# A parser for raw and identification mass-spectrometry data

Bernd Fischer, Steffen Neumann, Laurent Gatto and Qiang Kou

#### 1 May 2024

#### Package

mzR 2.38.0

# 1 Introduction

The mzR package aims at providing a common, low-level interface to several mass spectrometry data formats, namely, `mzXML` (Pedrioli et al. 2004), `mzML` (Martens et al. 2010) for raw data, and `mzIdentML` (Jones et al. 2012), somewhat similar to the Bioconductor package affyio for affymetrix raw data. No processing is done in `r BiocStyle::Biocpkg("mzR")`, which is left to packages such as `r BiocStyle::Biocpkg("xcms")` (Smith et al. 2006, Tautenhahn:2008) or `r BiocStyle::Biocpkg("MSnbase")` (Gatto and Lilley 2012). These packages also provide more convenient, high-level interfaces to raw and identification. data

Most importantly, access to the data should be fast and memory efficient. This is made possible by allowing on-disk random file access, i.e. retrieving specific data of interest without having to sequentially browser the full content nor loading the entire data into memory.

The actual work of reading and parsing the data files is handled by the included C/C++ libraries or backends. The C++ reference implementation for the `mzML` is the proteowizard library (Kessner et al. 2008) (pwiz in short), which in turn makes use of the boost C++ (http://www.boost.org/) library. More recently, the proteowizard (http://proteowizard.sourceforge.net/) (Chambers et al. 2012) has been fully integrated using the `mzRpwiz` backend for raw data, and is not the default option. The `mzRnetCDF` backend provides support to `CDF`-based formats. Finally, the `mzRident` backend is available to access identification data (`mzIdentML`) through pwiz.

The mzR package is in essence a collection of wrappers to the C++ code, and benefits from the C++ interface provided through the Rcpp package (Eddelbuettel and François 2011).

IMPORTANT New developers that need to access and manipulate raw mass spectrometry data are advised against using this infrastucture directly. They are invited to use the corresponding `MSnExp` (with on disk mode) from theMSnbase package instead. The latter supports reading multiple files at once and offers access to the spectra data (m/z and intensity) as well as all the spectra metadata using a coherent interface. The MSnbase infrastructure itself used the low level classes in mzR, thus offering fast and efficient access.

# 2 Mass spectrometry raw data

All the mass spectrometry file formats are organized similarly, where a set of metadata nodes about the run is followed by a list of spectra with the actual masses and intensities. In addition, each of these spectra has its own set of metadata, such as the retention time and acquisition parameters.

## 2.1 Spectral data access

Access to the spectral data is done via the `peaks` function. The return value is a list of two-column mass-to-charge and intensity matrices or a single matrix if one spectrum is queried.

## 2.2 Chromatogram access

Access to the chromatogram(s) is done using the `chromatogram` (or `chromatograms`) function, that return one (or a list of) data.frames. See `?chromatogram` for details. This functionality is only available with the `pwiz` backend.

## 2.3 Identification result access

The main access to identification result is done via `psms`, `score` and `modifications`. `psms` and `score` will return the detailed information on each psm and scores. `modifications` will return the details on each modification found in peptide.

## 2.4 Metadata access

Run metadata is available via several functions such as `instrumentInfo()` or `runInfo()`. The individual fields can be accessed via e.g. `detector()` etc.

Spectrum metadata is available via `header()`, which will return a list (for single scans) or a dataframe with information such as the `basePeakMZ`, `peaksCount`, … or, for higher-order MS the `msLevel` and precursor information.

Identification metadatais available via `mzidInfo()`, which will return a list with information such as the `software`, `ModificationSearched`, `enzymes`, `SpectraSource` and other information for this identification result.

The availability of this metadata can not always be guaranteed, and depends on the MS software which converted the data.

# 3 Example

## 3.1 `mzXML`/`mzML` files

A short example sequence to read data from a mass spectrometer. First open the file.

    library(mzR)

    ## Loading required package: Rcpp

    library(msdata)

    mzxml <- system.file("threonine/threonine_i2_e35_pH_tree.mzXML",
                         package = "msdata")
    aa <- openMSfile(mzxml)

We can obtain different kind of header information.

    runInfo(aa)

    ## $scanCount
    ## [1] 55
    ##
    ## $lowMz
    ## [1] 50.0036
    ##
    ## $highMz
    ## [1] 298.673
    ##
    ## $dStartTime
    ## [1] 0.3485
    ##
    ## $dEndTime
    ## [1] 390.027
    ##
    ## $msLevels
    ## [1] 1 2 3 4
    ##
    ## $startTimeStamp
    ## [1] NA

    instrumentInfo(aa)

