Chem-805 Identification of organic and inorganic compounds by spectroscopy Mass Spectrometry NMR ...

Chem-805Chem-805Identification of organic and inorganic Identification of organic and inorganic

compounds by spectroscopycompounds by spectroscopy

Mass SpectrometryMass SpectrometryNMRInfrared

Mass SpectrometryMass Spectrometry

• It subjects vaporized molecules to bombardment by a stream of high-energy electrons, converting these molecules to ionsions

• These ions are then accelerated in an electric field• The accelerated ions are then separated according to

their mass-to-charge ratiomass-to-charge ratio in a magnetic or electric magnetic or electric fieldfield

• The ions that have a particular mass-to-charge ratio are then detected by a device that counts the number of device that counts the number of ions striking itions striking it

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

Mass Spectrometry: Mass Spectrometry: IntroductionIntroduction

MSMS is concerned with the separation of matter according to atomic and separation of matter according to atomic and molecular massmolecular 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 Daltonsmass up to as high as 200,000 Daltons.

MSMS uses magnetic and electric fieldsmagnetic and electric fields to exert forces on charged particlescharged particles (ionsions) in a vacuumvacuum. Therefore, a compound must be charged or ionized to be analyzed by a mass spectrometer.

Mass SpectrometryMass Spectrometry

Atomic and molecular weight are expressed in

AAtomic MMass UUnit (amu)(amu).

The amuamu is based on a relative scale in which

1212CC is assigned exactly 12 amu12 amu. Also called daltondalton

Each isotope has specific exact mass. MS is interested in exact mass up to 3-4 figures after decimal point3-4 figures after decimal point (high resolution HRhigh resolution HR).

Chemical atomic weight is the average weight taking into account the mass of each isotope and their natural abundance.

Components of a mass spectrometerComponents of a mass spectrometer

System System InletInlet

Ion Ion SourceSource

Mass Mass AnalyzerAnalyzer

DetectorDetector

VacuumVacuumSystem System

Signal Signal processorprocessor

m/zm/z

1010-5-5 to 10 to 10-8 -8 TorrTorr

MS : MS : IonizationIonization

1.1. Gas Phase:Gas Phase:1. Electron ionization (EIEI)2. Chemical ionization (CICI)3. Field Ionization (FIFI)

2.2. DesorptionDesorption 1. Field Desorption (FDFD)2. Fast Atom Bombardment (FABFAB)3. Secondary Ion Mass Spectrometry (SIMSSIMS)4. Laser Desorption (LDLD)5. Plasma Desorption (PDPD)6. Thermal Desorption7. Thermospray ionization (TSTS)8. Electrospray (ESES)

Mass analyzers: Magnetic sectorMass analyzers: Magnetic sector

In Magnetic Field (HH) an ion of charge zz and mass mm experiences centripal force HHzzvv (where vv is the velocityvelocity of the ion). At the same time, any particle moving on a circle of radius rradius r experiences centrifugal force of mmvv22//rr When these two forces are equal the ion travel on a circle and

HHzzvv = mmvv22//rr mm//zz = = HrHr//vv

We could measure v,v, the velocityvelocity of the ion to determine the massmass but this is very difficult experimentally

Mass SpectrometerMass 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 -> curvature)

Ions with reater m/em/e have larger curve

Instruments has fixed curve => only particules with correct m/em/e can reach detector

Magnetic field is varied to detect all ionsMagnetic field is varied to detect all ions

1 mv2 = eV2

r = mv eH

m = H2 r2

e 2v

Components of MS: Components of MS: MassMass AnalyzersAnalyzers

Mass Mass AnalyzerAnalyzer

Dispersion of the ions is based on mass-to-charge ratiomass-to-charge ratio

There are several type of mass analyzers.

1.1. Magnetic sectorMagnetic sector

2.2. Electrostatic and Magnetic sectorElectrostatic and Magnetic sector

3.3. Quadrupole MS filterQuadrupole MS filter

4.4. Ion trap analyzersIon trap analyzers

5.5. TOF – Time-of-FlightTOF – Time-of-Flight

6.6. FT-MSFT-MS

Components of MS: Components of MS: DetectorDetector, , VacuumVacuum

DetectorDetectorConvert the beam of ions in an electrical signal that can be processed, stored, displayed and recorded in many ways.

VacuumVacuumSystem System

MS require the high vacuum is maintained in all spectrometer components (except signal processing)

Electron Multiplier (most commonly used)Electron Multiplier (most commonly used)

Faraday cupFaraday cupPhotographic platesPhotographic platesScintillation type Scintillation type

Other:

Amplifying signalAmplifying signal

Continuous Dynode electron multiplierContinuous Dynode electron multiplier

Provide high gain and nanosecond response time.

