Welcome to Chemistry 421

•Use of Physical Methods to determine structures in Organic Chemistry

Dr. Charles DeBrosse

232 Beury Hall (across from Dept. office)

NMR lab 001 Beury Hall

215-204-1082

Course Organization• Lectures Tuesday evenings 6-9pm. Attendance is strongly

encouraged. You should read the assigned material and do the homework problems prior to the lecture.

• Textbook, Organic Structural Spectroscopy, by Lambert, Shurvell, Lightner and Cooks

• Grade based on 3 In-class quizzes, homework sets and Final exam

• Course notes will be available on Blackboard generally the Monday before the lecture.

• I will reserve Monday 10-11am as office hour, and am accessible other times in my lab or office

Spectroscopy in Organic Chemistry….

• The Chemists Eyes, Ears and Nose

• How do we know what we have?

• Read labels (sometimes labels lie)

• Whose word do we have to take on it?

• Check it out for yourself! (Get a spectrum!)

• A spectroscopist. (never the hero, just the hero’s best friend)

Chemistry 421• Goal of the course is to give you the tools to answer

parts of these questions.• Organic Structure determination by spectroscopic

methods• NMR (nuclear magnetic resonance spectroscopy)• Mass Spectrometry (MS)• Infrared Spectroscopy (IR and other vibrational

classes like Near IR, Raman)• Electronic Spectra (Ultraviolet and chiroptical

methods)

Structural Features we can address Spectroscopically

• Molecular weight• Chemical Formula• Functional groups• Skeletal Connectivity, structural isomers• Spatial-geometric arrangements, stereoisomerism,

symmetry• Presence and location of chromophores• Chirality issues• Some of these are more central than others. Sometimes

we can stop when the answer is fit to purpose.

Techniques we will study in this Course

• NMR--looks atb atoms by means of their nuclei. Connectivity pathways, spatial arrangements of atoms and 1:1 correspondence between signals and atoms

• Mass Spec--measures molecular weight, most fundamentally useful for unknowns. Controlable fragmentation can distinguish among rival possibilities

• IR and Raman--Vibrations characteristic of bonds, particulary for functional group identification. Excellent “fingerprint”

• UV--reports on conjugation and multiple bonds. Provides entre’ to chiroptical probes to assymetric configurations

Chemistry 421• Goal is for you to gain a conversational

level of knowledge• Base level of theory, as pictoral as possible

should help you realize the scope and limitations of these methods.

• Survey of representative data and how to interpret it.

• Applying the right tool to the right question

Integrating data from various tools and sources (i.e. we should be able to explain all our observations. Also we should be able to observe features we predict, knowing our chemistry)

Complementary Spectroscopies•There are strengths and weaknesses in all the various spectroscopy methods

•“Blind Spots” (see story of three blind men describing an elephant)

•NMR e.g.; has “NMR silent” nuclei; fully substituted carbons as “blocks”; blind to inorganics, nearly blind to polymeric mtls.

•IR e.g.; robust for functional groups; great if you have a compound match in a library (fingerprint); shaky on quantitative response, same spectrum might explain multiple compounds; subject to selection rules governed by dipole moment in vibration (complement somewhat with Raman)

•UV; needs chromophores

•Mass Spec; compounds differ in ionizability.

A smart chemist will be attuned to possible blind spots before making conclusions. Best solution is to “marry up” complementary data. All the data needs to “agree” or at least, not conflict withing the various methods.

Total Synthesis as a Structural Proof

• Use of all our knowledge• Generally the product of a reaction is

rationally related to the ingredients we have used.

• Non-ambiguous route from known compounds

• Oxidative degradation to known compounds and history of chemistry

The Electromagnetic Spectrum•Light comes in different colors

•No matter what part of range, there are some features in common, that you should know.B

A

Propagation of e,b fields, time

Am

plitude

PhotonE= h

is frequency, Hz, 1/sec

= c/; |c=3x1010 cm/sec

h = Planck’s constant = 6.624 x10-34 J•sec

/c = 1/ = wavenumber, cm-1

1 wavelength, is a distance….

Peaks per time; frequency

Chemical Properties as related to the different colors of light

Log , meters

lower E,lower larger

higher E,higher smaller

-15 -10 -5 0 5

NMR

nuclear spin flips

X-Ray

Spacings between atomic nuclei in crystals Micro

waves

rotational states

IR+Raman

vibrations of bonds

UV

Electron orbital transitions

100 kcal/mol

10 kcal/mol

10-6 kcal/mol

What about Mass Spec?