    ## $manufacturer
    ## [1] "Thermo Scientific"
    ##
    ## $model
    ## [1] "LTQ Orbitrap"
    ##
    ## $ionisation
    ## [1] "electrospray ionization"
    ##
    ## $analyzer
    ## [1] "fourier transform ion cyclotron resonance mass spectrometer"
    ##
    ## $detector
    ## [1] "unknown"
    ##
    ## $software
    ## [1] "Xcalibur software 2.2 SP1"
    ##
    ## $sample
    ## [1] ""
    ##
    ## $source
    ## [1] ""

    header(aa,1)

    ##   seqNum acquisitionNum msLevel polarity peaksCount totIonCurrent retentionTime
    ## 1      1              1       1        1        684     341427000        0.3485
    ##   basePeakMZ basePeakIntensity collisionEnergy ionisationEnergy   lowMZ  highMZ
    ## 1    120.066         211860000              NA                0 50.3254 298.673
    ##   precursorScanNum precursorMZ precursorCharge precursorIntensity mergedScan
    ## 1               NA          NA              NA                 NA         NA
    ##   mergedResultScanNum mergedResultStartScanNum mergedResultEndScanNum
    ## 1                  NA                       NA                     NA
    ##   injectionTime filterString                                 spectrumId
    ## 1             0         <NA> controllerType=0 controllerNumber=1 scan=1
    ##   centroided ionMobilityDriftTime isolationWindowTargetMZ
    ## 1       TRUE                   NA                      NA
    ##   isolationWindowLowerOffset isolationWindowUpperOffset scanWindowLowerLimit
    ## 1                         NA                         NA              50.3254
    ##   scanWindowUpperLimit
    ## 1              298.673

Read a single spectrum from the file.

    pl <- peaks(aa,10)
    peaksCount(aa,10)

    ## [1] 317

    head(pl)

    ##            mz intensity
    ## [1,] 50.08176  6984.858
    ## [2,] 50.62267  7719.419
    ## [3,] 50.70530  7185.290
    ## [4,] 50.73298  7509.140
    ## [5,] 50.83848  9366.624
    ## [6,] 50.88303  8012.808

    plot(pl[,1], pl[,2], type="h", lwd=1)

One should always close the file when not needed any more. This will release the memory of cached content.

    close(aa)

## 3.2 `mzIdentML` files

You can use `openIDfile` to read a `mzIdentML` file (version 1.1), which use the pwiz backend.

    library(mzR)
    library(msdata)

    file <- system.file("mzid", "Tandem.mzid.gz", package="msdata")
    x <- openIDfile(file)

`mzidInfo` function will return general information about this identification result.

    mzidInfo(x)

    ## $FileProvider
    ## [1] "researcher"
    ##
    ## $CreationDate
    ## [1] "2012-07-25T14:03:16"
    ##
    ## $software
    ## [1] "xtandem x! tandem CYCLONE (2010.06.01.5) "
    ## [2] "ProteoWizard MzIdentML 3.0.21263 ProteoWizard"
    ##
    ## $ModificationSearched
    ## [1] "Oxidation"       "Carbamidomethyl"
    ##
    ## $FragmentTolerance
    ## [1] "0.8 dalton"
    ##
    ## $ParentTolerance
    ## [1] "1.5 dalton"
    ##
    ## $enzymes
    ## $enzymes$name
    ## [1] "Trypsin"
    ##
    ## $enzymes$nTermGain
    ## [1] "H"
    ##
    ## $enzymes$cTermGain
    ## [1] "OH"
    ##
    ## $enzymes$minDistance
    ## [1] "0"
    ##
    ## $enzymes$missedCleavages
    ## [1] "1"
    ##
    ##
    ## $SpectraSource
    ## [1] "D:/TestSpace/NeoTestMarch2011/55merge.mgf"

`psms` will return the detailed information on each peptide-spectrum-match, include `spectrumID`, `chargeState`, `sequence`. `modNum` and others.

    p <- psms(x)
    colnames(p)

    ##  [1] "spectrumID"               "chargeState"
    ##  [3] "rank"                     "passThreshold"
    ##  [5] "experimentalMassToCharge" "calculatedMassToCharge"
    ##  [7] "sequence"                 "peptideRef"
    ##  [9] "modNum"                   "isDecoy"
    ## [11] "post"                     "pre"
    ## [13] "start"                    "end"
    ## [15] "DatabaseAccess"           "DBseqLength"
    ## [17] "DatabaseSeq"              "DatabaseDescription"
    ## [19] "spectrum.title"           "acquisitionNum"

The modifications information can be accessed using `modifications`, which will return the `spectrumID`, `sequence`, `name`, `mass` and `location`.

