EAL

VN: Vitamin B-12 (2011)

Citation:

Study Design:

Class:

- Click here for explanation of classification scheme.

Quality Rating:

Research Purpose:

To investigate the Heptocorrin (HC) concentrations in a control population and pre- and post- B12 treatment in three populations:

Vitamin B₁₂ deficient vegan men
Patients with suspected B₁₂ deficiency
A group of individuals with normal B₁₂ status with new methods to further study the relationship between HC concentrations and vitamin B₁₂ status.

Inclusion Criteria:

Healthy men and women were included in the control population
The vegan population consisted of only males
The population with suspected deficiency was based on methylmalonic acid (MMA) concentrations of ≥0.28μmol/L within the past four years
The non-deficient population was part of the Western Norway B-vitamin Intervention Trial.

Exclusion Criteria:

Vegan population: Exclusion criteria consisted of a history of major gastrointestinal disease, presence of parietal cell or IF antibodies, liver disease, diabetes mellitus, vitamin B₁₂ injections or medication for psychiatric disorders
Suspected deficiency population: Exclusion criteria consisted of persons with MMA <0.40 or >0.200μmol/L, plasma creatinine >120μmol/L (females) or >133μmol/L (males), unable to cooperate or currently practicing in other clinical studies or receiving vitamin B₁₂ injections
Non-deficient population: Exclusion criteria consisted of participation in other studies, malignant disease, alcohol abuse, mental illness or unwillingness to do long-term follow-up.

Description of Study Protocol:

Recruitment

Participants were selected base on previously mentioned inclusion and exclusion criteria and were recruited through different cohorts or groups. Written informed consent was obtained from all patients.

Design

This is a non-randomized controlled trial

Blinding used

None

Intervention

Vegan population: A subgroup received oral vitamin B₁₂ treatment
Suspected deficient population: A subgroup received intramuscular injections of vitamin B₁₂
Non-deficient population: A subgroup was allocated to receive oral vitamin B₁₂ treatment.

Statistical Analysis

Associations were assessed by Spearman ranks correlation coefficients. Analysis was performed with the Wilcoxon matched-pairs signed-rank test.

Data Collection Summary:

Timing of Measurements

For control population the overall reference interval for total HC was reported from a previous study performed by the author
For the vegan population, 63 individuals were treated with a high oral dose of vitamin B₁₂ for three months
For the suspected deficient population, a total of 140 patients were randomized to receive weekly intramuscular injections or placebo for four weeks
For the non-deficient population one group received daily oral treatment with cyanocobalamin and folic acid either with our without vitamin B₆ or placebo
Samples from all groups were used from the following collection dates: Baseline, three, 14, 28 and 84 days after the start of the study from the non-deficient placebo group.

Dependent Variables

If HC is altered on treatment with vitamin B₁₂

Independent Variables

Supplementation of vitamin B₁₂ on several different populations (control, vegan, suspected deficient and non-deficient) with oral and intramuscular injections.

Description of Actual Data Sample:

Initial N:
- Control population: N=148
- Vegan population: N=174
- Suspected deficient population: N=140
- Non-deficient population: N=45
Attrition (final N):
- Control=148
- Vegan population=174
- 137 of 140 suspected deficient population
- 42 of 45 non-deficient were revisited
Age:
- For control groups:
  - 36 women≤50 years
  - 37 women>50
  - 35 men≤50
  - 40 men>50 years
- For the vegan population median age=44 (range=22-78)
- For suspected deficient population median age=74 (range=20-92)
- For non-deficient population median age=61 (range 38-79)
Ethnicity: No information included
Other relevant demographics:
- Control group 71 were female and 75 were male
- All vegan group was male (n=63)
- Suspected deficient group 72% female
- Non-deficient group 20% were female
Anthropometrics: Height, weight and other key anthropometrics were not reported
Location: Samples were located from many locations such as the Oxford cohort (London) of the European Prospective Investigation into Cancer and Nutrition and Western Norway (non-deficient population).

Summary of Results:

Key Findings

Concentrations in Three Groups Selected for B12 Treatment: Pre- and Post-Treatment Values

Median (range)

	Vegan	Suspected Deficiency	Non-deficient
Dosage (cyanocobalamin)	5mg per day	1mg per week for four weeks	0.4mg per day
Administration Route	Oral	Intramuscular	Oral
Number in Group	Pre=63, Post=63	Pre=69, Post=66	Pre=45, Post=42
Cobalamin, pmol/L	Pre=97 (32-386) Post=1,016 (142-2,896)*	Pre=281 (143-1,348 Post=960 (296- 10,404)*	Pre=350 (216-697) Post=527 (343-936)*
MMA, μmol/L	Pre=0.7 (0.1-4.3) Post=0.1 (0.1-0.3)*	Pre=0.5 (0.3-2.0) Post=0.2 (0.1-0.7)*	Pre=0.1 (0.07-0.3) Post=0.2 (0.1-0.3)
Total HC, pmol/L	Pre=330 (118-616) Post=377 (143-684)*	Pre=373 (168-848) Post=390 (162-796)	Pre=365 (235-725) Post=361 (263-870)
HoloHC, pmol/L	Pre=286 (101-626) Post=365 (135-700)*	Pre=336 (138-716 Post=377 (162-780)*	Pre=330 (225-674) Post=337 (234-830)

*Significantly different from pre-treatment levels

Other Findings

Total HC and holoHC increased 30pmol/L for every 100pmol/L increase in cobalamin compared to baseline in three study populations
For the vegan population, total HC and holoHC increased significantly (P<0.0001) after treatment with vitamin B₁₂
Only holoHC increased in the population with suspected deficiency, whereas no significant changes were seen with the non-deficient population.

Author Conclusion:

Reference interval compares well with previously published values obtained by similar methods
Higher values for holoHC over cobalamin in the vegan population and the population with suspected deficiency were noted suggesting that HC is partly saturated with analogs
A positive association between total HC and cobalamin were noted when analyzing the entire study cohort of 402 people and when analyzing three groups separately
It was noted that it is "impossible" to demonstrate differences in sex, age, clinical condition on vitamin B₁₂ treatment among the three treated populations and if these influenced the results
In summary, the results from the study indicate a complex relationship between HC and cobalamin. The HC concentration seems to be regulated by vitamin B₁₂ status, but there is also a correlation between HC and cobalamin in the cobalamin-replete and B₁₂-treated individuals.

Funding Source:

University/Hospital:	Aarus University Hospital, Denmark
Other:	EU Demonstration Project on the diagnostic utility of holoTC

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?	???
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?	???
	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?		No
	3.1.	Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT)	N/A
	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.)	No
	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?		No
	4.1.	Were follow-up methods described and the same for all groups?	???
	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?	No
	4.5.	If diagnostic test, was decision to perform reference test not dependent on results of test under study?	???
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?	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?	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?	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?	No
	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)?	???
	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?		N/A
	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