In this video, we're going to talk about mass spectrometry. Most of you are already somewhat familiar with mass spectrometry from your previous organic chemistry courses. In this video, we're going to review what mass spectrometry is and how it typically operates. Mass spectrometry, or MS, is a technique that can be applied to a wide variety of molecules, including proteins. Mass spectrometry ionizes, quantifies, and separates molecules based on their mass-to-charge ratios, or their m/z ratios for short. The m/z ratio is a unique property that can be used to identify a molecule and to get structural and chemical information about that molecule. Since the z here in the m/z ratio is the charge, and it's almost always equal to just 1 unit, that means that the m/z ratio is often considered to just be the mass of the molecule, as indicated by the word 'mass' in mass spectrometry. The rest of these bullet points here follow are dedicated to explaining how a typical mass spectrometer operates. Note that it's broken down into 3 general steps, numbered 1, 2, 3. Look at the example below, we have an image of a typical mass spectrometer, and the numbers 1, 2, and 3 in the image correspond with the numbers in the text above.
For our first step in how a typical mass spectrometer works, we have an already purified peptide that first needs to be converted ionized in a vacuum. The ionization occurs via controlled bombardment with electrons or a noble gas such as helium. The ionization typically leads to random fragmentation or breakdown of the peptide molecule. Usually, the bonds being broken are the peptide bonds that fragment the molecule. Later in our course, we will discuss more details about how a peptide is fragmented during mass spectrometry. For now, let's consider our example below. Notice on the far left, we have an already purified peptide that is relatively small and is being entered into a vacuum where it's going to be converted into a gas and ionized with the use of electrons, which are shown as these blue dots. After the first step is complete, we've generated all these orange dots, representing fragmented peptides in gas ion form.
Moving on to step number 2, these ionized gas peptide fragments are going to be exposed to an electric field or a magnetic field. The electric field essentially deflects the ionized gas fragments. The paths these ionized gas fragments end up taking is a direct result of their m/z ratio. Fragments that have a smaller m/z ratio are going to be deflected a lot more than fragments that have a larger m/z ratio because fragments with a larger m/z ratio have a larger mass and it requires more force to deflect them.
Quickly moving on to step number 3, there's a detector that's going to measure the fragments. Let's clear up steps 2 and 3. The fragmented gas peptide ions are directed into a mass filter where there's an electric or magnetic field. The paths that these ions take result from their deflection related to their m/z ratio. Molecules with a small m/z ratio are deflected a lot more and hit the detector at a different point than fragments with a larger m/z ratio, which take more to deflect. You can see that the detector is able to measure the abundance and the m/z ratio for each ionized gas fragment that hits the detector. The detector then translates this information into a data plot known as a mass spectrum, which we'll discuss in more detail in our next lesson video.
Note that on the y-axis we have the relative abundance of each peak, and on the x-axis, the m/z ratio of each peak. Note that the three ionized gas peptide fragments we showed in our image to the left, which have m/z ratios of 290, 430, and 570, are pointed out directly. Even though in our image we only showed three peptide fragments, in a typical mass spectrometry of a peptide, there will be many more peptide fragments shown. One of the biggest differences to note here is the m/z ratio axis and the scaling of that axis. Typically, in your previous organic chemistry courses, when you covered mass spectrometry, you were looking at relatively small molecules, where a single peak might actually represent a single functional group with a small number of atoms. Here, with mass spectrometry of an entire peptide, each peak represents an entire peptide fragment that contains many atoms, resulting in a larger mass. That's why the m/z ratio has a larger scaling than what you're probably used to, ranging from 200 up to 1400 units for the m/z ratio. We're going to talk a lot more about mass spectrums in our next lesson video. For now, this concludes our review of mass spectrometry, and we'll get a bit of practice in our next video. See you there.
