CHMC11
What samples can be studied by FT-IR?
Practically all materials have some vibrational absorption Solids, liquid films, bulk solutions, and gas can be studied with FT-IR
How do you determine the molecule with IR spectrum?
Produce the spectrum, run against database, find specific region spectrum peaks
What does a large transition energy change mean?
The larger the E change the more the population between the higher and lower states will vary, leading to increased sensitivity
What are wavelengths for IR?
780 nm to 300 000 nm (0.78 -300 um)
What is the Beer's Law equation?
A = Ebc
Define chromophore
Light-Absorbing compounds or molecules that are attractors of specific wavelengths
What compounds and at what temperature are in the cryomagnet of an NMR?
Liquid He: 4 K (-270 C) Liquid N2: 77 K (-196 C)
longest visible light wavelength
red
What is UV-VIS spectroscopy good and not good for?
good for: quantitative analysis (seeing how much of a given species is in a sample) not good: qualitative (identify components)
What causes a chemical shift?
Chemical shift is caused by small magnetic fields generated by electrons (electron cloud) as they circulate around nuclei• These fields oppose the applied field (B0). • We call this opposition by a cloud of electrons shielding Electron density around each nucleus in a molecule varies according to the types of nuclei and bonds and this causes the frequency at which they resonate to change or shift
On what basis does the IR Spectroscopic Process works?
19An infrared spectrometer is a device that scans through the infrared part of the electromagnetic spectrum and measures the amount of radiation absorbed by the sample at each frequency Normally most covalent bonds stretch and bend naturally. This causes a change in a spatial dipole (i.e. charge changing over space) This changing dipole moment causes an electromagnetic field to be generated If the frequency of the IR radiation exactly matches the natural vibrational frequency of the molecule, absorption of the radiation takes place changes the amplitude of the molecular vibration
What nuclei types are NMR active?
1H, 13-C
Whsat are the boundaries of visible light?
380-780 nm
What are two main types of UV-VIS detectors?
1) Scanning UV-VIS 2) Diode Array Detector
What are the advantages of Fourier Transform Infrared Spectroscopy (FT-IR)?
1. All frequencies are scanned at once so no time is wasted trying to scan individual frequencies separetely 2. Multiple "scans" can be done at once over a very short period of time giving an increased "signal to noise" (better resolution) 3. Prisms and diffraction gratings are not needed to filter the light before it hits the sample. (prisms and diffraction gratings are often inefficient- remove unwanted frequencies but reduce the intensity of the selected frequency as well)
List ways to measure chemical interactions by NMR- the conventional methods
1. Chemical Shift Perturbations 2. Relaxation Studies (very long, but often used because it's easiest to perform) 3. Saturation Transfer Difference 4. Diffusion Order Spectroscopy (DOSY) 5. Studies involving Overhausser Effects
What NMR can be used to answer the question: What components in a mixture is my contaminant bound to and what bonds are touching the "receptor" ?
3D DOSY-NOESY
What are the differences between Hydrogen and Carbon NMR spectra?
1. There is no splitting in carbon spectra. • In order to get splitting you would have to have a 13C nucleus neighbouring another 13C nucleus. • As only 1 in 100 atoms are 13C, the chances of getting two adjacent 13C nuclei is 1/100 * 1/100 = 1/10,000. • Therefore 13C couplings are too weak to appear in 13C NMR spectra 2. The chemical shift range for crabon spectra is much greater than for H NMR 3. The signal in carbon NMR are often "sharper" and there is less likely to be overlap in very complex molecules
What are the boundaries of UV light?
10-380 nm
How much is 1 ppm in Molarity?
1 mg/L
How does diffusion measurement NMR work?
