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Determination of aromatic amines in human urine using comprehensive multi-dimensional gas chromatography mass spectrometry (GCxGC-qMS).

Lamani X, Horst S, Zimmermann T, Schmidt TC - Anal Bioanal Chem (2014)

Bottom Line: The measurements were carried out in full scan mode with conservatively estimated limits of detection (LOD) in the range of several ng/L and relative standard deviation (RSD) less than 20 %.More than 150 aromatic amines have been identified in the urine of a smoking person, including alkylated and halogenated amines as well as substituted naphthylamines.Also in the urine of a non-smoker, a number of aromatic amines have been identified, which suggests that the detection of biomarkers in urine samples using a more comprehensive analysis as detailed in this report may be essential to complement the approach of the use of classic biomarkers.

View Article: PubMed Central - PubMed

Affiliation: Instrumental Analytical Chemistry, University of Duisburg-Essen, Universitätstrasse 5, 45141, Essen, Germany.

ABSTRACT
Aromatic amines are an important class of harmful components of cigarette smoke. Nevertheless, only few of them have been reported to occur in urine, which raises questions on the fate of these compounds in the human body. Here we report on the results of a new analytical method, in situ derivatization solid phase microextraction (SPME) multi-dimensional gas chromatography mass spectrometry (GCxGC-qMS), that allows for a comprehensive fingerprint analysis of the substance class in complex matrices. Due to the high polarity of amino compounds, the complex urine matrix and prevalence of conjugated anilines, pretreatment steps such as acidic hydrolysis, liquid-liquid extraction (LLE), and derivatization of amines to their corresponding aromatic iodine compounds are necessary. Prior to detection, the derivatives were enriched by headspace SPME with the extraction efficiency of the SPME fiber ranging between 65 % and 85 %. The measurements were carried out in full scan mode with conservatively estimated limits of detection (LOD) in the range of several ng/L and relative standard deviation (RSD) less than 20 %. More than 150 aromatic amines have been identified in the urine of a smoking person, including alkylated and halogenated amines as well as substituted naphthylamines. Also in the urine of a non-smoker, a number of aromatic amines have been identified, which suggests that the detection of biomarkers in urine samples using a more comprehensive analysis as detailed in this report may be essential to complement the approach of the use of classic biomarkers.

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(a) Reconstructed ion chromatogram (RIC) of a urine sample of a smoker showing typical m/z associated with homologous alkylated aromatic iodine compounds and a separation of aminoacetophenone and C3-aniline derivatives with the same mass-to-charge ratio, m/z 246. The compounds selected are derivatives of C1-anilines (m/z 218), C2-anilines (m/z 232), C3-anilines/aminoacetophenone (m/z 246), and C4-anilines (m/z 260), and the RIC is generated by summing up signals at these four m/z; (b) and (c) show mass spectra of iodinated derivatives of aminoacetophenone and C3-aniline, respectively, with fragmentation patterns and loss of m/z 127 (iodine)
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Fig5: (a) Reconstructed ion chromatogram (RIC) of a urine sample of a smoker showing typical m/z associated with homologous alkylated aromatic iodine compounds and a separation of aminoacetophenone and C3-aniline derivatives with the same mass-to-charge ratio, m/z 246. The compounds selected are derivatives of C1-anilines (m/z 218), C2-anilines (m/z 232), C3-anilines/aminoacetophenone (m/z 246), and C4-anilines (m/z 260), and the RIC is generated by summing up signals at these four m/z; (b) and (c) show mass spectra of iodinated derivatives of aminoacetophenone and C3-aniline, respectively, with fragmentation patterns and loss of m/z 127 (iodine)

Mentions: Urine samples of a tobacco smoker and a non-smoker were hydrolyzed, extracted, derivatized, and analyzed by GCxGC-qMS. The analyses were carried out in full-scan mode so that all extractable and GC-suitable compounds were detected. Figure 4a and b show the total ion chromatogram (TIC) of the aromatic iodine compounds that are proposed to be present in the urine of non-smoking and smoking donors, respectively. In both Fig. 4a and b, not all the compounds can be visualized because of low intensity, especially in the urine of a non-smoker, but their identification is possible. The mass spectra of all derivatives showed the typical iodine fragment ion signal with 127 m/z; as a consequence, aromatic amines present in the samples could be identified by monitoring the iodine fragment ion. Since intensity of the mass fragment 127 is typically small, it would be beneficial to monitor instead the neutral loss of iodine. However, so far there is no opportunity in GCxGC software to do so. For each compound, the isomers (viz. blobs) were counted to obtain the total sum of analytes present in the sample. This is summarized in Table 3 with the names of the anilines and the fragment ions of the iodinated derivatives. In the urine of a donor exposed to cigarette smoke, ca. 150 aromatic amines were tentatively identified. Among the substances found, less than 10 are described in literature in relation with tobacco smoking–bladder cancer [2, 4, 8]. In the urine of a person without known exposure to cigarette smoke, many anilines (ca. 120) were still identified; nonetheless, regarding the number and peak intensities of occurring iodinated derivatives, the sample was less burdened. Amongst the detected compounds, alkylated anilines were the largest group (n = 39), as shown in Fig. 5a in groups of isomeric homologues (C1- to C4-anilines). Here, the number of isomers rapidly increases with larger or more numerous alkyl substituents and the intensities vary substantially among the isomers. In addition, compounds of the same nominal mass, such as aminoacetophenone (m/z 246) and C3-anilines (m/z 246), derivatives may elute at similar retention time in the first dimension but as in this case could be well separated in the second dimension.Fig. 4


Determination of aromatic amines in human urine using comprehensive multi-dimensional gas chromatography mass spectrometry (GCxGC-qMS).

