 Good morning! Welcome back! Okay, so good morning, good afternoon, good evening, whatever time you're watching this video discussion. Good morning to you. I hope you're having a great day. So again, welcome back. This is Jomar Adams and I thank you for tuning in again with me for another video and another lesson that we will be discussing and I'm very excited again to share to you what we have in store for today. So I hope you'll be able to enjoy learning today. So let's dig into our discussion. So what we're going to talk about is all about your fluorometry. Yes, this is a part of the instrumentation in the clinical chemistry that I am discussing. So let's go to your fluorometry. So before we actually have finished discussing about your spectrophotometry, your atomic absorption spectrophotometry. So if you haven't watched that, please go to the video. I'll also be providing you the link down below. But for this morning, for today rather, we will be discussing your fluorometry. So to give you a wrap up of what we're going to do and discuss for today, we'll be talking about luminescence. So I opted to discuss luminescence because it is foundational for you to know that before we jump into fluorometry. And eventually when we go to your fluorometry, we'll be discussing the principle behind it. And of course, the different components alongside with it. And of course, after that, I'll be sharing to you what is quenching phenomenon. All right. Are you excited? So let's dig in. So first is luminescence. When we talk about luminescence, what does it mean? So luminescence is based on an energy exchange process that occurs when a certain compound absorbs an electromagnetic radiation. So the moment that particular substance or compound absorb that particular electromagnetic radiation, it will now be from being on its ground state, it will now become excited. And as it actually returns to its original level or as it returned to its ground state, it will be returning to an energy level lower than or equal to their original level. And as it undergoes that process, it will actually be emitting a light or it will also be emitting an energy as we pertain now as your luminescence. So when we talk about luminescence, we actually have three types of luminescence, which are very important for us to know. The first one is actually your fluorescence, which is actually the main thing that you will be discussing in your fluorometry. Aside from that, we have your phosphorescence and we also have your chemiluminescence. So for the first two, your fluorescence and your phosphorescence, both of them are actually aided by the principle of photoluminescence. They are photoluminescence. So what do we mean by fluorescence now? So again, it is a type of photoluminescence whereby the emission is basically an immediate and therefore generally only visible for a short period of time for as long as the light source is continuously on. So what do we mean by that is that when a molecule is exposed to a particular light source, a particular wavelength of light, it will undergo, it will actually be exciting the molecule and at the same time, you will actually be observing the fluorescence immediately. You'll actually be observing the emission of light in a short period of time compared to your phosphorescence, which is also a type of photoluminescence. Although this is now characterized when materials can actually store absorbed light energy for quite some time and release it later in time. So compared to your fluorescence and phosphorescence, both of them are photoluminescence. It's just that your fluorescence, when it absorbs the light, it absorbs the light and immediately in a short period of time, it will also be emitting that light in a short period of time. Unlike your phosphorescence, it can actually store up your light and eventually it can actually gradually or even slowly release the light that it actually absorbed. That is for your fluorescence and your phosphorescence. On the other hand, we have your chemiluminescence. A while back, we were talking that when a material is a fluoropore or can undergo phosphorescence, they should be exposed to light first. So we need to be having a particular light source that will excite them. Unlike your chemiluminescence, your chemiluminescence is the emission of light created when there is a particular chemical or electrochemical reaction that took place in the reaction medium. So it does not absorb any electromagnetic energy. The only thing here that for as long as there is a reaction, be it oxidation, reduction, or hydrolysis or whatsoever chemical process that may be or chemical reaction that is, it will produce your light. Phosphorescence, fluorescence, and chemiluminescence. And our main topic for today is all about your fluorescence and that is your fluorometry. So what is fluorometry all about? So fluorometry in your according to your henries, according to macpherson in pinkos, it is also known as your molecular luminescence spectroscopy. So if you hear me say molecular luminescence spectroscopy or your fluorometry, it's one and the same. Are we clear? It's one so what is fluorometry all about? So fluorometry measures the amount of light emitted by a particular molecule, okay your fluorophores, we call them fluorophores. So a material that can fluoresce is called fluorophores. And aside from that, it measures again the amount of light emitted by a molecule after excitation by electromagnetic radiation, meaning there is a light source in it. So it is 1000 times more sensitive than your spectrophotometry. Okay, comparing now to your spectrophotometry, it actually is more sensitive. But when we are talking about which one is the most widely used, it's your spectrophotometry. Aside from that, it is very specific. It's more specific than your spectrophotometry. Take for example, you have two compounds in a solution. Both of them can actually absorb the same light. Take for example, we are in 340 nanometer, so that is an ultraviolet light. So two compounds, take for example, you have compound A, you have compound B, you have compound A and compound B, both are absorbing the UV light 340 nanometer. But you can actually discriminate one from the other when they start to fluoresce because they will be fluorescing in terms of the light that they will be fluorescing, it will be different. So one would be this one would be A, one would be B, and the system can easily discriminate that. You can easily identify which is which. And in a nutshell, in its most simplest term, fluorometry is the measurement of the fluorescence. How much does a material or a molecule is fluorescing? So that is about fluorometry. And we are talking about fluorometry. This is actually an example. So I took this in an article online. So you can actually see how a material fluoresce. They can actually fluoresce for as long as they are also is exposed to a particular light source. So this is a basic setup of your fluorometry or your fluorometer. On your left, you can actually see this in your Henrys. So you have your light source here. You have your attenuator here. You have your primary filter, your secondary filter. Those are actually monochromators as well. And you have here your sampled holder. You have your photo multiplier tube and your readout device. Okay, let's dig in. So on the other side, you also have your on our on your right, you also