Contents

1 Background

Quality Control (QC) has been considered as an essential step in the metabolomics platform for high reproducibility and accuracy of data. The repetitive use of the same QC samples is more and more accepted for correcting the signal drift during the sequence of MS run order, especially beneficial to improve the quality of data in multi-block experiments of large-scale metabolomic study. statTarget is an easy use tool to provide a graphical user interface for quality control based signal shift correction, integration of metabolomic data from multi-batch experiments, and comprehensive statistic analysis in non-targeted or targeted metabolomics. This document is intended to guide the user to use statTargetGUI to perform metabolomic data analysis. Note that this document will not describe the inner workings of statTarget algorithm.

1.1 System requirements

Dependent on R (>= 3.3.0)

1.2 Opening the GUI

Load the package with biocLite():

source("https://bioconductor.org/biocLite.R")
#> Bioconductor version 3.7 (BiocInstaller 1.30.0), ?biocLite for help
biocLite("statTarget")
#> BioC_mirror: https://bioconductor.org
#> Using Bioconductor 3.7 (BiocInstaller 1.30.0), R 3.5.0 (2018-04-23).
#> Installing package(s) 'statTarget'

For mac PC, the package statTargetGUI requires X11 support (XQuartz). Download it from https://www.xquartz.org.

2 GUI overview

An easy to use tool providing a graphical user interface (Figure 1) for quality control based signal correction, integration of metabolomic data from multiple batches, and comprehensive statistic analysis for non-targeted and targeted approaches. (URL: https://github.com/13479776/statTarget)

2.1 What does statTarget offer statistically

The main GUI of statTarget has two basic sections. The first section is Shift Correction. It includes quality control-based robust LOESS signal correction (QC-RLSC) that is a widely accepted method for quality control based signal correction and integration of metabolomic data from multiple analytical batches (Dunn WB., et al. 2011; Luan H., et al. 2015). The second section is Statistical Analysis. It provides comprehensively computational and statistical methods that are commonly applied to analyze metabolomics data, and offers multiple results for biomarker discovery.

statTargetGUI

Section 1 - Shift Correction provide QC-RLSC algorithm that fit the QC data, and each metabolites in the true sample will be normalized to the QC sample. To avoid overfitting of the observed data, LOESS based generalised cross-validation (GCV) would be automatically applied, when the QCspan was set at 0.

Section 2 - Statistical Analysis provide features including Data preprocessing, Data descriptions, Multivariate statistics analysis and Univariate analysis.

Data preprocessing : 80-precent rule, glog transformation, KNN imputation, Median imputation and Minimum values imputation.

Data descriptions : Mean value, Median value, Sum, Quartile, Standard derivatives, etc.

Multivariate statistics analysis : PCA, PLSDA, VIP, Random forest.

Univariate analysis : Welch’s T-test, Shapiro-Wilk normality test and Mann-Whitney test.

Biomarkers analysis: ROC, Odd ratio.

2.2 Running Shift Correction from the GUI

Pheno File

Meta information includes the Sample name, class, batch and order. Do not change the name of each column. (a) Class: The QC should be labeled as NA. (b) Order : Injection sequence. (c) Batch: The analysis blocks or batches with ordinal number,e.g., 1,2,3,…. (d) Sample name should be consistent in Pheno file and Profile file. (See the example data)

Profile File

Expression data includes the sample name and expression data.(See the example data)

NA.Filter

NA.Filter: Removing peaks with more than 80 percent of missing values (NA or 0) in each group. (Default: 0.8)

QCspan

The smoothing parameter which controls the bias-variance tradeoff. The common range of QCspan value is from 0.2 to 0.75. If you choose a span that is too small then there will be a large variance. If the span is too large, a large bias will be produced. The default value of QCspan is set at ‘0’, the generalised cross-validation will be performed for choosing a good value, avoiding overfitting of the observed data. (Default: 0)

degree

Lets you specify local constant regression (i.e., the Nadaraya-Watson estimator, degree=0), local linear regression (degree=1), or local polynomial fits (degree=2). (Default: 2)

Imputation

Imputation: The parameter for imputation method.(i.e., nearest neighbor averaging, “KNN”; minimum values for imputed variables, “min”; median values for imputed variables (Group dependent) “median”. (Default: KNN)

2.3 Running Statistical Analysis from the GUI

Stat File

Expression data includes the sample name, group, and expression data.

