Non-Traditional Approaches to MALDI- TOF Mass Spectrometry Analysis of Low Molecular Weight Polymers
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1 Non-Traditional Approaches to MALDI- TOF Mass Spectrometry Analysis of Low Molecular Weight Polymers Justin R. Engle and S. Kim R. Williams Laboratory for Advance Separation Technologies Department of Chemistry and Geochemistry Colorado School of Mines Golden, Colorado
2 MALDI-TOF sample preparation Analyte Matrix Polystyrene (PS) 8 n H M w 826 M n HO OH O OH 2,5-dihydroxybenzoic acid (DHB) + Salt Polyethylene glycol (PEG) 7 H O n OH M w 7 M n 645 Deposition
3 MALDI-TOF schematic Perspetive Biosystem Voyager DE-STR with delay extraction option Applied Biosystems. Perspetive Biosystems Voyager DE-STR. Verison
4 Laser desorption/ionization process Matrix absorbs laser energy Multiple ionization processes at ablation site and in plume
5 Traditional sample preparation Matrix 1 mg/ml Analyte 5 mg/ml Salt 5 mg/ml PS-Ag + ; PEG-Na + Mixed in 1:1:1 (v/v/v).5 μl spotted on plate Instrument settings Acceleration voltage: 25, volts Delay extraction time: 1 ns Grid voltage: 96% Laser intensity: 1926 units
6 DHB + PS 8 + AgTFA Stainless Steel MALDI Plate n=7 PS 8 + Ag + end groups: t-butyl and H + PS 8 + K + end groups: t-butyl and OH Gold MALDI Plate PS 8 + Ag + (n = 4,5 ) end groups: t-butyl and H PS 8 + Au + (n = 3,4 ) end groups: t-butyl and H + PS 8 + Cu + (n= 4,5) end groups: t-butyl and H
7 DHB + PEG 7 + NaTFA 3 Stainless Steel MALDI Plate PEG 7 + Na + end groups: O H - + H + n=14 44 m /z PEG 7 + K + end groups: O H - and H + PEG 7 + H + end groups: O H - and H + PEG 7 + Li + end groups: O H - and H + PEG 7 + Ag + end groups: O H - and H m /z Gold MALDI Plate 44 m /z n=16 PEG 7 + Na + end groups: OH - and H + PEG 7 + Li + end groups: OH - and H + PEG 7 + Ag + end groups: OH - and H
8 Traditional approach results Messy mass spectra Multiple unexplainable mass peaks Result from fragmentation of organic matrix Incomplete mass series for PS PS on Stainless Steel PS on Gold PEG on Stainless Steel PEG on Gold Calc M w 945 ± ± ± ± 35 Calc M n 862 ± ± ± ± 28
9 Objectives Eliminate use of organic matrix Use of metal substrates for desorption/ionization Provide good cleaning process for limited mass peaks Determine reproducibility using metal substrates Obtain accurate molecular weights
10 Non-traditional laser desorption/ionization Metal substrate and/or cationizing salt absorbs laser energy Fewer particle interactions
11 Non-Traditional sample preparation Metal substrates: Commercial grade aluminum, copper, brass, stainless steel MALDI plate, gold MALDI plate Non-traditional approach Analyte 5 mg/ml Salt 5 mg/ml PS-Ag + ; PEG-Na + Mixed in 1:1 (v/v).5 μl spot volumes Instrument settings Acceleration voltage: 25, volts Delay extraction time:1 ns Grid voltage: 96% Laser intensity: 1926 units
12 Substrate Cleaning Washings in order of tetrahydrofuran, 6M nitric acid, DI water, and acetone 7 Dirty copper substrate 5 Clean copper substrate Relative Intensity units
13 Metal substrate mass spectra Copper substrate 375 Brass substrate Stainless steel MALDI plate Aluminum substrate Gold MALDI plate
14 Aluminum substrate 7 Ag+ Ag + 2 Polystyrene Ag + 3 n H 14 n=8 PS 8 + Ag + end groups: t-butyl and H Polyethylene glycol H O n OH 44 n=13 PEG 7 + Na + end groups: OH - and H + PEG 7 + K + end groups: OH - and H + PEG 7 + Li + end groups: OH - and H
15 Copper substrate Polystyrene 6 Ag+ n H PS 8 + Cu + end groups: t-butyl and H + Ag + 2 Ag + 3 n=7 14 PS 8 + Ag + end groups: t-butyl and H Polyethylene glycol H O n OH 44 m /z n = 1 2 fo r C u n = 1 3 fo r N a PEG 7 + Cu + end groups: O H - and H + PEG 7 + Na + end groups: O H - and H + PEG 7 + Li + end groups: O H - and H m /z
16 Reproducibility: Al substrate m /z
17 Calculated Molecular Weights PS M w = 826 M n = 726 Aluminum Brass Copper SS MALDI plate Gold MALDI plate M w 936 ± ± ± ± ± 29 M n 831 ± ± ± ± ± 19 PEG M w = 7 M n = 645 Aluminum Brass Copper SS MALDI plate Gold MALDI plate M w 66 ± ± ± ± 19 N/A M n 564 ± ± ± ± 22 N/A
18 Conclusions Successful elimination of organic matrix for desorption/ionization of low molecular weight polymers Metal substrates provided means of desorption/ionization Good cleaning procedure Baseline for MALDI plates Mass peaks below 125 Da for commercial grade metal substrates Good reproducibility seen in mass spectra Calculated molecular weights different when using different metal substrates Possible selective ionization of polymers
19 Acknowledgments NSF-CHE Williams Group Dr. Dean Lee Dr. Ilyong Park Dr. Claudia Lohmann J. Ray Runyon Adam Goering Ed Dempsey (CSM)
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