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Sheetal Gangula, Institute of Food chemistry

Quantitative analysis of complex oligomeric mixtures by MS

8 May 2014 | Sheetal Gangula |TU Braunschweig 2

WHAT

Distribution of substituents along the polymer chaino random distribution?o high and low substituted areas?

WHY

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

HOW

quantitative mass spectrometry (MS) of oligomers

Quantitative Analysis of Cellulose Ethers

8 May 2014 | Sheetal Gangula |TU Braunschweig 3

Quantitative MS of Methyl Cellulose

methyl cellulose

CH3

CD3

permethylationwith CD3I

partial degradation to oligomers

mass spectrometric analysis

R. Adden et al., Macromol. Chem. Phys., 207 (2006) 954-965

number of

[M+Na]+

[M+Na]+

8 May 2014 | Sheetal Gangula |TU Braunschweig

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

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8 May 2014 | Sheetal Gangula |TU Braunschweig

For example different compounds at equimolar concentrations give different peak intensities

Analysis by MS

So for quantitative evaluation behaviour of different compounds with respect to chemistry and instrumental parameters has to be studied

Mass spectrum from ESI-IT-MS

m/z-

m/z-

O

O

H3CO OCH3

OCH3

OH

O

HO

H3CO OCH3

OCH3

O

O

C2H5O

OC2H5

OH

OHO

C2H5OOC2H5

OC2H5

DP2-Me

DP2-Et

The samples are seen as sodium adducts M+Na+ in MS

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8 May 2014 | Sheetal Gangula |TU Braunschweig

Our current study

preparation of a roughly equimolar

oligosaccharide mixtures

determination of the

exact composition

Evaluation of sodium adducts in mass spectrum

ESI-IT MS MALDI-ToF-MS

Samples measured under defined conditions

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8 May 2014 | Sheetal Gangula |TU Braunschweig

Sample PreparationO

OH3CO

OCH3

OCH3

O

O

H3CO

H3CO

OCH3

O

OH3CO

H3CO

OCH3

O

OOCH3

H3CO

H3CO

O

O

OCH3

OCH3

H3CO

O

OOCH3

OCH3

H3CO

2

O

OD3CO

OCD3

OCD3

O

O

D3CO

D3CO

OCD3

O

OD3CO

D3CO

OCD3

O

OOCD3

D3CO

D3CO

O

O

OCD3

OCD3

D3CO

O

OOCD3

OCD3D3CO

2

O

OC2H5O

OC2H5

OC2H5

O

O

C2H5O

OC2H5

O

O

OC2H5

O

OC2H5O

C2H5O

O

O

OC2H5

OC2H5

C2H5O

O

O

OC2H5

OC2H5C2H5O

2

OC2H5

C2H5O

C2H5O

C2H5O

2,3,6-O-Me-β-CD 2,3,6-O-Med3-β-CD 2,3,6-O-Et-β-CD

Reductive amination

Mixture of oligosaccharides (DP1-DP7) are obtainted

from each sample

Mixture of oligosaccharides (DP1-DP7) after

reductive amination

LC/UV

Partial hydrolysis

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8 May 2014 | Sheetal Gangula |TU Braunschweig

Types of mixtures

Three types of mixtures are presented :

Methylated and deuteromethylated oligomeric mixtures (Me,Med3)

Methylated and ethylated oligomeric mixtures (Me,Et)

Deuteromethylated and ethylated oligomeric mixtures (Med3,Et)

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8 May 2014 | Sheetal Gangula |TU Braunschweig

ESI-IT-MS (electrospary ionization mass spectrometry)

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8 May 2014 | Sheetal Gangula |TU Braunschweig

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,Med3 mixture Me,Et mixture Med3,Et mixture

Results from ESI-IT-MS

RI0

1

2

3

4

5

6

1.09 0,920.96000000000

0001

Relative ion intensity Med3/Me

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP3 DP4DP2

RI0

1

2

3

4

5

6

3.37

2.68 2.69

Relative ion intensity Et/Me

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP3 DP4DP2 1 2 30

1

2

3

4

5

6

3.25

2.53 2.48

Relative ion intensity Et/Med3

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP2 DP3 DP4

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8 May 2014 | Sheetal Gangula |TU Braunschweig

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)

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2,5-dihydroxybenzoic acid

O

OH

HO

OH

2-hydroxy-5-methoxybenzoic acid

O

OH

O

OHMolecular Weight: 154,12

Molecular Weight: 168,15

sDHB

8 May 2014 | Sheetal Gangula |TU Braunschweig

Selection of laser power

Me,Et

273.3

381.2

467.2680.5

857.4

893.7

1061.6

1106.91320.0

1533.2

0_N2\1: +MS

273.3

381.2

467.2

680.4

857.4

893.6

1061.5

1106.8

1320.0

1533.2

0_N2\1: +MS

273.3

381.2

467.2

680.4

857.4

893.7

1061.6

1106.9

1320.1

1533.2

0_N2\1: +MS

0

1000

2000

3000

Intens.

0

1

2

3

4

5

4x10

0.0

0.2

0.4

0.6

0.8

1.0

5x10

200 400 600 800 1000 1200 1400 m/z

Me,Med3

LP 40 %

LP 45 %

LP 50 %

287.3

375.0449.3

533.4

653.5

779.7

857.7

1025.91272.1

1518.4 1746.6

0_D4\1: +MS

287.3

449.2

533.4

653.5

779.7

857.6

1025.9

1272.11470.1

1746.6

0_D4\1: +MS

287.3

449.2

533.4 653.4

779.6 857.6 1025.9

1272.2

1470.2

1765.7

0_D4\1: +MS

0

1

2

3

4

4x10Intens.

0.0

0.2

0.4

0.6

0.8

5x10

0.0

0.2

0.4

0.6

0.8

5x10

200 400 600 800 1000 1200 1400 1600 1800 m/z

LP 50 %

LP 55 %

LP 60 %

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8 May 2014 | Sheetal Gangula |TU Braunschweig

Results from MALDI-ToF-MS

RI0

1

2

3

4

5

6

1.03 0.99 0.99

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP3 DP4DP2RI

0.00

1.00

2.00

3.00

4.00

5.00

6.00

1.591.96

1.64

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP3 DP4DP2 RI0

1

2

3

4

5

6

1.862.29

1.91

Ave

rag

e re

lati

ve i

on

in

ten

sity

DP3 DP4DP2

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,Med3 mixture Me,Et mixture Med3,Et mixture

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8 May 2014 | Sheetal Gangula |TU Braunschweig

SUMMARY

Under optimal conditions for both instruments...

Me and Med3 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 Med3-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.

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8 May 2014 | Sheetal Gangula |TU Braunschweig

Thank you for your kind attention

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