The properties measured do not directly bear on absorption of light

More properly termed mass spectrometry

We do however in quadrupole mass spectrometry scan an electric field that induced different curve paths for different masses

Radiofrequency also does show up in FT mass spec (ion cyclotron resonance)

Sensitivity of the methods we will study

• NMR is worst, typically needing 10s of g to 10s of mg for 13C.

• Mass Spec is generally about 103-104 x more sensitive than NMR

• UV is about 100 x more sensitive than NMR

Questions, Questions….• What’s in this flask?• What have you just synthesized?• Did my reaction Work?• What is present in that sample?• How could this reaction have possibly Failed?• Isolated materials?• Purity? Mixture? What kind of mixture?• Is this material suitable for the next step?• Best to Ask Yourself these questions! (Better than making

your Boss ask, or not to be able to answer a “Customer”

Answers!• Bad Answer: “…because this always works in our

lab…”• Bad Answer #2 “..because my Professor (or some

slightly older grad student, postdoc) said so..”• Slightly less Bad Answer #3 “…this was done in

the literature…”• Good Answer “…because all the spectral and

other analytical data agree with me”• Analytical Chemistry. If you do it right, nobody

has to “take your word” on the answers!

Features in Common for all Spectroscopy

Measuring Scheme

Some Physical Property

Energy Transitions

Light, (Frequency Dispersion)

Data

Wisdom (or Progress)

KnowledgeA key for us here is, we use instruments to Disperse energy across a scale appropriate to a chemical property

Analysis, Interpretation

Therefore we have Precise Analytical Instruments that can

Disperse Energy•Accurate, precise, reproducible

•Combine the energy dispersion scheme with a detection scheme.

•Generally the sample sits physically between the source and the detector.

•Detector provides selectivity in response, usually generates a voltage. We record voltage responses as “DATA”

Detector

The dispersion is easy to achieve with ordinary light

Light sourceMonochromator rotates

Prism or diffraction grating

Spacings on grating appropriate to wavelength

Schemes use slits to admit a select region of spectrum

Pretty ineffective for radio waves

RecorderSample

Spectroscopy “Spreads out Vision”All the techniques we will discuss have some features in common

Data will have a running variable (x-axis) that is in some sense, a “energy” scale. (not at least directly, a time axis. Therefore a snapshot in time of a molecule)

The response variable (an absorption or other intensity) is related to the chemical preponderance of some feature that cause the response.

Energy

Inst

rum

ent

Res

pons

e

There is a peak hereBut not here

The position informs us about some chemical property in the sample

The peak height informs us about how much of that property is in the sample

Fourier Transform MethodsAn alternative to Energy

Dispersive methods •All modern NMR and IR is done this way

•Measures all frequencies at same time. More efficient at signal-gathering in a give time (better S/N)

•The frequencies present are deconvoluted (or dispersed) after data is collected.

•Fourier Analysis is the mathematical method for doing this. It is based on the theory that any complex periodic (repeats over time) wave can be decomposed into a linear combination of sinusoids

To get the measurement, we collect a detector response as a

function of time

Lots of different frequencies present from the sample

Their voltages “beat” against each other making interference pattern (interferogram)

Interference is periodic, because the frequencies are constant w.r.t each other

Inst

rum

ent

Res

pons

e

time

An Oscillating voltage is interpreted as a Frequency

This is a Frequency Axis. Think Hz!

The process is similar to the way a sound wave is digitized to make e.g. a music CD

Key to this is sampling at exactly equal time intervals

Interfering Sinusoids are Represented in a decaying trace

Key to the process is a very precisely defined time base (the x axis) that the FT algorithm uses to count

Space is frequency 1 Space is beating of frequency 2 vs 1 (1 - 2)

A human being could compute this FT, counting beats per time unit

Interference patterns--Almost able to Transform by Hand…

-1.5

-1

-0.5

0

0.5

1

1.5

2

2.5

0 200 400 600 800 1000

Added together

Time(ms)Time(ms)

-1

-0.5

0

0.5

1

0 200 400 600 800 1000

Damped Sines of 2.77, 28, 34 Hz

But its really the Fast Fourier Transforms and fast computers that make this practical!

Adding accumulating scans from the detector into memory of computer

Signals are coherent and adding the scans causes signal to grow linearly with number of scans.

