[Effects of titanium dioxide nanoparticles on fecal metabolome in rats after oral administration for 90 days]

Abstract

Objective: To explore the effects and related mechanisms of oral exposure titanium dioxide nanoparticles (TiO₂ NPs) for 90 days on the intestinal and the gut microbiota of rats, through fecal metabolomics.

Methods: Twelve 4-week-old clean-grade Sprague Dawley (SD) rats were randomly de-vided into 2 groups by body weight, treated with TiO₂ NPs at dose of 0 or 50 mg/kg body weight everyday respectively for 90 days. The solution of each infection was freshly prepared and shocked fully by ultrasonic. Characterization of the particle size, crystal form, purity, and specific surface area of TiO₂ NPs was conducted. And the fresh feces of the rats were collected on the 90th day. After lyophilized and hydrophilic phase extraction, ultra performance liquid chromatography-Q-exactive orbitrap-high-resolution mass spectrometry system (UPLC-QEMS) was utilized for non-targeted determination of fecal meta-bolites. The metabolites were identified and labeled through Compound Discoverer 3.0 software, and used for subsequent metabolomics analysis. Bioinformatics analysis was carried out including unsupervised principal component analysis and supervised orthogonal projection to latent structure discriminant analysis for the differential metabolites between the two groups. The differential metabolites were followed-up for Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis.

Results: Compared with the control group, the body weight of the rats was significantly reduced (P<0.05) in the treatment group. A total of 22 metabolites in fecal metabolomics showed significant changes. Among them, xanthine, 1-methyladenine, 3-hydroxypyridine, methionine sulfoxide, pyridoxine, 1,5-isoquinolinediol, N-acetylornithine, N-acetyl-D-galactosamine, L-citrulline, L-methionine, leucine, DL-tryptophan, L-ornithine, 4-methyl-5-thiazoleethanol, and L-glutamic acid totaled 15 metabolites increased significantly. N-acetylhistamine, D-pipecolinic acid, imidazolelactic acid, L-valine, 2,3,4,6-tetramethylpyrazine, caprolactam, and histamine totaled 7 metabolites decreased significantly. N-acetylhistamine, L-valine and methionine sulfoxide were changed more than 16 times. Analysis of KEGG pathway revealed that the two metabolic pathways arginine biosynthesis and aminoacyl-tRNA biosynthesis were significantly changed (false discover rate < 0.05, pathway impact > 0.1).

Conclusion: Oral exposure to TiO₂ NPs for 90 days could disrupt the metabolism of the intestine and gut microbiota, causing significant changes in metabolites and metabolic pathways which were related to inflammatory response, oxidative stress, glucose homeostasis, blood system and amino acid homeostasis in rat feces. It is suggested that the toxic effect of TiO₂ NPs on rats may be closely related to intestinal and gut microbiota metabolism.

Keywords: Feces; Metabolomics; Rats, Sprague-Dawley; Titanium dioxide nanoparticles.

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TiO₂ NPs(50 mg/kg)染毒组和对照组体质量变化 Changes in body weight of TiO₂ NPs (50 mg/kg) treatment and control group

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TiO₂ NPs染毒组与对照组代谢物浓度热图 Heat map of metabolite concentrations in the TiO₂ NPs treatment and control group

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TiO₂ NPs染毒组与对照组样品中代谢物的PCA得分图(A)和OPLS-DA得分图(B) PCA score (A) and OPLS-DA score (B) graphs of metabolites in samples of the TiO₂ NPs treatment group and the control group

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代谢物OPLS-DA模型的V得分图 V-score plot of the metabolite in OPLS-DA model

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差异代谢物的KEGG通路分析结果 KEGG pathway analysis results of differential metabolites