Metabolomics Fiehn Lab plasticizer

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Metabolomics Fiehn Lab plasticizer Powered By Docstoc
					                              Multiple Stage Tandem Mass Spectrometry
                     for identification of plastic contaminations in blood plasma
                                  using nano electrospray ionization




                                          Tobias Kind and Kwang-Hyeon Liu
                                              FiehnLab UC Davis 2009

                  http://fiehnlab.ucdavis.edu/staff/kind/Metabolomics/LipidAnalysis




File under: Anecdotal mass spectrometry
CC-by Free Reuse License granted; Tobias Kind FiehnLab 2009
Instrumentation – robotized chip based nano electrospray ionization
                         with linear iontrap




                                 +




              nanoESI chip              LTQ Linear IonTrap


                Advion NanoMate robot not shown
                     Full scan nanoESI positive mode with data dependent
                                  multiple stage tandem MS

Plasma_Pos_LowMS_60MSMS_CE45_01 #35-73 RT: 0.07-0.15 AV: 39 NL: 2.53E4
T: ITMS + p ESI Full ms [200.00-1200.00]
   140
   130                        We will fragment this ion with the awe-inspiring power of
   120                        MS/MS/MS/MS/MS/MS/MS/MS or MS8
   110
                     650.55
   100
    90
    80
    70                                                                                  703.64
    60
                                                         678.64
    50
                            654.18
    40                                                         680.27
    30                                               676.55         686.27 690.55            705.64
    20        648.55            655.27 664.55                                       701.64    706.55
                                  660.45   666.55
    10      644.45                                                                              707.55

     0
      640             650            660       670            680          690       700         710
                                                      m/z




         Slowly repeat with me: MS1..MS2..MS3..MS4..MS5..MS6..MS7..MS8...
                                                   What could it be?
                     Plasma_Pos_LowMS_60MSMS_CE45_01 #184 RT: 1.15 AV: 1 NL: 2.29E3
                     T: ITMS + c ESI d Full ms2 664.00@cid45.00 [170.00-675.00]
                                                                                                            607.32
                        100
                         95                                                                       -56.8
                         90
                         85
                                  MS^2
                         80
                         75
                                  precursor m/z 664.00 (ESI+)
                         70
                         65
                                                                                                551.24
Relative Abundance




                         60
                         55
                         50
                                                                                    -112.76
                         45
                         40
                         35
                         30
                         25
                         20
                         15
                                                                   -168.74
                                                                                     495.26
                         10
                          5                              359.24                                          604.28 631.35
                                                                                             540.52
                                212.44     283.20 325.27                 437.36
                          0
                               200       250    300      350      400         450      500        550       600      650
                                                                        m/z
Plasma_Pos_LowMS_60MSMS_CE45_01 #190 RT: 1.27 AV: 1 NL: 1.16E2
T: ITMS + c ESI d Full ms8 664.00@cid45.00 607.32@cid45.00 551.33@cid45.00 495.34@cid35.00 439.26 ...
                                                            233.03
     100
    95
    90
                                                           214.97
    85
    80
           MS^8 (ESI+)
    75     precursor m/z 327.00
    70
    65
    60
    55
    50
    45
    40
    35
    30
    25
    20                         153.09                               251.07
    15
                                        171.07                                  291.17
    10                                                                                   309.12

      5                                     174.99
      0
              100              150                   200            250              300
                                                           m/z
Plasma_Pos_LowMS_60MSMS_CE45_01 #184-190 RT: 1.15-1.27 AV: 7 NL: 2.99E3
T: Composite spectrum MS8 664.00,607.32,551.33,495.34,439.26,383.17,327.06
                                                                           551.29
    100
    95
    90
               MS^8 composite tandem mass spectrum
    85
               of precursor m/z 664.00 (ESI+)
    80
                                                                                            607.32
                                                                            495.31
    75
    70
    65
    60
    55
    50
    45
    40
    35
                                              327.08
                                                       383.17      439.26
    30
    25
                                                             56 Da              56 Da
    20                                            56 Da                56 Da             56 Da
    15
    10
     5                         233.03
                153.09             251.07         359.24     419.50    480.52   540.52            631.35
     0
         100             200                300              400              500           600
                                                       m/z


                                                                                                   Sample: KH Liu (FiehnLab)
              List of ions from precursor m/z 664.00 (ESI+)


