UV-Vis and Mass Spectrometry Flashcards
UV/Vis Spectrophotometry
An instrument for irradiating a sample with photons of UV or vis light of particular wavelength/s
Must be run against a sample blank, (same solution but no drug)
Liquid sample dispensed into a cuvette (solvent not abs. above 200nm) UV – quartz; vis – plastic, polystyrene
spectrophotometry measures either:
absorption - light not passing through sample transmission – light passing through sample
Molecular Orbitals (MO)
The region in which there is the highest probability of finding an electron of a molecule
Highest Occupied Molecular Orbital (HOMO):
Highest energy MO with any electrons in it
Lowest Unoccupied Molecular Orbital (LUMO):
The next highest energy orbital. It will be empty and so is the lowest energy orbital in which to place an electron or excite one into.
Ultraviolet Absorption Spectroscopy
Wavelengths of UV light absorbed by a molecule are determined by the electronic energy differences between the HOMO and LUMO.
Absorption is only possible if the molecule contains ____ ________ levels separated by energy (Transition from bonding to anti-bonding orbitals, 𝜋 to 𝜋, 𝜋 to σ
two molecular
Ultraviolet Absorption Spectroscopy - The molecule will now be in what state?
after energy will dissipate rapidly and return to what state?
excited
stable
graph for UV energy
electronic excited states
AE = (E excited - E ground) = hv
Lambhda max (A max) =
wavelength showing the greatest absorbance
What is Beer’s Law?
states that the absorptive capacity of a dissolved substance is directly proportional to its concentration in a solution
Absorbance is what we measure and has arbitrary units
The molar absorption coefficient is a standard
The concentration is in moldm-3
The path length is the width of the cuvette (normally 1 cm)
Beer’s Law equation
A = ecl
A - absorbance
e - molar absorption coefficient M-1cm-1
c - molar concentartion M
l - optical path length cm
How to interpret UV-Vis Spectra?
- Wavelength(s) of maximum absorbance
- Molar absorptivity e at each maximum
Explain the relationship between aromatic compounds and absorption of UV
highly conjugated
absorb strongly in UV
UV-Vis = Active Compounds
Chromophores - structural features pf a molecule that absorb specific wavelengths ( e.g. conjugation)
Auxochromes
auxiliary groups attached to a chromophore, unable to absorb, cause (often substantial) shifts
resonance
different electronic configurations of same structure
inductive effects
groups feeling in or pulling out e-
example: Primaquine free base ( what colour)
yellow liquid
primaquine diphosphate
red solid, due to bathochromic shift caused by auxochromes
Coloured Organic Compounds
white object reflects all of the light
coloured object absorbs some wavelengths and reflects the rest of the spectrum
colour we see - complete visible - absorbed portion
Mass Spectrometry
Provides Molecular Weight
Provides valuable information about the molecular formula
Does not use light (different from spectrometry)
Sample is destroyed
High-energy electrons fragment the molecules
Mass of the fragments is measured
Information used to “reconstruct” the molecule
Mass Spectrometry - step by step
- Negatively charged accelerator plate with narrow slit to allow ions to pass through
- Charged particle passes through magnetic field (bends path) – Curvature of the bend depends on the m/z ratio (mass / charge)
- Slit followed by detector – at any given magnetic field, only ions of one particular mass are bent exactly the right amount to pass through the detector. The detector signal is proportional to the number of ions striking it
Peaks are typically very sharp - often vertical lines
masses rounded to the nearest whole-number mass unit
Base peak does not necessarily correspond to the mass of the molecular ion (M+)
often very small peaks close to expected MW - isotope peaks
molecular ions (M+)
usually the particles of highest MW in the spectrum
m/z value - give the MW of the compound
if no M+ observed: use a gentle ionization
Determination of Molecular Formula by MS
look at masses - exact mass
molecular ion peaks (M+) what do they provide
information about the molecular formula
most elements contain:
heavier isotopes in varying amount
Heavier isotopes: small peaks at higher mass numbers than the M+ ion peak
M + 1 (ones mass unit heavier M+); M + 2 (two mass unit heavier M+)
Determination of Molecular Formula by MS
use of HEAVIER isotope peaks - recognisable elements in the mass spectrum
Compound with no S, Cl or Br: M + 1 peak small and M+2 peak non visible
Compound containing S: M+2 (34 S) larger than usual
(4% of M+ 33S)
Compound containing Cl:L M+2 (37Cl) 1/3 as
large M+ (25Cl)
Compund containing Br: M+2 (81 Br) as large as
M+ (79Br)
Compound containing I:
I+ iodonium ion at 127; large gap
Compound N:
odd M+ some even-numbered fragments
Stable compounds (C H O )
even M+, odd numbered fragments
Fragmentation pattern in MS
Fragmentation splitting out a small molecule
loss of small stable molecules:
indicated by a fragment peak with even mass numbers