1. DEPHASE: (signal pules) gradient magnetic field is applied in addition to the standard NMR magnetic field 2. Waiting diffusion time for protein to change position: Molecules in a sample move at different rates due to differences in size and shape. The gradient field imparts a unique phase to the NMR signal of each molecule, depending on its diffusion rate (The large molecule essentially does not move position as it have very slow molecular self-diffusion) 2. REPHASE (redoing rephase; Signal with a pulse field gradient) Molecules that do not move position are completely refocused at the end of the experiment (i.e.signal is unaffected). Result is: Signal from large molecule = unaffected; Small molecule = strongly attenuated 3. Read-out signal at the end The larger the gradient the smaller the distance between the "slices" in the NMR tube. Therefore by ramping the gradient strength over a number of experiments we can estimate how far different molecules move in the tube and therefore their diffusion coefficients.
How can we use NMR spectroscopy in Organic, Inorganic Chemistry & Biochemistry?
1. Determination of the structure of unknown compounds 2. Confirmation of the structure of synthesized compounds. F 3. Identifying molecular Interaction, binding etc. 4. Understanding the solution conformation and folding of proteins and DNA--> NOSY 5. Binding Sites for Glucose on Insulin---> NOESY
What are the types of limitations of Beer's Law?
1. Fundamental Beer's law is valid only for low concentrations of analyte (<0.01M) 2. Chemical A chemical deviation from Beer's law may occur for an equilibrium reaction HA + H2O--> H3O+ + A− At different concentrations the equilibrium may shift, if A- and HA both absorb then Beer's Law may deviate from linear 3. Instrumental Stray Radiation: Imperfections in instrument that allow light to enter the instrument and reach the detector without passing through the sample Monochromatic: Beer's law assumes that the radiation reaching the sample is of a single wavelength—that is, that the radiation is purely monochromatic.Even the best monochromators pass radiation with a small, but finite effective bandwidth
In 3 points explain how IR spectrometer works
1. Infrared radiation is emitted by a source. 2. Sample molecules absorb specific IR wavelengths. 3. A detector measures absorption, yielding a unique spectrum for analysis
List different IR sources
1. Nernst Glower (mid IR) 2. Globar (mid IR): heated SiC (silicon carbide) rod 3. Tungsten filament lamp (mid IR) 4. CO2 laser (mid IR): stimulated emission lines 9-11 μm
Name 3 IR sources and their properties
1. Nernst Glower (mid IR): heated rare earth oxiderod (yttrium oxide) (~1500 K) 1-10 μm2) 2. Globar (mid IR): heated SiC (silicon carbide) rod(~1500 K) 1-10 μm3) 3. Tungsten filament lamp (mid IR): (1100 K) 0.78-2.5 μm4) 4. Hg arc lamp (far IR): plasma >50 μm5) 5. CO2 laser (mid IR): stimulated emission lines 9-11 μm
What are advantages of IR spectroscopy?
1. Relatively cheap 2. Can provide great qualitative information 3. Relatively portable (TV remotes, and computers all have IR sources and detectors in them)
What can DOSY do, what is it useful for?
1. Tell us which component/components bind to a protein or partition between the free and bounds states with micelles of colloids (When drugs/contaminant bind to proteins/DOM there diffusivity will change and this can be monitored by DOSY NMR) 2. Can all provide information as to the major components in solution that interact/aggregate to form colloids
How to choose a correct container or solvent for FT-IR?
A medium that does not absorb strongly in the FT-Region should be used, e.g; KBr does not absorb strongly in the FT-IR region. Consequently it is often used in the construction of sample holders for FT-IR spectroscopy
How are gases tested in FT-IR?
Fill gas cell: • Transparent windows (NaCl/KBr) • Long path length (10 m) - few molecules
List advantages and disadvantages of Diode Array Detector
Advantages: 1. ideal for flow based detection (chromatography) 2. full spectra at 160 times per second (greater sensitivity) Disadvantages: 1. More expensive than Scanning 2. It is difficult to incorporate a reference cell inside DAD 3. Faster data- DAD can record full spectra instantly VS scanning takes 1 min for all wavelengths
What are advantages and disadvantages of ATR?
