{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST001430","ANALYSIS_ID":"AN002391","VERSION":"1","CREATED_ON":"July 24, 2020, 10:46 am"},

"PROJECT":{"PROJECT_TITLE":"Metabolic dynamics and prediction of gestational age and time to delivery in pregnant women","PROJECT_SUMMARY":"Metabolism during pregnancy is a constantly changing yet precisely programmed process, the failure of which may have devastating consequences for the fetus. To capture in high resolution the sequence of metabolic events underlying the normal human pregnancy, we carried out an untargeted metabolome investigation on 784 weekly blood samples (3 outlier samples are removed) collected from 30 Danish pregnant women. The study revealed extensive metabolome alterations over the course of normal pregnancy: of 9,651 detected metabolic features, 4,995 were significantly changed (FDR < 0.05). Many metabolic changes were timed precisely according to pregnancy progression so that the overall metabolic profile demonstrated a highly choreographed pattern. Using machine-learning methods, we were able to build a linear models with five metabolites (four steroids and one phospholipid) that predicts gestational age with high accuracy (Pearson correlation coefficient, R = 0.95).","INSTITUTE":"Stanford University","LAST_NAME":"Liang","FIRST_NAME":"Liang","ADDRESS":"Alway M339, 300 Pasteur Drive, Palo Alto, California, 94305, USA","EMAIL":"liangtro@stanford.edu","PHONE":"8167852490"},

"STUDY":{"STUDY_TITLE":"Metabolic dynamics and prediction og gestational ange and time to delivery in pregant women","STUDY_SUMMARY":"Metabolism during pregnancy is a constantly changing yet precisely programmed process, the failure of which may have devastating consequences for the fetus. To capture in high resolution the sequence of metabolic events underlying the normal human pregnancy, we carried out an untargeted metabolome investigation on 784 weekly blood samples collected from 30 Danish pregnant women. The study revealed extensive metabolome alterations over the course of normal pregnancy: of 9,651 detected metabolic features, 4,995 were significantly changed (FDR < 0.05). Many metabolic changes were timed precisely according to pregnancy progression so that the overall metabolic profile demonstrated a highly choreographed pattern. Using machine-learning methods, we were able to build a linear models with five metabolites (four steroids and one phospholipid) that predicts gestational age with high accuracy (Pearson correlation coefficient, R = 0.95).","INSTITUTE":"Stanford University","LAST_NAME":"Liang","FIRST_NAME":"Liang","ADDRESS":"Alway M339, 300 Pasteur Drive, Palo Alto, California, 94305, USA","EMAIL":"liangtro@stanford.edu","PHONE":"+1 8167852490"},

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"COLLECTION":{"COLLECTION_SUMMARY":"To capture the highly dynamic pregnancy process, we established a multi-year single-center Danish normal pregnancy cohort with a unique design of high-density blood sampling. Consented female participants submitted weekly blood draws beginning in week 5 of pregnancy until the postpartum period. A total of 30 women with weekly blood samples were assigned to a discovery (N=21) and a validation (Validation-1, N=9) cohort , whose samples were analyzed in two separated years.","SAMPLE_TYPE":"Blood (plasma)","STORAGE_CONDITIONS":"-80℃"},

