Chem805 Identification of organic and inorganic compounds by spectroscopy

1
Chem-805Identification of organic and inorganic
compounds by spectroscopy

• Mass Spectrometry
• NMR
• Infrared

2
Mass Spectrometry
Simplest form of mass spectrometer performs 4
essential functions (under vacuum 10-6 mm Hg)

• It subjects vaporized molecules to bombardment by
  a stream of high-energy electrons, converting
  these molecules to ions
• These ions are then accelerated in an electric
  field
• The accelerated ions are then separated according
  to their mass-to-charge ratio in a magnetic or
  electric field
• The ions that have a particular mass-to-charge
  ratio are then detected by a device that counts
  the number of ions striking it

3
Mass Spectrometry Introduction
MS is concerned with the separation of matter
according to atomic and molecular mass. With
recent development and improvement of
instrumentation and techniques it can be used in
the analysis of organic compounds of molecular
mass up to as high as 200,000 Daltons.
MS uses magnetic and electric fields to exert
forces on charged particles (ions) in a vacuum.
Therefore, a compound must be charged or ionized
to be analyzed by a mass spectrometer.
4
Mass Spectrometry
Atomic and molecular weight are expressed in
Atomic Mass Unit (amu).
The amu is based on a relative scale in which
12C is assigned exactly 12 amu. Also called
dalton
Each isotope has specific exact mass. MS is
interested in exact mass up to 3-4 figures after
decimal point (high resolution HR).
Chemical atomic weight is the average weight
taking into account the mass of each isotope and
their natural abundance.
5
Components of a mass spectrometer
10-5 to 10-8 Torr
System Inlet
Ion Source
Mass Analyzer
Detector
Signal processor
Vacuum System
m/z
6
?
7
(No Transcript)
8
(No Transcript)
9
MS Ionization

• Gas Phase
• Electron ionization (EI)
• Chemical ionization (CI)
• Field Ionization (FI)
• Desorption
• Field Desorption (FD)
• Fast Atom Bombardment (FAB)
• Secondary Ion Mass Spectrometry (SIMS)
• Laser Desorption (LD)
• Plasma Desorption (PD)
• Thermal Desorption
• Thermospray ionization (TS)
• Electrospray (ES)

10
Mass analyzers Magnetic sector
In Magnetic Field (H) an ion of charge z and mass
m experiences centripal force Hzv (where v is the
velocity of the ion). At the same time, any
particle moving on a circle of radius r
experiences centrifugal force of mv2/r When these
two forces are equal the ion travel on a circle
and Hzv mv2/r m/z Hr/v
We could measure v, the velocity of the ion to
determine the mass but this is very difficult
experimentally
11
(No Transcript)
12
Mass Spectrometer

• If a molecule absorb an electron (creating a
  Negative Ion), It will be absorbed by the
  repeller plate
• The repeller plate (Positively charged) directs
  the ions through a series of accelerating plates
  Kinetic Energy
• In Magnetic Field (H) ions describe a curved Path
  (r -gt curvature)
• Ions with reater m/e have larger curve
• Instruments has fixed curve gt only particules
  with correct m/e can reach detector
• Magnetic field is varied to detect all ions

1 mv2 eV2
m H2 r2 e 2v
r mv eH
13
Components of MS Mass Analyzers
Dispersion of the ions is based on mass-to-charge
ratio
Mass Analyzer
There are several type of mass analyzers.

• Magnetic sector
• Electrostatic and Magnetic sector
• Quadrupole MS filter
• Ion trap analyzers
• TOF Time-of-Flight
• FT-MS