Advantages: • Faster sampling with no preparation. • Excellent sample-to-sample reproducibility • Minimal operator-induced variations. Disadvantages: Sensitivity of the technique can be low as the FT-IR energy may only penetrate some samples by a very small amount (1-2 μm)
The pi-system of alkenes, aromatic compounds and carbonyls strongly de-shield attached protons and move them "downfield" to higher ppm values - Why ?
Alkenes and aromatic compounds contain pi bonds (double bonds in alkenes and the delocalized pi electrons in aromatic rings). These pi electrons create regions of increased electron density in the vicinity of the protons. Carbonyl groups (like those in ketones and aldehydes) contain a highly electronegative oxygen atom bonded to carbon. This oxygen atom withdraws electron density from the adjacent carbon, which in turn deshields the protons on that carbon.
How are chemical shifts assigned?
All chemical shifts are expressed in relative terms. • The shift, is expressed in parts per million (ppm) versus some reference compounds. • This reference compound is normally tetramethylsilane (TMS), (CH3)4Si. • This peak is assigned a chemical shift of zero.• All other peaks are referenced to it.
What does ATR stands for? Why is it relevant?
Attenuated Total Reflectance It's a new modern approach for IR spectrometry where a sample is treated under pressure in a Diamond ATR crystal. An IR light source emits light onto the sample from which it is reflected to a detector
Why is carrot organe?
Because Beta carrotene absorbs blue and reflects orange
Why do some bonds absorb UV-VIS?
Bonds can absorb UV-VIS light when the energy of the incident photons matches the energy required to promote electrons from their ground state to an excited state. The majority of UV spectroscopy involves wavelengths longer than 185 nm Both n to π* & π to π* transitions are common in molecules that contain unsaturations (double bonds, aromatics, carbonyl, ester)
Out of CH3, CH2, CH, which group will have the highest chemical shift? Why?
CH will have the highest because
What peaks up and what down in the Distortionless Enhancement by PolarizationTransfer (DEPT) edited carbon spectrum?
CH3 and CH up CH2 down quaternary C don't appear
What solvents are used in NMR and why?
D2O (Deuterated Water), CDCl 3 (Deuterated Chloroform) This is because deuterium does not show up in the 1H NMR spectrum so it does not complicate the spectrum or hide peaks. Note it appears in carbon spectrum (see previous slide) but dissapears in the DEPT spectrum as no protons are attached The Dueterium in the solvent acts as a "lock". The NMRinstrument locks onto the known solvent frequency. This actsas a reference. This allows the NMR to adjust for changes inthe magnetic field during the experiment, so that theexperimental conditions stay constant
What two types of lamps are used in UV-VIS?
Deuterium Lamp (most light sources use a deuterium lamp to generate UV-VIS, 190-370 nm) Tungsten Filament (tungsten Filament with a little iodine added, Visible Region, 320-1100nm
What are light sources for UV-VIS spectroscopy and what are they properties?
Deuterium lamp (UV and VIS; 190-370 nm) and tungsten filament (VIS only, 320-1100 nm, can have iodine added)
What are two main types of UV-VIS detectors?
Diode array detector and scanning UV-VIS detector
What is the meaning behind "double beam instrument" in Scanning UV-VIS detector?
Double beam instrument refers to the design of most UV-VIS detectors that contain two lamps; one for solvent + sample one for solvent only
Provide equation of molar absorptivity
E = A / (cl) where l is the pathway length in cm c is concentration in mol/L A is absorbance E is in L/cmM
What is the effect of electron-withdrawing groups on the chemical shift?
Electronegative groups are "deshielding" and tend to move NMR signals from neighboring protons further "downfield" (to higher ppm values).
Give 3 examples of qualitative application of IR
Environmental Monitoring CO 2 levels in greenhouses, SO 2 emissions from factories, or volatile organic chemicals (VOCs) in the air. Finger Print Regions help identify compounds via library matches.Spectrometers are so small they can be handheld : • Rapid Compound identification anywhere • Easier and Cheaper for Security Screening Airports
What changes time domain signal innto frequency signal?
Fourier Transform
List regions of electromagnetic spectrum
Gamma < X-ray < UV < VIS <IR < mico < macro
Amplitude
Height of a wave/distance from rest to peak position
What are two general categories of 2D NMR?
