Department of Organic Chemistry, IMM, Radboud University Nijmegen … · 2007-11-08 · Department of Organic Chemistry, IMM, Radboud University Nijmegen Instrumental Analysis in

Department of Organic Chemistry, IMM, Radboud University Nijmegen

Instrumental Analysis in Molecular Chemistry 1

Fragmentation in Mass Spectrometry

• Introduction• Ionization• Separation• Fragmentation• Isotope Effects• High Resolution



Identification of Molecular Ion (1)

As we have seen, it is expected that a chemically pure compoundcontains a mixture of isotopes, based on the natural abundance. Molecular ion assigned on the basis of the most abundant isotope(s).

Conditions for the assignment of M+.:

a) M+. should be the highest m/z in the spectrum, apart from weaksatellite peaks that result from other isotopes.Problems with unstable M+. or chemical impurities.

b) M+. should be a radical cation in EI as it is ionized by removing 1 e.In formula CxHyNzOn (where C is any 4-valent element, H any monovalent,N any 3-valent, and O any divalent)the number of DBE: x – ½.y + ½.z + 1 should be an integer.

c) Value of M+. is even unless there is an odd number of N.



Identification of Molecular Ion (2)

d) If M+. is correctly assigned, the other peaks at high m/z can be logically explained by loss of neutral parts or molecules

OK: loss of CH3 (M-15), H2O (M-18), CH3O (M-31), CH3C=O (M-43) etc.

Illogical: M-2 till M-14M-21 till M-25M-33, M-37, M-38

If M+. cannot be satisfactorily assigned, look for an other assignment,or try another technique: CI, FAB, MALDI, electrospray …



Important Factors for Fragmentationa) Energy of the molecular ion and the fragments formed from it

b) Stability of the bonds in the ions

c) For rearrangements: steric factors.It is easier to move an H than a whole group

d) Stability of the formed ions or neutral particlesresonance stabilization such as in an acylium ion

Stevenson’s Rule:Upon dissociation of AB+. → A+ + B. or A. + B+

A+ will be formed if it has the lower ionization energy

In branched radical cations, the largest group is preferentially lost

CH

C4H9

CH3C2H5

+.CH+CH3

C2H5+HC C4H9

CH3C+ C4H9C2H5 C

+C4H9

CH3C2H5> >

H>



Stevenson’s Rule, Energy DiagramEn

ergy

ABA. + B.

A+ + B.A. + B+

DAB

IAIB

IAB

AB+.APA+

APB+

Ionization Energy (I), the energy required to convert radical into cation

Dissociation Energy (D), energy for homolytic dissociation into radicals

Appearance Potential (AP), energy required for cation to appear in MS



Ionization Energies for Common Hydrocarbons (1)Energy of Formation of Aliphatic and Olefinic Radicals

and Carbocations from RH and RCl

0

50

100

150

200

250

tBu iPr Et Me Bn All vinyl Ph

R

Ener

gy (k

cal/M

ol)

cationradical



Ionization Energies for Common Hydrocarbons (2)Formation Energies of Radicals & Carbocations < RH & RCl

0

50

100

150

200

250

300

350

tBu iPr Bn All Et vinyl Ph Me H

Ener

gy (k

cal/M

ol)

cationradical



Loss of Largest Group from Branched Radical IonStevenson’s Rule:

Upon dissociation of AB+. → A+ + B. or A. + B+

A+ will be formed if it has the lower ionization energyIn branched radical cations, the largest group is preferentially lost

Do not confuse ionization potential and cation/radical stability

CH

C4H9

CH3C2H5

+.CH+CH3

C2H5

+HC C4H9

CH3

C+ C4H9C2H5

C+

C4H9

CH3C2H5

H

.C4H9

.C2H5

.CH3

.H

+

+

+

+

sec. cation + tert. radical

sec. cation + sec. radical

sec. cation + prim. radical

tert. cation + .H



Metastable Peaks

Metastable ions: arise because of fragmentation after the ion hasleft the ionization chamber

M1+ → M2

+ + (M1 - M2)

The kinetic energy of such a M2+ is smaller than when it would have

been formed in the ionization chamber and accelerated there, and it willbe detected at a lower m/z than expected, and moreover broadened.