NA.Filter

Removing peaks with more than 80 percent of missing values (NA or 0) in each group. (Default: 0.8)

Imputation

The parameter for imputation method.(i.e., nearest neighbor averaging, “KNN”; minimum values for imputed variables, “min”; median values for imputed variables (Group dependent) “median”. (Default: KNN)

Glog

Generalised logarithm (glog) transformation for Variance stabilization
(Default: TRUE)

Scaling Method

Scaling method before statistic analysis (PCA or PLS). Pareto can be used for specifying the Pareto scaling. Auto can be used for specifying the Auto scaling (or unit variance scaling). Vast can be used for specifying the vast scaling. Range can be used for specifying the Range scaling. (Default: Pareto)

M.U.Stat

Multiple statistical analysis and univariate analysis (Default: TRUE)

Permutation times

The number of random permutation times for PLS-DA model (Default: 20)

PCs

PCs in the Xaxis or Yaxis: Principal components in PCA-PLS model for the x or y-axis (Default: 1 and 2)

nvarRF

The number of variables in Gini plot of Randomforest model (=< 100). (Default: 20)

Labels

To show the name of sample in the Score plot. (Default: TRUE)

Multiple testing

This multiple testing correction via false discovery rate (FDR) estimation with Benjamini-Hochberg method. The false discovery rate for conceptualizing the rate of type I errors in null hypothesis testing when conducting multiple comparisons. (Default: TRUE)

Volcano FC

The up or down -regulated metabolites using Fold Changes cut off values in the Volcano plot. (Default: > 2 or < 1.5)

Volcano Pvalue

The significance level for metabolites in the Volcano plot.(Default: 0.05)

3 Investigating the results

Download the statTarget tutorial and example data .

Once data files have been analysed it is time to investigate them. Please get this info. through the GitHub page. (URL: https://github.com/13479776/statTarget)

3.1 Results of Shift Correction (ShiftCor)

  • The output file: 
statTarget -- shiftCor 
-- After_shiftCor # The corrected results including the loplot using statTarget
-- Before_shiftCor # The raw results using statTarget
-- RSDresult # The RSD analysis
  • The Figures:

Loplot (left): the visible Figure of QC-RLS correction for each peak.

The RSD distribution (right): The relative standard deviation of peaks in the samples and QCs

  • The status log (Example data):
#############################
# Shift Correction function #
#############################

Data File Checking Start..., Time:  Thu Jan  5 18:58:09 2017 

217 Pheno Samples vs 218 Profile samples

The Pheno samples list (*NA, missing data from the Profile File)
  [1] "QC1"              "QC2"              "QC3"              "QC4"             
  [5] "QC5"              "A1"               "A2"               "A3"              
  [9] "A4"               "A5"               "A6"               "A7"              
 [13] "A8"               "A9"               "A10"              "QC6"             
 [17] "A11"              "A12"              "A13"              "A14"             
 [21] "A15"              "B16"              "B17"              "B18"             
 [25] "B19"              "B20"              "QC7"              "B21"             
 [29] "B22"              "B23"              "B24"              "B25"             
 [33] "B26"              "B27"              "B28"              "B29"             
 [37] "B30"              "QC8"              "C31"              "C32"             
 [41] "C33"              "C34"              "C35"              "QC9"             
 [45] "QC10"             "QC11"             "QC12"             "QC13"            
 [49] "C36_120918171155" "C37"              "C38"              "C39"             
 [53] "C40"              "QC14"             "C41"              "C42"             
 [57] "C43"              "C44"              "C45"              "D46"             
 [61] "D47"              "D48"              "D49"              "D50"             
 [65] "QC15"             "D51"              "D52"              "D53"             
 [69] "D54"              "D55"              "D56"              "D57"             
 [73] "D58"              "D59"              "D60"              "QC16"            
 [77] "E61"              "E62"              "E63"              "E64"             
 [81] "E65"              "E66"              "E67"              "E68"             
 [85] "E69"              "E70"              "QC17"             "E71"             
 [89] "E72"              "E73"              "E74"              "E75"             
 [93] "F76"              "F77"              "F78"              "F79"             
 [97] "F80"              "QC18"             "F81"              "F82"             
[101] "F83"              "F84"              "F85"              "F86"             
[105] "F87"              "F88"              "F89"              "F90"             
[109] "QC19"             "QC20"             "QC21"             "QC22"            
[113] "QC23"             "QC24"             "a1"               "a2"              
[117] "a3"               "a4"               "a5"               "a6"              
[121] "a7"               "a8"               "a9"               "a10"             
[125] "QC25"             "a11"              "a12"              "a13"             
[129] "a14"              "a15"              "b16"              "b18"             
[133] "b19"              "b20"              "QC26"             "b21"             
[137] "b22"              "b23"              "b24"              "b25"             
[141] "b26"              "b27"              "b28"              "b29"             
[145] "b30"              "QC27"             "c31"              "c32"             
[149] "c33"              "c34"              "c35"              "QC28"            
[153] "QC29"             "QC31"             "QC32"             "c36"             
[157] "c37"              "c38"              "c39"              "c40"             
[161] "QC33"             "c41"              "c42"              "c43"             
[165] "c44"              "c45"              "d46"              "d47"             
[169] "d48"              "d49"              "d50"              "QC34"            
[173] "d51"              "d52"              "d53"              "d54"             
[177] "d55"              "d56"              "d57"              "d58"             
[181] "d59"              "d60"              "QC35"             "e61"             
[185] "e62"              "e63"              "e64"              "e65"             
[189] "e66"              "e67"              "e68"              "e69"             
[193] "e70"              "QC36"             "e71"              "e72"             
[197] "e73"              "e74"              "e75"              "f76"             
[201] "f77"              "f78"              "f79"              "f80"             
[205] "QC37"             "f81"              "f82"              "f83"             
[209] "f84"              "f85"              "f86"              "f87"             
[213] "f88"              "f89_120921102721" "f90"              "QC38"            
[217] "QC39"            

