Presentation - Isobaric PEG systems

Travelling Wave Ion Mobility Studies of Polymer

Microstructure

Jim Scrivens

Challenges in characterising polymer

formulations• Extremely complex mixtures• Variation of starting materials• Poorly controlled reactions• Molecular weight range• Sold on properties not structure• Chromatographic separation

difficult

Requirement

• Rapid analysis• High information content• Molecular weight and structural

information• Ability to differentiate small

differences in complex formulations

Ion mobility platforms

• Drift cell– Currently predominately academic based

• Differential mobility spectroscopy (DMS)– Includes FAIMS– Theory challenging

• Travelling wave– Commercially available– Theory challenging

Ion mobility issues

• Sensitivity• Speed • Selectivity• Ease of use• Resolution• Availability

• Information content

• Reproducibility• Calibration• Cost• Data analysis

References

• Ion mobility–mass spectrometry– Abu B. Kanu, Prabha Dwivedi, Maggie Tam, Laura Matz and Herbert H.

Hill Jr.– J. Mass Spectrom. 2008; 43: 1–22

• Differential Ion Mobility Spectrometry: Nonlinear Ion Transport And Fundamentals Of FAIMS– Alexandre A Shvartsburg– CRC Press, ISBN: 9781420051063, 2008

Travelling Wave References

• An investigation of the mobility separation of some peptide and protein ions using a new hybrid quadrupole/travelling wave IMS/oa-ToF instrument– Steven D. Pringle , Kevin Giles , Jason L. Wildgoose , Jonathan P.

Williams , Susan E. Slade , Konstantinos Thalassinos , Robert H. Bateman , Michael T. Bowers and James H. Scrivens

– International Journal of Mass Spectrometry, 261, 1-12, 2007

• Applications of Travelling Wave Ion Mobility-Mass Spectrometry – Konstantinos Thalassinos and James H Scrivens– Practical Aspects of Trapped Ion Mass Spectrometry Volume 5, 2009

• Special issue of IJMS on Ion Mobility– Edited by Richard Yost, James Scrivens– IJMS, 2010

Pringle, S. D. et al., International Journal of Mass Spectrometry, 261, 1-12, 2007Thalassinos K and Scrivens J H, “Applications of Travelling Wave Ion Mobility-Mass Spectrometry”, Practical Aspects of Trapped Ion Mass Spectrometry Volume 5

Schematic of Synapt G1

Features of Synapt

• Ease of use

• Rapid analysis (typically 200 spectra in 18ms)

• High sensitivity (fmole)

• Can acquire MS, MS/MS with accurate mass data

• Estimated relative cross-sections can be obtained by use of calibration against known standards

Aspirations

• Higher mobility resolution

• Better dynamic range

• Higher resolution mass spectrometry

• No compromise in: -– Sensitivity– Speed– Ease of use

Schematic of Synapt G2

TOF developments

QuanTof improvements— High field pusher — Dual stage reflectron — Hybrid ion detection system — compatible with UPLC separations— compatible with HDMS analysis

Performance— Resolution – over 40,000 FWHM— Mass Measurement – 1ppm RMS— Dynamic Range – up to 105

— Speed - 20 Spectra/sec

Mobility Cell improvements

Second generation Triwave device— Increased ion mobility

resolution (over 40 Ω/ΔΩ) IMS cell 40% longer Higher gas pressure in IMS T-

Wave (2.5mb versus 0.5mb) Modified T-Wave pattern - use of

Higher T-Wave pulse amplitudes/fields

Helium cell balances N2

pressure in Maximizes transmission of ions on entry into the mobility cell

Rabbit haemoglobin peptide Synapt G1

20 40 60 80 100 120 140 160 180 200Scan

99

84

115

81

76 m/z 977

m/z 857

m/z 1037

m/z 1134

Rabbit haemoglobin peptide Synapt G2

20 40 60 80 100 120 140 160 180 200Scan

102

89

110

86

84 m/z 977

m/z 857

m/z 1037

m/z 1134

Rabbit haemoglobin peptide ATD comparison

20 40 60 80 100 120 140 160 180 200Scan

99

84

81

76

102

89

86

84

Synapt G2

m/z 977

m/z 857

m/z 1037

m/z 1134

Positive ion [M+Na]+ ESI mass spectrum of N-glycans released from chicken

ovalbumin

Ion mobility separations of positive ions [M+Na]+ of N-glycans released from chicken ovalbumin with