Noise being random and incoherent grows with √no.of scans

From this, the Signal-to-Noise ratio (S/N) grows proportionally to the square root of number of scans

E.g., a spectrum acquired with 100 scans will be 10x better than one with 1 scan only.

Since the time-acquisition is fast and efficient it is easy to Signal-Average

Some Features common to all Spectra

Using computerized Data systems adds an additional limitation on our resolution problem

More data points are better but usually at a cost to expt efficiency

See here 2 identical peaks, digitized differently

Bears directly on our ability to determine the position of Imax

Linewidth. Usually measured at 0.5 Imax

Can be limited by the instrumentation, or be limited by nature.

Nature, here exerts herself as uncertainty to to slight chemical variation, or inability to measure energy precisely.

Units of linewidth are same as axis, e.g. Hz, cm-1

Resolution (ability to distinguish line from closely spaced neighboring line is related to linewidth

Noise, the curse of Science•All measurements, especially those we carry out with instruments, generate Noise.

•Detectors of all sorts generate electrical noise

•Noise is bad. It is random and incoherent and does not possess information. We go to tremendous expense and effort to eliminate, suppress, and finesse our way past noise.

•Signals are good. They give us information.

•Noise limits our ability to even observe very weak signals or to quantify somewhat weak signals. The Signal-to-Noise Ratio is an important parameter is assessing our ability to interpret data.

•Noise is superimposed on top of peaks

Signal-to-Noise (S/N) ratios

Noise(rms) is 0.707 x peak to peak

Measure height

S/N=6.3/2*0.707

=4.45

So this peak is reliably detectable, but not reliably quantitatable

Typical rule of thumb:

Limit of detection, S/N=3

Limit of Quantitation, S/N=10

Chemistry 421--Structure Determination

• Interpretable Connection between Structural Features and Spectroscopic signals

• We will interpret spectra to learn about structures.

• The Interpretation “paradigm” consists of charting:

Observation Inference

IR shows a C=O at 1730cm-1

NMR shows a 3-proton singlet at 2.2

My molecule contains a ketone or aldehyde

I may have a methyl group adjacent to C=O

May need more experiments or to look further in the data!

A way of Thinking……

“Partial Unknowns”

New compound?

Side reaction product

Wrong starting material, carry through known synthetic steps

Can we track known compounds?

Peaks from precursor compounds may have “descendents”

Total Unknowns

Isolated natural products

Unrelated impurities, contaminants

Single component, vs mixture?

Known compounds

(verify structure)

All predicted signals present?

Agreement with literature?

Impurities present?

Fingerprint?

????????

A Strategy for Handling Unknown Structures

• Complementary 1H NMR, 13C NMR, Mass Spec, UV--any features stand out?

• Get the Molecuar Weight from MS

• Heavy Atoms? (ratio of M to M+1, M+2)

• If heavy atoms are identified, subtract from MW

• Consult various molecular formula DBs (Merck, CRC etc). Write out Molecular Formula

• Use the DBE (sites of unsaturation) rule

• Infrared-- Functional groups present? Identify as possibly subtract from formula (retain the need to incorporate at end)

• Inventory 13C NMR and classify the C,H groups present. Tabulate fragments of structure. Reconcile MS fragments.

• Assemble possible structures

Molecular Weight and Molecular Formulas

•Absolutely critical to Stucture determination

•Centrality of Mass Spectrometry to modern Chemistry

•Molecular weight must agree with the structure. Note well, that a given nominal MW generally is consistent with several possible formulas.

•The “nitrogen rule”. A compound with an even-numbered molecular weight has 0, 2 or an even number of nitrogens.

•Very Important: Learn the rule for sites of unsaturation (double-bond equivalents, DBE) as a predictive tool for multiple bonds and/or rings. These are based on the standard valencies for ordinary atoms.

• Aim to reduce a formula to CNH2N+2

• Take formula and cross off Oxygen atoms

• Replace all halogen atoms with hydrogen

• Cross off all Nitrogen atoms, and for each N remove one H atom.

• Sulfur treat like Oxygen (? Use care if there are a lot of oxygens, possible O=S=O type groups, similar issues with Phosphorus)

• Subtract your newly reduced formula (looks like CxHx, from

CxH2x+2 number H (even number)

• Divide this answer by 2. Result is DBE.