Homonuclear Experiments - Interactions between nuclei of the same type. For example, interactions between protons• (e.g COSY, TOCSY) Heteronuclear Experiments - Interactions between nuclei of different type. For example, interactions between protons and carbons (e.g HSQC, HMBC) +specialized experiments: DOSY
How do free vs bonded hydroxyl groups manifest on IR spectra?
Hydrogen bonds are more prevalent at higher concentrations. • Presence of hydrogen bonds leads to broadening of the O-HIR peaks • This band broadening can be minimized by either acquiring the IR spectra in the gas phase or by creating a very dilute solution of the sample bonded hydroxyl group--> broad, long stretch at around 3600 cm-1 free has a mich smaller, sharper signal
What is the difference in visual look between IR Absorbance Spectra and IR Transmittance spectra? Which one is more common?
IR Absorbance Spectra are peaks up whereas IR Transmittance spectra are peaks down IR Transmittance Spectra are more common
What is the structure of IR spectometer?
IR source, sample, rotating table, prims/gretting, detector
What happens if we want to test in UV-VIS spectrometer a compound that does not absorb energy? Give an example of such a compound
If compounds of interest do not absorb UV-VIS, they can be derivatized into compounds that do Sugars (colourless)---> adding CYSTEINE--->Yellow Product
How should we prepare a liquid sample for FT-IR if we wanted to use water as solvent?
If water has to be used as the solvent then windows are commonly made of materials such as polycrystalline calcium fluoride, polycrystalline zinc selenide, or polyethylene. The path length has to be very short (10-30 um) and the analyte at high concentration
How can O-H absorption in IR be easily recognized?
In concentrated solutions, hydrogen bonding in larger compounds may be sterically hindered leading to a sharp O-H signal around 3600 cm-1
What happens in the Infrared Bending mode?
In this mode, the two bonds bend up and down in a coupled mannerInfrared Bending Mode
How are liquids tested in FT-IR?
Liquids fill the liquid cell Short path length (0.015-1 mm) - because solvents absorb Window is often made of KBr or NaCl. Water cannot be used as it would dissolve the windows + water absorbs very strongly in the IR region and is an undesirable solvent
Why some inorganic compounds are colored?
Ions & complexes of elements in the first two transition series absorb visible radiation in at least one of their oxidation states and are thus colored • Absorption involves transitions between filled & unfilled d-orbitals • The energy difference between these d-orbitals (and absorption max) depend onoxidation state and the nature of the ligand bonded to it
What is transition energy? How can we calculate it?
It is the difference in energy between levels. transition energy= (yhB)/(2*pi)
What is a "time domain signal" also called "interferogram"?
Modern-day Fourier Transform (FT)-IR instruments do not "scan" wavelengths. • Infrared light spanning the full frequency range is passed through the sample at the same time, and all frequencies are detected at once. • A digital detector samples the intensity of IR over time. • This results in what we call a "time domain signal" or an "interferogram". Time domain--> changed to: Frequency domain (achieved by Fourier Transform)
Which compounds cannot absorb IR radiation?
No net change in dipole moment occurs during the vibration of homo-nuclear species like O 2, N2 or Cl 2
What does NMR stands for?
Nuclear Magnetic Resonance Spectroscopy
What are Relaxation Studies for? How can we calculate for them?
Relaxation studies calculate how much of the contaminant is binding in a mixture
How does Saturation Difference NMR work?
SD NMR is a technique used to selectively identify and analyze specific types of nuclei in a complex mixture by taking advantage of the different relaxation times of the nuclei of interest 1. Selective saturation pulse is applied on the receptor using a weak radio frequency field 2. Spin diffusion propagates saturation over the entire receptor so that the receptor gets "coated" with the saturation 3. Saturation is subsequently transferred to the bound ligand 4. Ligand protons closest to the receptor receive the most saturation 5. The saturation data obtained helps to create an epitope map • Protons that interact the strongest are set to 100% • Other protons expressed relative to this
How to find out which molecule of which part of a molecule binds to a "receptor"? What NMR can be used?
