Urinary Metabolomics of Plastic Manufacturing Workers: A Pilot Study

Abstract

The plastic manufacturing industry has a crucial role in the global economy with a significant impact in a wide range of fields. The chemical risk to which workers are potentially exposed is difficult to characterize and strictly related to both the products and processes adopted. Among the chemicals used, we can cite styrene, phenol, butadiene and phthalates, but nano- and microplastic particles can also be released in the work environment. In this pilot study, we present for the first time an NMR-based metabolomic approach for assessing urinary profiles of workers employed in a plastic manufacturing company. Urine samples from twelve workers and thirteen healthy volunteers were collected and analyzed by NMR spectroscopy. Forty-six urinary metabolites belonging to different chemical classes were univocally identified and quantified. The dataset so obtained was then subjected to multivariate statistical analysis to characterize each profile and highlight any differences. An alteration in some metabolites involved in several pathways, such as amino acid metabolism and NAD metabolism, was found, and a strong impact on gut microflora was also speculated. Ultimately, our work has the objective of adding a tile to the knowledge of biological effects possibly related to occupational exposure even if it is below the threshold limit values.

Keywords: NMR-based metabolomics; multivariate statistical analysis; occupational exposure; plastic manufacturing; urinary profile.

Figure 1

Representative ¹H-NMR spectrum of human urine of both aromatic and aliphatic regiorns (to allow for better visualization, the portion of the spectrum containing the water and urea signals was removed).

Figure 2

PLS-DA scores (A), weights on CV1 (B) plots for the comparison between the CTRL (red) and workers (blue). Only variables whose confidence interval bounds do not cross the threshold of 0 are considered significant. According to the described method, leucine (Leu), valine (Val), isoleucine (Ile), 3-hydroxyisobutyrate (3-HIBA), threo 2,3 dihydroxybutyrate (Threo 2,3 DHBA), 3-hydroxy-3-methylbutyrate (3-H-3MBA), alanine (Ala), acetate (AA), N-acetylglutamine (N-AcGln), citrate (CA), sarcosine (Sar), methyl-guanidine (MG), creatine (Crt), trimethylamine-N-oxide (TMAO), 4-hydroxyphenylacetate (4-HPAA), tyrosine (Tyr), phenylacetylglycine (PAG), hippurate (Hipp), pseudouridine (PSI), 4-hydroxybenzoate (4-HbzA), formate (FA) and 1-methylnicotinate (1-MNA) were found significant for CTRL, while pyro-Glutamate (pyro-Glu), glutamine (Gln), glycine (Gly), furoylglycine, trigonelline (Trig) and the unknown compound 3 (U03) were significant for the Exposed group.

Figure 3

Metabolites found significant between healthy volunteers (CTRL) and exposed workers (Exposed) with a confidence level of 95% in the univariate statistical analysis. Details are reported in the Statistical Analysis section. The black dots represent the actual distribution of data for each variable. All data are expressed as µmol/mmol of urinary creatinine.