14
Components of MS Detector, Vacuum
Detector
Convert the beam of ions in an electrical signal
that can be processed, stored, displayed and
recorded in many ways.
Electron Multiplier (most commonly used)
Faraday cup Photographic plates Scintillation
type
Other
Vacuum System
MS require the high vacuum is maintained in all
spectrometer components (except signal processing)
15
Amplifying signal
Continuous Dynode electron multiplier
Provide high gain and nanosecond response time.
16
Mass Spectrum
Ionization
m/z 32
Molecular ion
Fragmentation
m/z 31
CH3 HO ?
CHO H2
m/z 15
m/z 29
B ? Base Peak ? 100
Tabular presentation Bar graph presentation
17
Benzamide EI
B
M
77
44
- NH2?
- CO
C6H5CO
C6H5
m/z 105
m/z 77
105
18
3-methyl-6-i-PropylCyclohex-2-ene-1-one
MW 152
Isotopic cluster M, M1, M2
19
Natural Abundance of a few elements
Isotope Atomic weight Mass Abundance
1H 1.008 1.00783 99.99 2H 2.01410 0.016
12C 12.011 12.0000 (std) 98.89 13C
13.00336 1.11
14N 14.0067 14.0031 99.64 15N 15.0001
0.36
16O 15.9994 15.9949 99.76 17O 16.9991
0.04 18O 17.9992 0.20
20
Natural Abundance of a few elements
Isotope Atomic weight Mass Abundance
19F 18.998 18.9984 100.0
28Si 28.0855 27.9769 92.17 29Si 28.9765
4.71 30Si 29.9738 3.12
32S 32.066 31.9721 99.64 33S 32.9715
0.76 34S 33.9679 4.20
35Cl 35.4527 34.9689 75.77 37Cl
36.9659 24.23
21
Natural Abundance of a few elements
Isotope Atomic weight Mass Abundance
31P 30.9738 30.9738 100.0
79Br 79.9094 78.9183 50.52 81Br
80.9163 49.48
126I 126.9045 126.9045 100.00
The Atomic weight
Average atomic weight of all isotopes with their
natural abundances
Atomic weight Br
79Br x 78.9183 81Br x 80.9163
50.52 x 78.9183 49.48 x 80.9163 79.9094
22
Natural Abundance of a few elements
23
Molecular formula
M? (molecular ion) ? m/z
M ? mass of the most abundant isotope
e.g C7H7NO
7 x 12C 84 7 x 1H 7 1 x 14N 14 1 x 16O
16
m/z ? 121
Isotope peaks ( respect to M)
M1 due to (13C) or 15N
(17O) or 2H are negligeable)
(M1) (1.1 C) (.38 N) 8.08
(M2) (1.1 C)2/200 (.2 O) 0.5
For C, H, N,O, F, P composition
24
Molecular formula in general
Molecular ion
Even Mass can contain C, H, O , halogen, even
N
Odd Mass can contain C, H, O , halogen, odd N
Fragmentation ions
Even Mass From even mass M comes from
rearrangement or 2 bond breaking
Odd Mass From even mass M comes from single
bond breaking
even Mass from Odd Mass M comes from single
bond breaking
25
Molecular formula RULE of 13
Consider CH unit ? 13 amu
If we divide the mass by 13 we can establish
easily possible formula
Example Molecular ion gt 152
152 / 13 carbons 11
Mass 12 11 132 therefore H 152 132 20
Basic formula C11 H20
If Oxygen is present mass 16 ? remove CH4 If
Nitrogen is present mass 14 ? remove CH2
If one oxygen C11 H20 CH4 O C10 H16 O
If second oxygen C10 H16 O CH4 O C9 H12 O2
If third oxygen C9 H12 O2 CH4 O C8 H8 O3
26
Molecular formula RULE of 13
After establishing the basic formula with only
Carbon/hydrogen, Other element can be introduced
by substracting the proper hydrocarbon value
16O gt CH4
14N gt CH2
19F gt CH7
1H12 gt C
28Si gt C2H4
31P gt C2H7
32S gt C2H8
35Cl gt C2H11
79Br gt C6H7
127I gt C10H7
27
Nitrogen rule
Molecular ion gt 26
Molecular ion gt 27 odd!
26 / 13 carbons 2
27 / 13 carbons 2
Mass 12 2 24 therefore H 2