DBE Rules

So some “Decision-Tree” thinking is possible

Synthetic Product

Correlations to protons

Carbon Survey

Noe for stereochemistry

Proton coupling pattern Assess Purity

Do I need more information?What do I need to find out

Worth more spectroscopy?Back to the Lab!

1H NMRCould it be what I want?

Quick Inventory of signals

Data

InformationContent higher

“granularity” of questions

The NMR branch.

Integrate at higher level with other techniques

Separations methods, Feedback to synthesis.

NO

YESNeed Assignments?

YES

NO

Nuclear Magnetic Resonance(NMR)

Tonight’s Subjects

I. How do the spectrometers work?

II. The NMR measurable quantities

What is NMR Spectroscopy?

• Nuclear Magnetic Resonance

• Radio Frequency Absorption Spectra of atomic nuclei in substances subjected to magnetic fields.

• Spectral Dispersion is Sensitive to the chemical environment via “coupling” to the electrons surrounding the nuclei.

• Interactions can be interpreted in terms of structure, bonding, reactivity

The Fundamental NMR equations

• Spinning nuclei produce a magnetic field that is proportional to its magnetic moment . The proportionality constant is ; = hI

• An active nucleus in a magnetic field B0 has an energy w.r.t. zero field of:

E (= h= h) = - • B0 where is the component of the magnetic moment colinear with B0

This gives for IZ= ±1/2; E = ± 1/2 (h B0)

E = h B0 and in angular units = B0

Origin of the NMR Effect• Nuclei with other than A(#protons+neutrons) and Z(#protons)

both even numbers, possess net spin and associated angular momenta

• Reveals itself only in magnetic field. As usual, such momenta are quantized

• States have different energies, populated according to Boltzmann distribution

• States are 1/2, 3/2, 5/2…for A= odd number and integer if A= even number and Z= odd number

• Transitions of individual nuclei between spin states is possible (both directions) leading to an equilibrium of populations

• Number of states is 2I + 1

Pictoral View of Spin

Precession of nuclear magnet--Units of Torque

Z

X

Y

M

Because we are forced into observing the group behavior, we have the mathematic equivalent of the simple picture on the right

Averages out in x,y plane; small net resultant vector along the z axis

Direction of the Applied Magnetic Field

X

Y

Z

Nucleus Moments precess about magnetic field.

Quantized either with or oppossed to field

The Boltzmann excess of low over high energy state is very small, 1 in 106

Resonance--A general phenomenon for energy pumping

Imagine a kid on a swing…

The period (frequency of the swing is determined (g, r(length), ).

Lets say the natural period is 3 seconds, or the frequency is 0.33If the Daddy gives a push every 3 seconds, the kid will go higher and energy will be absorbed.

Every 2 seconds and the motion will get stalled and “interfered” with.

Every 1.5 seconds and the energy will get absorbed but not as efficiently. The Daddy will get tired.

This general principal applies in NMR among other kinds of measurement, and holds whether we scan through the applied frequency or multiplex all at once

NMR-What is it Good For?(absolutely everything!)

• Solving structures of compounds like synthetics, impurities, natural products

• Identifying metabolites

• Stereochemical determination

• Follow reactions

• Validating electronic theory; trends within series of compds.

• Kinetics

• Extended structure, e.g. protein nmr

• Molecular interactions e.g. ligand binding

• Acid-base questions

• Purities

• Mechanisms, e.g. isotope distributions, other effects

• Questions about the solid state

• Imaging

And Besides that…

• You get your sample back!

• Not so for mass spec

• Try recovering your compound from a KBr pellet or nujol mull

But on the other Hand…

• NMR is one of the least sensitive analytical methods

• Characterized by long relaxation time constants, limiting experimental efficiency in real time

• Sometimes too much information. Can be demanding on interpretation skill

• Relatively Expensive compared with other analytical methods

• As with other methods NMR has “blind spots” and cannot serve as an analytical panacea

What Do I Hope you will Learn?• Enough theory to make you conversant in the area….

• NMR with respect to how the effects arise and can be predicted; connection with experiments and limitations of these; survey of how the instruments work.

• Basis of the experiments

• Data processing considerations, at level to appreciate what may have been done to give your result.

• A basic toolbox of experiments, what they do and how to use them in your work

• A working knowledge of organic chemical shifts and influence of symmetry on signal counting

• Spin coupling, coupling networks and connectivity, use of J-coupling constants in chemistry

Why NMR?

