Analysis of Complex Molecules and Their Reactions on Surfaces by Means of Cluster-Induced Desorption/Ionization Mass Spectrometry

Desorption ionization induced by neutral SO2 clusters which we call in short DINeC, is a soft and efficient desorption ionization method. We use it among others for mass spectrometry of biomolecules, such as peptides and proteins. As DINeC, desorption molecules intact from the sample surfaces, subtle chemical changes in these molecules are easily detected in the mass spectra.

The process is illustrated in the following simulation. The incoming clusters shatters through in-cluster surface impact. The surface isotope dipeptide is dissolved in one of the cluster fragments and desorpted by this process.

In the experiment, the beam of SO2 clusters is produced via supersonic expansion from a pulsed nozzle. The molecules that are dissolved and ionized during cluster surface impact are transferred into the ion trap for mass spectrometry. Typical mass spectra only show peaks at the MO set values of the intact molecule.

Demonstrating the procedure will be Karolin Bomhardt, and Pascal Schneider, two PhD students in my laboratory. For standard samples, cut the substrates from silicon wafers with thickness of approximately 0.5 to one millimeters in pieces of one by one square centimeter. Clean the silicon substrates in an ultrasound bath of ethanol and acetone for 15 minutes each.

Dry the substrates in a stream of dry nitrogen gas. Then, drop cast five to 30 microliters of the sample solution on the substrate. Depending on vapor pressure of the solvent, let the sample dry under ambient conditions or in a desiccator until all solvent has been evaporated and a dry film has been formed.

Consider the simplest preparation scheme. As an example, for the investigation of highlighter ink, just draw a dot on the substrate surface. Mount the samples on the sample holder with sticky tape or clamps tightened by screws.

Additionally mount a reference sample such as a micrometer thick film of Angiotensin II on the sample holder. Then, press Vent load lock to vent the load lock system. Wait for five minutes, or until atmospheric pressure has been reached.

Open the load lock and mount the sample holder. Close the load lock and pump down the load lock chamber to a pressure below two times 10 to the negative five millibar. Open the valve to the DINeC chamber and transfer the sample holder with the transfer rod to the main manipulator.

Attach the sample holder to the manipulator. Retract the transfer rod and close the valve between the load lock and the DINeC chamber. Open the valve between the gas cylinder and the nozzle.

Adjust the pressure of the sulfur dioxide and helium gas mixture at the outline of the gas mixing system to 15 bar. Set the position of the manipulator to the reference sample's position. For measuring cationic mass spectra, set the sample and grid bias to plus 40 volts, and plus seven volts respectively.

To drive the pulsed nozzle and the ion trap mass spectrometer, set the external function generator to two Hertz. With the delay generator, set the time delay between the clear trap signal from the ion trap and the trigger signal for the pulsed nozzle to five milliseconds. In the Bruker control software, in the Mode page of the main dialogue window, select Enhanced Resolution for Scan mode.

Type in M over Z 50 to 3000 for range, type in 0.1 milliseconds for accumulation time, type in 10 cycles for average, and for polarity, select positive for the measurement of cationic mass spectra and negative for the measurement of anionic spectra. Once a pressure below three times 10 to the negative six millibar has been reached in the DINeC chamber, start the measurement. Set the M over Z value to the respective mass in order to follow the time dependent chromatogram signal.

Press Operate in the control software. Start recording the measurements by pressing the Record button. Measure test spectrum from a reference sample, such as Angiotensin II for about 300 seconds.

Optimize signal intensity by adjustment of the time delay between the clear trap signal and the signal triggering the pulsed nozzle. With the time delay optimized, move the manipulator to the position of the sample to be measured. Click on the Record button to acquire mass spectra over the time span of interest.

After measurement is finished, load the respective data set in the Data Analysis program as shown here for the reference measurement. Select the time span of interest in the chromatogram with the right button of the mouse. The average spectrum is displayed in a separate window.

Click on Export as ASCII file to export the spectrum as a data file for further processing in a program of choice. The DINeC mass spectrum from an Angiotensin II sample changed with time as the sample was heated to about 140 degrees Celsius. Once the temperature reached the final value, the peak of the attacked protinated molecule at M over Z equalling 1047 dropped and new peaks were observed.

The process is summarized in these three graphs. The additional peaks at M over Z equaling 932, 1012, and 1029 were well observed. Analysis of the reaction kinetics demonstrates that the entity with M over Z equaling 1029 is an intermediate which was further decomposed into smaller fragments as the intensity first increased and then decreased.

The most important features of DINeC are the soft nature of the desorption process, the efficiency, and the high surface sensitivity. They all come together with the low requirements with respect to sample preparation. DINeC can thus be applied to a wide variety of different samples ranging from organic bark materials to complex surface adsorbates in the sub-monolayer region.

In all cases, full chemical information can be obtained in real time.