Basic formula C2 H3
Basic formula C2 H2
One Nitrogen C2 H3 - CH2 N
28
Nitrogen rule
Molecular ion gt 100
Molecular ion gt 99 odd!
100 / 13 carbons 7
Basic formula C7 H16
Mass 12 7 84 therefore H 16
1 Nitrogen C7 H16 CH2 N
Basic formula C7 H16
C6 H14 N
1 Oxygen C7 H16 - CH4 O
C6 H12 O
29
Calculating M1 and M2
30
Isotope peaks
Usually M2 peak is very small
Except for
M M1 M2
Sulfur 32S 100 34S 4.4
Silicon 28Si 100 29Si 5.2 30Si 3.35
Chlorine 35Cl 100 37Cl 32.5
Bromine 79Br 100 81Br 98
31
Isotope peaks CH3Br
12CH379Br
12CH381Br
M - H
13CH379Br
12CH279Br
12CH281Br
13CH381Br
32
Bromine
79Br
81Br
79Br
81Br
2 intense peaks for the molecular ion, spaced by
2 daltons. 79Br and 81Br
33
Chlorine
35Cl (P .75) 37Cl (P .25) Ratio 31
Probability M / M 2 0.75 / 0.25 3 / 1
100 33
34
Calculation of isotope pattern 2 Cl
C2H235Cl2
1 Cl
C2H235Cl 37Cl
C2H237Cl 35Cl
C2H2 37Cl2
2 Cl
P (2 35Cl) (0.75 )2 0.563
P (35Cl 37Cl) P (37Cl 35Cl ) (0.75 ) (0.25 )
(0.25 ) (0.75 ) 0.375
P (2 37Cl) (0.25 )2 0.063
M / M 2 / M 4 100 / 66 / 11
35
Calculation of isotope pattern ClBr
35Cl gt .75 37Cl gt .25
79Br gt 51 81Br gt 49
RClBr can exist as 4 isotopic forms
35Cl 79Br M 0.75 x 0.51 38
35Cl 81Br M2 0.75 x 0.49 37
50
37Cl 79Br M2 0.25 x 0.51 13
37Cl 81Br M4 0.25 x 0.49 12
At very high resolution, the 2 (M 2) peaks can
be distinguished (separated by 0.001 Dalton)
36
Chlorine and Bromine
The Molecular ion M is always the lowest mass
peak in the ion cluster (regardless of its
relative intensity)
37
Isotopic abundances for Carbon containing
compounds
M gt p(M) p(n 12C) (0.989)n
The probability of finding 1 13C among n carbons
is
M1 gt p(M1) p(n -1) 12C 1 13C n pn-1
12C x p(13C)
n(0.989)n-1 (0.011)
Relative ratio M1 / M n(0.989)n-1
(0.011) / (0.989)n
n (0.011) / (0.989)
n (0.0111)
In percentage n x 1.1
38
Isotopic abundances for Carbon containing
compounds
When comparing calculated abundances with
observed intensities We see that the ratio is
not identical due to experimental error The
relative abundance of the larger peaks are
reproducible to 10 For smaller peaks, relative
intensity is larger
Generally speaking, The size of the M1 ion
can be used to figure out the number of carbons
in a molecule
For example, if molecular ion is observed at m/z
118 and the ion at m/z 119 has an intensity of
about 9
There are probably 8 carbons (1.1 x 8 8.8 )
39
Isotopic abundances for Carbon containing
compounds
Due to the relatively large error on peak
intensity ( 10) determination of the number of
carbons is sometime ambiguous For example M1
gt intensity 22.5 2.3 Range covers 20.2 to
24.8 gt 20 2 carbons
40
Isotopic abundances for other common nuclei
For 15N M1 / M gt n x 0.36
For 33S M1 / M gt n x 0.80
For 18O M2 / M gt n x 0.20
For 34S M2 / M gt n x 4.42
41
Calculating Peak Intensities from Isotopic
abundances
42
Isotopic abundances for Silicon
Often observed is TMS compounds
M2 larger than usual
Often observed either from GC column Or septum
43
Isotopic abundances for Sulfur
SO
M2
M2
For 18O M2 / M gt n x 0.20
For 34S M2 / M gt n x 4.42
M2 4.42 (2 x 0.20) 4.82
44
isotope pattern 2 Cl (example)
When there are many carbons, isotopic pattern for
13C adds up to chlorine pattern
45
Steps in the Identification of Unknown