Mass SpectrumMass SpectrumIonization: Ionization:

CHCH33OH + OH + ee-- CHCH33OHOH++ + + 2e2e--

m/z 32m/z 32

Molecular ionMolecular ion

Fragmentation: Fragmentation: [CH[CH33OH]OH]++ [CH[CH22OH]OH] + + + H + H

CHCH33++ + HO + HO

m/z 15m/z 15

m/z 31m/z 31

CHOCHO++ + H+ H22

B B Base Peak Base Peak 100% 100%

Tabular presentationTabular presentationBar graph presentationBar graph presentation

m/z 29m/z 29

C

O

NH2

105

77 44

- NH- NH22 CC66HH55C=O C=O ++

m/z 105m/z 105

- CO- CO CC66HH55++

Benzamide: EIBenzamide: EIB+B+

MM++

m/z 77m/z 77

3-methyl-6-3-methyl-6-i-i-PropylCyclohex-2-ene-1-onePropylCyclohex-2-ene-1-one

MW = 152MW = 152

Isotopic cluster Isotopic cluster MM, , M+1M+1, , M+2M+2

11HH 1.0081.008 1.007831.00783 99.9999.9922HH 2.014102.01410 0.0160.016

Natural Abundance of a few elementsNatural Abundance of a few elements

IsotopeIsotope Atomic weight Atomic weight MassMass % Abundance% Abundance

1212CC 12.01112.011 12.0000 (std)12.0000 (std) 98.8998.891313CC 13.0033613.00336 1.11 1.11

1414NN 14.006714.0067 14.0031 14.0031 99.6499.641515NN 15.000115.0001 0.36 0.36

1616OO 15.999415.9994 15.9949 15.9949 99.7699.761717OO 16.999116.9991 0.04 0.041818OO 17.999217.9992 0.20 0.20

1919FF 18.99818.998 18.998418.9984 100.0100.0



2828SiSi 28.085528.0855 27.9769 27.9769 92.1792.172929SiSi 28.976528.9765 4.71 4.713030SiSi 29.973829.9738 3.12 3.12

3232SS 32.06632.066 31.9721 31.9721 99.6499.643333SS 32.971532.9715 0.76 0.763434SS 33.967933.9679 4.20 4.20

3535ClCl 35.452735.4527 34.9689 34.9689 75.7775.773737ClCl 36.965936.9659 24.2324.23

3131PP 30.973830.9738 30.9738 30.9738 100.0100.0



7979BrBr 79.909479.9094 78.9183 78.9183 50.5250.528181BrBr 80.916380.9163 49.4849.48

126126II 126.9045126.9045 126.9045 126.9045 100.00100.00

The Atomic weight:The Atomic weight: Average atomic weight of all isotopes with their Average atomic weight of all isotopes with their natural abundances natural abundances

Atomic weight Atomic weight BrBr: : [% [% 7979Br Br x x 78.918378.9183] + [% ] + [% 8181Br Br x x 80.916380.9163]]

[[50.52 50.52 x x 78.918378.9183] + [] + [49.48 49.48 x x 80.916380.9163]] == 79.9094 79.9094


Molecular formulaMolecular formula MM++ ( (molecular ionmolecular ion) ) m/zm/z

MM mass of the most abundant isotope

e.g Ce.g C77HH77NO :NO : 7 x 7 x 1212C = 84C = 847 x 7 x 11H = 7H = 71 x 1 x 1414N = 14N = 141 x 1 x 1616O = 16O = 16

m/z m/z 121 121

Isotope peaks: (% respect to M)Isotope peaks: (% respect to M)

M+1 due to (M+1 due to (1313C) or C) or 1515NN ((1717O) or O) or 22H are negligeable)H are negligeable)

%(M+1) = (1.1 * #C) + (.38 * #N) = 8.08 %%(M+1) = (1.1 * #C) + (.38 * #N) = 8.08 %

%(M+2) = (1.1 * #C)%(M+2) = (1.1 * #C)22/200 + (.2 * #O) = 0.5%/200 + (.2 * #O) = 0.5%

For C, H, N,O, F, P compositionFor C, H, N,O, F, P composition

Molecular formula in generalMolecular formula in general

Even Mass : Even Mass : can containcan contain C, H, O , halogen,C, H, O , halogen, even # N even # N

Molecular ion:Molecular ion:

