Cardiovascular Disease and Micronutrients

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

To study the relative effect of tocotrienol supplements of different compositions (mixed alpha- plus gamma-, high gamma- or P25-complex tocotrienol) on blood lipids, fasting blood glucose and the excretion of 8-iso-prostaglandin F2alpha in healthy hypercholesterolemic men and women.

Inclusion Criteria:
  • Male or female adult
  • Healthy (determined based on blood chemistry analysis and as the subjects reported on a medical history questionnaire)
  • Initial baseline serum total cholesterol concentration was 5.17mmol to 7.76mmol per L
  • Initial baseline LDL-cholesterol was greater than 3.36mmol per L
  • Initial baseline triacylglycerol was less than or equal to 4.52mmol per L.
Exclusion Criteria:
  • Subjects currently (or within the previous three months) taking prescription medication known to have an effect on blood lipids
  • Subjects currently (or within the past 30 days) taking bulk laxatives or dietary supplements to reduce blood cholesterol concentrations
  • Women taking hormone replacement therapy or oral contraceptives for less than three months prior to the study
  • Women taking hormone replacement therapy or oral contraceptives for more than three months prior to the study with varying dosages.
Description of Study Protocol:
  • Recruitment: Not described
  • Design: Randomized, double-blind parallel design
  • Blinding used: Supplements were sealed in bottles and labeled with a masked study number.

Intervention

  • Subjects were randomly assigned to receive one of three tocotrienol supplements or placebo
  • Additionally, they followed a NCEP Step 1 diet and followed a meal pattern including a regular daily lunch meal and consumed all capsules at the midday meal.

Statistical Analysis

  • Subjects were ranked in descending order, blocked by four and randomly assigned to four groups according to their baseline LDL-cholesterol concentrations. Data collected at baseline and during the run-in-phase and supplementation phase were examined to test the assumption of normally-distributed residuals with the use of the Shapiro-Wilk test
  • Comparison of the groups for values obtained during the run-in and supplementation phases and the calculated percentage change from the run-in phase to the supplementation phase was first carried out with two-way repeated-measures analysis of variance
  • Because an interaction (group-by-time) was significant, one-way ANOVA was used to analyze the run-in and supplementation phases and the percentage change data, if the assumption of non-normally distributed residuals was met at each time point
  • Variables for which the residuals showed evidence of being non-normally distributed at greater than or equal to one time point were ranked and one-way ANOVA was carried out on the rank data (Kruskal-Wallis one-way layout test)
  • When significant group effects were detected, a post-hoc Tukey's honestly significant difference test was used to identify the pair that caused the significant difference
  • Spearman's rank correlation coefficients were calculated to compare blood lipid and tocotrienol data and the correlation between blood lipid and urine data
  • Demographic variables were summarized with frequencies and percentages.
Data Collection Summary:

Timing of Measurements

  • Fasting blood samples from subjects were taken once at screening, on Days 19 and 21 during the Step 1 diet run-in phase and on Days 47 and 49 during the supplementation phase
  • Urine samples (overnight; 12 hours plus morning void) were collected by subjects twice, on Day 19 or 21 during the Step 1 diet run-in phase and on Day 47 or 49 during the supplementation phase.

Dependent Variable

  • Total cholesterol, measured enzymatically with the use of a validated method on a diagnostic autoanalyzer
  • LDL-cholesterol, calculated with the use of the Friedwald's equation, where LDL equals total cholesterol, HDL-cholesterol (triacylglycerols divided by five)
  • HDL-cholesterol, measured enzymatically with the use of a validated method on a diagnostic autoanalyzer; measured after the the precipitation of apolipoprotein B with the use of dextran sulfate
  • Triglycerides, measured enzymatically with the use of a validated method on a diagnostic autoanalyze
  • Glucose, measured with an automated glucose hexokinase enzymatic assay on a diagnostic autoanalyzer
  • 8-iso-prostaglandin F2alpha, measured by radioimmunoassay after alkaline hydrolysis.

Independent Variables

Tocotrienol supplements:

  • Mixed alpha plus gamma analyzed by normal-phase HPLC with the use of a Diol column
  • High gamma analyzed by normal-phase HPLC with the use of a Diol column
  • P25-complex tocotrienol analyzed by normal-phase HPLC with the use of a Diol column.
Description of Actual Data Sample:
  • Initial N: 68 healthy subjects (39 men, 29 females)
  • Attrition (final N): 67 healthy subjects (38 men, 29 females)
  • Age: 25 to 65 years
  • Ethnicity: Not described
  • Location: Not described.
Summary of Results:

Significance Levels Between Fasting Serum Levels and Treatment Group

 

Alpha- Plus Gamma-

Gamma-

P25-Complex

Total Cholesterol

Not significant

Not significant

Not significant

LDL-Cholesterol

Not significant

Not significant

Not significant

HDL-Cholesterol

Not significant

Not significant

Not significant

Triacylglycerols

Not significant

Not significant

Not significant

Glucose

Not significant

Not significant

Not significant

8-iso-prostaglandin
F2alpha

Not significant

Not significant

Not significant

Other Findings

When values for total cholesterol, HDL-cholesterol and glucose were expressed as a percentage change from the concentration during the run-in phase the increases in total and LDL-cholesterol were significantly greater (P<0.05) in the group consuming the alpha- plus gamma- tocotrienol supplement than those consuming the P25-complex supplement.

 

Author Conclusion:

Supplementation with 200mg tocotrienols per day from three commercially-available sources had no beneficial effect on key cardiovascular disease risk factors in highly compliant adults with elevated blood lipid concentrations.

Funding Source:
Industry:
Ross Products Division
Pharmaceutical/Dietary Supplement Company:
Reviewer Comments:
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) 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? 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? N/A
  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? 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)? N/A
  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