Warning: The sample size in Profile File is larger than Pheno File! 

Pheno information:
  Class No.
1     1  30
2     2  29
3     3  30
4     4  30
5     5  30
6     6  30
7    QC  38
  Batch No.
1     1 108
2     2 109

Profile information:
                No.
QC and samples  218
Metabolites    1312

statTarget: shiftCor start...Time:  Thu Jan  5 18:58:11 2017 

Step 1: Evaluation of missing value...

The number of NA value in Data Profile before QC-RLSC: 2280

The number of variables including 80 % of missing value : 3

Step 2: Imputation start...

The number of NA value in Data Profile after the initial imputation: 0

Imputation Finished!

Step 3: QC-RLSC Start... Time:  Thu Jan  5 18:58:12 2017

Warning: The QCspan was set at '0'.

The GCV was used to avoid overfitting the observed data

  |===============================================================================| 100%

High-resolution images output...

Calculation of CV distribution of raw peaks (QC)...

            CV<5%    CV<10%   CV<15%   CV<20%   CV<25%   CV<30%   CV<35%   CV<40%
Batch_1 0.6875477  7.944996 23.98778 37.58594 46.98243 54.39267 61.19175 67.99083
Batch_2 4.0488923 25.821238 45.76012 57.44843 64.40031 70.51184 76.39419 80.29030
Total   0.3819710  6.722689 21.08480 33.38426 44.38503 51.87166 59.20550 64.55309
          CV<45%   CV<50%   CV<55%   CV<60%   CV<65%   CV<70%   CV<75%   CV<80%   CV<85%
Batch_1 72.80367 77.92208 80.97785 84.11001 87.16578 88.69366 89.45760 90.67991 91.59664
Batch_2 83.34607 86.40183 88.31169 90.52712 92.58976 93.43010 94.42322 95.64553 96.18029
Total   69.36593 74.56073 78.53323 81.51261 82.96409 85.10313 87.39496 89.53400 91.36746
          CV<90%   CV<95%  CV<100%
Batch_1 92.66616 93.35371 94.57601
Batch_2 96.48587 97.17341 97.40260
Total   92.89534 94.27044 94.95798


Calculation of CV distribution of corrected peaks (QC)...

           CV<5%   CV<10%   CV<15%   CV<20%   CV<25%   CV<30%   CV<35%   CV<40%   CV<45%
Batch_1 18.25821 45.98930 64.40031 72.72727 78.45684 83.72804 86.17265 88.54087 89.76318
Batch_2 20.24446 51.48969 68.06723 78.22765 84.56837 88.23529 90.75630 92.36058 93.50649
Total   15.73720 44.46142 64.62949 73.18564 80.36669 84.79756 87.31856 88.69366 89.68678
          CV<50%   CV<55%   CV<60%   CV<65%   CV<70%   CV<75%   CV<80%   CV<85%   CV<90%
Batch_1 91.06188 91.90222 92.58976 93.04813 93.43010 94.04125 94.65241 95.11077 95.56914
Batch_2 94.11765 94.88159 95.49274 96.18029 96.63866 96.86784 97.09702 97.40260 97.70817
Total   90.75630 91.97861 93.20092 93.96486 94.57601 95.33995 95.87471 96.10390 96.63866
          CV<95%  CV<100%
Batch_1 95.95111 96.02750
Batch_2 98.09015 98.31933
Total   96.71505 97.09702