Odd Mass :Odd Mass : can containcan contain C, H, O , halogen,C, H, O , halogen, odd # Nodd # N

Fragmentation ions:Fragmentation ions:

Even MassEven Mass : From even mass M : From even mass M++ comes from rearrangement or 2 bond breakingcomes from rearrangement or 2 bond breaking

even Masseven Mass :: from Odd Mass Mfrom Odd Mass M++ comes fromcomes from single bond breakingsingle bond breaking

Odd MassOdd Mass : From even mass M : From even mass M++ comes from single bond breakingcomes from single bond breaking

Molecular formula: Molecular formula: RULE of 13RULE of 13

Consider CHCH unit 1313 amu

If we divide the mass by 13divide the mass by 13 we can establish easily possible formula:

Example: Molecular ionMolecular ion =>=> 152152

152 152 / 13 = / 13 = # carbons# carbons = = 1111

Mass = 12 * 11 = 132 therefore H = 152 – 132 = 20

Basic formula Basic formula CC1111 HH2020

If OxygenOxygen is present : mass = mass = 1616 remove CHremove CH44

If NitrogenNitrogen is present : mass = mass = 1414 remove CHremove CH22

If one oxygen : CC1111 HH2020 – – CCHH44 + + OO = = CC1010 HH1616 OO

If second oxygen : CC1010 HH16 16 OO – – CCHH44 + + OO = = CC99 HH1212 OO22

If third oxygen : CC99 HH12 12 OO22 – – CCHH44 + + OO = = CC88 HH88 OO33

Molecular formula: Molecular formula: RULE of 13RULE of 13

After establishing the basic formula with only Carbon/hydrogen,Other element can be introduced by substracting the proper hydrocarbonvalue

1616OO => => CHCH44

1414NN => => CHCH22

1919FF => => CHCH77 11HH1212 => => CC2828SiSi => => CC22HH44

3131PP => => CC22HH77

3232SS => => CC22HH88

3535ClCl => => CC22HH1111

7979BrBr => => CC66HH77

127127II => => CC1010HH77

Nitrogen ruleNitrogen rule

Molecular ionMolecular ion =>=> 2626

26 26 / 13 = / 13 = # carbons# carbons = = 22

Mass = 12 * 2 = 24 therefore H = 2


C C HH

Molecular ionMolecular ion =>=> 27 odd!27 odd!

27 27 / 13 = / 13 = # carbons# carbons = = 22


One One NitrogenNitrogen CC22 HH3 3 - - CCHH2 2 + + NNC NH


Molecular ionMolecular ion =>=> 100100

100 100 / 13 = / 13 = # carbons# carbons = = 77

Mass = 12 * 7 = 84 therefore H = 16


1 1 OxygenOxygen: : CC77 HH16 16 - - CCHH4 4 + + OO

CC66 HH12 12 OO

Molecular ionMolecular ion =>=> 99 odd!99 odd!


1 1 NitrogenNitrogen: : CC77 HH16 16 – – CCHH2 2 + + NN

CC66 HH14 14 NN

OH NH2

Calculating M+1M+1 and M+2M+2

Isotope peaksIsotope peaksUsually [M+2] peak is very smallUsually [M+2] peak is very small

Except for:

Sulfur : Sulfur : 3232S : 100S : 100 3434S : 4.4S : 4.4

Chlorine : Chlorine : 3535Cl : 100Cl : 100 3737Cl : 32.5Cl : 32.5

Bromine : Bromine : 7979Br : 100Br : 100 8181Br : 98Br : 98

Silicon : Silicon : 2828Si: 100Si: 100 2929Si: 5.2 Si: 5.2 3030Si : 3.35Si : 3.35

MM M+1 M+1 M+2 M+2

Isotope peaks : CHIsotope peaks : CH33BrBr

[[1212CHCH337979Br]Br]++

[[1212CHCH338181Br]Br]++

[[1212CHCH227979Br]Br]++

[[1212CHCH228181Br]Br]++

[[1313CHCH337979Br]Br]++

M - HM - H

[[1313CHCH338181Br]Br]++

BromineBromine

2 intense peaks for the molecular ion, spaced by 2 daltons.