  Plasma_Pos_LowMS_60MSMS_CE45_01.raw
  Friday, April 03, 2009

  MS2 precursor 664.00
  |__MS3 precursor 607.32
  | |__MS4 precursor 551.33
  | | |__MS5 precursor 495.34
  | | | |__MS6 precursor 439.26
  | | | | |__MS7 precursor 383.17
  | | | | | |__MS8 precursor 327.06
  | | | | | | |__Composite spectrum MS8 664.00,607.32,551.33,495.34,439.26,383.17,327.06
  | | | | | | |__Single spectrum MS8 664.00,607.32,551.33,495.34,439.26,383.17,327.06 (190)
  | | | | | |__Composite spectrum MS7 664.00,607.32,551.33,495.34,439.26,383.17
  | | | | | |__Single spectrum MS7 664.00,607.32,551.33,495.34,439.26,383.17 (189)
  | | | | |__Composite spectrum MS6 664.00,607.32,551.33,495.34,439.26
  | | | | |__Single spectrum MS6 664.00,607.32,551.33,495.34,439.26 (188)
  | | | |__Composite spectrum MS5 664.00,607.32,551.33,495.34
  | | | |__Single spectrum MS5 664.00,607.32,551.33,495.34 (187)
  | | |__Composite spectrum MS4 664.00,607.32,551.33
  | | |__Single spectrum MS4 664.00,607.32,551.33 (186)
  | |__Composite spectrum MS3 664.00,607.32
  | |__Single spectrum MS3 664.00,607.32 (185)
  |__Single spectrum MS2 664.00 (184)




List of product ions:
m/z 664 , 607 ,551 ,495 ,439 ,383 ,327
accurate masses see above
 Solutions – well Plasma – well Lipids use LipidMaps tools!
CE(18:2(Ep))               18    2           663.5711   0.4289     C45H75O3+
CE(18:2(Ke))               18    2           663.5711   0.4289     C45H75O3+
CE(18:3(OH))               18    3           663.5711   0.4289     C45H75O3+
CE(18:1(Ep))-cyclo         18    1           663.5711   0.4289     C45H75O3+
CE(18:1(Ke))-cyclo         18    1           663.5711   0.4289     C45H75O3+
CE(18:2(OH))-cyclo         18    2           663.5711   0.4289     C45H75O3+
CE(16:2(Ke,Ep2))           16    2           663.4983   0.5017     C43H67O5+
CE(16:2(Ke2,Ep))           16    2           663.4983   0.5017     C43H67O5+
CE(16:3(Ep,OO))            16    3           663.4983   0.5017     C43H67O5+
CE(16:3(Ke,OO))            16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH,Ep2))           16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH,Ke,Ep))         16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH,Ke2))           16    3           663.4983   0.5017     C43H67O5+
CE(16:4(OOH,Ep))           16    4           663.4983   0.5017     C43H67O5+
CE(16:4(OOH,Ke))           16    4           663.4983   0.5017     C43H67O5+
CE(16:4(OH,OO))            16    4           663.4983   0.5017     C43H67O5+
CE(16:4(OH2,Ep))           16    4           663.4983   0.5017     C43H67O5+
CE(16:4(OH2,Ke))           16    4           663.4983   0.5017     C43H67O5+
CE(16:5(OH,OOH))           16    5           663.4983   0.5017     C43H67O5+
CE(16:5(OH3))              16    5           663.4983   0.5017     C43H67O5+
CE(16:1(Ke,Ep2))-cyclo     16    1           663.4983   0.5017     C43H67O5+
CE(16:1(Ke2,Ep))-cyclo     16    1           663.4983   0.5017     C43H67O5+
CE(16:2(Ep,OO))-cyclo      16    2           663.4983   0.5017     C43H67O5+
CE(16:2(Ke,OO))-cyclo      16    2           663.4983   0.5017     C43H67O5+
CE(16:2(OH,Ep2))-cyclo     16    2           663.4983   0.5017     C43H67O5+
CE(16:2(OH,Ke,Ep))-cyclo   16    2           663.4983   0.5017     C43H67O5+
CE(16:2(OH,Ke2))-cyclo     16    2           663.4983   0.5017     C43H67O5+
CE(16:3(OOH,Ep))-cyclo     16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OOH,Ke))-cyclo     16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH,OO))-cyclo      16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH2,Ep))-cyclo     16    3           663.4983   0.5017     C43H67O5+
CE(16:3(OH2,Ke))-cyclo     16    3           663.4983   0.5017     C43H67O5+
CE(16:4(OH,OOH))-cyclo     16    4           663.4983   0.5017     C43H67O5+
CE(16:4(OH3))-cyclo        16    4           663.4983   0.5017     C43H67O5+




 Current mass accuracy for unit mass resolution LTQ:
 78 ppm (external calibration) or 0.046 Da @ 594.46 Da for reference purposes
 LipidMaps Tools used with M+H
                         Solutions – well Plasma – well Lipids!


