Component Description
The NHANES program suspended field
operations in March 2020 due to the coronavirus disease 2019 (COVID-19)
pandemic. As a result, data collection for the NHANES 2019-2020 cycle was not
completed. Data collected in 2019-March 2020 can be accessed as
convenience samples through the NCHS Research Data Center (RDC). Any analyses based solely on the
2019-March 2020 data would not be generalizable to the U.S. civilian
non-institutionalized population. Please refer to the Analytic Notes section
for more details on the use of the data.
The
analysis of red blood cell (RBC) fatty acids (FA) is an indicator of long term
(4 months) FA status. Evaluation of
RBC fatty acid content has become increasingly favored as a measure of n-3 polyunsaturated fatty acids (PUFA) intake. PUFA alter membrane physical
characteristics and the activity of membrane-bound proteins. In membranes, they
interact with ion channels and can be converted into bioactive eicosanoid
(Harris, 2008).
The
American Heart Association recommendation for those with no history of coronary
heart disease (CHD) is to eat two fatty fish servings per week and for those
with a history of CHD to consume 1 g/day of two omega-3 fatty acids,
eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid (Kris-Etherton, et al.,
2002). The Omega-3 Index is a measure of EPA (C20:5 n-3) and DHA (C22:6 n-3) in the
blood, and expressed as a percent of 21 fatty acids, in specifically the red
blood cell membranes (Harris, et al., 2004). The Omega-3 Index
has been suggested for use to identify those at risk for death from coronary
heart disease (Harris, et. al., 2004).
Eligible Sample
Examined participants aged 6 years and older in the
NHANES 2019-March 2020 convenience sample were eligible.
Description of Laboratory Methodology
Esterified FA are
hydrolyzed from washed RBC membranes using sequential treatment with mineral
acid and base in the presence of heat. Using a modification of Lagerstedt et.
al., total FA are hexane-extracted from the matrix (100 µL 50:50 saline diluted RBC)
along with an internal standard solution containing thirteen stable
isotopically-labeled FA to account for recovery (Lagerstedt et. al., 2001). The
extract is derivatized with pentafluorobenzyl bromide (PFBBr) in the presence
of triethylamine to form pentafluorobenzyl esters. The reaction mixture is
injected onto a capillary gas chromatograph column to resolve individual FA of
interest from other matrix constituents. FA are detected using electron capture
negative-ion mass spectrometry within 34 minutes. Six saturated FA (SFA), four monounsaturated
FA (MUFA), and eleven polyunsaturated FA (PUFA) (21 FA in total) are measured
using selected ion monitoring. Quantitation is accomplished by comparing the
peak area ratio of the analyte to internal standard in the unknown sample with
the peak area ratio of a known amount of analyte to internal standard in a
calibrator solution.
Refer to the
Laboratory Method Files section for a detailed description on the laboratory
methods used.
This
is a new component in the 2019-2020 survey cycle.
Laboratory Method Files
Fatty Acids - RBC
(July 2023)
Laboratory Quality Assurance and Monitoring
Washed red blood
cell specimens are 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 the National Center for Environmental Health for
testing.
The NHANES quality assurance and quality control (QA/QC) protocols meet the
1988 Clinical Laboratory Improvement Amendments. 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 QA evaluation during unscheduled visits to
evaluate both the quality of the laboratory work and the QC 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.
Analytical
Laboratories
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 QC protocol for all the contract laboratories, which outlined the
use of Westgard rules (Westgard, et. al. 1981) when testing 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’ QA/QC performance criteria
for accuracy and precision, similar to the Westgard rules (Caudill, et. al.
2008).
Data Processing and Editing
The data were
reviewed. Incomplete data or improbable values were sent to the performing
laboratory for confirmation.
Analytic Notes
The COVID-19 pandemic required
suspension of NHANES 2019-2020 field operations in March 2020 after data were
collected in 18 of the 30 survey locations in the 2019-2020 sample. Data
collection was cancelled for the remaining 12 locations. Calculation of survey
weights for this partial cycle is not possible due to incomplete data
collection. Therefore, data from survey components that were only collected in
2019-March 2020 are made available as convenience samples through NCHS's Research Data Center (RDC) because
unbiased estimates for the NHANES target population cannot be produced with
these samples.
For survey components conducted
in both 2017-2018 and 2019-2020 cycles, data collected from 2019 to March 2020
were combined with data from 2017 to 2018 to form a nationally representative
sample of NHANES 2017-March 2020 pre-pandemic data. Please see the NHANES
2017-March 2020 pre-pandemic data page for detailed
information on this combined sample.