• Identify Molecular ion M.
• Determine Molecular Formula (odd / even mass)
• Analyze heteroatom (M1 and M2 )
• S, Si, Cl, Br, .
• Use rule of 13 to determine Carbons (M1 and
  M2 )
• Compare with 13C-NMR ( carbons) with APT
  experiment (J-MOD) ( protons)
• Compare with proton NMR ( protons)
• Identify base peak (note if even / odd)
• One or two bond fragmentation
• Test your conclusions in lab make derivatives
  (TMS or Na or K complexes ? mass shift)

46
Solving problems in MS

• Try to identify the Molecular Ion or decide if it
  is present (most critical step in solving a
  structure)
• Check if M1 ion is too large to accommodate
  reasonable number of carbons. (the M1 ion
  might be the very small M instead!)
• Determine the first loss from proposed molecular
  ion. Some loss are impossible (e.g 12, 14, 23
  daltons)
• Does the spectrum appear dirty? (lots of small
  peaks even at high mass)
• If GC of the comopund Is available, compare
  retention time
• Is the molecular weight even or odd?
• An odd mass can be associated with an odd number
  of Nitrogen
• An even mass means no Nitrogen or an even number
  of Nitrogen
• This Rule is applicable only to Molecular ion and
  to odd-electron ions
• Examine ion cluster for isotopic natural
  abundance (look for special heteroatom pattern).
  Try to calculate number of carbons

47
Solving problems in MS

• From the overall appearance is it a fragile
  compound? Is it likely to be aromatic or
  aliphatic?
• Look in the low mass ions. Do you see any clues
  of the family of compounds that you might be
  dealing with?
• Make a list of suggested losses from the
  molecular ion and try to make a pattellsrn from
  them.
• Look for intense odd-electron ions in the
  spectrum this is almost impossible in compounds
  containing Nitrogen! These provides clues for
  rearrangements (retro Diels Alder, McLafferty)
• Speculate on the structure using all that
  information

Index of Hydrogen deficiency
CxHyNzOn
Index x ½ y ½ z 1
48
Neutral losses and Ion series
M-30 NO ? (Nitro compounds), H2 CO ? (anisoles)
M-1 H?
M-15 CH3 ?
M-16 O ? (rare) , NH2 ?
M-31 CH3O ?
M-17 OH ?, NH3 (rare)
M-32 CH3OH
M-18 H2O
M-35 Cl ?
M-19 F ?
M-36 HCl
M-20 HF (very rare)
M-42 CH2CO, CH2CH-CH3
M-26 HCCH , CN
M-43 CH3CO ? , C3H7 ?
M-27 HCN
M-44 CO2
M-28 H2CCH2 , CO
M-45 CH3CH2O ? , CO2H ?
M-29 CH3CH2 ? , HCO ?
49
Neutral losses and Ion series
Figuring out which peak is molecular ion can be
supported by identifying what fragment is lost.
There can be sometimes 2 consecutive loss In
steroid, M-33 is often observed comes from the
loss of Me and H2O
The ions loss are only useful from molecular ion
There is no fragment in organic compounds between
M-1 and M-15
Loss of M-14 is never observed!
Other gaps in mass loss are between 21-25,
33-34, 37-41
Ions in these areas should be viewed
suspiciously either compound is not pure or
postulated molecular ion is wrong
50
Neutral losses and Ion series
Among the losses most common are Loss of H?, CH3
? , H2O (from some oxygenated compounds), HC?CH
(from aromatic compounds), HC?N (from aromatic
compounds containing Nitrogen), C?O and CH2CH2
(both at 28! Difficult to tell which one is lost)
Ethyl radical (29) Methoxy radical (31) Cl and
HCl (35, 36) Acetyl (43) accompanied by m/z 43
prominent and propyl (43) radical
51
Resolution
52
Exact Masses of isotopes
53
Mass Defects
By definition the atomic weight of 12C is 12.0000
Daltons All other elements are determined
relative to 12C Their difference to nearest
integral is called Mass Defect
Light element has small positive mass defect
Vast majority of elements have substantial
negative mass defect
54
Mass Defects
The Mass Defect of a C23H35O2Si is shown here on
a low resolution MS (/- 0.1)
Mass defect for H 1.008-1.000 .008 Mass
defect for O 15.995-16.000 -.005 Mass defect
for Si 27.977 -28.000 -.003
(35 x 0.008) (2 x -.005) (-.003) .27
Mass defect for organic compounds is mainly due
to Hydrogen due to their large number
55
Mass Defects
Mass defect brings the maximum of the C23H35O2Si
peak not at 371 but at 371.25 If the Mass
spectrometer measure the mass with a resolution
of 1, the sensitivity is reduced.
Greatest application of Mass Defect is surely
very high resolution, which provide mass with
.001 to .0001 accuracy
56
High resolution
Exact Mass can provide Molecular Formula
With sufficient accuracy, unique molecular
formula can be determine
e.g. Distinguish CO, N2, CH2N and C2H4 (all
having m/z 28)
57
Exact Mass can provide Molecular Formula
Consider the following 2 formulas which have m/z
287
C15H10NO3Cl
C14H8N2O3Cl
Mass Defect of C15H10NO3Cl (10 x .0078)
(.0031) (3 x -.0051 ) (-.0311 ) .0347
Mass Defect of C14H8N2O3Cl (8 x .0078) (2 x
.0031) (3 x -.0051 ) (-.0311 ) .0202
With sufficiently high resolution MS, it is
possible to propose a unique empirical formula
for an ion.
58
Index of Hydrogen deficiency
Nitrogen rule
M? even ? even of N 0, 2, 4,
M? odd ? odd of N 1, 3, 5,
Index C H/2 X/2 N/2 1
e.g. C7H7NO ? Index 7 - 3.5 1 5
Hydrogen deficiency can be unsaturation (multiple
bonds) or cyclic structure
R-C?N
I2
I4 (3 DB cycle)
I1 (1 DB cycle)
59
Index of hydrogen deficiency General
Index IV (I/2) (III/2) 1
IV ? any tetravalent atom (e.g. C, Si..)
I ? any monovalent atom (e.g. H, D,Cl, Br, F )
III ? any trivalent atom (e.g. N, P, )
II ? any divalent atom (e.g. S, O,).
These count for 0 in the formula
Higher valences uses valence shells ?lewis octet
rule
phosphine
60
Fragmentation in different isomers
Molecular ion almost absent
61
Ionization Techniques
Electron Impact (EI)
Under these conditions, very energetic Ions are
produced gt Fragmentation M. is often weak
e.g. ROH often loose H2O
Intensity of M. Depends on the stability of the
Ion
Most Stable
M. is often absent in
62
Other Ionization Techniques
Under EI (Electron Impact), the molecular ion may
be weak or absent
In such case, the best solution is to run CI
Chemical Ionization Which result in intense M1
ion with little fragmentation
CI Chemical Ionization