Correction Finished! Time:  Thu Jan  5 19:00:51 2017

3.2 Results of statistic analysis (statAnalysis)

  • The output file: 
statTarget -- statAnalysis 
-- PCA_Data_Pareto # Principal Component Analysis
-- PLS_DA_Pareto # Partial least squares Discriminant Analysis
-- Univariate# The RSD analysis 
   ----- BoxPlot
   ----- Fold_Changes
   ----- Mann-Whitney_Tests # For non-normally distributed variables
   ----- oddratio # odd ratio
   ----- Pvalues # Intergation pvalues from Welch_test and MWT_test 
   ----- RForest # Random Forest
   ----- ROC # receiver operating characteristic curve
   ----- Shapiro_Tests 
   ----- Significant_Variables # The Peaks with P-value < 0.05 
   ----- Volcano_Plots
   ----- WelchTest  # For normally distributed variables
  • The Figures:

  • The status log (Example data):
#################################
# Statistical Analysis function #
#################################

statTarget: statistical analysis start... Time:  Fri Jan  6 11:57:48 2017 

Step 1: Evaluation of missing value...

The number of NA value in Data Profile: 0

The number of variables including 80 % of missing value : 0

Step 2: Imputation start... Time:  Fri Jan  6 11:57:50 2017

The number of NA value in Data Profile after the initialimputation: 0

Imputation Finished!

