R=0.187
Erythrocyte zinc and RPM latency for for males P<0.05 R=0.155
Five-choice reaction time and erythrocyte zinc for females P<0.05 R=0.164
Serum zinc and five-choice reaction time for males P<0.01 R=0.206

Thus there was no strong evidence of any consistent relationships between zinc intake or blood and urine levels of zinc and cognitive measures at baseline.

Age Χ Treatment Χ Time Analysis of Variance

For each dependent measure, there was a highly significant effect of age group (all P<0.001). Younger adults were more accurate (PRM, SWM, spatial span), faster (PRM, MTS, five-choice reaction time, five-choice movement time) and more efficient (SWM strategy) than older adults for overall means.

There were significant main effects of time for:

PRM accuracy F(2, 760) 3.67 P=0.026
PRM latency F(1.8, 681) 29.07 P<0.001
SWM total errors F(1.9, 561) 35.21 P<0.001
SWM strategy F(2, 580) 9.52  P<0.001
 MTS latency F(2, 736) 15.49 P<0.001

These differed significantly between younger and older adults (age Χ time interactions) for:

PRM accuracy F(2, 760) 3.09 P=0.046
SWM total errors F(1.9, 561) 6.5 P=0.002
MTS latency F(2, 736) 4.5 P=0.012
  • For PRM, accuracy fell slightly between months zero and three for younger adults only ranging 88%, 86%, 86% for months zero, three and six for younger adults; 82% for months zero, three and six for older adults. PRM latency decreased from 2,703 to 2,462 to 2,382ms from months zero to three to six. SWM errors decreased over time, but not more so for older adults (an overall drop of twelve errors over six months) than for younger adults (a drop of six). SWM strategy decreased over time, particularly for younger adults (an overall decrease of 408ms from months zero to six) compared with older adults (a decrease of 134ms).
  • There were significant treatment Χ time interactions for SWM errors [F(3.9, 561) 2.74; P=0.03) and MTS latency (F3.9, 736) 3.15; P=0.015]. Separate ANOVA on months zero to three and on months three to six both revealed significant treatment Χ time interactions (P=0.046 and 0.014, respectively). For MTS latency, the significant interaction was due to greater improvement over the six months for the zero and 30mg per day treatment. There was also a marginal interaction for SWM strategy [F(4, 580) 2.31; P=0.057] with the 30mg-per-day group showing less improvement form zero to three months than the other groups, but this may reflect a ceiling effect as the 30mg per day group had the best strategy score of the three groups at baseline.
  • There were no other significant effects. In particular, there were no three-way (age Χ treatment Χ time) interactions indicating that the effects of zinc supplementation did not differ significantly between younger and older adults.
Author Conclusion:
  • Zinc intake at baseline was similar in younger and older adults (mean ages of 62 and 74 years, respectively) but erythrocyte zinc was higher and urinary zinc was lower in younger adults than in older adults, with no age difference for serum zinc. There were almost no significant associations between either zinc intake or status at baseline and measures of cognitive function. Supplementation was effective in that serum and urinary zinc levels both increased and more so for the higher doses than the lower dose with similar trends for erythrocyte zinc. For the cognitive measures, younger adults outperformed older adults on all tests. There were only two significant interactions indicating effects of zinc supplementation on cognitive function with a beneficial effect (but at only three months) of 15 and 30mg per day for SWM errors and a detrimental effect of 15mg per day for MTS latency.
  • Zinc intake was unaffected by age, which is different from previous literature. Urinary zinc was higher in older adults than in younger adults, suggesting changes in zinc metabolism with age. Erythrocyte but not serum zinc decreased with age. 
  • The complete absence of any relationships between zinc intake and cognitive measures (MMSE and CANTAB scores) at baseline contrasts with the results of a previous study, which found a significant but weak link between zinc intake and MMSE score that accounted for a less than 2% variance. Age-related deficits were observed on all CANTAB measures as expected from previous studies. 
  • Although the present study revealed evidence of only one beneficial effect of zinc supplementation on cognitive function, they also showed only one adverse effect. This is important because older adults may be taking zinc supplements for non-cognitive reasons, such as to prevent macular degeneration progression, based on results from the Age-Related Eye Disease Study Research Group. It is also reassuring in view of the growing availability and use of both fortified foods and dietary supplements. 
Funding Source:
Government: European Commission 'Quality of Life and Management of Living Resources' Fifth Framework Programme
Reviewer Comments:

The authors note the following limitations:

There were few significant effects of zinc supplementation on cognitive function. This may have been related to the following aspects of the study:

  • The participants were not sufficiently deficient in zinc to gain much benefit from supplements
  • They were a highly selective sample with 49% of the initial 842 volunteers being excluded from the study at screening on the basis of cognitive impairment, depression, pathological conditions, medications and so on
  • It is not know whether the present results would generalize to the less healthy half of the population who were excluded from the study
  • Thus it remains possible that the results of zinc supplementation on cognitive function would be more positive in other, more vulnerable populations.
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? Yes
  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? Yes
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? 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? Yes
  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? Yes
  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? Yes
  6.6. Were extra or unplanned treatments described? ???
  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? 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)? Yes
  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? N/A
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