Lamani X, Horst S, Zimmermann T, Schmidt TC - Anal Bioanal Chem (2014)

(a) Reconstructed ion chromatogram (RIC) of a urine sample of a smoker showing typical m/z associated with homologous alkylated aromatic iodine compounds and a separation of aminoacetophenone and C3-aniline derivatives with the same mass-to-charge ratio, m/z 246. The compounds selected are derivatives of C1-anilines (m/z 218), C2-anilines (m/z 232), C3-anilines/aminoacetophenone (m/z 246), and C4-anilines (m/z 260), and the RIC is generated by summing up signals at these four m/z; (b) and (c) show mass spectra of iodinated derivatives of aminoacetophenone and C3-aniline, respectively, with fragmentation patterns and loss of m/z 127 (iodine)
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Related In: Results  -  Collection

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Fig5: (a) Reconstructed ion chromatogram (RIC) of a urine sample of a smoker showing typical m/z associated with homologous alkylated aromatic iodine compounds and a separation of aminoacetophenone and C3-aniline derivatives with the same mass-to-charge ratio, m/z 246. The compounds selected are derivatives of C1-anilines (m/z 218), C2-anilines (m/z 232), C3-anilines/aminoacetophenone (m/z 246), and C4-anilines (m/z 260), and the RIC is generated by summing up signals at these four m/z; (b) and (c) show mass spectra of iodinated derivatives of aminoacetophenone and C3-aniline, respectively, with fragmentation patterns and loss of m/z 127 (iodine)
Mentions: Urine samples of a tobacco smoker and a non-smoker were hydrolyzed, extracted, derivatized, and analyzed by GCxGC-qMS. The analyses were carried out in full-scan mode so that all extractable and GC-suitable compounds were detected. Figure 4a and b show the total ion chromatogram (TIC) of the aromatic iodine compounds that are proposed to be present in the urine of non-smoking and smoking donors, respectively. In both Fig. 4a and b, not all the compounds can be visualized because of low intensity, especially in the urine of a non-smoker, but their identification is possible. The mass spectra of all derivatives showed the typical iodine fragment ion signal with 127 m/z; as a consequence, aromatic amines present in the samples could be identified by monitoring the iodine fragment ion. Since intensity of the mass fragment 127 is typically small, it would be beneficial to monitor instead the neutral loss of iodine. However, so far there is no opportunity in GCxGC software to do so. For each compound, the isomers (viz. blobs) were counted to obtain the total sum of analytes present in the sample. This is summarized in Table 3 with the names of the anilines and the fragment ions of the iodinated derivatives. In the urine of a donor exposed to cigarette smoke, ca. 150 aromatic amines were tentatively identified. Among the substances found, less than 10 are described in literature in relation with tobacco smoking–bladder cancer [2, 4, 8]. In the urine of a person without known exposure to cigarette smoke, many anilines (ca. 120) were still identified; nonetheless, regarding the number and peak intensities of occurring iodinated derivatives, the sample was less burdened. Amongst the detected compounds, alkylated anilines were the largest group (n = 39), as shown in Fig. 5a in groups of isomeric homologues (C1- to C4-anilines). Here, the number of isomers rapidly increases with larger or more numerous alkyl substituents and the intensities vary substantially among the isomers. In addition, compounds of the same nominal mass, such as aminoacetophenone (m/z 246) and C3-anilines (m/z 246), derivatives may elute at similar retention time in the first dimension but as in this case could be well separated in the second dimension.Fig. 4

Bottom Line: The measurements were carried out in full scan mode with conservatively estimated limits of detection (LOD) in the range of several ng/L and relative standard deviation (RSD) less than 20 %.More than 150 aromatic amines have been identified in the urine of a smoking person, including alkylated and halogenated amines as well as substituted naphthylamines.Also in the urine of a non-smoker, a number of aromatic amines have been identified, which suggests that the detection of biomarkers in urine samples using a more comprehensive analysis as detailed in this report may be essential to complement the approach of the use of classic biomarkers.

View Article: PubMed Central - PubMed

Affiliation: Instrumental Analytical Chemistry, University of Duisburg-Essen, Universitätstrasse 5, 45141, Essen, Germany.

ABSTRACT
Aromatic amines are an important class of harmful components of cigarette smoke. Nevertheless, only few of them have been reported to occur in urine, which raises questions on the fate of these compounds in the human body. Here we report on the results of a new analytical method, in situ derivatization solid phase microextraction (SPME) multi-dimensional gas chromatography mass spectrometry (GCxGC-qMS), that allows for a comprehensive fingerprint analysis of the substance class in complex matrices. Due to the high polarity of amino compounds, the complex urine matrix and prevalence of conjugated anilines, pretreatment steps such as acidic hydrolysis, liquid-liquid extraction (LLE), and derivatization of amines to their corresponding aromatic iodine compounds are necessary. Prior to detection, the derivatives were enriched by headspace SPME with the extraction efficiency of the SPME fiber ranging between 65 % and 85 %. The measurements were carried out in full scan mode with conservatively estimated limits of detection (LOD) in the range of several ng/L and relative standard deviation (RSD) less than 20 %. More than 150 aromatic amines have been identified in the urine of a smoking person, including alkylated and halogenated amines as well as substituted naphthylamines. Also in the urine of a non-smoker, a number of aromatic amines have been identified, which suggests that the detection of biomarkers in urine samples using a more comprehensive analysis as detailed in this report may be essential to complement the approach of the use of classic biomarkers.

Show MeSH
Related in: MedlinePlus