Step 3: Statistic Summary Start... Time:  Fri Jan  6 11:57:50 2017

Step 4: Glog PCA-PLSDA start... Time:  Fri Jan  6 11:58:19 2017

PCA Model Summary

217 samples x 1309 variables

Variance Explained of PCA Model: 
                             PC1      PC2       PC3       PC4       PC5       PC6
Standard deviation     0.1471269 0.143504 0.1286476 0.1217399 0.1087545 0.1029451
Proportion of Variance 0.0743800 0.070770 0.0568700 0.0509300 0.0406400 0.0364200
Cumulative Proportion  0.0743800 0.145150 0.2020200 0.2529500 0.2935900 0.3300100
                              PC7        PC8        PC9       PC10       PC11       PC12
Standard deviation     0.09463045 0.09204723 0.08859019 0.08179698 0.07815861 0.07343806
Proportion of Variance 0.03077000 0.02911000 0.02697000 0.02299000 0.02099000 0.01853000
Cumulative Proportion  0.36078000 0.38989000 0.41686000 0.43985000 0.46085000 0.47938000
                             PC13       PC14       PC15       PC16      PC17       PC18
Standard deviation     0.06927193 0.06884729 0.06481461 0.06338068 0.0625105 0.05918608
Proportion of Variance 0.01649000 0.01629000 0.01444000 0.01380000 0.0134300 0.01204000
Cumulative Proportion  0.49587000 0.51216000 0.52659000 0.54040000 0.5538200 0.56586000
                             PC19      PC20       PC21       PC22       PC23      PC24
Standard deviation     0.05852846 0.0565814 0.05494036 0.05354714 0.05199812 0.0514794
Proportion of Variance 0.01177000 0.0110000 0.01037000 0.00985000 0.00929000 0.0091100
Cumulative Proportion  0.57763000 0.5886300 0.59901000 0.60886000 0.61815000 0.6272600
                             PC25       PC26       PC27       PC28       PC29       PC30
Standard deviation     0.05023623 0.05002373 0.04918839 0.04848824 0.04719809 0.04592107
Proportion of Variance 0.00867000 0.00860000 0.00831000 0.00808000 0.00765000 0.00725000
Cumulative Proportion  0.63593000 0.64453000 0.65284000 0.66092000 0.66858000 0.67582000
                             PC31       PC32     PC33       PC34       PC35       PC36
Standard deviation     0.04495383 0.04433005 0.043467 0.04273003 0.04211339 0.04168549
Proportion of Variance 0.00694000 0.00675000 0.006490 0.00627000 0.00609000 0.00597000
Cumulative Proportion  0.68277000 0.68952000 0.696010 0.70229000 0.70838000 0.71435000
                             PC37      PC38       PC39      PC40       PC41       PC42
Standard deviation     0.04074753 0.0399799 0.03970366 0.0395391 0.03887607 0.03829039
Proportion of Variance 0.00571000 0.0054900 0.00542000 0.0053700 0.00519000 0.00504000
Cumulative Proportion  0.72006000 0.7255500 0.73097000 0.7363400 0.74153000 0.74657000
                             PC43       PC44       PC45       PC46       PC47       PC48
Standard deviation     0.03757011 0.03717074 0.03680406 0.03627876 0.03578231 0.03561238
Proportion of Variance 0.00485000 0.00475000 0.00465000 0.00452000 0.00440000 0.00436000
Cumulative Proportion  0.75142000 0.75617000 0.76082000 0.76535000 0.76975000 0.77410000
                             PC49       PC50       PC51       PC52       PC53       PC54
Standard deviation     0.03500362 0.03466778 0.03451624 0.03404736 0.03367672 0.03328364
Proportion of Variance 0.00421000 0.00413000 0.00409000 0.00398000 0.00390000 0.00381000
Cumulative Proportion  0.77831000 0.78244000 0.78654000 0.79052000 0.79442000 0.79822000
                            PC55       PC56       PC57       PC58       PC59       PC60
Standard deviation     0.0329035 0.03283489 0.03263074 0.03220013 0.03174905 0.03127306
Proportion of Variance 0.0037200 0.00370000 0.00366000 0.00356000 0.00346000 0.00336000
Cumulative Proportion  0.8019400 0.80565000 0.80931000 0.81287000 0.81634000 0.81970000
                             PC61       PC62       PC63       PC64      PC65      PC66
Standard deviation     0.03099456 0.03067399 0.03047579 0.03027017 0.0299977 0.0293092
Proportion of Variance 0.00330000 0.00323000 0.00319000 0.00315000 0.0030900 0.0029500
Cumulative Proportion  0.82300000 0.82623000 0.82942000 0.83257000 0.8356600 0.8386100
                             PC67       PC68       PC69       PC70      PC71       PC72
Standard deviation     0.02910743 0.02891644 0.02871973 0.02851117 0.0284027 0.02802608
Proportion of Variance 0.00291000 0.00287000 0.00283000 0.00279000 0.0027700 0.00270000
Cumulative Proportion  0.84153000 0.84440000 0.84723000 0.85003000 0.8528000 0.85550000
                             PC73      PC74       PC75      PC76       PC77       PC78
Standard deviation     0.02767845 0.0274821 0.02707466 0.0270495 0.02689331 0.02669644
Proportion of Variance 0.00263000 0.0026000 0.00252000 0.0025100 0.00249000 0.00245000
Cumulative Proportion  0.85813000 0.8607300 0.86325000 0.8657600 0.86824000 0.87069000
                             PC79       PC80       PC81       PC82      PC83       PC84
Standard deviation     0.02641874 0.02597847 0.02569734 0.02537066 0.0252014 0.02514993
Proportion of Variance 0.00240000 0.00232000 0.00227000 0.00221000 0.0021800 0.00217000
Cumulative Proportion  0.87309000 0.87541000 0.87768000 0.87989000 0.8820700 0.88425000
                             PC85       PC86       PC87       PC88      PC89       PC90
Standard deviation     0.02505431 0.02457329 0.02445747 0.02427666 0.0240513 0.02389179
Proportion of Variance 0.00216000 0.00207000 0.00206000 0.00203000 0.0019900 0.00196000
Cumulative Proportion  0.88641000 0.88848000 0.89054000 0.89256000 0.8945500 0.89651000
                             PC91       PC92       PC93       PC94       PC95       PC96
Standard deviation     0.02381883 0.02338754 0.02322625 0.02314694 0.02290039 0.02280068
Proportion of Variance 0.00195000 0.00188000 0.00185000 0.00184000 0.00180000 0.00179000
Cumulative Proportion  0.89846000 0.90034000 0.90219000 0.90403000 0.90584000 0.90762000
                             PC97       PC98       PC99      PC100      PC101     PC102
Standard deviation     0.02259418 0.02254316 0.02231249 0.02210274 0.02203839 0.0219603
Proportion of Variance 0.00175000 0.00175000 0.00171000 0.00168000 0.00167000 0.0016600
Cumulative Proportion  0.90938000 0.91112000 0.91283000 0.91451000 0.91618000 0.9178400
                            PC103      PC104      PC105      PC106      PC107      PC108
Standard deviation     0.02173771 0.02144804 0.02114322 0.02103189 0.02086914 0.02065242
Proportion of Variance 0.00162000 0.00158000 0.00154000 0.00152000 0.00150000 0.00147000
Cumulative Proportion  0.91946000 0.92104000 0.92258000 0.92410000 0.92560000 0.92706000