• Vaporized Sample is introduced with excess gas
  (usually methane)
• The gaz is ionized producing Primary ions that
  react with excess gaz to produce secondary
  ions

CH4. CH4 ? CH5 CH3.CH3 CH4 ? C2H5 H2

• Secondary ions react with sample M

CH5 M ? MH CH4C2H5 M ? MH C2H4
Less energy than CI gt less fragment
63
Chemical Ionization techniques
In Electron Impact (EI) and chemical ionization,
vaporization of the Sample is a prerequisite.
Therefore the study of non-volatile or thermally
fragile compound is not possible with these
techniques.
Several methods have been developed to go around
these limitation
(methods that enable ions to be obtained from
liquid/solid)
1- Field Desorption ions are desorbed by strong
electric field
In this and other techniques,, the aim is to
transfer energy to the Sample, causing transfer
of molecules/ions to gas phase
2- SIMS other source of energy is a beam of
ions (Secondary Ion Mass Spectroscopy)
Typically Argon (FAB)
Field Desorption methods give a high
concentration of Molecular ion gt Particularly
useful for identification of unknown
64
Fast Atom Bombardment (FAB)
The sample is dissolved in a liquid matrix such
as glycerol, thioglycerol, m-nitrobenzyl alcohol,
or diethanolamine and a small amount (about 1
microliter) is placed on a target. The target is
bombarded with a fast atom beam (for example, 6
keV xenon atoms) that desorb molecular-like ions
and fragments from the analyte.
e-
Xe? Xe
Xe
Xe?
Is accelerated to 6-10 keV, and pass through Xe
Xe Xe?
Xe
(Xenon with Kinetic energy) FAB
Cluster ions from the liquid matrix are also
desorbed and produce a chemical background that
varies with the matrix used.
65
FAB Benefits
Sample introduction can be through direct
insertion probe or LC/MS (continuous-flow FAB).