    CE(18:2(Ep))              18       2       663.5711   0.4289   C45H75O3+
    CE(18:2(Ke))              18       2       663.5711   0.4289   C45H75O3+
    CE(18:3(OH))              18       3       663.5711   0.4289   C45H75O3+
    CE(18:1(Ep))-cyclo        18       1       663.5711   0.4289   C45H75O3+
    CE(18:1(Ke))-cyclo        18       1       663.5711   0.4289   C45H75O3+
    CE(18:2(OH))-cyclo        18       2       663.5711   0.4289   C45H75O3+

    Sphingosine               CerP     18:1    644.5013   0.5013   C36H71NO6P+
    Sphingosine               HexCer   12:0    644.5096   0.5096   C36H70NO8+
    Sphinganine               Cer      24:4    644.5976   0.5976   C42H78NO3+




Problem: Our mass accuracy is 0.046 Da but the solutions are 0.5 Da difference

Without Google Scholar we would be lost to search single m/z values
or numbers. Still its a horrible way to do mass spectral library search.
Has anybody a Google API example?
                        Solutions – well Plasma – well Google It!


      On the lipid composition of human meibum and tears:
      Comparative analysis of nonpolar lipids
      Igor A. Butovich
      http://www.iovs.org/cgi/content/abstract/iovs.08-1889v1

"However, rushing to this conclusion would have been a mistake because subsequent fragmentation
of ion m/z 663 in sequential MSn experiments gave us the following chain of product ions
differing by 56 amu: 663 (MS1) -> 607 (MS2) -> 551 (MS3) -> 495 (MS4) -> 439 (MS5) -> 383 (MS6) -> 327 (MS7)
(not shown). Note that the first four ions (663, 607, 551, 495) were also easily detectable in a simple MS1 experiment
(Fig. 11A) . These transformations were indicative of a sequential loss of several up to six) t-butyl groups and
were formerly described for oxidized form of Irgafos 168 (C42H63O4P, Fig. 11A , inset) a
polymer additive present in polyethylene and polypropylene and its derivatives."




 Researcher working with tears = The Dacriologist (Book by Djerassi)
 Dakryology, the neologism for the science of tears
 10.1007/BF00137944
 also CRYING: The Natural and Cultural History of Tears By Tom Lutz Norton
              Solutions – well Plasma – well Plasticizers!



    Analysis of Additives in Polyethylene with Desorption
    Chemical Ionization/Tandem Mass Spectrometry

    S. W. CHEN and G. R. HER*
    Appl. Spectrosc. 47, 844-851 (1993)
    http://www.opticsinfobase.org/abstract.cfm?URI=as-47-6-844




          Picture Source: S. W. CHEN and G. R. HER Appl. Spectrosc. 47, 844-851 (1993)



Also Oxidized Naugard 524 - http://dx.doi.org/10.1016/0003-2670(95)00183-Z
                                        Solution – probably Irganox / Irgafos contamination




                                           Plasma_Pos_LowMS_60MSMS_CE45_01 #184-190 RT: 1.15-1.27 AV: 7 NL: 2.99E3
                                           T: Composite spectrum MS8 664.00,607.32,551.33,495.34,439.26,383.17,327.06




                                                                                                                      551
                                                  100

                                                    90




                                                                                                                             607
                                                                                                                495
                                                    80

                                                    70
  Relative Abundance




                                                    60

                                                    50

                                                    40


                                                                                        327




                                                                                                          439
                                                                                               383
                                                    30

                                                    20                   Our MS^8
                                                    10                   "Lipid"
                                                      0
                                                            100         200       300               400         500         600
                                                                                              m/z


Picture Source: Analysis of Additives in Polyethylene with Desorption
                                                                         Problem solved, "lipid" is oxidized plasticizer.
Chemical Ionization/Tandem Mass Spectrometry
S. W. CHEN and G. R. HER*
Appl. Spectrosc. 47, 844-851 (1993)
http://www.opticsinfobase.org/abstract.cfm?URI=as-47-6-844
                        Is the problem really solved?




                                    No, because for the formula = C42H63O4P
                                    the exact mass = 662.446397016.

                                    The [M+H]+ would be 663.4542 Da
                                    and not 664.0

                                    In this case an ultrazoom investigation or
                                    infusion into FT-ICR-MS with resolving power
         CID 14572930               50,000 to 100,000 would probably help.

InChIKey: AZSKHRTUXHLAHS-UHFFFAOYSA-N




MS/MS on isotopic peak, ion suppression of isobars, overlap with
multiple ceramides or cholesterol and oxycholesterol species possible
            Final question – How can we avoid rediscovery (reinventing the wheel)?

             • sharing experiments (free)
             • sharing mass spectra (free or sell)
             • putting publications into OA for easier searching


             How can we prevent plasticizer contaminations?

             • running solvent blanks with each experiment
             • running methods blanks with each experiment
             • using only (baked) glassware

             Other solutions?

             • library search (but compound is not in NIST)
             • remember the awesome power of MS/MS/MS/MS/MS/MS/MS/MS
             • modern iontraps are currently not used to the full extend Data Dependent
             • exclusion libraries for unwanted "contaminations"



See: XLS supplement from Keller, B.O.; Sui, J.; Young, A.B.; Whittal, R.M.
Interferences and contaminants encountered in modern mass spectrometry.

				
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