                            PC109      PC110      PC111      PC112      PC113      PC114
Standard deviation     0.02033067 0.02023229 0.02004202 0.01989872 0.01975983 0.01957412
Proportion of Variance 0.00142000 0.00141000 0.00138000 0.00136000 0.00134000 0.00132000
Cumulative Proportion  0.92848000 0.92989000 0.93127000 0.93263000 0.93397000 0.93529000
                            PC115      PC116      PC117      PC118      PC119      PC120
Standard deviation     0.01944685 0.01934141 0.01919089 0.01906135 0.01896053 0.01881113
Proportion of Variance 0.00130000 0.00129000 0.00127000 0.00125000 0.00124000 0.00122000
Cumulative Proportion  0.93659000 0.93787000 0.93914000 0.94039000 0.94162000 0.94284000
                           PC121      PC122      PC123      PC124      PC125      PC126
Standard deviation     0.0187546 0.01861762 0.01844026 0.01822854 0.01801426 0.01786499
Proportion of Variance 0.0012100 0.00119000 0.00117000 0.00114000 0.00112000 0.00110000
Cumulative Proportion  0.9440500 0.94524000 0.94641000 0.94755000 0.94866000 0.94976000
                            PC127      PC128      PC129      PC130      PC131      PC132
Standard deviation     0.01785116 0.01775044 0.01756618 0.01746211 0.01721473 0.01709386
Proportion of Variance 0.00110000 0.00108000 0.00106000 0.00105000 0.00102000 0.00100000
Cumulative Proportion  0.95086000 0.95194000 0.95300000 0.95405000 0.95506000 0.95607000
                            PC133      PC134      PC135      PC136      PC137      PC138
Standard deviation     0.01705175 0.01684786 0.01671747 0.01664932 0.01648871 0.01640131
Proportion of Variance 0.00100000 0.00098000 0.00096000 0.00095000 0.00093000 0.00092000
Cumulative Proportion  0.95707000 0.95804000 0.95900000 0.95996000 0.96089000 0.96181000
                            PC139      PC140      PC141      PC142      PC143     PC144
Standard deviation     0.01632371 0.01600625 0.01580221 0.01571107 0.01562155 0.0155936
Proportion of Variance 0.00092000 0.00088000 0.00086000 0.00085000 0.00084000 0.0008400
Cumulative Proportion  0.96273000 0.96361000 0.96447000 0.96532000 0.96616000 0.9669900
                            PC145     PC146      PC147      PC148      PC149    PC150
Standard deviation     0.01537886 0.0152965 0.01517045 0.01506012 0.01501493 0.014774
Proportion of Variance 0.00081000 0.0008000 0.00079000 0.00078000 0.00077000 0.000750
Cumulative Proportion  0.96780000 0.9686100 0.96940000 0.97018000 0.97095000 0.971700
                            PC151     PC152      PC153      PC154      PC155      PC156
Standard deviation     0.01463149 0.0144876 0.01434792 0.01433986 0.01424448 0.01412483
Proportion of Variance 0.00074000 0.0007200 0.00071000 0.00071000 0.00070000 0.00069000
Cumulative Proportion  0.97244000 0.9731600 0.97387000 0.97457000 0.97527000 0.97596000
                            PC157      PC158      PC159      PC160      PC161      PC162
Standard deviation     0.01410833 0.01396131 0.01392313 0.01371741 0.01365028 0.01356903
Proportion of Variance 0.00068000 0.00067000 0.00067000 0.00065000 0.00064000 0.00063000
Cumulative Proportion  0.97664000 0.97731000 0.97798000 0.97862000 0.97926000 0.97990000
                            PC163      PC164      PC165      PC166      PC167      PC168
Standard deviation     0.01349251 0.01332544 0.01327065 0.01325211 0.01294927 0.01286386
Proportion of Variance 0.00063000 0.00061000 0.00061000 0.00060000 0.00058000 0.00057000
Cumulative Proportion  0.98052000 0.98113000 0.98174000 0.98234000 0.98292000 0.98349000
                            PC169      PC170      PC171      PC172      PC173      PC174
Standard deviation     0.01276056 0.01258496 0.01257078 0.01251062 0.01231667 0.01228647
Proportion of Variance 0.00056000 0.00054000 0.00054000 0.00054000 0.00052000 0.00052000
Cumulative Proportion  0.98404000 0.98459000 0.98513000 0.98567000 0.98619000 0.98671000
                            PC175      PC176      PC177      PC178      PC179      PC180
Standard deviation     0.01217565 0.01199646 0.01196251 0.01171993 0.01152939 0.01141553
Proportion of Variance 0.00051000 0.00049000 0.00049000 0.00047000 0.00046000 0.00045000
Cumulative Proportion  0.98722000 0.98771000 0.98821000 0.98868000 0.98913000 0.98958000
                            PC181      PC182     PC183      PC184      PC185      PC186
Standard deviation     0.01128977 0.01124502 0.0110693 0.01099837 0.01089239 0.01081447
Proportion of Variance 0.00044000 0.00043000 0.0004200 0.00042000 0.00041000 0.00040000
Cumulative Proportion  0.99002000 0.99045000 0.9908800 0.99129000 0.99170000 0.99210000
                            PC187      PC188      PC189      PC190      PC191      PC192
Standard deviation     0.01072824 0.01050272 0.01043874 0.01033548 0.01013789 0.01006607
Proportion of Variance 0.00040000 0.00038000 0.00037000 0.00037000 0.00035000 0.00035000
Cumulative Proportion  0.99250000 0.99288000 0.99325000 0.99362000 0.99397000 0.99432000
                             PC193       PC194       PC195       PC196       PC197
Standard deviation     0.009888063 0.009794131 0.009684863 0.009511312 0.009457666
Proportion of Variance 0.000340000 0.000330000 0.000320000 0.000310000 0.000310000
Cumulative Proportion  0.994650000 0.994980000 0.995310000 0.995620000 0.995920000
                             PC198       PC199       PC200       PC201      PC202
Standard deviation     0.009313158 0.009225365 0.009156228 0.008910202 0.00886853
Proportion of Variance 0.000300000 0.000290000 0.000290000 0.000270000 0.00027000
Cumulative Proportion  0.996220000 0.996520000 0.996800000 0.997080000 0.99735000
                             PC203       PC204       PC205       PC206      PC207
Standard deviation     0.008698389 0.008673122 0.008489401 0.008365039 0.00825881
Proportion of Variance 0.000260000 0.000260000 0.000250000 0.000240000 0.00023000
Cumulative Proportion  0.997610000 0.997860000 0.998110000 0.998350000 0.99859000
                             PC208       PC209       PC210       PC211       PC212
Standard deviation     0.008037357 0.007981711 0.007688713 0.007528352 0.007244244
Proportion of Variance 0.000220000 0.000220000 0.000200000 0.000190000 0.000180000
Cumulative Proportion  0.998810000 0.999030000 0.999230000 0.999430000 0.999610000
                             PC213       PC214       PC215      PC216        PC217
Standard deviation     0.007139559 0.005997918 0.004271462 0.00305905 1.082174e-16
Proportion of Variance 0.000180000 0.000120000 0.000060000 0.00003000 0.000000e+00
Cumulative Proportion  0.999780000 0.999910000 0.999970000 1.00000000 1.000000e+00