"TREATMENT":{"TREATMENT_SUMMARY":"No treatment."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"784 normal pregnancy samples (3 outlier samples were removed) were completely randomized within each cohort (Discovery and Validation - 1) and analyzed in 12 batches across two years. 200 μL plasma was extracted by mixing 800 μL 1:1:1 acetone: acetonitrile: methanol with the internal standard mixture. The extraction mixture was vortexed and mixed for 15 min at 4 C and incubated at -20 C for 2 hours to allow protein precipitation. The supernatant was collected after centrifugation and evaporated to dryness under nitrogen (Biotage Turbovap). The dry extracts were reconstituted with 200 μL 1:1 methanol: water before analysis."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"Chromatographic conditions RPLC separation was performed using Zorbax SB columns (2.1 X 50mm, 1.8 Micron, 600 Bar; 827700-914) purchased from Agilent Technologies (Santa Clara, CA, USA). Mobile phases for RPLC consisted of 0.06% acetic acid in water (phase A) and 0.06% acetic acid in MeOH (phase B). Metabolites were eluted from the column at a flow rate of 0.6 mL/min, leading to a backpressure of 220– 280 bar at 99% phase A. A linear 1%–80% phase B gradient was applied over 9–10 min. The oven temperature was set to 60C, and the sample injection volume was 5 mL.","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Thermo Dionex Ultimate 3000","COLUMN_NAME":"Agilent Zorbax Eclipse Plus C18 (100 x 2.1mm, 1.8 um)"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Plus Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"POSITIVE","MS_COMMENTS":"MS acquisition Metabolic extracts were analyzed by reversed-phase liquid chromatographic (RPLC)-mass spectrometry (MS) in both positive and negative ionization modes. Thermo Q Exactive Hybrid Quadrupole-Orbitrap plus and Q Exactive mass spectrometers (Xcalibur, Thermo Scientific, San Jose, CA, USA) were operated in full MS-scan mode for data acquisition (acquisition from m/z 500 to 2,000) with a scan rate of approximately 4 Hz and a resolution set at 30,000 (at m/z 400). The MS/MS spectra of the QC sample were acquired under different fragmentation energy (25 NCE and 50 NCE) of the top 10 parent ions. The resulting mass spectra were exported into Progenesis QI Software (Nonlinear Dynamics, Durham, NC, USA) for further processing. Section 1: Metabolomics Data Processing Metabolomic features were extracted with a unique mass/charge ratio and retention time, then aligned and quantified with the Progenesis QI software (Nonlinear Dynamics, Durham, NC, USA, http://www.nonlinear.com/progenesis/qi/). Peak deconvolution ll e2 Cell 181, 1680–1692.e1–e5, June 25, 2020 Resource was performed under default settings in Progenesis QI. Acquired data were processed using an analysis pipeline written in R (https:// www.R-project.org). Progenesis QI output was then processed by removing all metabolites that were quantified in less than 30% of the samples or had a median intensity of less than twofold signal over the noise threshold (S/N < 2). The noise threshold was estimated by using the median signal across all the blank runs (if no quantitation was reported in any of the blank runs, the feature was also included in the analysis, as it likely had good S/N characteristics). Then the data were log-transformed and normalized. For each run, the median of all features was centered to correct for variation in the sample amount. Then for each analyte, a linear correction was applied per batch to correct for any linear decrease or increase in abundance during the acquisition of a batch. In short, for each analyte and each batch, a linear model was fitted with the log-abundance of the analyte as the dependent variable and the acquisition number [run order (randomized)] as the independent variable. The model prediction was interpreted as an underlying drift in mass spectrometric sensitivity and subtracted from the analyte level to yield within-batch normalized abundances. Finally, for each analyte, the abundances were median centered by batch to correct for sensitivity differences between batches. The positive- and negative-mode features were then concatenated for downstream analysis. In total, 9,651 features were included in the final analysis. In addition, for samples with more than 50% of the values missing, the sample was removed (one sample in total). The remaining missing values were imputed by the nearest 10 neighbors using the k-Nearest Neighbor algorithm (Altman, 1992). Note that Discovery and Test Set 1 were normalized together, while samples of Test Set 2 were normalized independently. We applied principal component analysis (PCA) to examine the overall distribution of the sample data (with all 9,651 features) and check the run quality. The gestational ages (based on first-trimester ultrasound measurements) were superimposed to facilitate the analysis. During the analysis, the vast majority of the samples were separated by pre- and postpartum in PCA space defined by two components, which explained the largest variations (PC1 and 2, Figure 1B), while two samples of a same subject (last two in her collection, before and after childbirth) displayed irregular behavior in PCA and unsupervised clustering analysis. The two samples were treated as outliers and excluded from further analysis. We also performed partial least-squares discriminant analysis (PLSDA) according to the categories of gestational age (by the mixOmics package).","MS_RESULTS_FILE":"ST001430_AN002391_Results.txt UNITS:peak area Has m/z:Yes Has RT:Yes RT units:Minutes"}

}