STD can be combined with 2D NMR
How to choose a solvent for UV-VIS spectroscopy?
Solvent should not absorb in the same region as the substance of interest. Avoid polar solvent if at all possible. In a polar solvent, hydrogen bonding forms a solute-solvent complex, and the fine structure disappears
Characterize vibrations of bonds
Stretching: change in distance interactions of a bond between two atoms symmetric (bonds vibrate in a coupled manner such that both shorten or lengthen together) asymmetric (bonds vibrate in a coupled, yet opposite manner; i.e., one shortens while the second coupled vibration lengthens) Bending: change in angle interactions between two bonds in-plane rocking, in-plane scissoring
How does magnetic field strength influence chemical shift?
The chemical shift for the same proton is the same irrespective of the magnetic field of the spectrometer
What is the fingerprint region in IR spectroscopy?
The range on a spectrum between 600 - 1200 cm-1 (single bonds region) Small differences in the structure & and constitution of a molecule result in significant changes in absorption maxima in the region Thus the fingerprint region of a given compound is unique and, therefore, can be used to distinguish between compounds
In terms of protons, what makes the NMR experiment successful?
The success of an NMR experiment depends on a small excess of lower-energy protons The number of low-energy nuclei is linearly related to the strength of the magnetic field. Therefore, stronger magnetic fields produce more intense NMR spectra (improved NMR sensitivity)
How does NMR work?
The whole NMR station is composed of the magnet, spectrometer console, RF Transmitter and the workstation where the readout is issued (e.g computer) 1. Some nuclei align with the magnetic field(low energy state) while others align in the opposite direction (high energy state) On average slightly more nuclei align with the magnetic field (low energy) state 2. In NMR, nuclear spins align with a magnetic field, and an RF pulse is applied at a specific frequency to induce resonance. 3. This causes nuclear spins to absorb energy and flip from low-energy to high-energy states 4. Nuclear spins gradually return to their low-energy states, releasing energy as radiofrequency signals. These signals are detected and converted into a Free Induction Decay (FID). 5. Through data processing, the FID is transformed into an NMR spectrum. Peaks in the spectrum correspond to resonant frequencies of the nuclei in the sample, providing information about the molecular structure. Peak position and intensity reveal the type and connectivity of atoms, while peak splitting patterns convey details about neighboring atoms 6. NMR is a non-destructive and versatile tool used in chemistry, biology, and materials science to determine the composition and structure of molecules
How are solids tested in FT-IR?
There are two methods of testing solids in FT-IR: pelleting and mulls • Pelleting: Make semi-transparent disc of KBr - Grind sample with KBr and then use a press to create a near transparent disc. • Mulls: Grind and mix with nujol (hydrocarbon oil) to forma mull. One drop between NaCl plates. The nujol absorbs IR. However the method is quick and easy so is often used
What is Chemical Shift Anisotropy? For what molecules does it occur?
These delocalized electrons can produce small magnetic fields of their own, which enhance the externally applied magnetic field causing nearby nuclei to resonate at a higher chemical shift Anisotropy is very common in molecules that have delocalized electrons and is most common for molecules with double bonds or aromatic rings anisotropy is more evident for aromatic rings than double bonds)
Are time domain signal meaningful? What is meaningful?
They are not, frequency spectrums are meaningful (x axis is not time)
What is the application of UV-VIS spectrospcopy?
To quantify compounds containing absorbing groups or chromophores
Why are double beam instruments used instead of detectors with just one lamp?
To reduce problems with fluctuations of source output, drift in transducer and amplifier. Are ideal for continuous recording of spectra
What are the basic elements of UV-VIS spectrometre?
UV-VIS Scanning: lamp, lens, monochromator, sample cuvette, detector, applicator, signal readout (DAD) Diode Array Detector: W and D2 lamps, flow cell, diffraction grating, photodiode Array,
How can we calculate the distribution of protons?