The following observations are calculated as outliers: 
[1] "a3"  "a6"  "B22" "F84" "F86" "F88"

PLS(-DA) Two Component Model Summary

217 samples x 1309 variables

Cumulative Proportion of Variance Explained: R2X(cum) = 12.86179%

Cumulative Proportion of Response(s):
                Y1        Y2        Y3         Y4          Y5        Y6        Y7
R2Y(cum) 0.1969126 0.2318528 0.1584459 0.10698318 0.020843739 0.3107075 0.7235089
Q2Y(cum) 0.1341826 0.1893128 0.1428475 0.08146868 0.007217958 0.2840960 0.6905388

Warning: More than two groups, permutation test skipped!

Warning: VIP was only implemented for the single-response model!

Step 5: Univariate Test Start...! Time:  Fri Jan  6 11:58:33 2017

P-value Calculating...

*P-value was adjusted using Benjamini-Hochberg Method

Odd.Ratio Calculating...

ROC Calculating...

*Group.G1 Vs. Group.G2
  |===============================================================================| 100%

*Group.G1 Vs. Group.G3
  |===============================================================================| 100%

*Group.G1 Vs. Group.G4
  |===============================================================================| 100%

*Group.G1 Vs. Group.G5
  |===============================================================================| 100%

*Group.G1 Vs. Group.G6
  |===============================================================================| 100%

*Group.G1 Vs. Group.QC
  |===============================================================================| 100%

*Group.G2 Vs. Group.G3
  |===============================================================================| 100%

*Group.G2 Vs. Group.G4
  |===============================================================================| 100%

*Group.G2 Vs. Group.G5
  |===============================================================================| 100%

*Group.G2 Vs. Group.G6
  |===============================================================================| 100%

*Group.G2 Vs. Group.QC
  |===============================================================================| 100%

*Group.G3 Vs. Group.G4
  |===============================================================================| 100%