compositions of Hex3GlcNAc2 Hex3GlcNAc3 (two isomers) and Hex3GlcNAc4

Ion mobility separations of positive ions [M+Na]+ of N-glycans released from chicken ovalbumin with compositions of Hex3GlcNAc2 Hex3GlcNAc3 (two

isomers) and Hex3GlcNAc4

Positive ion [M+Na]+ ion mobility MS/MS spectra of the first and second N-glycan isomers of m/z 1136

from chicken ovalbumin

EESI of aerosol formulations

Sample

Sample headspace

Solvent

Desolvation Gas plus analyte

Sample molecules

Charged solvent droplets

Desolvation Gas plus analyte

Sample Container

Mass Spectrometer

Carbomethoxypyridines

N

O

CH3

O

N

O

CH3

O

N

O

CH3

O

Mobility separation of isomers

Methyl Picolinate 1/200 dilution 138 MS/MS transfer, 2000WV

Scan10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200

%

0

100

10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200

%

0

100

10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200

%

0

100

WAR112409_09_dt_01 Sm (SG, 2x3) TOF MSMS ES+ 137.6_138.5

1.70e4100

WAR112409_14_dt_01 Sm (SG, 2x3) TOF MSMS ES+ 137.9_138.2

2.21e499

WAR112409_11_dt_01 Sm (SG, 2x3) TOF MSMS ES+ 138_138.1

2.27e493

Para

Meta

Ortho

ATD for isomers

OrthoPara

Meta

Isobaric PEG systems

• Oligomers of di-hydroxyl end-capped PEG & PEG monooleate have same nominal mass-to-charge ratio – Different number of moles of ethylene oxide (EO)

• Resolution required to separate oligomers is ~6300

• Difference in m/z for two oligomers is 0.0880

– m/z 553.3411

– m/z 553.4292

12

O CH2CH2 OHH

6

CH3 (CH2)6 CH2 CH CH CH2 (CH2)5 CH2 C O CH2CH2 O H

O

Synapt G1 mobility separation – m/z 553

20 40 60 80 100 120 140

Scan

100 200 300 400 500m/z

100 200 300 400 500m/z

12

O CH2CH2 OHH6

CH3 (CH2)6 CH2 CH CH CH2 (CH2)5 CH2 C O CH2CH2 O H

O

[M+Li]+

[M+Li]+

Hilton G. R., et al,. Anal. Chem., 2008, 80 (24), 9720-9725

Synapt G1 mobility separation – m/z 861

50 60 70 80 90 100110120130140150Scan

100 200 300 400 500 600 700 800 900m/z 100 200 300 400 500 600 700 800 900m/z

19O CH2CH2 OHH

13CH3 (CH2)6 CH2 CH CH CH2 (CH2)5 CH2 C O CH2CH2 O H

O

[M+Li]+

[M+Li]+

Hilton G. R., et al,. Anal. Chem., 2008, 80 (24), 9720-9725

Synapt G2: Ion mobility separation – m/z 1126

Scan120 130 140 150 160 170 180 190 200

m/z200 400 600 800 1000

18O CH2CH2 OHH

25

m/z200 400 600 800 1000

12CH3 (CH2)6 CH2 CH CHCH2 (CH2)5 CH2 C O CH2CH2 O H

O

19

[M+Li]+[M+Li]+

Precursor ion resolution8434

Driftscope separation G2

PEG 1000

PEG mono oleate

Synthesis of Tween 20

[C2H4O]nO+

- H2O +

IsosorbideSorbitan

- H2O

Sorbitol

Structures of Tween formulations

Formulation Structure Indicated purity

Tween 20 Polyoxyethylene (20) sorbitan monolaurate

50%

Tween 40 Polyoxyethylene (20) sorbitan monopalmatate

90%

Tween 60 Polyoxyethylene (20) sorbitan monostearate

50%

Tween 80 Polyoxyethylene (20) sorbitan monooleate

70%

Structures of major products

Sorbitan polyethoxylate [SPE]Isosorbide polyethoxylate [SPE]

Polysorbate monoester [PME]

O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OCR

H2CH2COHO

W

X

Y

Z

O

O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OH

H2CH2COHO

W

X

Y

Z

O

O

OCH2CH2

H2CH2CO

OH

HO

M

P

Tween 20 overall averaged spectrum

m/z200 400 600 800 1000 1200 1400

%

0

100617.4

595.4573.3

410.2

223.2

141.1

388.2

686.4

708.5

752.5

774.5

775.5819.5821.5

Major species Tween 20

Series 1

686.4 + n*22Li2 [2+]R = C11H23 [laurate]

