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Quantitative analysis of complex oligomeric mixtures by MS

Sheetal Gangula, Institute of Food chemistry

Quantitative Analysis of Cellulose Ethers

WHAT

 Distribution of substituents along the polymer chain o o random distribution?

high and low substituted areas?

WHY

 structure-property relationships o E.g. thermo-reversible gelation

HOW

 quantitative mass spectrometry (MS) of oligomers 8 May 2014 | Sheetal Gangula |TU Braunschweig 2

Quantitative MS of Methyl Cellulose

methyl cellulose

[M+Na] + number of permethylation with CD 3 I

CH 3 CD 3 R. Adden

et al

.,

Macromol. Chem. Phys.,

207 (2006) 954-965

partial degradation to oligomers mass spectrometric analysis [M+Na] +

8 May 2014 | Sheetal Gangula |TU Braunschweig 3

Analysis by MS QUALITATIVE ANALYSIS

Peak Identified by m/z value

QUANTITATIVE ANALYSIS

Depends on peak intensity But this is not simple as peak intensity varies depending on chemistry,instrumental parameters and mass differnce 8 May 2014 | Sheetal Gangula |TU Braunschweig 4

Analysis by MS

For example different compounds at equimolar concentrations give different peak intensities m/z Mass spectrum from

ESI-IT-MS DP2 Me DP2 Et

m/z The samples are seen as sodium adducts M+Na + in MS So for quantitative evaluation behaviour of different compounds with respect to chemistry and instrumental parameters has to be studied 8 May 2014 | Sheetal Gangula |TU Braunschweig 5

Our current study

preparation of a roughly equimolar

oligosaccharide mixtures

determination of the exact composition

Samples measured under defined conditions ESI-IT MS MALDI-ToF-MS Evaluation of sodium adducts in mass spectrum

8 May 2014 | Sheetal Gangula |TU Braunschweig 6

Sample Preparation

2,3,6-O-

Me

β-CD Partial hydrolysis 2,3,6-O-

Med

3

-

β-CD Reductive amination Mixture of oligosaccharides (DP1-DP7) are obtainted from each sample 8 May 2014 | Sheetal Gangula |TU Braunschweig Mixture of oligosaccharides (DP1-DP7) after reductive amination 2,3,6-O-

Et-

β-CD

LC/UV

7

Types of mixtures

Three types of mixtures are presented :  Methylated and deuteromethylated oligomeric mixtures

( Me

,

Med

3

)

 Methylated and ethylated oligomeric mixtures

( Me

,

Et )

 Deuteromethylated and ethylated oligomeric mixtures

( Med

3,

Et )

8 May 2014 | Sheetal Gangula |TU Braunschweig 8

ESI-IT-MS (electrospary ionization mass spectrometry)

8 May 2014 | Sheetal Gangula |TU Braunschweig 9

Results from ESI-IT-MS

Results are expressed as

relative ion intensities

. This is obtained by dividing the intensity of one compound in mass spectra by another compound w.r.t a certain DP in a mixture.

Me

,

Med

3

mixture Relative ion intensity

4 3 6 5 2 1 1,09 0

DP2 Med 3

0,92

/ Me

0,96

DP4 Me

,

Et mixture Relative ion intensity

6 5 1 0 4 3,37 3 2

DP2 Et

2,68

/ Me DP3

RI 2,69

DP4 Med

3 ,

Et mixture

6 5 4 1 0 3 2

Relative ion intensity Et / Med 3

3,25 2,53 2,48

DP3

8 May 2014 | Sheetal Gangula |TU Braunschweig 10

COST action FP1105 STSM

MALDI-ToF-MS:

Measurements were performed at: The institute of Polymer Technology at KTH Royal Institute of Technology, Stockholm Instrument used: Bruker Daltonics Ultraflex 8 May 2014 | Sheetal Gangula |TU Braunschweig

MALDI-ToF-MS (matrix assisted laser desorption mass spectrometry)

sDHB 8 May 2014 | Sheetal Gangula |TU Braunschweig 11

Selection of laser power

Me , Med

3

Intens.

3000 381.2

0_N2\1: +MS

LP 40 %

2000 1000 467.2

273.3

0.6

0.4

0.2

0.0

x10 0 5 1.0

0.8

2 1 x10 0 4 5 4 3 200 273.3

273.3

381.2

381.2

400 467.2

467.2

600 680.5

680.4

680.4

800 893.7

857.4

893.6

857.4

893.7

857.4

1000 1106.9

1061.6

1106.8

1061.5

1106.9

1061.6

1200 1320.0

1320.0

1400 1533.2

0_N2\1: +MS

LP 45 %

1533.2

LP 50 %

1320.1

0_N2\1: +MS 1533.2

m/z

Me , Et

0.8

0.6

0.4

0.2

0.0

Intens.

x10 4 4 3 x10 0 5 0.8

0.6

0.4

0.2

0.0

x10 5 2 1 287.3

287.3

287.3

200 375.0

449.3

533.4

653.5

779.7

857.7

1025.9

449.2

449.2

533.4

653.5

779.7

857.6

1025.9

400 533.4

653.4

779.6 857.6

1025.9

600 800 1000 1200 1272.1

1272.1

1272.2

1400 1470.1

1470.2

1518.4

1600 0_D4\1: +MS

LP 50 %

1746.6

0_D4\1: +MS

LP 55 %

1746.6

0_D4\1: +MS

LP 60 %

1765.7

1800 m/z 8 May 2014 | Sheetal Gangula |TU Braunschweig 12

Results from MALDI-ToF-MS

Results are expressed as

relative ion intensities

. This is obtained by dividing the intensity of one compound in mass spectra by another compound w.r.t a certain DP in a mixture.

Me

,

Med

3

mixture

4 3 6 5 2 1 1,03 0

DP2

0,99

DP3

RI 0,99

DP4 Me

,

Et mixture

6,00 5,00 4,00 3,00 2,00 1,59 1,00 0,00

DP2

1,96

DP3

RI 1,64

DP4 Med

3 ,

Et mixture

6 5 4 1 0 3 2 1,86

DP2

2,29 1,91

DP4

8 May 2014 | Sheetal Gangula |TU Braunschweig 13

SUMMARY

Under optimal conditions for both instruments...

 Me and Me

d 3

oligosaccharides are similar in polarities, hence their relative ion intensities in mass spectra are almost 1:1 with a little scattering for various DP. The scattering of data is lees in MALDI-ToF-MS compared to ESI-IT-MS.

 Polarity of Et oligosaccharides is lower than that of Me-derivatives and Me

d 3 -

derivatives, these compounds in ESI-IT-MS:  show 3-4 times higher intensities, but this factor is decreasing with DP MALDI-ToF-MS:  the factor is scattering around 1.5-2.5, but it has no clear trend. May be since there is difference in polarities of the compounds they may have different interaction with matrix so the scattering is higher. This has to be further investigated.

8 May 2014 | Sheetal Gangula |TU Braunschweig 14

Thank you for your kind attention

8 May 2014 | Sheetal Gangula |TU Braunschweig 15