For fragmentation between electrostatic analyser and magnetic sector:

M* = M22/M1

e.g. C5H9+ (m/z = 69) → C3H5

+ (41) + C2H4 (28)

Calculated m* = (41)2/69 = 24.36, observed m* = 24.4

(It is more difficult to solve this problem in the other direction)



Types of Fragmentation Reactions (1)

R+.CR3 R. + +CR3

R+.I R. + +I

Alkanes:

Low IE elements

McLafferty & Turecek:

1) Sigma electron ionisation (σ):

2) Radical site initiation (alpha cleavage, α), homolytic dissociation:N > S, R, O, π > Cl, Br > H

Donates an electron,forms new bond to anadjacent atomconcomitant withcleavage of other bond to that atom,moves . site, loss of largestalkyl favoured

R CR2 YR+.

R2C Y+RR. +

Y+R CH2 CH2.

CH2 CH2+YR+.

Saturated

Unsaturatedheteroatom

Allylic

R CR Y RC Y++.R.

R CH2 CH2 CH2+.

+

H2C CH +CH2R. +

R -eR

+

.Retro-Diels-Alder(double α-cleavage)

α2

α,i chargemigration

RHCH2C

++.

RC2H3+. + C4H6



Types of Fragmentation Reactions (2)

R Y R R+ + .YR+.

OE+. :

R YH2 R+ + .YH2+

EE+ :

McLafferty & Turecek:

3) Charge site initiation (inductive effect, i), heterolytic dissociation:Cl, Br, NO2 > O, S >> N, C

Attracts e pair,cleaves bond,moves + site,most stable + formed

4) Rearrangements (r):Example with Hand unsaturatedreceptor site in ring

Also with saturatedreceptor site,2H, displacement (rd)and elimination (re)

YH rH

+.Y+H.

Y+H.

α

i.+

+

+

HY

HY+

.Charge retention, ion gains H

Charge migration, ion loses H

or



Fragmentation Reactions(Hesse, Meier, & Zeeh)

Alpha splitting

Benzyl- and Allyl splitting

Splitting of non-activated bonds

Retro-Diels-Alder reaction

McLafferty rearrangement

Onium reaction

Loss of CO



α-Splitting (Homolytic Dissociation) (1)

2-butanone

The acylium ion formed by lossof the largest alkyl radical ispreferred

O O+

orO+. .

CH3. + CH3CH2.+H3C

O+O+

CH3(15) m/z 57 m/z 43 (29)



α-Splitting (Homolytic Dissociation) (2) 100

0

50

75

25

20 25 30 35 40 45 50 55 60 65 70

45

3127 59294341

19 28 57

CH C 3HC 2HC3H

OH•

C 2HCH

C3H

OH

CHC3H C 2H

C 3H

CHC3H

OH

C 2H

OH

100

0

50

75

25

15 20 25 30 35 40 45 50 55 60 65 70

59

31

41 43 15 29 5727 39 60

C 3H

OH•

C 3H

C3HC

CC 3H

OH

C 3H

CC3H

C 3H

C 3H

2-butanol

splittingof Et.preferred

t-butanol

only Me.splittingpossible




2-AminoethanolIonization by loss of n-electronfrom N(less electronegative than O)which is also a likely startingpoint for radical-site initiatedfragmentation

H2N OH- e

H2N OH+.m/z 61

H2C N+H2

m/z = 30

.CH2OH(31)




α-splitting, mainly of Me, but also of H,is the main mechanism of fragmentationin amines because of the electron-donatingability of N



α-Splitting in MS of Protected Steroid

O

O

Me

Me

+.

