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  • >> All right so listen today what I want to talk

  • about is something that's more conceptual.

  • We've been sort of talking nuts and bolts

  • and today practical stuff, interpretive stuff.

  • Today I want to talk a little bit more

  • about how some stuff works in NMR spectroscopy.

  • It's not going to be anything too fancy

  • and it gives you a little perspective on my take

  • and understanding of NMR experiments,

  • which is very different say

  • than Professor Shaka's [phonetic] take

  • or interpretation or Professor Martin's take or interpretation.

  • These are NMR development people and my group

  • and you will be users of NMR spectroscopy as a research tool,

  • but one of the things is you can't just take NMRs

  • as a black box.

  • As you're already seeing when you're going

  • down using the instrument you kind of have to keep your head

  • about you, oh, what am I doing, why am I locking it,

  • why am I shimmying it, what's the shimmy doing,

  • what does it do if I don't do this?

  • And what I would like to be doing is giving you a feel

  • for some parameters and things

  • that you may not be understanding in spectra.

  • So we've already, for example, had instances where people look

  • at their own DEPT spectra and say wait a second why do I see

  • in the DEPT 90 a little peak for say a methyl group

  • or a little peak for a CH2 group?

  • A lot of this has to do with parameters

  • and so that's what I want to give you a feeling about.

  • We're going to start with 1 experiment that's kind

  • of like a DEPT but a lot less fancy

  • and I think it's one that's easy to understand.

  • It's called the APT.

  • So it was a precursor.

  • That stands for attached proton test.

  • [ Writing on board ]

  • And the more NMR spectroscopic name for the technique

  • which actually is going to make a lot of sense to you

  • when we start to talk about how it works is called the J

  • modulated spin echo technique.

  • [ Writing on board ]

  • And then we'll talk a little bit about DEPT.

  • We won't go, and understand it as well,

  • but you'll see how the parameters that we talked

  • about for the APT relate to the DEPT experiment.

  • DEPT if you don't know is Distortionless Enhancement

  • Bipolarization Transfer.

  • [ Writing on board ]

  • From a practical point of view what the depth does

  • as you all know is allows you to identify the CHs, CH2s and CH3.

  • So in other words, the methines, the methylenes

  • and the methyl groups.

  • What the APT does is it's a less sophisticated experiment.

  • It gives you your Cs, your quats and CH2s and distinguishes them

  • so I'll say distinguishes let me maybe put it that way.

  • [ Writing on board ]

  • Quat and CH2 from CH from your methine and methyl.

  • So in a way, the DEBT experiment is more

  • of a Litmus test experiment.

  • So we're going to start to get a feel

  • for complex pulse sequences.

  • I'm going to introduce some concepts like the rotating frame

  • and the effect of various pulses.

  • We talked a little bit about this

  • when we introduced NMR spectroscopy

  • and we'll see a spin echo experiment.

  • So I just want to review some of the spin physics that we learned

  • when we started to talk about NMR spectroscopy.

  • So remember you start and you've got this difference

  • in population between the alpha and beta states and that leads

  • to a net magnetic vector along the X axis and remember

  • that vector is a bunch of vectors that are precessing

  • and we talked before about the concept

  • of applying a 90-degree pulse.

  • So remember 90-degree X pulse and you think

  • about the right hand rule

  • that means you have a quail along the X axis you're applying

  • essentially a 4 sign to the vector,

  • which is bringing the vector down into the X, Y plane.

  • That brings it along the Y axis.

  • Now remember what that means.

  • Having the net magnetization in the X, Y plane means

  • that we've now equalized the alpha and beta state populations

  • by having a differentiated population of alpha

  • and beta states you have net magnetization along the Z axis,

  • but when the alpha states are equal to the beta states,

  • then you have no net magnetization along the Z axis.

  • What you do have is magnetization on the X, Y plane

  • and because you've done this as a pulse all

  • of your vectors start out initially along the Y axis

  • and then remember you have your precession that leads to motion

  • in the X, Y plane of the vectors and that gets picked

  • up by the coil and gives you a cosine wave

  • that when you Fourier transform gives you a peak.

  • So the other pulse I'll give you an idea on is just remember

  • if we apply instead of a 90-degree pulse,

  • 180-degree pulse along the X axis,

  • now that flips the population of the alpha and beta states.

  • So, instead of dragging the magnetization down into the X,

  • Y plane you dragged it all the way along

  • to the negative Z axis.

  • I'm trying to represent things in 3 dimensions here.

  • So that brings our magnetization to the negative Z axis.

  • If you think about it, this means now

  • that you've inverted the population of alpha

  • and beta states so if we had more nuclei in the alpha state

  • than the beta state before then after more nuclei in the spin

  • down state in the, if you've had more nuclei in the alpha state

  • than the beta state, you flipped the population over here.

  • So, that's basically all the spin physics that we need to get

  • to a point to really think our way through an experiment

  • that ultimately is going to allow us

  • to distinguish our quats and our methylenes

  • from our methyls and methines.

  • So now I want to develop that idea.

  • [ Writing on board ]

  • So let's take this situation here and I'm just going

  • to project the X, Y plane into the plane of the blackboard

  • because it's going to help us to look down on the system rather

  • than for me to try to keep making these horrible 3

  • dimensional drawings.

  • So, okay, here we are in this situation

  • where we've got our X axis, we've got our Y axis

  • and now we're looking down the Z axis

  • and we have our magnetization along the Y axis.