can see the the fluorometry set up from coming from the shop. Okay, coming from the shop. And let's move on. Okay, so we have seven. Some of these are actually very much familiar to you already because we discussed this in your spectrophotometer and even your atomic absorption spectrophotometry. So we have your light source. We have your attenuator. We have your primary or your excitation monochromator. You have your qubit. We have your secondary or your emission monochromator. You have your photo detector. And you also have now your readout device. Okay, let's move on. We have your light source. So your light source, it is now a source of light. Obviously, yes, what I want to be specific, this is a source of light that is short, short wave, short wavelength, but high energy light. So the reason why it is a short way, a shorter wave, but of higher energy, because your goal again is to excite the molecule that is your goal. And usually we have your light source like your mercury arc clamp or your xenon arc clamp. So that is the most common light source that we use. Again, thing that I want you to remember here is that the light that we are, the light that is being emitted by your light source is the short wave with high energy. Short wave with high energy because this would be very important when we discuss now your fluorescence eventually. So we also have your attenuator. So the attenuator intensifies the light coming from your light source. So it further intensify the light. It further intensify your light. So it is very important. It is very important. So later on, I'll try to differentiate your spectrophotometry, your spectrophotometer from your atomic absorption spectrophotometry in your fluorometer. So I'm actually thinking of doing a review video regarding those things. And if you are interested with that, just comment it down below so that I know that you would want that content so I can shoot it immediately. So we have your light source going back. We have your light source. We have your attenuator. On the other hand, this is very important. I am actually discussing it in order as the system goes. But we have here your primary. In some textbook, in some textbook, you can actually see this as the primary filter. But in Henry's, it actually was called as your excitation monochromator. Okay, your excitation monochromator selects the wavelength that will best be absorbed by the solution that is being measured. So whether that is UV, whether that is invisible region or in the infrared region, your primary monochromator or your primary filter aims one thing that is your excitation monochromator. Okay, that is your excitation monochromator. So after that, the light will now pass through your attenuator going to your primary excitation monochromator and it will now reach your cuvette. Your cuvette or your sample cell holds the solution or the specimen that is being measured. So usually, similarly to spectrophotometry, our cuvette is actually more of a square shape rather than round. So again, going back to the band pass, the plant that needs to be traveled by your light. But going back to your cuvette, your cuvette contains again your sample cell. So after passing through the primary or your excitation monochromator, it will now reach your sample. Comes after that is your secondary emission or your secondary filter or your emission monochromator. And I want us to be specific because it is placed on a right angle. So unlike most of this instrumentation that we discussed, most are actually linear from the light source, from the entrance lead monochromator and then your exit lead to your cuvette going to your photo detector and then eventually to your readout device for your spectrophotometer that is just linear. You also have your atomic absorption where you have your light source, your beamchaper and then eventually your nebulizer, your atomizer, your flame or your graphite furnace. And then comes after that is your monochromator and then your photo detector and eventually leading to your readout device. So unlike other instrumentation, your fluorometer and your fluorometry, the secondary filter or the emission monochromator is always placed on the right angle of the cuvette to avoid incidence light come from reaching your detector. So what you want to only detect is the fluorescence of your compound and not the one coming from your light source. So what you do is to place your monochromator on a right angle, on a 90 degree angle. So eventually the same thing your photo detector is also placed there so it converts now the light energy to its equivalent electrical energy. It detects the fluorescence light in short. So your photo multiplier tube is again the most commonly used photo detector. Again, why? Why photo multiplier tube because it is sensitive and it can actually detect quick burst of light and low intensity light. So that is for your photo detector and of course you also have your readout device that displays your measurement either by an LED or a galvanometer or an amperometer. So so much about that we go to your quenching phenomenon which is the last part of this discussion. What is quenching phenomenon all about? So this phenomenon happens when the excited molecule actually interacts with other compounds within the system and what happen is that the molecule loses some of its energy because of interacting with other components on the reaction system. So it's more likely to have an analogy. Ganito siya, na sayo na kayo na pero nung nag-interact ka sa maraming, kung na re, di ba, ikaw at yung jawa mo, akalam mo kayo na, nakipag-interact ka sa maraming tao at dun mo na realize na unti-unti siyang nawawala sa yung hanggang tolu yan na ang nawawala dahil inagaw na ng iba na nandun din sa iisang lugar. So technically that is quenching phenomenon when your molecule loses its energy because other compounds within the system took that energy when interacting with that molecule. So that is quenching phenomenon o di ba pati dito may hugot pa rin tayo. So so much about that we were able to discuss fluorometry and that is very quick. We have your luminescence. We have the different types of luminescence. We have your fluorescence, your phosphorescence, and your chemiluminescence. We also have your fluorometry, the basic components from the light source to your attenuator to your primary or your excitation monochromator to your sample cell or your quvet to your secondary filter or your emission monochromator to your photo detectors specifically your photo multiplier tube and then your readout device and of course nakahugot din tayo when discussing your quenching phenomenon. So i guess that would be all for today. So thank you so much again i'd leave you with a quote from zigzaglar. It is your attitude more than your aptitude that will determine your altitude and at the end of the day attitude over aptitude because that will determine your altitude. Thank you so much for keeping up with me. This has been Jomar Adams again thank you if you have any questions you can find me on my youtube channel just comment down below or email sandbian email and thank you so much this has been Jomar Adams please do like this video if you learned something share this video and please do not do not forget to subscribe to my channel to be updated for the latest happening and uploads that i will be making. Thank you so much and have a great day.