现代化学研究方法与技术—2

HO HO
HO
O O
OH
O OH
O
HO
O
N
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HO
CN
O
OH
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2,5-dihydrobenzoic acid, sinapinic acid, nicotinic acid, a-cyano-4-hydroxycinnamic acid
etc..
MALDI is typically used in conjunction with time-of-flight analyzer and can be used to measure the molecular weights of proteins in excess of 100,000 daltons.
1-2. Mass Spectrometry
Refs: Handbook of Spectroscopy (Wiley-2003)
1-2-1 Introduction
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What we can get from the MS?
1. Accurate mass measurements can be used to match empirical formulae.
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Instrumentation:
All mass spectrometer systems
have 5 main components
Sample introduction
Ionization (source)
Ion Analyser
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Ion Detection
> [AH+H]+ + A- - 'self chemical ionization'
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a * superscript are in high energy sates.
a • superscript are radicals.
Species with a " superscript are short lived intermediates which are not seen in the spectra.
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Field Desorption (FD)
FD is another technique used for involatile compounds. FD works best for compounds of low polarity, and thus complements FAB. FD uses the same emitter wires used in FI, but the samples are loaded directly on the wire by dipping into a solution of the sample. The sample for FD must be soluble in some solvent (usually 2 mg in 100 microliter).
R + MH (R-2H) + MH
R + (M-H)
R+ M
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The CI spectra of lavanduyl
acetate using
methane, isobutane
and
ammonia.
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11Field Ioniza Nhomakorabeaion (FI)
There are three practical differences between CI and FI: there is less fragmentation in FI, there is no high-resolution FI, and FI is less sensitive. Sensitivity is not an issue unless there is an extremely small amount of sample.
4. Common peaks observed in a spectrum can give useful information regarding functional groups.
5. Relative isotope abundance's are used to get information regarding the elements making up a compound.
when glycerol as a
matrix.
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FAB Ionization Mechanism
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Matrix-assisted Laser Desorption/Ionization (MALDI)
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Matrixes commonly used in MALDI
Display/ Analysis
4
Ionization
Hard and Soft
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Electron ionization or
Electron impact
ionization (EI)
Mechanisms of ion formation
Consider the ionization of the analyte species AB: 1. AB + e-* -----> A+ + B- + e 2. AB + e-* -----> A+ + B·+ 2e 3. AB + e-* ---> [AB+·*] + 2e-followed by [AB+·*] --->
6. Complex mixtures can be analysed via 'hyphenated' techniques such as GC-MS and HPLC-MS,
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What kind of Mass spec (MS) samples can be run?
a) pure organic or inorganic compounds b) complex mixtures e.g. environmental
samples c) biological samples e.g. proteins, nucleic
acids, blood,urine, whole cells, (hot new area “proteomics” - use of MS to study total protein content of a cell).
1 and 2 are the highest abundance and are termed instantaneous fragmentation. This is the reason why EI is considered a hard ionization process.
3 is fairly high abundance and is the process responsible for the molecular ion formation. Unfortunately the highly energetic radical intermediate [AB+°*] tends to undergo fragmentation (or rearrangement) as a stabilizing process, this is responsible for the lower mass fragment ions present in the spectra.
2. Fragmentation fingerprints (specific to each compound) can be used to identify samples by comparison to fragment databases.
3. Controlled fragmentation (through MS/MS and MSn) can be used for structural elucidation of novel compounds.
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Fast Atom Bombardment (FAB)
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FAB is comparatively soft ionization technique, and is thus well suited to the analysis of low volatility species, typically producing large peaks for the pseudomolecular ion species [M+H]+ and [M-H]-, along with structurally significant fragment ions and some higher mass cluster ions and dimers
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4 is a very low abundance process, but theoretically it can occur.
5 is a process which can occur at higher pressures (self Chemical Ionization), this is especially problematic in the ionization of alcohols and amines where you may find that the dominant ionization process is proton exchange between two analyte molecules, leading to the formation of the [M+H]+ pseudo-molecular ion.