7979BrBr and 8181BrBr

7979BrBr

7979BrBr 8181BrBr

8181BrBr

ChlorineChlorine

3535Cl Cl (P=(P= .75 .75))3737Cl Cl (P=(P= .25 .25)) RatioRatio 3: 3:11

Probability : Probability : [M[M++]] / / [M + 2][M + 2] = = 0.750.75 / / 0.250.25 = = 33 / / 11

100100 : : 3333

Calculation of isotope pattern: 2 ClCalculation of isotope pattern: 2 Cl

1 Cl1 Cl

2 Cl2 Cl

CC22HH223535ClCl22

CC22HH223535Cl Cl 3737ClCl

CC22HH22 3737ClCl22

P (P (2 2 3535ClCl) = () = (0.750.75 ) )22 = 0.563 = 0.563

P (P (3535Cl Cl 3737ClCl) + P () + P (3737Cl Cl 3535Cl Cl ) = () = (0.750.75 ) ( ) (0.250.25 ) + ( ) + (0.250.25 ) ( ) (0.750.75 ) = 0.375 ) = 0.375

P (P (2 2 3737ClCl) = () = (0.250.25 ) )22 = 0.063 = 0.063

[M[M++]] / / [M + 2][M + 2] / / [M + 4][M + 4] = = 100100 / / 6666 / / 1111

CC22HH223737ClCl 3535ClCl

Calculation of isotope pattern: ClBrCalculation of isotope pattern: ClBr

3535Cl => .75Cl => .75 3737Cl => .25Cl => .25

7979Br => 51%Br => 51% 8181Br => 49%Br => 49%

[[RRClBr]ClBr]++ can exist as 4 isotopic forms: can exist as 4 isotopic forms:

3535Cl Cl 7979BrBr MM 0.750.75 x x 0.51 0.51 = 38%= 38%

3535Cl Cl 8181BrBr M+2M+2 0.750.75 x x 0.490.49 = 37%= 37%

3737Cl Cl 7979BrBr M+2M+2 0.250.25 x x 0.51 0.51 = 13%= 13%

3737Cl Cl 8181BrBr M+4M+4 0.250.25 x x 0.490.49 = 12%= 12%

50%50%

At very high resolution, the 2 (M + 2M + 2) peaks can be distinguished (separated by 0.001 Dalton0.001 Dalton)

Chlorine and Chlorine and BromineBromine

The Molecular ionMolecular ion MM++ is always the lowest mass peak in the ion cluster (regardless of it’s relative intensity)

Isotopic abundances for Carbon containing compoundsIsotopic abundances for Carbon containing compounds

MM++ => p(M) = p( => p(M) = p(nn 1212CC) = () = (0.9890.989))nn

[M+1][M+1]++ => p(M+1) = p( => p(M+1) = p(nn -1-1) ) 1212CC + + 1 1 1313CC = n p[= n p[n-1n-1] ] 1212C C x p(x p(1313CC))

The probability of finding 1 1 1313CC among n n carbons is:

= n(= n(0.9890.989))n-1n-1 ((0.0110.011))

Relative ratio: [M+1][M+1]++ / / [MM++ ] ] == nn((0.9890.989))n-1n-1 ((0.0110.011) / () / (0.9890.989))nn

== n n ((0.0110.011) / () / (0.9890.989))

== n n ((0.01110.0111))

In percentage: n n x x 1.1 %1.1 %


When comparing calculated abundancescalculated abundances with observed intensitiesobserved intensities:

We see that the ratio is not identicalratio is not identical: due to experimental errorexperimental error

The relative abundance of the larger peaks are reproducible to ±±10%10%

For smaller peaks, relative intensity is larger

Generally speaking, The size of the [M+1][M+1]++ ion can be used to figure out the number of carbonsnumber of carbons in a molecule

For example, if molecular ion is observed at m/z 118m/z 118

and the ion at m/z 119m/z 119 has an intensity of about 9%9%

There are probably 8 carbons8 carbons ((1.1%1.1% x x 88 = = 8.8%8.8% ) )


Due to the relatively large error on peak intensity ( ±±10%10%) determination of the number of carbons is sometime ambiguous

For example: [M+1][M+1]++ => intensity=> intensity 22.522.5 ± ± 2.3%2.3%

Range covers 20.220.2 to 24.824.8 => 20 ± 2 carbons

Isotopic abundances for other common nucleiIsotopic abundances for other common nuclei

For For 1515N: [M+1]N: [M+1]++ / [M] / [M]++ => n x 0.36% => n x 0.36%

For For 3333S: [M+1]S: [M+1]++ / [M] / [M]++ => n x 0.80% => n x 0.80%

For For 1818O: [M+2]O: [M+2]++ / [M] / [M]++ => n x 0.20% => n x 0.20%


Calculating Peak Intensities from Isotopic abundancesCalculating Peak Intensities from Isotopic abundances