    m <- modifications(x)
    head(m)

    ##   spectrumID                 sequence
    ## 1   index=12  LCYIALDFDEEMKAAEDSSDIEK
    ## 2   index=12  LCYIALDFDEEMKAAEDSSDIEK
    ## 3  index=285   KDLYGNVVLSGGTTMYEGIGER
    ## 4   index=83   KDLYGNVVLSGGTTMYEGIGER
    ## 5   index=21 VIDENFGLVEGLMTTVHAATGTQK
    ## 6  index=198          GVGGAIVLVLYDEMK
    ##                                            peptideRef            name    mass
    ## 1 LCYIALDFDEEMKAAEDSSDIEK_15.9949@M$12;_57.0215@C$2;_ Carbamidomethyl 57.0215
    ## 2 LCYIALDFDEEMKAAEDSSDIEK_15.9949@M$12;_57.0215@C$2;_       Oxidation 15.9949
    ## 3              KDLYGNVVLSGGTTMYEGIGER_15.9949@M$15;__       Oxidation 15.9949
    ## 4              KDLYGNVVLSGGTTMYEGIGER_15.9949@M$15;__       Oxidation 15.9949
    ## 5            VIDENFGLVEGLMTTVHAATGTQK_15.9949@M$13;__       Oxidation 15.9949
    ## 6                     GVGGAIVLVLYDEMK_15.9949@M$14;__       Oxidation 15.9949
    ##   location
    ## 1        2
    ## 2       12
    ## 3       15
    ## 4       15
    ## 5       13
    ## 6       14

Since different software will use different scoring function, we provide a `score` to extract the scores for each psm. It will return a data.frame with different columns depending on software generating this file.

    scr <- score(x)
    colnames(scr)

    ## [1] "spectrumID"          "X.Tandem.expect"     "X.Tandem.hyperscore"

# 4 Future plans

Other file formats provided by HUPO, such as `mzQuantML` for quantitative data (Walzer et al. 2013) are also possible in the future.

# 5 Session information

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Eddelbuettel, Dirk, and Romain François. 2011. “Rcpp: Seamless R and C++ Integration.” Journal of Statistical Software 40 (8): 1–18. http://www.jstatsoft.org/v40/i08/.
Gatto, L, and K S Lilley. 2012. “MSnbase – an R/Bioconductor Package for Isobaric Tagged Mass Spectrometry Data Visualization, Processing and Quantitation.” Bioinformatics 28 (2): 288–89. https://doi.org/10.1093/bioinformatics/btr645.
Jones, A R, M Eisenacher, G Mayer, O Kohlbacher, J Siepen, S J Hubbard, J N Selley, et al. 2012. “The mzIdentML Data Standard for Mass Spectrometry-Based Proteomics Results.” Mol Cell Proteomics 11 (7): M111.014381. https://doi.org/10.1074/mcp.M111.014381.
Kessner, Darren, Matt Chambers, Robert Burke, David Agus, and Parag Mallick. 2008. “ProteoWizard: Open Source Software for Rapid Proteomics Tools Development.” Bioinformatics 24 (21): 2534–36. https://doi.org/10.1093/bioinformatics/btn323.
Martens, Lennart, Matthew Chambers, Marc Sturm, Darren Kessner, Fredrik Levander, Jim Shofstahl, Wilfred H Tang, et al. 2010. “mzML - a Community Standard for Mass Spectrometry Data.” Molecular and Cellular Proteomics : MCP. https://doi.org/10.1074/mcp.R110.000133.
Pedrioli, Patrick G A, Jimmy K Eng, Robert Hubley, Mathijs Vogelzang, Eric W Deutsch, Brian Raught, Brian Pratt, et al. 2004. “A Common Open Representation of Mass Spectrometry Data and Its Application to Proteomics Research.” Nat. Biotechnol. 22 (11): 1459–66. https://doi.org/10.1038/nbt1031.
Smith, C A, E J Want, G O’Maille, R Abagyan, and G Siuzdak. 2006. “XCMS: Processing Mass Spectrometry Data for Metabolite Profiling Using Nonlinear Peak Alignment, Matching, and Identification.” Anal Chem 78 (3): 779–87. https://doi.org/10.1021/ac051437y.
Walzer, M, D Qi, G Mayer, J Uszkoreit, M Eisenacher, T Sachsenberg, F F Gonzalez-Galarza, et al. 2013. “The mzQuantML Data Standard for Mass Spectrometry-Based Quantitative Studies in Proteomics.” Mol Cell Proteomics 12 (8): 2332–40. https://doi.org/10.1074/mcp.O113.028506.