O

O

Me

Me

+.

or

O+

O

Me

O+

O

Me

H. MeO+

O

Me.O+

O

m/z 99

H. Me

O+

O

Me

.Me

O+

O

Me.H

MeO+

O

Me

.

O+

Om/z 125



Formation of Stable Allylic Carbocation

R CH2 CH2 CH2+.

H2C CH +CH2R. +

+

EI of 1-heptene

EI of 4-methyl-1-hexene

m/z 41



Formation of Stable Benzylic Carbocation (1)

EI of butylbenzene

.+

or .+ .C4H9

+

(57)

+

m/z = 77m/z = 51C2H2

α

.C3H7(43)

+

+

+ +

m/z = 65m/z = 91

(26)

C2H2(26)



Formation of Stable Benzylic Carbocation (2)

The EI-MS of o-Cl-toluene andbenzyl chloride are similar, bothare dominated by the tropyliumion



σ-Electron Ionization, Non-activated Bonds

EI-MS of an unbranchedalkane shows an almostrandom fragmentationpattern (homologousseries, differing in theno. of CH2 groups), everyσ-bond is as likely to breakas the other one.

In the branched alkane,the possibility to haverelatively stable secondarycarbocations as opposedto primary ones influencesthe fragmentation pattern.

Stevenson’s rule,loss of largest alkyl



Alkyl halide spectra (1)

C7H14 F+.H

C7H14 F+

H. HF(20)(118)

C7H14+.

O.E. ion

variousfragments

n-heptyl F: strong C-F bonds,electronegativity of Fmakes + on F unattractive,HF formation attractiveNo M+. observed

n-heptyl I: less electronegative, less basic,I+ (m/z 127) or I. (with R+, m/z 99) feasible,but base peak at m/z 57 (C4H9+) ?!

R+.I R. + +I (m/z127)

or R+ (m/z 99) + .Im/z 226

σ



Alkyl halide spectra (2)

X+.

m/z = 57

+ + X.

X+. .

43

X++

m/z = M - 43

(X = F, 67)X = Cl, 91/93X = Br, 135/137(X = I, 183)

(X = F, 53)(X = Cl, 77/79)(X = Br, 121/123)(X = I, 169)

α

i

Fragments containing Br, Clrecognized by satellite peaksAn α splitting with cyclization isimportant (base peak for Cl)An i splitting with cyclization isalso important (base peak forI, Br)



Retro-Diels-Alder (1)

Retro-Diels-Alder, not pericyclic

Charge retention or migration,depending on R

NB: α followed by α or i gives another OE ion

See the example of4-phenyl-cyclohexene

chargemigration

R = Ph, 100 %

HCH2C

++.

R -eR

+

.Retro-Diels-Alder

R

+

.or

R

+

.α

α i

HCH2C

+R R .

+

chargeretention

R = H, 80 %



Retro-Diels-Alder (2)

O

α-ionone

O

β-iononeO.+ O

+

+ .CH3

See the example of the iononesα-ionone gives the expected Retro-DAβ-ionone has a competing loss of methyl group



1,2,3,4-Tetrahydrocarbazole

HN+.

C2H4

HN

HN +.

HN .+ H

N .+

C2H4

m/z 143

m/z 171



5,7-dihydroxy-4’-methoxyisoflavonone



McLafferty Rearrangement

YH rH

+.Y+H.

Y+H.

α

i.+

+

+

HY

HY+


Charge migration, ion loses H

or

CH2CH2

OH rH

+.O+H.

α+

HO+


CH3

C2H5

H3C

C2H5

H3C

C2H5(42)

m/z 6220 %

m/z 104

In McLafferty rearrangement(H rearrangement with unsaturatedreceptor site) a H from the carbon inγ position relative to a C=Y double bond istransferred to the Y atom, accompanied bya β-splitting. This can be with chargeretention (α) or charge migration (i).

