CSA analysis by IDSL.CSA
Composite Spectra Analysis (CSA) was devised as an analytical methodology for the aggregation of correlating MS1-only peaks extracted from IDSL.IPA peaklists. The objective of the CSA workflow is to streamline the examination of extensive mass spectrometry data through the grouping of related MS1-only peaks into a composite spectra. This approach facilitates the identification and interpretation of the biological information inherent presented by peaks only in MS1 level, and offers a more efficient and coherent means of analyzing complex mass spectrometry datasets. There are two approaches represented by CSA0012 in the CSA tab of the IDSL.CSA parameter spreadsheet to detect co-occurring fragmentation peaks.
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The peakList approach is based on solely utilizing the inter-sample correlation among IDSL.IPA peaks to provide a straightforward method to analyze individual samples. This approach is generally beneficial when the aligned table information is not available.
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The alignedTable approach groups related IDSL.IPA peaks based on their previously established correlation on the aligned peak height table prior to inter-sample corrolation analyses. This appraoch is particularly advantageous when the samples being analyzed are complex, and strong co-elution of MS1 ions is present to eliminate randomly co-occurring ions that are present in only a limited number of samples.
It is important to note that the CSA analysis is not able to determine precursor values, and the exported MSP fragmentation spectra do not contain PrecursorMZ entries. Therefore, precursor matching option should be ignored to annotate the MSP files generated from the CSA analysis.
MS1-only chromatogram deconvolution for Pyridoxine in the MSV000090242 study using the CSA approach.
Aligned smoothed EICs:
Annotation:
Three points about this example:
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In few instances, the intensity of the aligned EIC fgiures may not be proportionate to the intensity of ions during the annotation step. This is due to two factors: Firstly, the LOESS smoothing process applied to the EICs can result in a reduction of overall peak heights. Secondly, during the MSP exportation for annotation, the cumulated raw intensity of the chromatogram is used which is not proportionate to the intensity of the aligned EIC figures in few cases.
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The precursor value was not utilized in the annotation process despite the availability in the reference library highlighting the strength of the IDSL.FSA annotation workflow which allows optional precursor match.
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This compound in the same sample was also annotated using MS2 SWATH-MS data at a higher collision energy, demonstrating the utility of lower collision energy libraries in the CSA annotation process.
The IDSL.CSA software utilizes a novel approach of grouping individual Composite Spectra Analysis (CSA) peaks at the level of the aligned table generated by the IDSL.IPA workflow. This allows for the utilization of statistical power on the aligned table level to cluster the related CSA spectra. In instances where a peak (m/z-RT) may be present in multiple CSA clusters, Tanimato coefficients are calculated for each aligned CSA spectra. Then, the peaks (m/z-RT) with the greatest Tanimato coefficients are clustered into an aligned CSA spectra. Subsequently, two networks, comprised of the most abundant CSA and integrated CSA spectra respectively, are created and can be visualized through the use of Cytoscape. Additionally, the CSA spectra networks can be annotated using an FSDB to facilitate interpretation of the visualized networks.
Example: Most abundant CSA spectra for Glutathione (oxidized) on the aligned table was isolated from one sample from ST002263 study.
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Example: Aligned integarted CSA spectra for Glutathione (oxidized) from ST002263 study.
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