• Component Description
• Eligible Sample
• Description of Laboratory Methodology
• Laboratory Quality Assurance and Monitoring
• Data Processing and Editing
• Analytic Notes
• References
• Codebook
  • SEQN - Respondent sequence number
  • WTSSCL2Y - Surplus specimen CLTY 15-16 2 yr weights
  • SSCL1Y - 3-chlorotyrosine (Cl-Tyr) (ng/mL)
  • SSCL1YL - 3-chlorotyrosine (Cl-Tyr) Comment Code
  • SSCL2Y - 3,5-dichlorotyrosine (Cl2-Tyr) (ng/mL)
  • SSCL2YL - 3,5-dichlorotyrosine (Cl2-Tyr) Comt Code

National Health and Nutrition Examination Survey

2015-2016 Data Documentation, Codebook, and Frequencies

Chlorinated Tyrosine – Serum (Surplus) (SSCLTY_I)

Data File: SSCLTY_I.xpt

First Published: January 2023

Last Revised: NA

Component Description

Molecular chlorine (Cl2) is a highly reactive toxic industrial chemical with a history of use as a chemical warfare agent (Everts, 2015; OPCW, 2015). Low dose exposure (1-50 ppm) causes irritation of the airways while exposure to higher doses (>500 ppm) are fatal (Winder, 2001). When inhaled, chlorine rapidly hydrolyzes to form hypochlorous acid, (HOCl), hydrochloric acid (HCl), and hypochlorite (-OCl) (White et. al., 2010). Tyrosine residues are oxidized by hypochlorite at the ortho positions resulting in two stable chlorine adducts, 3-chlorotyrosine (Cl-Tyr) and 3,5-dichlorotyrosine (Cl2-Tyr) that can be used as biomarkers of chlorine gas exposure (Hureiki et. al., 1994; Sochaski et. al., 2008). However, hypochlorite produced by neutrophil myeloperoxidase (Domigan, 1995) or found in disinfecting products (e.g., bleach) can produce low levels of Cl-Tyr and possibly Cl2-Tyr.

Measuring Cl-Tyr and Cl2-Tyr is useful for determining potential exposure to chlorine. However, low levels of Cl2-Tyr coupled with elevated levels of Cl-Tyr, are found in some persons with chronic inflammatory disease (Chapman et. al., 2000; Mocatta et. al., 2007; Hazen et. al., 1997). In order to distinguish acute exposure to chlorine gas from neutrophil myeloperoxidase activity or low-level exposure to bleach products, the value must be compared to a reference range. Currently, population-based reference ranges of Cl-Tyr and Cl2-Tyr do not exist but would be useful in the event of a public incident involving suspected chlorine exposure. 

Eligible Sample

Examined participants aged 12 years and older from a one-third sample were eligible.

Description of Laboratory Methodology

This method required 10 microliters of serum and utilizes liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) and isotope dilution for quantification. Briefly, calibrators, quality controls, a matrix blank, blind QCs, and NHANES samples were prepared by performing a protein digestion using pronase to release the chlorinated tyrosine biomarkers. After digestion, any undigested proteins were removed by protein precipitation. Samples were then separated via liquid chromatography and detected via tandem mass spectrometry as previously reported (Pantazides et. al., 2021). This method allowed for rapid detection of Cl-Tyr and Cl2-Tyr in human serum with limits of detection in the low parts per billion (ppb or ng/mL) range.

Laboratory Quality Assurance and Monitoring

The analytical measurements were conducted following strict quality control/quality assurance CLIA guidelines. Along with the study samples, each analytical run included high-, mid-, and low-concentration quality-control materials (QCMs) and matrix blanks to assure the accuracy and reliability of the data. The concentrations of the QCMs were evaluated using standard statistical probability 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

Refer to the 2015-2016 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. Additionally, availability of specimens for surplus projects is lower than for other laboratory tests performed on NHANES participants. The specimen availability can also vary by age or other population characteristics. 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 details on the use of sample weights and analytic issues.