• Benefits
• rapid, simple
• relatively tolerant of variations in
  sampling
• good for a large variety of compounds
• Useful fragentation pattern
• strong ion currents -- good for
  high-resolution measurements

66
FAB Limitations

• Limitations
• high chemical background defines detection limits
  
• may be difficult to distinguish
  low-molecular-weight compounds from chemical
  background
• analyte must be soluble in the liquid matrix
• no good for multiply charged compounds with more
  than 2 charges
• requires a high concentration of the organic
  liquid matrix (typically 80 to 95 glycerol)
  which limits sensitivity
• Mass range
• Moderate Typically 300 Da to about 6000 Da.

67
Comparing different techniques
68
Classification according to method of separating
charged particule

• Magnetic Field Deflection
• Magnetic Field only Mass 12-500 in seconds
• Double Focussing High resolution to 4
  decimalsr 60,000
• Quadrupole Mass Spectrometer
• Quadrupole Mass Filter Mass Scanning by varying
  RF DC Frequencies
• Quadrupole Ion Storage (Ion Trap)-
  Compact-Inexpensive-Very Sensitive-GC/MS

69
Classification according to method of separating
charged particule

• Time of Flight- Need Fast Electronic (10-7 s)-
  Used With Sophisticated Ionization Methods
  (FAB, Laser Desorption .)
• FT-ICR (Fourier Transform Ion Cyclotron
  Resonance)- Very High Precision most expensive
• MS/MS (Tandem Mass Spectro.)-Specific Ions are
  Separated in First MS- Pass one at a time in a
  collision chamber- Second MS produce Daughter
  Ions-used for large molecule and resolution of
  mixture

70
Time-of-Flight analyzer (TOF)
In TOF instruments, positive ions are produced
periodically using brief pulses of electrons,
secondary ions or laser generated photons
?
These pulses have typically a frequency of 10-50
kHz and lifetime 0.25ms
The ions produced are then accelerated by
electric field pulse (103-104 V) that has the
same frequency as ionization pulse but lags
behind.
The accelerated ions then pass into a field-free
drift tube (about 1 m long)
71
Time-of-Flight analyzer (TOF)
MALDI ionization, Time-of-flight Mass Spectrometer
72
Tandem-MS
73
Comparing Mass Spectrum of a compound in
different type of mass spectrometers
Magnetic sector
Quadrupole
Time of Flight
105
MW 164
74
Organometallic compounds in MS
It is often possible to determine molecular
weight of a compound by MS
For example Manganese carbonyl gt m/z 390
Mn2(CO)10
As Mn gt 55 and CO gt 28 ?
Another example Iridium complex
NMR and IR can give a lot of information but
without MS it is very difficult to show the
presence of Cl With MS, It is easy to show that 4
Chlorine are present
Molecular ion is usually present with Laser
desorption. However, ions produced That way
comes from condensed phase (solid/liquid) and
structure in these phase Might be very different
from the one in gaz phase.
With FAB most intense cation peak is the
protonation ion (M1) peak. Anion (M-1) can also
be formed.
75
Isotope Abundance patterns for some atoms and
group of atoms
Can be diagnostic for some isotopes
76
Calculation of isotope pattern
Abundances can be calculated by multiplying the
abundances of the Constituent isotopes
ReBr can exist as 4 isotopic forms
185Re79Br 264 m/z 0.37 x 0.51 19
185Re81Br 266 m/z 0.37 x 0.49 18.1
50.2
187Re79Br 266 m/z 0.63 x 0.51 32.1
187Re81Br 268 m/z 0.63 x 0.49 30.9
77
Metastable ions
Some ions have so short lifetimes that they
dissociate while moving through the spectrometer.

• An ion of mass m1 is accelerated after initial
  ionization
• But a different ion m2 (daughter ion) passes the
  magnetic analyzer
• The resulting peak is neither m1 or m2 but appear
  at m (metastable)

m (m2)2/m1
These metastable ions are formed during 10-5 s
(time spent between electrostatic and magnetic
analyzer) are quite broad but provide direct
information about ion reactions
78
Platinum
79
Tungsten
Index
MS-fragmentation