Hello Hiroshi,
Thanks so much for developing MS-dial, it is such a great program!
I am currently analysing metabolomics data using version 4.6.
After processing the raw data and scrolling through the ion table I opened up the chromatogram and visualised the individual peaks for each sample. I noticed that lots of those samples had a peak intensity of 0, despite there being a clear clean peak there (right top table, picture 1). I learned then that I can manually modify those peaks by following what is indicated on the left table (picture 1). So after updating I then got an intensity for all peaks (picture 2).
I was wondering:
- Why are so many peaks not detected and assigned an intensity of 0? I unticked the gap-filling option so I wondered if this was the reason?
- If that was not the reason, should I perform this manual adjustment for all metabolites detected?
Prior to this I would replace all values by 1/10 of the minimum peak intensity during the export option. However, I can really see how this could bias my results, especially when that 1/10 assigned would not approximate the actual intensity of those peaks. On the other side, I could never manually change all of them as I detected over 19'000, so if I only did it for a subset of more interesting ones, I would also be biasing my final result. What do you suggest?
I couldn't find any reference to this functionality on the tutorial page, so I wanted to check directly with you.
Thanks in advance for your input.
Giulia.
Could you please provide some detail about your parameter setting, e.g., peak detection and alignment tab.
Best,
Sukis
Hi Sukis,
sure, below are all of the parameters I used. Same both for positive and negative mode. Thanks!
MS-DIAL ver. 4.60
#Project
MS1 Data type Profile
MS2 Data type Profile
Ion mode Negative
Target Metablomics
Mode ddMSMS
#Data collection parameters
Retention time begin 0
Retention time end 32
Mass range begin 50
Mass range end 1000
MS2 mass range begin 50
MS2 mass range end 1000
#Centroid parameters
MS1 tolerance 0.002
MS2 tolerance 0.002
#Isotope recognition
Maximum charged number 2
#Data processing
Number of threads 2
#Peak detection parameters
Smoothing method LinearWeightedMovingAverage
Smoothing level 3
Minimum peak width 5
Minimum peak height 100000
#Peak spotting parameters
Mass slice width 0.05
Exclusion mass list (mass & tolerance)
#Deconvolution parameters
Sigma window value 0.5
MS2Dec amplitude cut off 0
Exclude after precursor True
Keep isotope until 0.5
Keep original precursor isotopes False
#MSP file and MS/MS identification setting
MSP file E:\Metabolomics\Database\MSMS-Neg-MassBank.msp
Retention time tolerance 0.1
Accurate mass tolerance (MS1) 0.002
Accurate mass tolerance (MS2) 0.002
Identification score cut off 80
Using retention time for scoring False
Using retention time for filtering False
#Text file and post identification (retention time and accurate mass based) setting
Text file E:\Metabolomics\Database\Edited_STd.txt
Retention time tolerance 0.1
Accurate mass tolerance 0.002
Identification score cut off 85
#Advanced setting for identification
Relative abundance cut off 0
Top candidate report True
#Adduct ion setting
[M-H]-
[M+Na-2H]-
[M+Cl]-
#Alignment parameters setting
Reference file E:\Metabolomics\MSDial abf converted data\28-QC_2.abf
Retention time tolerance 0.1
MS1 tolerance 0.003
Retention time factor 0.5
MS1 factor 0.5
Peak count filter 0
N% detected in at least one group 0
Remove feature based on peak height fold-change True
Sample max / blank average 5
Sample average / blank average 5
Keep identified and annotated metabolites False
Keep removable features and assign the tag for checking False
Gap filling by compulsion False
#Tracking of isotope labels
Tracking of isotopic labels FALSE
#Ion mobility
Ion mobility data FALSE
Hi Giacono,
Is it possible that you have too small MS1 tolerance for alignment?
Best,
Sukis
Hi Giacono,
>> I unticked the gap-filling option so I wondered if this was the reason?
Yes, this is the reason. Did you try the processing with the gap-filling option?
Hiroshi