Isotopic abundances for SiliconIsotopic abundances for Silicon

M+2 larger than usualM+2 larger than usual

Often observed is TMS compounds

Often observed either from GC columnOr septum

Isotopic abundances for SulfurIsotopic abundances for Sulfur

M+2M+2

For For 1818O: [M+2]O: [M+2]++ / [M] / [M]++ => n x 0.20% => n x 0.20%


[M+2][M+2]++ = = 4.42 4.42 + (+ (2 2 xx 0.20 0.20) = ) = 4.82%4.82%

SOSO++

M+2M+2

isotope pattern: 2 Cl (example)isotope pattern: 2 Cl (example)

When there are many carbons, isotopic pattern for 13C adds up to chlorine pattern

Steps in the Identification of UnknownSteps in the Identification of Unknown

• Identify Molecular ion MM..++

• Determine Molecular FormulaMolecular Formula (oddodd / eveneven mass)

• Analyze heteroatomheteroatom (M+1M+1 and M+2M+2 …)

– S, Si, Cl, Br, ….

• Use rule of 13 to determine # Carbons (M+1M+1 and M+2M+2 …)

• Compare with 1313C-NMRC-NMR (# carbons(# carbons) with APTAPT experiment (J-MOD) ( # protons# protons)

• Compare with proton NMRproton NMR ( # protons# protons)

• Identify base peakbase peak (note if eveneven / oddodd)

– One or two bond fragmentation

• Test your conclusions: in lab make derivatives (TMS … or Na or K complexes mass shift)

Solving problems in MSSolving problems in MS

• Try to identify the Molecular Ion or decide if it is present (most critical step in solving a structure)– Check if [M+1]+ ion is too large to accommodate reasonable number of

carbons. (the [M+1]+ 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

Solving problems in MSSolving 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 deficiencyIndex of Hydrogen deficiency

CCxxHHyyNNzzOOnn IndexIndex = x – = x – ½ y½ y + + ½ z½ z +1+1

Neutral losses and Ion series Neutral losses and Ion series

M-1M-1 HH

M-15M-15 CHCH33

M-16M-16 O O (rare) , NH(rare) , NH2 2

M-17M-17 OH OH , NH, NH33 (rare) (rare)

M-18M-18 HH22O O

M-19M-19 F F

M-20M-20 HF (very rare)HF (very rare)

M-26M-26 HCCH , CNHCCH , CN

M-27M-27 HCNHCN

M-28M-28 HH22C=CHC=CH22 , CO , CO

M-29M-29 CHCH33CHCH2 2 , , HCO HCO

M-30M-30 NO NO (Nitro compounds), (Nitro compounds), HH22 CO CO (anisoles)(anisoles)

M-31M-31 CHCH33O O

M-32M-32 CHCH33OH OH

M-35M-35 Cl Cl

M-36M-36 HCl HCl

M-42M-42 CHCH22=C=O, CH=C=O, CH22=CH-CH=CH-CH33

M-43M-43 CHCH33CO CO , C, C33HH77

M-44M-44 COCO22

M-45M-45 CHCH33CHCH22O O , CO, CO22H H

Neutral losses and Ion series Neutral losses and Ion series Figuring out which peak is molecular ionmolecular ion can be supported by identifying what fragment is lost.fragment is lost.

There can be sometimes 2 consecutive loss:

In steroid, M-33M-33 is often observed: comes from the loss of Meloss of Me and HH22OO

The ions lossions loss are only useful from molecular ionmolecular ion

There is no fragment in organic compounds between M-1M-1 and M-15M-15

Loss of M-14M-14 is never observed!

Other gaps in mass loss are: between 21-2521-25, 33-3433-34, 37-4137-41

IonsIons in these areasin these areas should be viewed suspiciouslyviewed suspiciously: either compound is not pure or postulated molecular ion is wrongmolecular ion is wrong

Neutral losses and Ion series Neutral losses and Ion series

Among the losses: most common are

Loss of HH, , CHCH33 , , HH22OO (from some oxygenated compounds), (from some oxygenated compounds),

HCHCCHCH (from aromatic compounds), (from aromatic compounds),

HCHCN N (from aromatic compounds containing Nitrogen), (from aromatic compounds containing Nitrogen),

CCOO and and CHCH22=CH=CH22 (both at 28! Difficult to tell which one is lost) (both at 28! Difficult to tell which one is lost)

EthylEthyl radical ( radical (2929))

MethoxyMethoxy radical ( radical (3131))

ClCl and and HClHCl ( (3535, , 3636))