Note that McLafferty rearrangements are relatively exceptional(cf. retro Diels-Alder, CO loss) in producing another OE ion.

Example: a relatively long ketone



EI-MS of methyl butanoate

CH2CH2

CH2

OH rH

+.O+H.

α+

HO+

.OCH3 OCH3OCH3(28)

m/z 74

m/z 102

or

CH3CH2

CH2

O+.

OCH3

α

CH3O.(31)

O+

OCH3m/z 59

CO(28)

CH3O+

m/z 31

or

CH3CH2

CH2

O+.

OCH3

α

CH3CH2CH2.(43)

O+

m/z 71 CO(28)

CH3CH2CH2+

m/z 43

The alkyl chain of the acid partof the ester is long enough togive a McLafferty rearrangement,in addition to α-fragmentationand subsequent loss of CO.



Loss of COO

H O

HH H H

+.

CO

+.

H.+

m/z 94 m/z 66m/z 65



Some Loss of COO

O

+.

m/z 192

+.O+.

m/z 164 m/z 136

C O

+.+

m/z 65m/z 96

+.m/z 68



No Loss of COO+.

m/z 98

O+

H. Me

O+

.O+

CH2.C3H7 m/z 55

+O

CH2m/z 57

H



Cyclohexylamine

+N

CH2m/z 84

Et



Onium Reaction

YRH+.

HYR+

. HYR+.

YRH+

HYR+ HYR

+

In the Onium reaction(H rearrangement withsaturated receptor site)a H is transferred fromsomewhere in an alkylchain to the heteroatomof initial ionization.

Y can be O (oxonium),N (nitronium), etc.

In OE ions, H is transferredwith one e; in EE ions it is ahydride shift, H as H- with 2 e.



Onium reaction, Ether

O+.

O+

H

m/z 102

m/z 87

m/z 31

C4H8 (56)

.CH3

α

Onium

or+O

H Me

m/z 45O+

Me CH2

m/z 59O+

CH2

HO+CH2

m/z 31

.C3H7

HO+CH2

α

McLafferty C3H6 (42) Onium

C2H4 (28)



Onium reaction, Amine

N

MeMe

Me+.

N+

Me

Me

N+H

Me

Me

H

m/z 129

m/z 114

m/z 58

C4H8 (56)

.CH3

α

Onium

orN+

HMe

Me

Me

McLafferty

m/z 72Me

N+Me CH2

m/z 86Me

N+Me CH2

Me

Onium

H+N

Me CH2

m/z 44

.C3H7

C3H6 (42) C3H6 (42)

α



Standard Interpretation Procedure(McLafferty & Turecek) (1)

1) Study all available information (spectroscopic, chemical, samplehistory). Give explicit directions for obtaining spectrum.Verify m/z assignments.

2) Using isotopic abundances, where possible deduce the elementalcomposition of each peak in the spectrum; calculate rings plusdouble bonds.

3) Test molecular ion identity; must be highest peak in mass spectrum,odd-electron ion, and give logical neutral losses. Check with CI or othersoft ionization.

4) Mark ‘important’ ions: odd electron and those of high abundance,highest mass, and/or highest mass in a group of peaks.



Standard Interpretation Procedure(McLafferty & Turecek) (2)

5) Study general appearance of the spectrum; molecular stability,labile bonds.

6) Postulate and rank possible structural assignments for:- important low-mass ion series;- important primary neutral fragments from M+. indicated by

high-mass ions (loss of largest alkyl favoured) plus those fromcollision activation;

- important characteristic ions.

7) Postulate molecular structures; test again reference spectrum,against spectra of similar compounds, or against spectra predictedfrom mechanisms of ion decompositions.

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Department of Organic Chemistry, IMM, Radboud University Nijmegen … · 2007-11-08 · Department of Organic Chemistry, IMM, Radboud University Nijmegen Instrumental Analysis in