Characterization of Synthetic Polymers via Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI-TOF) Mass Spectrometry

The overall goal of this MALDI-TOF MS tutorial is to teach users about sample preparation, acquisition, and analysis of polymers during characterization by MALDI-TOF MS.This method can help answer key questions in the polymer characterization field, such as polymer molecular weight, dispersity, and end group identity. The main advantage of this technique is that it allows for determination of homopolymer end groups. First, prepare a series of solutions by mixing previously prepared matrix, analyte, and cation solutions while varying the relative proportions of the components such that nine unique sample mixtures are made.

For this experiment, combine 15 microliters of poly(L-lactide)solution with 15 microliters of 2, 5-dihydroxybenzoic acid solution and one microliter of sodium trichloroacetate solution. Pipette one microliter of each solution mixture onto an individual sample well on a MALDI target plate. Add the samples incrementally in small portions to prevent the sample from flowing out of the sample well, allowing each aliquot to evaporate to dryness before adding additional sample.

To initiate data acquisition, first, open the data acquisition software, Flexcontrol. Eject the platform to enable the loading of the target plate by pressing the eject button. Gently place the target plate with the loaded calibrant and analyte samples on the platform in the appropriate orientation.

Next, use the acquisition software to inject the target plate on the platform by pressing the eject button again. Select an appropriate data acquisition method by pressing File and choosing Select Method. Before acquiring data, make sure an appropriate mass range for collecting data is selected by clicking on the Detection tab and viewing Mass Range.

From the acquisition software, select the position on the target plate which corresponds to the desired analyte. To initiate data collection, press Start and move the laser target around the sample to maximize the signal. Using the slide bar on the left side of the camera window adjust the laser power such that the minimum power necessary to achieve isotopic resolution is achieved.

Zooming in on an individual peak in the middle of the mass range of interest, optimize the resolution by adjusting the difference in acceleration voltages by varying the IS2 value in the Spectrometer tab. Finally, optimize the laser power by reducing it as low as possible while still generating a reasonable signal to noise ratio. Once the acquisition parameters are optimized, save the method by selecting File and then Save Method As.Make sure that any existing calibration is voided or in a position to be overwritten by pressing Invalidate Calibration under the Spectrometer tab.

Using the same acquisition parameters, move the laser over to the sample well containing the calibrant by selecting the corresponding well with the cursor and acquire a spectrum by pressing Start. Once a sufficient signal has been acquired, press Start to finish acquiring the data. Once a mass spectrum of the calibrant has been acquired, select the Mass Control List dropdown menu in the Calibration tab that corresponds to that calibration standard.

Before matching the corresponding reference peak to each selected calibrant peak, ensure that an appropriate peak picking protocol is being used by selecting the Processing tab. Apply the reference mass from the Mass Control List to the corresponding signal for the calibrant mass spectrum by selecting the area to the left of the peak of interest and then clicking on the corresponding mass in the control list to apply. Continue this process for the remaining calibrant peaks.

Reacquire the analyte spectrum once the mass scale for the optimized acquisition parameters have been calibrated. Open the Analyte Spectrum in the data analysis software. Zoom in on a peak to identify if isotopic resolution has been achieved by selecting the Zoom in X range button.

Now, press Mass List and choose Find to select peaks. If the monoisotopic peak is resolved, select this first peak in the isotopic distribution to determine its mass using a monoisotopic peak picking protocol. MALDI-TOF MS confirms the narrow distribution of poly(L-lactide)thiol terminated and analysis software was used to calculate the polymer characteristics shown here.

The observed mass for the 26-mer is 1973.62, which is minus 0.07 daltons different than the theoretical value. A smaller signal at 2045.74 is indicative of transesterification during the ring opening polymerization of lactic acid. A small peak at 2057.73 is likely the consequence of initiation from water during the ring opening polymerization of the lactide monomer.

MALDI-TOF MS confirms the narrow distribution of the poly(L-lactide)thiol terminated after a thiol-ene reaction with maleimide and analysis software was used to calculate the polymer characteristics shown here. The theoretical mass of the 26-mer of poly(L-lactide)thiol terminated corresponds to 2070.56891, which is 0.03 daltons different than the observed mass. The same species ionizing with potassium is also observed at 2086.49.

A small peak at 2167.58 is indicative of the same impurity from water initiation that was observed in the starting material. The same shift in mass observed for the thiol-ene reaction products does not occur, which indicates that this carboxylic acid terminated compound lacked the thiol end group to undergo the functionalization reaction. After watching this video, you should have a good understanding of how to optimize sample preparation and data acquisition parameters, as well as analyze data generated by MALDI-TOF mass spectrometry characterization of polymers.