<i>IGF2</i> DMR methylation in the child according to maternal periconceptional folic acid use of 400 µg per day.

<p>Linear Mixed Model analysis. Independent absolute methylation of the CpG dinucleotides without adjustments is presented in mean and (standard error).</p

<i>IGF2</i> DMR methylation in the child and independent factors of the mother and the child.

<p>Linear Mixed Model analysis. Data are presented in percentage (standard error) of mean change in relative methylation. For independent quantitative parameters the change in relative methylation is given per SD-change in that parameter. The p-value of the significant association of periconceptional folic acid use and <i>IGF2</i> DMR methylation was additionally adjusted for maternal education. The p-value for the significant association between maternal SAM and IGF2 DMR methylation was also adjusted for maternal education and the SAM concentration of the child. The p-value for the significant association between IGF2 DMR methylation and birth weight was additionally adjusted for periconceptional folic acid use and gestational age at delivery.</p

Validation results of PPD in human urine.

<p>Inter- and intraday accuracy and precision measurements (<i>n = 3</i> for each value). The mean relative error of each concentration is expressed as % bias and the reproducibility is depicted as the coefficient of variation (% CV). Intraday accuracy and precision were also determined on the HPLC-UV system.</p

Validation results of PPD in human urine.

<p>MALDI-MS/MS assay: linearity, lower limit of quantification (LLOQ), recovery (at 130, 400 and 800 µmol/L) and stability after addition of formic acid.</p

Calibration curve of PPD (A) on the MALDI-MS/MS system (50–1000 µmol/L) and (B) on the HPLC-UV system (150–1000 µmol/L).

<p>Graphs shows the mean values of (<b>A</b>) 5 and (<b>B</b>) 3 independent measurements and the corresponding <i>r²</i> value. Mean +/− SEM.</p

Wet chemistry assay for spiked blank and clinical urine samples.

<p>(<b>A</b>) <i>Left row bottom to top:</i> urine blank and 5 different spiked concentrations (25, 50, 100, 250 and 500 µmol/L). <i>Right row top three vials:</i> three clinical samples, which show a positive test result for PPD. (<b>B</b>) Concentration dependent transmittance (%) from 25–250 µmol/L on a spectrophotometer at 450 nm.</p

TOF spectrum (pos. mode) for the identification of two PPD metabolites MAPPD (<i>m/z</i> [M+H]⁺ 151) and DAPPD (<i>m/z</i> [M+H]⁺ 193) in patient urine.

<p>Comparative control urine did not show any signals at the corresponding <i>m/z</i> values.</p

MALDI-MS/MS measurements of urine samples from patients with suspected PPD intoxication and controls.

<p>(<b>A</b>) Measured PPD concentrations in clinical urine samples (<i>n = 15</i>); dotted lines show the LLOQ of the MALDI-MS/MS and the HPLC-UV application, resp. (<b>B</b>) Box-and-whisker blot with the 5–95 percentiles for the comparison of PPD, MAPPD and DAPPD peak areas in drug free control urine (<i>n = 5</i>) and clinical samples of intoxication (<i>n = 15</i>). (<b>C</b>) Data correlation (<i>r²</i> = 0.7618) of PPD and DAPPD in patient urine (<i>n = 15</i>). Respective p-values are given in the graphs.</p

Chemical structures and respective multiple reaction monitoring (MRM) transitions of (A) PPD and its metabolites (MAPPD and DAPPD) and (B) the internal standard (IS) 2-amino-5-nitropyridine.

<p>Chemical structures and respective multiple reaction monitoring (MRM) transitions of (A) PPD and its metabolites (MAPPD and DAPPD) and (B) the internal standard (IS) 2-amino-5-nitropyridine.</p

Manhattan plot.

<p>Association between MTHFR 677C>T (rs1801133) and genome-wide DNA methylation in 9,894 samples, with 35 <i>cis</i>-meQTLs at chromosome 1 (black/grey) and 1 <i>trans</i>-meQTL at chromosome 6 (green) with FDR<0.05.</p