Subsample Weights

The analytes included in this dataset were measured in a one-third subsample of participants aged 12 years and older. Special sample weights are required to analyze these data properly. Specific sample weights for this subsample, WTSSCL2Y, are included in this data file and should be used when analyzing these data. The sample weights created for this file used the examination sample weight, i.e., WTMEC2YR, as the base weight. The base weight was adjusted for additional nonresponse to these lab tests and re-poststratified to the population total using sex, age, and race/ethnicity. Participants who were part of the eligible population but who did not provide a serum specimen, or did not have sufficient volume of biospecimens, or who did not give consent for their specimens to be used for future research are included in the file, but they have a sample weight assigned “0” in their records.

Demographic and Other Related Variables

The analysis of NHANES laboratory data must be conducted using the appropriate survey design and demographic variables. The NHANES 2015-2016 Demographics 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.

This laboratory data file can be linked to the other NHANES data files 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. Two variables are provided for each of these analyte. The variable name ending in “L” (ex., SSCL1YL) 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 (ex., SSCL1Y) provides the analytic result for that analyte. For analytes with analytic results below the lower limit of detection (ex., SSCL1YL=1), an imputed fill value was placed in the analyte results field. This value is the lower limit of detection divided by square root of 2 (LLOD/sqrt[2]). The lower limit of detection (LLOD in ng/mL) for SSCL1Y and SSCL2Y:

References

• Caudill S.P., Schleicher R.L., Pirkle J.L. Multi-rule quality control for the age-related eye disease study. Stat. Med. (2008) 27(20):4094-4106.
• Chapman A.L., Senthilmohan R., Winterbourn C.C., Kettle A.J. Comparison of mono- and dichlorinated tyrosines with carbonyls for detection of hypochlorous acid modified proteins. Arch. Biochem. Biophys. (2000) 377(1):95-100.
• Domigan N.M., Charlton T.S., Duncan M.W., Winterbourn C.C., Kettle A.J. Chlorination of tyrosyl residues in peptides by myeloperoxidase and human neutrophils. The J. Biol. Chem. (1995) 270(28):16542-16548.
• Everts S. When chemicals became weapons of war. Chem. Eng. News (2015) 93(8):8-21.
• Hazen S.L., Crowley J.R., Mueller D.M., Heinecke J.W. Mass spectrometric quantification of 3-chlorotyrosine in human tissues with attomole sensitivity: A sensitive and specific marker for myeloperoxidase-catalyzed chlorination at sites of inflammation. Free Radic Biol Med 1997;23(6):909-916.
• Hureiki L., Croué J.P., Legube B. Chlorination studies of free and combined amino acids. Water Res. (1994) 28(12):2521-2531.
• Mocatta T.J., Pilbrow A.P., Cameron V.A., et. al. Plasma concentrations of myeloperoxidase predict mortality after myocardial infarction. J Am Coll Cardiol (2007) 49(20):1993-2000.
• OPCW. Organisation for the Prohibition of Chemical Weapons fact finding mission: "Compelling confirmation" that chlorine gas used as weapon in syria. (2014) Available from: http://www.opcw.org/news/article/opcw-fact-finding-mission-compelling-confirmation-that-chlorine-gas-used-as-weapon-in-syria/ (accessed 24 February 2015).
• Pantazides B.G., Crow B.S., Quiñones-González J., et al. Development of a clinical assay to measure chlorinated tyrosine in hair and tissue samples using a mouse chlorine inhalation exposure model. Anal. Bioanal. Chem. (2021) 413(6):1765-1776.
• Sochaski M.A., Jarabek A.M., Murphy J., Andersen M.E. 3-Chlorotyrosine and 3,5-Dichlorotyrosine as biomarkers of respiratory tract exposure to chlorine gas. J Anal. Toxicol. (2008) 32(1):99-105.
• White C.W., Martin J.G. Chlorine gas inhalation: human clinical evidence of toxicity and experience in animal models. Proc. Am. Thorac. Soc. (2010) 7(4):257-263.
• Winder C. The toxicology of chlorine. Environ Res (2001) 85(2):105-114.

Codebook and Frequencies