AcetylAcetyl ( (4343) accompanied by ) accompanied by m/z 43m/z 43 prominent and prominent and propylpropyl ( (4343) radical) radical

ResolutionResolution

Exact Masses Exact Masses of isotopesof isotopes

Mass DefectsMass DefectsBy definition the atomic weight of 1212C is 12.0000 DaltonsC is 12.0000 Daltons

All other elements are determined relative to 1212C C

Their difference to nearest integral is called Mass DefectMass Defect

Light element has small Light element has small positive mass defectpositive mass defect

Vast majority of elements Vast majority of elements have substantial negative have substantial negative mass defectmass defect

Mass DefectsMass DefectsThe Mass DefectMass Defect of a CC2323HH3535OO22SiSi is shown here on a “low resolution” MS (+/- 0.1)

Mass defect for HH : 1.008-1.000 = .0081.008-1.000 = .008

Mass defect for OO : 15.995-16.000 = -.00515.995-16.000 = -.005

Mass defect for SiSi : 27.97727.977 -28.000 = -.003-28.000 = -.003

((35 35 xx 0.008 0.008) + () + (2 2 x x -.005-.005) + () + (-.003-.003) = ) = .27.27

Mass defectMass defect for organic compounds is mainly due to Hydrogenmainly due to Hydrogen due to their large number

Mass DefectsMass Defects

Mass defectMass defect brings the maximum of the CC2323HH3535OO22Si Si peak not at 371371 but at 371.25371.25

If the Mass spectrometer measure the mass with a resolution of 1resolution of 1, the sensitivity is reduced.

Greatest application of Mass Defect is surely very high resolution, which provide mass with .001 to .0001 accuracy

High resolution:High resolution:

With sufficient accuracy, unique molecular formula can be determineWith sufficient accuracy, unique molecular formula can be determine

e.g. Distinguish COCO, NN22, CHCH22NN and CC22HH44 (all having m/z 28m/z 28)

COCO 1212C C 12.000012.00001616O O 15.994915.9949

27.994927.9949

NN221414N N 14.003114.00311414N N 14.003114.0031

28.006228.0062

CHCH22NN1212C C 12.000012.000011H x2 H x2 2.0156 2.0156 1414N N 14.0031 14.0031

28.018728.0187

CC22HH441212C x2 C x2 24.000024.000011H x4 H x4 4.0312 4.0312 28.031228.0312

Exact Mass can provide Molecular FormulaExact Mass can provide Molecular Formula

Exact Mass can provide Molecular FormulaExact Mass can provide Molecular Formula

Consider the following 2 formulas which have m/z 287m/z 287:

CC1515HH1010NONO33ClCl CC1414HH88NN22OO33ClCl

Mass Defect of Mass Defect of CC1515HH1010NNOO33ClCl::

((10 10 x x .0078.0078) + (.) + (.00310031) + () + (3 3 x x -.0051-.0051 ) + ( ) + (-.0311-.0311 ) = ) = .0347.0347

Mass Defect ofMass Defect of CC1414HH88NN22OO33ClCl::

((8 8 x x .0078.0078) + () + (22 x . x .00310031) + () + (3 3 x x -.0051-.0051 ) + ( ) + (-.0311-.0311 ) = ) = .0202.0202

With sufficiently high resolution MS, it is possible to propose a unique With sufficiently high resolution MS, it is possible to propose a unique empirical formula for an ion.empirical formula for an ion.

Index of Hydrogen deficiencyIndex of Hydrogen deficiency


MM+ + even even even # of N 0, 2, 4, … even # of N 0, 2, 4, …

MM+ + odd odd odd # of N 1, 3, 5, … odd # of N 1, 3, 5, …

Index = C – H/2 –X/2 + N/2 + 1Index = C – H/2 –X/2 + N/2 + 1

e.g. Ce.g. C77HH77NO NO Index = 7 - 3.5 + 1 = 5 Index = 7 - 3.5 + 1 = 5

Hydrogen deficiency can be unsaturation (multiple bonds)Hydrogen deficiency can be unsaturation (multiple bonds) or cyclic structure or cyclic structure

I=4 (3 DB + cycle)I=4 (3 DB + cycle)

R-CN

I=2I=2I=1 (1 DB + cycle)I=1 (1 DB + cycle)

Index of hydrogen deficiency : GeneralIndex of hydrogen deficiency : General

Index = Index = IVIV – ( – (II/2) + (/2) + (IIIIII/2) +1/2) +1

IVIV any tetravalent atom (e.g. any tetravalent atom (e.g. CC, , SiSi..)..)