  • Now, remember the magnetization along the Y axis is precessing

  • and so if you just think of this as a single line,

  • it's going to precess at an angular velocity omega.

  • [ Writing on board ]

  • Like if it's 500 megahertz you're going to have precession

  • at 500 million times per second in the X, Y plane.

  • That's this angular velocity if you're interested.

  • It's just my way of representing that it's moving.

  • So in other words as you wait, if we're precessing

  • around in the X, Y plane as you wait, sometime T now

  • when we look at the X, Y plane now your vector is over here

  • and it's continuing to precess.

  • Remember this is what gives rise to your signal that's your FID

  • where you get this cosine wave

  • over here that's basically the coil picking

  • up your precessing vector and generating electricity,

  • generating alternating current in the coil

  • that ultimately you amplify and use the analog

  • to digital converter on and then Fourier transform.

  • All right what I want to do at this point though is

  • to introduce a concept that NMR spectroscopists use

  • to make their lives easier and the idea is the rotating frame

  • and it's basically saying let's just have our axes precess

  • for conceptual purposes actually works

  • out for the detection purposes

  • with various electronic techniques,

  • but let's have our axes rotate at the angular velocity.

  • So that means we'll have the axes rotate at omega

  • and so we'll have our rotating frame and we'll call

  • that X prime, Y prime.

  • So now if you think about it if you're in the rotating frame,

  • if your frame is rotating, your frame of reference is rotating

  • with angular velocity omega and we wait time T

  • from the rotating frame how does our magnetization look

  • at time T?

  • Exactly the same and that's the whole big concept

  • of the rotating frame is simply we go ahead and we adjust

  • so we're not spinning around it 5 million times per second.

  • All right now we're ready to introduce the concept

  • of the spin echo and that's going to be the basis

  • for differentiating our methines and methylenes, our quats

  • and methylenes from our methines and methyls

  • in the DEBT experiment.

  • Okay, so the idea behind the spin echo is as follows.

  • We're going to be it the rotating frame and so we start

  • with a 90-degree pulse and I'll call it a 90-degree X prime

  • pulse just to show that we're working in the rotating frame.

  • Imagine for a moment now that I have some vectors

  • and they start along the Y prime axis, but now let's say

  • for a moment that some of those vectors are moving a little bit

  • faster than the Larmor frequency than the velocity omega and some

  • of them are moving a little bit slower.

  • So, in other words, we've got our frame rotating

  • at 500 million cycles per second

  • but some vectors are going a little faster

  • and some vectors are going a little slower.

  • So now if you imagine on this thought experiment

  • that you wait time T, now what's going to happen after time T

  • to after some time to those vectors?

  • The ones that are rotating slower will appear

  • to go clockwise and the ones that are going, right,

  • clockwise and counterclockwise.

  • So now our vectors are going to fan out with respect

  • to the rotating frame and those

  • that are going one direction are going to be going this way.

  • I think it's actually counter clockwise in this axis,

  • axis system, but the point is

  • that they're diverging in velocity.

  • So now we've got some that are headed this way and some

  • that are headed this way.

  • Now, imagine for a moment we now at this point

  • after time T apply a 180-degree X prime pulse.

  • In other words, now we do the same thing that we did before

  • and we apply a 180-degree pulse.

  • Now, when you do that, again just think right hand rule

  • and remember any component

  • of your magnetization that's along the X prime axis

  • when you apply a pulse to it right hand rule,

  • it doesn't do anything.

  • Any component of your vector that's along the Y prime axis

  • flips over 180 degrees.

  • Does that make sense?

  • So when we take this vector

  • and we apply our 180-degree pulse the Y prime component

  • comes on over to the negative Y prime like so and we do

  • that to all of these different vectors and they've all flipped

  • around the X prime axis and they've all gone

  • to the negative Y prime axis and yet they are still precessing

  • at the rotating frame but now they're headed inwards

  • and that's the key to this whole thing.

  • So because they're still going

  • in the same direction these ones are a little bit faster,

  • these ones a little bit slower

  • than the rotating frame they converge inward

  • and if we wait another time T, that same time increment

  • so I'll say T again now what's our picture at this point.

  • They're all on the negative Y axis so they are all

  • like so along the negative Y axis.

  • This is the basis for many, many sorts of NMR experiments.

  • This is called the spin echo and one of the reasons

  • that people invariably do spin echoes is in addition

  • to what we're talking about here which I'll show you in a second

  • which is J modulation, you've also got T2 relaxation

  • where vectors fan out in the X, Y plane

  • and what the spin echo experiment does,

  • what the spin echo does is a refocusing of those vectors.

  • So the ones that are inadvertently due

  • to T2 effects moving a little faster and the ones

  • that are moving a little slower

  • by applying this 180-degree pulse halfway

  • through they refocus and so almost every experiment you see,

  • the ones I'm talking about now

  • but also 2D experiments are going to involve some sort

  • of thing that involves a weight and then an equal weight.

  • So if I write out this simple pulse sequence,

  • what it is is we apply a 90-degree pulse, we wait,

  • we apply 180-degree pulse, we have an equal weight and at

  • that point the spin is refocused.

  • [ Writing on board ]

  • And so if you've already started to notice as you're working some

  • of the 2D experiments, there are various delays

  • and you may see this in the handout about various delays,

  • some of the delays you've learned

  • about in the 1D experiments may be relaxation delays

  • where it's just for your magnetization to return