These data will be used to estimate deficiencies and toxicities of specific nutrients in the population and subgroups, to provide population reference data, and to estimate the contribution of diet, supplements, and other factors to serum levels of nutrients. Data will be used for research to further define nutrient requirements as well as optimal levels for disease prevention and health promotion.
Examined participants aged 20 years and older were eligible.
Methylmalonic acid (MMA) is analyzed by LC-MS/MS as dibutylester (Mineva et al.). MMA is extracted from serum (75 µL) along with an added internal standard (d3-MMA) via liquid-liquid extraction with tert-butylmethylether/H+. The extracted acid is then derivatized with butanol to form a dibutylester. The butanol is evaporated under vacuum and the derivatized sample is reconstituted in acetonitrile/water. The sample preparation takes ~4 h for a run containing 96 samples (calibrators, QC’s and unknowns). MMA is chromatographically separated from other compounds as well as the isobaric succinic acid (same molecular mass as MMA), using isocratic mobile phase conditions within 5.9 min and measured by LC-MS/MS using multiple reaction monitoring. Quantitation is based on peak area ratios interpolated against a six-point aqueous calibration curve.
Increased concentrations of MMA in plasma or serum and excessive urinary excretion of MMA are believed to be direct measures of tissue stores of cobalamin (vitamin B12) and to be the first indication of cobalamin deficiency. The concentration of MMA in plasma or serum was found to be a useful indicator of cobalamin deficiency, especially in patients with few or no hematological abnormalities, normal results for the Schilling test, or normal or only slightly depressed serum cobalamin concentrations. In folate deficiency, MMA is normal. MMA may be elevated due to impaired renal function and as a result of an inborn error of metabolism (methylmalonic aciduria).
Refer to the Laboratory Method Files section for a detailed description of the laboratory methods used. There were changes to the lab method and lab equipment, but not to the lab site for this component in the NHANES 2011-2012 cycle.
Methylmalonic acid (MMA) (October 2018)
Serum samples were processed, stored, and shipped to the Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA for analysis.
Detailed instructions on specimen collection and processing are discussed in the NHANES Laboratory Procedures Manual (LPM). Vials are stored under appropriate frozen (–30°C) conditions until they are shipped to National Center for Environmental Health for testing.
The NHANES quality assurance and quality control (QA/QC) protocols meet the 1988 Clinical Laboratory Improvement Act mandates. Detailed QA/QC instructions are discussed in the NHANES LPM.
Mobile Examination Centers (MECs)
Laboratory team performance is monitored using several techniques. NCHS and contract consultants use a structured competency assessment evaluation during visits to evaluate both the quality of the laboratory work and the quality-control procedures. Each laboratory staff member is observed for equipment operation, specimen collection and preparation; testing procedures and constructive feedback are given to each staff member. Formal retraining sessions are conducted annually to ensure that required skill levels were maintained.
NHANES uses several methods to monitor the quality of the analyses performed by the contract laboratories. In the MEC, these methods include performing blind split samples collected during “dry run” sessions. In addition, contract laboratories randomly perform repeat testing on 2% of all specimens.
NCHS developed and distributed a quality control protocol for all CDC and contract laboratories, which outlined the use of Westgard rules (Westgard, et al. 1981) when running NHANES specimens. Progress reports containing any problems encountered during shipping or receipt of specimens, summary statistics for each control pool, QC graphs, instrument calibration, reagents, and any special considerations are submitted to NCHS quarterly. The reports are reviewed for trends or shifts in the data. The laboratories are required to explain any identified areas of concern.
All QC procedures recommended by the manufacturers were followed. Reported results for all assays meet the Division of Laboratory Sciences’ quality control and quality assurance performance criteria for accuracy and precision, similar to the Westgard rules (Caudill, et al. 2008).
The data were reviewed. Incomplete data or improbable values were sent to the performing laboratory for confirmation.
Refer to the 2011 - 2012 Laboratory Data Overview for general information on NHANES laboratory data.
Demographic and Other Related Variables
The analysis of NHANES laboratory data must be conducted using the appropriate survey design and demographic variables. The NHANES 2011–2012 Demographic File contains demographic data, health indicators, and other related information collected during household interviews as well as the sample design variables. The recommended procedure for variance estimation requires use of stratum and PSU variables (SDMVSTRA and SDMVPSU, respectively), in the demographic data file.
The Fasting Questionnaire File includes auxiliary information, such as fasting status, the length of fast and the time of venipuncture.
This laboratory data file can be linked to the other NHANES data files using the unique survey participant identifier (i.e., SEQN).
The detection limits were constant for all of the analytes in the data set. Two variables are provided for each of these analytes. The variable name ending in “LC” (ex., LBDMMALC) indicates whether the result was below the limit of detection: the value “0” means that the result was at or above the limit of detection, “1” indicates that the result was below the limit of detection. The other variable prefixed LBX (ex., LBXMMASI) provides the analytic result for that analyte. For analytes with analytic results below the lower limit of detection (ex., LBDMMALC=1), an imputed fill value was placed in the analyte results field. This value is the lower limit of detection divided by the square root of 2 (LLOD/sqrt).
The lower limit of detection (LLOD in nmol/L) for MMA:
|Variable Name||SAS Label||LLOD|
|LBXMMASI||Methylmalonic Acid||22.1 nmol/L|
No Correction Needed for Methylmalonic Acid Results for NHANES 2011-2012:
The MMA method changed in 2011 from a GC/MS method (used 1999–2004) to an LC-MS/MS method. The LC-MS/MS method showed excellent correlation (n=326, r=0.99) and no bias (Deming regression, Bland-Altman analysis) compared to the GC/MS method (Mineva et al.).
|Code or Value||Value Description||Count||Cumulative||Skip to Item|
|32.6 to 3020||Range of Values||4880||4880|
|Code or Value||Value Description||Count||Cumulative||Skip to Item|
|0||At or above the detection limit||4880||4880|
|1||Below lower detection limit||0||4880|