Refer to the 2019-2020 Laboratory Data Overview
for general information on NHANES laboratory data.
There are over 800 laboratory
tests performed on NHANES participants. However, not all participants provided
biospecimens or enough volume for all the tests to be performed. The specimen
availability can also vary by age or other population characteristics. For
example, in 2019-2020, approximately 71% of children aged 1-17 years who were
examined in the MEC provided a blood specimen through phlebotomy, while 94% of
examined adults aged 18 and older provided a blood specimen. Analysts should
evaluate the extent of missing data in the dataset related to the outcome of
interest as well as any predictor variables used in the analyses to determine
whether additional re-weighting for item non-response is necessary.
Please refer to the
NHANES Analytic Guidelines and
the on-line NHANES Tutorial for
further details on the use of sample weights and other analytic issues.
Demographic and Other Related Variables
The
analysis of NHANES laboratory data may require additional demographic
variables. The NHANES 2019-March 2020 Demographics File contains demographic data, health indicators, and
other related information collected during household interviews.
This
laboratory data file can be linked to the Demographics file and other NHANES
data files in the 2019-March 2020 convenience sample using the unique
survey participant identifier (i.e., SEQN).
Detection Limits
The detection limits were constant for all of the analytes in the data set. The lower limit of detection (LLOD, in mg/L) for the RBC fatty acids are shown below.
| Variable Name |
Analyte Description |
LLOD |
| LBXPAN |
alpha-Linolenic acid (C18:3n3) (%) |
0.03 |
| LBXP1A |
Arachidic acid (C20:0) (%) |
0.30 |
| LBXPRA |
Arachidonic acid (C20:4n6) (%) |
0.08 |
| LBXPDA |
Docosanoic acid (C22:0) (%) |
0.10 |
| LBXPHA |
Docosahexaenoic acid (C22:6n3) (%) |
0.20 |
| LBXPD3 |
Docosapentaenoic acid 3 (C22:5n3) (%) |
0.07 |
| LBXPD6 |
Docosapentaenoic acid 6 (C22:5n6) (%) |
0.08 |
| LBXPTA |
Docosatetraenoic acid (C22:4n6) (%) |
0.05 |
| LBXPED |
11,14-Eicosadienoic acid (C20:2n6) (%) |
0.05 |
| LBXP1E |
11-Eicosenoic acid (C20:1n9) (%) |
0.06 |
| LBXPPE |
Eicosapentaenoic acid (C20:5n3) (%) |
0.40 |
| LBXPLG |
gamma-Linolenic acid (C18:3n6) (%) |
0.07 |
| LBXPGH |
homo-gamma-Linolenic acid (C20:3n6) (%) |
0.06 |
| LBXP1G |
Tetracosanoic acid (C24:0) (%) |
0.10 |
| LBXPNL |
Linoleic acid (C18:2n6) (%) |
0.30 |
| LBXPMR |
Myristic acid (C14:0) (%) |
1.00 |
| LBXPNR |
15-Tetracosenoic acid (C24:1n9) (%) |
0.05 |
| LBXPOL |
Oleic acid (C18:1n9) (%) |
0.30 |
| LBXPPL |
Palmitoleic acid (C16:1n7) (%) |
0.05 |
| LBXPPM |
Palmitic acid (C16:0) (%) |
3.00 |
| LBXPST |
Stearic acid (C18:0) (%) |
9.00 |
This component does not report data as mass concentrations (mg/L). Instead, data are reported as weight percentages of total fatty acids (see section on Calculating Percent RBC Fatty Acid). Two variables are provided for each of these analytes. The variable name ending in “LC” (ex., LBDPANLC) indicates whether the mass concentration result used in the weight percentage calculation 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., LBXPAN) provides the analytic weight percentage result for that analyte. For analytes with mass concentration results below the lower limit of detection (ex., LBDPANLC=1), the weight percentage values were set to missing because the weight percentage data cannot be compared to the LLOD. Ten mass concentration results in the dataset were below the LLOD and corresponding weight percentage values were set to missing and their comment variables set to 1.
Calculating Percent RBC Fatty Acid
Fatty acid concentrations measured in mg/L are summed for the total FA concentration (mg/L). Individual fatty acid weight percentages are calculated by dividing the individual fatty acid concentrations (mg/L) by the total FA (mg/L). The fatty acid data is expressed as weight percentages of total fatty acids.