II any monovalent atom (e.g. any monovalent atom (e.g. HH, , DD,,ClCl, , BrBr, , FF …) …)

IIIIII any trivalent atom (e.g. any trivalent atom (e.g. NN, , PP, …), …)

IIII any divalent atom (e.g. any divalent atom (e.g. SS, , OO,…). ,…). These count for 0 in the formulaThese count for 0 in the formula

Higher valences uses valence shells Higher valences uses valence shells lewis octet rulelewis octet rule

CHCH33--SS-CH-CH33

OO

++

-I = 0I = 0

CHCH33--NNOO

O-O-

+

I = 1I = 1

phosphinephosphineCHCH33--PP-CH-CH33+

OO-

CHCH33

I = 0I = 0

Fragmentation in different isomersFragmentation in different isomers

Molecular ion almost absentMolecular ion almost absent

Ionization TechniquesIonization Techniques

Electron Impact (EI)Electron Impact (EI)

Under these conditions, very energetic Ions are produced

=> FragmentationFragmentation MM..++ is often weak is often weake.g. ROH often loose H2O

Intensity of MM..++ Depends on the stability of the IonIon

Most StableMost StableAromaticConjugated AlkenesCyclic compoundsSulfidesShort Linear AlkanesMercaptans

Least StableLeast Stable

MM..++ is often absent in: is often absent in:

KetonesAminesEsterEtherAcid ~ Amides ~ Aldehydes

Alcohols,

Nitriles, Nitrates, Nitro, Nitriles {{

Other Ionization TechniquesOther Ionization Techniques

Under EIEI (Electron ImpactElectron Impact), the molecular ion may be weak or absent

In such case, the best solution is to run CI – Chemical IonizationCI – Chemical IonizationWhich result in intense M+1M+1 ion with little fragmentation

CI – Chemical IonizationCI – Chemical Ionization

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

CHCH44..++ + CH+ CH44 CHCH55

++ + CH + CH33..

CHCH33++ + CH+ CH44 CC22HH55

++ + H + H22

CHCH55++ + + MM [M+H][M+H]++ + CH + CH44

CC22HH55++ + + MM [M+H][M+H]++ + C + C22HH44

Less energy than CI => less fragment

• Secondary ionsSecondary ions react with sample MM:

Chemical Ionization techniquesChemical Ionization techniquesIn Electron Impact (EI) and chemical ionization,

vaporization of the Sample is a prerequisitevaporization 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 Desorption1- Field Desorption: ions are desorbed by strong electric field

2- SIMS :2- SIMS : other source of energy is a beam of ions (SSecondary IIon MMass SSpectroscopy): Typically Argon (FABFAB)

In this and other techniques,, the aim is to transfer energy to the Sample, causing transfer of molecules/ions to gas phase

Field DesorptionField Desorption methods give a high concentration of Molecular ionhigh concentration of Molecular ion=> Particularly useful for identification of unknown

Fast Atom Bombardment (Fast Atom Bombardment (FABFAB))

The sample is dissolved in a liquid matrixsample is dissolved in a liquid matrix such as glycerolglycerol, thioglycerolthioglycerol, m-nitrobenzyl alcoholm-nitrobenzyl alcohol, or diethanolaminediethanolamine and a small amount (about 1 microliter) is placed on a target.

The target is bombarded with a fast atom beamfast atom beam (for example, 6 keV 6 keV xenon atomsxenon atoms) that desorb molecular-like ions and fragmentsdesorb molecular-like ions and fragments from the analyte.

Cluster ions from the liquid matrix are also desorbed and produce a Cluster ions from the liquid matrix are also desorbed and produce a chemical background that varies with the matrix usedchemical background that varies with the matrix used.

XeXe ee--

XeXe++XeXe

XeXe++ Is accelerated to ~6-10 keV, and pass through XeXe

Xe + Xe + XeXe++XeXe (Xenon with Kinetic energy) FABFAB

FABFAB: Benefits: Benefits

Sample introduction can be through direct insertion probe or LC/MS (continuous-flow FAB).

BenefitsBenefits

• 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 measurementsgood for high-resolution measurements

FABFAB: Limitations: Limitations

LimitationsLimitations

• 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 rangeMass range

Moderate Typically ~300 Da to about 6000 Da~300 Da to about 6000 Da.