686*2 = 13721372 – 14 [Li2] = 13581358 – 164 [sorbitan] = 11941194 – 182 [RCOOH – H2O] = 10121012/44 [CH2CH2O] = 23

W + X + Y + Z = 23


O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OCR

H2CH2COHO

W

X

Y

Z

O


Series 2

573.3 + n*22Li2 [2+]

573*2 = 11461146 – 14 [Li2] = 11321132 – 164 [sorbitan] = 968968/44 [CH2CH2O] = 22

W + X + Y + Z = 22

Sorbitan polyethoxylate [SPE]

O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OH

H2CH2COHO

W

X

Y

Z


Series 3

322 + n*22Li2 [2+]

322*2 = 644644 – 14 [Li2] = 630630 – 146 [isosorbide] = 484484/44 [CH2CH2O] = 11

P + M = 11

Isosorbide polyethoxylate [SPE]

O

O

OCH2CH2

H2CH2CO

OH

HO

M

P

Tween 20 mobility separationWAR112409_28A.raw : 1

Tween 20 mobility separation

WAR112409_28A.raw : 1

Tween 20 mobility separationWAR112409_28A.raw : 1

Tween 20 MALDI spectrum

Isosorbide polyethoxylate [SPE]Sorbitan polyethoxylate [SPE]


Folahan O Ayorinde et al Rapid Comm. Mass Spectrom, 14, 2116, (2000)

m/z400 600 800 1000 1200 1400

%

0

100692.5

670.4

595.4

537.4

485.3

410.2

736.5

758.5

759.0

781.0

803.0

875.6

921.6


Major series Tween 40

Series 1

670.4 + n*22Li2 [2+]R = C15H31 [palmitate]

670*2 = 13401340 – 14 [Li2] = 13261326 – 164 [sorbitan] = 11621162 – 238 [RCOOH – H2O] = 924924/44 [CH2CH2O] = 21

W + X + Y + Z = 21


O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OCR

H2CH2COHO

W

X

Y

Z

O


Series 2

573.3 + n*22Li2 [2+]

573*2 = 11461146 – 14 [Li2] = 11321132 – 164 [sorbitan] = 968968/44 [CH2CH2O] = 22

W + X + Y + Z = 22


O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OH

H2CH2COHO

W

X

Y

Z


Series 3

322 + n*22Li2 [2+]

322*2 = 644644 – 14 [Li2] = 630630 – 146 [isosorbide] = 484484/44 [CH2CH2O] = 11

P + M = 11


O

O

OCH2CH2

H2CH2CO

OH

HO

M

P

WAR112409_29.raw : 1


Tween 40 extracted regionsWAR112409_29.raw : 1

A

B

Tween 40 conformational families

m/z500 550 600 650 700 750 800

%

0

100

%

0

100692.5

670.4

669.9

714.5

758.5780.5

802.5 824.5

573.3551.3

529.3507.3

595.4

617.9

639.9

647.4

A

B



O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OCR

H2CH2COHO

W

X

Y

Z

O

O

OCH2CH2 OH

OCH2CH2

OCH2CH2 OH

OH

H2CH2COHO

W

X

Y

Z

Tween 40 extracted regions

WAR112409_29.raw : 1

a

b

c

m/z600 700 800 900 1000 1100 1200 1300

%

0

100

%

0

100

%

0

1001025.8981.7

893.7849.7

1069.8

1157.81201.9

831.6743.5

699.5

615.5

875.6

963.6 1007.71201.8

1051.6963.6919.5

901.5900.5

1095.61183.7

1271.71315.8

c

b

a

Tween 40 conformational families


Polyisosorbide monoester [PME]

m/z400 600 800 1000 1200 1400

%

0

100595.4

451.1

443.3

428.6

692.5728.5

736.5

758.5

773.0

802.5817.0


WAR112409_30.raw : 1



m/z400 500 600 700 800 900 1000 1100 1200 1300

%

0

100x2x2

525.1

575.4 771.5604.7619.7

815.5

860.1

925.6

Tween 80 mobility separationWAR112409_31.raw : 1

Conclusions

• ESI mobility-separated spectra offer an excellent screening approach for complex polymer formulations

• A number of, previously unseen, conformational series may be observed and extracted

• Mobility-separated MS/MS data can provide more detailed structural information

• The ESI spectra show greater agreement with published compositions than those obtained using MALDI

BMSP research group

Funding

http://www.defra.gov.uk/

http://www.food.gov.uk/

http://www.mrc.ac.uk/

Presentation - Isobaric PEG systems

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