• Unmatched versatility as an Analytical technique• High on chemical information content• Significant interpretability• Interpretable at several levels of sophistication• Response related to molar preponderance• These attributes are true for solids, liquids,

mixtures, and to a small extent, gas phase• More than half the periodic table has at least one

NMR active isotope

What are the Measurables in NMR?

• Intensity (analytical parameter, proportional to molarity)

• Chemical Shift (the electronic surroundings)

• Couplings (scalar J and dipolar D; bond paths, angles connectivity and distances)

• Relaxation parameters (motions, distances)

How do we Generate, and Record NMR Spectra?

signal PreAmp

Probe in Magnet

receiver

transmitter

Pha

se lo

cked

loop

RF pulse signal

•Pulser•Frequency generation•Power Amplifier•Oscillator

FID with 90deg phase shift

Free Induction Decay

Host Workstation

Acquisition computer

Data file storage

NMR Acquisition commands

Timing control signals

•Superheterodyne (beat-down to AF)•Phase sensitive detection•A/D convertor

•User interface•Expt. Setup, control•Data processing, plotting

Network

Block Diagram for Spectrometer

Radio Frequency Transmit-Receive system

•Finely controlled RF pulses

•Microsecond control

•Precise control of timing, e.g pulses and delays

•Other precisely delivered RF for decoupling, selective excitation

•Gradient amp and generator, shielded in probe to avoid eddy currents

Modern Superconducting NMR Magnets

Lines of force project several feet into the room. They concentrate at the top and bottom. Magnets can grab iron objects and accelerate them.

Older Magnets (1970s) had opposed pole faces. High voltages and currents demanded heroic temperature control. Field ran side to side through sample

Supercon magnets have much larger fields, better homogeneity. Field runs up the axis of the sample.

New technology! Built in auxilliary magnet with reversed current acts as “active” shield, partly eliminating the projection into the room.

Note: Special superconducting alloys Niobium-Tantalum. Search goes on for higher temperature superconductors.

What’s the role of the magnet?• Bigger the field strength, the better. This is both

from a sensitivity and dispersion of signals point of view.

• Expressed in Hz, permits easiser math and trig as needed. Gauss would generate energies in ergs.

E= h H0Iz ener

gy

Field strength, H0

Remember the energy difference gives the population excess. Roughly H7/4 increase

The NMR Probe

Coil

Tuned Circuit

Matching to Tx network

Sample goes inside here

Usually there is a double tuned response for Deuterium lock

A second coil provides a decoupling, gradient or other RF

How Sensitive is NMR?

3 000

22 )1(HH

IkT

IIN

h

http://arrhenius.rider.edu/nmr/NMR_tutor/periodic_table/nmr_pt_frameset.htmlAnother good site is http://nmr.magnet.fsu.edu/resources/nuclei/table.htm

The Rider site, referenced below gives receptivity vs. 13C with clickable entries. These reflect natural abundance, , etc.

N=#spins

The answer to that question is…• Not all that Sensitive! • At any given time Mass spec is at least a 104 times more sensitive

• Compare with UV, IR at least 102x sensitive

• This is tied to the fact that NMR detects only the tiny Boltzmann excess. Any old molecule can fragment in MS or absorb a IR photon. Lots of research in NMR aimed at the sensitivity problem

€

NβNα

= eΔE

kT

k is Boltzmann constant 1.38x10-23J/molecule•K

Because the E is so small, the excess (which is what we detect) is miniscule

, are the short names for the upper, lower spin quantum states of a spin=1/2 nucleus

Most Important Nuclei in NMR•1H, (also 2H, 3H)•13C•31P•15N especially when labeled into proteins•19F•29Si

•Some isotopes of Sn, Cd. Pb, Ag, PtNo coincidence that these are the I=1/2 nuclei. Spin numbers higher possess nuclear quadrupole moment as well. This couples to, broadens and complicates the nuclear spin angular momentum. For the most part these are niche nuclei. Exception is 11B

Quadrupolar Nuclei

• Spin ≤ 1• Electric field at nucleus non-symmetrical• Effective relaxation mechanism, promotes loss of

NMR fine structure• “decouple” from attached spins. Can even wipe

out attached spin 1/2 signals.• Lines are broad, very challenging NMR• 35Cl, 11B, 17O, 14N, 7Li, etc.• Some redeem themselves, deuterium, 6Li