Comparing Comparing different different

techniquestechniques

Classification according to method of separating charged particuleClassification according to method of separating charged particule

1.1. MagneticMagnetic Field DeflectionField Deflection1. Magnetic Field only: Mass 12-500 in secondsMass 12-500 in seconds2. Double Focussing:

High resolution to 4 decimalsHigh resolution to 4 decimalsr = 60,000

• Quadrupole Mass SpectrometerQuadrupole Mass Spectrometer• Quadrupole Mass Filter: Mass Scanning by varying RF & DC

Frequencies• Quadrupole Ion Storage (Ion Trap)

- Compact-Inexpensive-Very Sensitive-GC/MS

Classification according to method of separating charged particuleClassification according to method of separating charged particule

3.3. Time of FlightTime of Flight- Need Fast Electronic (10-7 s)- Used With Sophisticated Ionization Methods (FAB, Laser Desorption ….)

4.4. FT-ICR (Fourier Transform Ion Cyclotron FT-ICR (Fourier Transform Ion Cyclotron Resonance)Resonance)- Very High Precision – most expensive

5.5. MS/MS (Tandem Mass Spectro.)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

Time-of-Flight analyzer (TOF)Time-of-Flight analyzer (TOF)

In TOFTOF instruments, positive ionspositive ions are produced periodically using

brief pulses of electronsbrief pulses of electrons,

secondary ionssecondary ions or

laser generated photonslaser generated photons

These pulses have typically a frequency of 10-50 kHzfrequency of 10-50 kHz and lifetime lifetime 0.250.25ss

The ionsions produced are then acceleratedaccelerated by electric field pulseelectric field pulse (103-104 V) that has the same frequencysame frequency as ionization pulse but lags behindbut lags behind.

The acceleratedaccelerated ionsions then pass into a field-free drift tubefield-free drift tube (about 1 m long)

Time-of-Flight analyzer (TOF)Time-of-Flight analyzer (TOF)

MALDIMALDI ionization, TTime-oof-fflight MMass SSpectrometer

Tandem-MSTandem-MS

Comparing Mass Spectrum of a compound in Comparing Mass Spectrum of a compound in different type of mass spectrometersdifferent type of mass spectrometers

105105

MW = 164MW = 164

Magnetic sectorMagnetic sector QuadrupoleQuadrupole

Time of FlightTime of Flight

Organometallic compounds in MSOrganometallic compounds in MS

It is often possible to determine molecular weight of a compound by MS

For example: Manganese carbonylManganese carbonyl => m/z 390m/z 390

As MnMn => 5555 and C=OC=O => 28 28 MnMn22(COCO)1010

Another example: Iridium complex Ir P

PEt3

PEt3

Cl

ClCl

OC

Cl

NMR and IR can give a lot of information but withoutMS it is very difficult to show the presence of ClWith 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 phaseMight be very different from the one in gaz phase.

With FAB most intense cation peak is the protonation ion (M+1) peak. Anion (M-1) can also be formed.

Isotope Abundance patterns for some Isotope Abundance patterns for some atoms and group of atomsatoms and group of atoms

Can be diagnostic Can be diagnostic for some isotopesfor some isotopes

Abundances can be calculated by multiplying the abundances of theConstituent isotopes

Calculation of isotope patternCalculation of isotope pattern

e.g. 185185Re => 37%Re => 37% 187187Re => 63%Re => 63%

7979Br => 51%Br => 51% 8181Br => 49%Br => 49%

[ReBr][ReBr]++ can exist as 4 isotopic forms: can exist as 4 isotopic forms:

185185ReRe7979BrBr 264 m/z264 m/z 0.370.37 x x 0.51 0.51 = 19%= 19%

185185ReRe8181BrBr 266 m/z266 m/z 0.370.37 x x 0.490.49 = 18.1%= 18.1%

187187ReRe7979BrBr 266 m/z266 m/z 0.630.63 x x 0.51 0.51 = 32.1%= 32.1%

187187ReRe8181BrBr 268 m/z268 m/z 0.630.63 x x 0.490.49 = 30.9%= 30.9%

50.2%50.2%

Metastable ionsMetastable ions

Some ions have so short lifetimes that they dissociate while moving through the spectrometer.

•An ion of mass m1mass m1 is accelerated after initial ionizationinitial ionization •But a different ion m2different ion m2 (daughter iondaughter ion) passes the magnetic analyzermagnetic analyzer•The resulting peak is neither m1m1 or m2m2 but appear at m* (metastable)m* (metastable)

m*m* = (m2m2)2/m1m1

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

PlatinumPlatinum

TungstenTungsten

Index MS-fragmentationMS-fragmentation

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Chem-805 Identification of organic and inorganic compounds by spectroscopy Mass Spectrometry NMR ...

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