*Group.G3 Vs. Group.G5
  |===============================================================================| 100%

*Group.G3 Vs. Group.G6
  |===============================================================================| 100%

*Group.G3 Vs. Group.QC
  |===============================================================================| 100%

*Group.G4 Vs. Group.G5
  |===============================================================================| 100%

*Group.G4 Vs. Group.G6
  |===============================================================================| 100%

*Group.G4 Vs. Group.QC
  |===============================================================================| 100%

*Group.G5 Vs. Group.G6
  |===============================================================================| 100%

*Group.G5 Vs. Group.QC
  |===============================================================================| 100%

*Group.G6 Vs. Group.QC
  |===============================================================================| 100%

RandomForest Calculating...

*Group.G1 Vs. Group.G2
  |===============================================================================| 100%

*Group.G1 Vs. Group.G3
  |===============================================================================| 100%

*Group.G1 Vs. Group.G4
  |===============================================================================| 100%

*Group.G1 Vs. Group.G5
  |===============================================================================| 100%

*Group.G1 Vs. Group.G6
  |===============================================================================| 100%

*Group.G1 Vs. Group.QC
  |===============================================================================| 100%

*Group.G2 Vs. Group.G3
  |===============================================================================| 100%

*Group.G2 Vs. Group.G4
  |===============================================================================| 100%

*Group.G2 Vs. Group.G5
  |===============================================================================| 100%

*Group.G2 Vs. Group.G6
  |===============================================================================| 100%

*Group.G2 Vs. Group.QC
  |===============================================================================| 100%

*Group.G3 Vs. Group.G4
  |===============================================================================| 100%

*Group.G3 Vs. Group.G5
  |===============================================================================| 100%

*Group.G3 Vs. Group.G6
  |===============================================================================| 100%

*Group.G3 Vs. Group.QC
  |===============================================================================| 100%

*Group.G4 Vs. Group.G5
  |===============================================================================| 100%

*Group.G4 Vs. Group.G6
  |===============================================================================| 100%

*Group.G4 Vs. Group.QC
  |===============================================================================| 100%

*Group.G5 Vs. Group.G6
  |===============================================================================| 100%

*Group.G5 Vs. Group.QC
  |===============================================================================| 100%

*Group.G6 Vs. Group.QC
  |===============================================================================| 100%

Volcano Plot and Box Plot Output...

Statistical Analysis Finished! Time: Fri Jan  6 12:42:04 2017

4 Session info

Here is the output of sessionInfo on the system on which this document was compiled:

sessionInfo()
#> R version 3.5.0 (2018-04-23)
#> Platform: x86_64-pc-linux-gnu (64-bit)
#> Running under: Ubuntu 16.04.4 LTS
#> 
#> Matrix products: default
#> BLAS: /home/biocbuild/bbs-3.7-bioc/R/lib/libRblas.so
#> LAPACK: /home/biocbuild/bbs-3.7-bioc/R/lib/libRlapack.so
#> 
#> locale:
#>  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
#>  [3] LC_TIME=en_US.UTF-8        LC_COLLATE=C              
#>  [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8   
#>  [7] LC_PAPER=en_US.UTF-8       LC_NAME=C                 
#>  [9] LC_ADDRESS=C               LC_TELEPHONE=C            
#> [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       
#> 
#> attached base packages:
#> [1] stats     graphics  grDevices utils     datasets  methods   base     
#> 
#> other attached packages:
#> [1] BiocInstaller_1.30.0 BiocStyle_2.8.0     
#> 
#> loaded via a namespace (and not attached):
#>  [1] Rcpp_0.12.16    bookdown_0.7    digest_0.6.15   rprojroot_1.3-2
#>  [5] backports_1.1.2 magrittr_1.5    evaluate_0.10.1 stringi_1.1.7  
#>  [9] rmarkdown_1.9   tools_3.5.0     stringr_1.3.0   xfun_0.1       
#> [13] yaml_2.1.18     compiler_3.5.0  htmltools_0.3.6 knitr_1.20

5 References

Dunn, W.B., et al.,Procedures for large-scale metabolic profiling of serum and plasma using gas chromatography and liquid chromatography coupled to mass spectrometry. Nature Protocols 2011, 6, 1060.

Luan H., LC-MS-Based Urinary Metabolite Signatures in Idiopathic Parkinson’s Disease. J Proteome Res., 2015, 14,467.

Luan H., Non-targeted metabolomics and lipidomics LC-MS data from maternal plasma of 180 healthy pregnant women. GigaScience 2015 4:16