Using the Boltzman equation: high energy protons/ low energy protons= exp (-transitional energy change/k*T)
Which vibrations are IR active?
Vibrations that net change the dipole moment of a molecule asymmetric stretching and bending change the net dipole moment changes BUT symmetric stretching vibration makes the molecule remain symmetrical and there is no change in the dipole and so no infrared absorption is observed for the symmetric stretch of e.g CO2
Give 3 examples of chromophores
ethene, ethanal, 1-hexyne, methyl, iodide, bromide
Why do some things appear colored?
When white light passes through or is reflected by a colored substance, a characteristic portion of the mixed wavelengths is absorbed. The remaining light will then assume the complementary color to the wavelength(s) absorbed. This relationship is demonstrated by the color wheel shown below
How does Two Dimensional (2D) NMR work?
coherence transfer allows transferring the magnetization from one nuclei to another nuclei within the same molecule
How to calculate chemical shift?
delta (chemical shift)= ((v-v reference)*10^6)/ v reference
Wavelength
distance between two adjacent peaks (throughs)
What is Diode Array Detector (DAD) perfect for?
flow based detection
UV, VIS, IR- What increases in that order and what decreases?
increases: wavelength, decrease: energy
What is the mid-infrared region and where it is on the electromagnetic spectrum?
it's the region of IR that is most useful for spectroscopy of organic compounds, 2500 to 16 000 nm (2.5-16 um)
What are the components of UV-VIS scanning detector:
lamp, lens, monochromator, cuvette sample, detector, amplified, readout
What is the structure of Diode Array Detector (DAD)?
light sources from W and D2 lamps ---> flow cells (where light from two lamps merges and is shone on the flow cell) ---> diffraction grating ---> photodiode arrays (light intensity converted into electrical signal corresponding to wavelengths)
Give an example of when IR is used for quantitative spectroscopy
niche applications such as CO2 sensors on plant growth chamber
Frequency
number of wave cycles that travel past a fixed point per unit time (in Hertz [cycles per second])
What does the wavenumber (cm-1 ) mean?
the number of wave cycles in one centimeter
shortest visible light wavelength
violet
How does a Fourier Transformation work?
• During NMR experiments, nuclei in a strong magnetic field are subjected to brief pulses of RFradiation • A time domain Radio frequency signal, called the free-induction decay (FID) is emitted by the excited nuclei as they relax. RF causes the nuclear magnetizations to switch to 90°to external magnetic field This FID signal is detected by a radio receiver coil surrounding the sample • Successive pulses are added ("scans") to improve the signal-to-noise of the NMR spectra • The resulting accumulated time-domain data are then converted to frequency-domain data by FT crude NMR signal --> Fourier Transform --> NRM Sceptrum
What are the uses of IR spectroscopy?
• Identification of unknown compounds - often achieved by searching a spectral database of reference compounds. • Remote sensing, night vision camera etc.• Quality Control in manufacturing.• Monitoring chemical reactions. • Environmental Monitoring CO 2 levels in greenhouses, SO 2 emissions from factories, or volatile organic chemicals (VOCs) in the air. - One example of quantitative application
What can qualitative analysis done by an IR Spectrometer do?
• Identify functional groups (group frequency region) (~1200cm-1 and higher) • Use the unique information in the fingerprint region to match unknowncompounds to library standards (~1200cm-1 and lower)
How does UV-VIS Spectroscopy work?
• In UV-VIS spectroscopy, an electron absorbs light and becomes excited. M +hv ---> M* (The wavelength of radiation absorbed, is a property of a group of atoms,rather than of electrons themselves) • The electron is promoted from an orbital with a lower energy to one with a higher energy. • The energy gap between the two orbital determines the frequency of light that is absorbed • Relaxation processes lead to de-excitation: M* ---> M + heat (relaxation might result in emission of fluorescence)
Why is quantittive analysis harder with IR than UV-VIS spectroscopy?
• complex spectra• instrumental limitations- relatively weak IR sources- low sensitivity of detector • solvent absorption
