Assessment of Exposure to Mycotoxins in Spanish Children through the Analysis of Their Levels in Plasma Samples

Abstract

In this study, we present, for the first time in Spain, the levels of 19 mycotoxins in plasma samples from healthy and sick children (digestive, autism spectrum (ASD), and attention deficit hyperactivity (ADHD) disorders) (n = 79, aged 2-16). The samples were analyzed by liquid chromatography-mass spectrometry (triple quadrupole) (LC-MS/MS). To detect Phase II metabolites, the samples were reanalyzed after pre-treatment with β-glucuronidase/arylsulfatase. The most prevalent mycotoxin was ochratoxin A (OTA) in all groups of children, before and after enzyme treatment. In healthy children, the incidence of OTA was 92.5% in both cases and higher than in sick children before (36.7% in digestive disorders, 50% in ASD, and 14.3% in ADHD) and also after the enzymatic treatment (76.6 % in digestive disorders, 50% in ASD, and 85.7% in ADHD). OTA levels increased in over 40% of healthy children after enzymatic treatment, and this increase in incidence and levels was also observed in all sick children. This suggests the presence of OTA conjugates in plasma. In addition, differences in OTA metabolism may be assumed. OTA levels are higher in healthy children, even after enzymatic treatment (mean OTA value for healthy children 3.29 ng/mL, 1.90 ng/mL for digestive disorders, 1.90 ng/mL for ASD, and 0.82 ng/mL for ADHD). Ochratoxin B appears only in the samples of healthy children with a low incidence (11.4%), always co-occurring with OTA. Sterigmatocystin (STER) was detected after enzymatic hydrolysis with a high incidence in all groups, especially in sick children (98.7% in healthy children and 100% in patients). This supports glucuronidation as a pathway for STER metabolism in children. Although other mycotoxins were studied (aflatoxins B1, B2, G1, G2, and M1; T-2 and HT-2 toxins; deoxynivalenol, deepoxy-deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol; zearalenone; nivalenol; fusarenon-X; neosolaniol; and diacetoxyscirpenol), they were not detected either before or after enzymatic treatment in any of the groups of children. In conclusion, OTA and STER should be highly considered in the risk assessment of mycotoxins. Studies concerning their sources of exposure, toxicokinetics, and the relationship between plasma levels and toxic effects are of utmost importance in children.

Keywords: child; digestive problems; human biomonitoring; mycotoxins; ochratoxin A; ochratoxin B; sterigmatocystin.

Figure 1

Superposed extracted chromatograms obtained for mycotoxin group I from a calibrator at 10× limit of quantification (LOQ) level (A,B) and plasma samples from a healthy child (PS37) (C,D) and a patient child (PD23) (E,F) before enzymatic treatment. (A,C,E) display the Q transition, and (B,D,F) the q transition, respectively.

Figure 2

Superposed extracted chromatograms obtained for mycotoxin group II from a calibrator at 10× LOQ level (A,B) and plasma samples from a healthy child (PS37) (C,D) and a patient child (PD23) (E,F) before enzymatic treatment. (A,C,E) display the Q transition, and (B,D,F) the q transition, respectively.

Figure 3

Superposed extracted chromatograms obtained for mycotoxin group I from a calibrator at 10× LOQ level (A,B) and plasma samples from a healthy child (PS37) (C,D) and a patient child (PD23) (E,F) after enzymatic treatment. (A,C,E) display the Q transition, and (B,D,F) the q transition, respectively.

Figure 4

Superposed extracted chromatograms obtained for mycotoxin group II from a calibrator at 10× LOQ level (A,B) and plasma samples from a healthy child (PS37) (C,D) and a patient child (PD23) (E,F) after enzymatic treatment. (A,C,E) display the Q transition, and (B,D,F) the q transition, respectively.

Figure 5

Global incidence (%) of each of the obtained mycotoxins (OTA, OTB, and STER) according to enzymatic treatment. AE: after enzymatic treatment. BE: before enzymatic treatment.

Figure 6

Comparison of OTA incidence before and after enzymatic treatment in healthy children and children with digestive disorders.

Figure 7

Percentage of STER incidence after the treatment. PS: healthy children. PD: children with digestive disorders.