MAAC: Analytical Spectrophometry 1

Question 1

What is analytical Spectroscopy?

Answer 1

The science of determining how much of a substance is present by accurately measuring the light absorbed or emitted by atoms or molecules within it

Question 2

What instrument is used for this? What does it do?

Answer 2

A Spectrophotomer - measues the intesity of light absorbed by molecules or atoms

Question 3

What are the key components of a spectrophotometer?

Answer 3

1. A source that generates a broad band of EM radiation
2. A dispersion device which selects a particular wavelength from the broad band of radiation from the source
3. A sample area - cuvette
4. One or more Detectors to measure the intensity of radiation

Question 4

How does a single beam Spectrophotometer work?

Answer 4

1. As the instrument is turned on the shutter is closed. The amount of light detected is called dark current control. This is background light.
2. The refence sample is inserted and range of wavelenth set
3. Absorbance is measured and transmission set to 100% - saying no light is absorbed.
4. Ref replaced with Sample and Sample is scanned
5. Ref - Sample gives absorbance reading

Question 5

Advantages and disadvantages of single beam

Answer 5

1. Cheap
2. Both the Ref and Sample scanned at different time therefore can get a source drift. The intensity of light can fluctuate as it heats up and gets waem during the day therefore light passing throufh the cell can differ - leads to inaccuracies and absorbcane

Question 6

What are the adv and disadvantages of double beam instument

Answer 6

Disadvantage - costs more as more parts e.g. more detectors

Advantages - V-shaped mirror splits light source into 2. One measuring Ref and one measuring sample. This therefore takes into account differences in light intesnities so prevents light source drift.

Question 7

What are the ideal properties of a light source

Answer 7

•Ideally the light source would provide a constant intensity over all wavelengths with low noise and long-term stability

• Constant intensity - A high intensity at one wavelnegth and low at another would effect the amount of light the sample interacts with therefore absorbance
• Low noise - Noise can interere with the detector and effect readings
• Long term stability - More benefical as cheaper as dont have to keep replacing bulb

Most instuments have 2 light sources to fill the requirements and also cover all wavelengths

Question 8

What are the 2 different light sources used? What wavelengths do they cover.

Why are these used?

Answer 8

1. Tungsten: 350 - 2000nm, suitable for colourimetry
2. Deuterium lamp - 200 - 370nm, UV spectroscopy

So the output from the deuterium lamp is maximized in the UV and the tungsten lamp is maximized in the visible and when you put the two together. You can then get a nice stable light source intensity across the full spectrum range.

Question 9

What must light be for most quantative analysis?

What is used for monochromative light in these instuements?

Answer 9

Monchromatic

prism or difraction grating

Question 10

What must all monochromators contain?

Answer 10

•All monochromators contain:

–An entrance slit

–A collimating mirror (or lens)

–A dispersing device (usually a prism or grating)

–A focusing mirror (or lens)

–An exit slit

Question 11

How does the Monochromator and diffraction grating work?

Answer 11

1. Polychromatic light enters through the enterance slit
2. The beam is collimated (means all light goes in one direction) - makes light parallel before hitting dispersion element and strikes dispersion element at an angle
3. Beam is split by compondent wavelength by grating
4. By moving dispersion element or exit slit, radiation of a particular wavelenth leaves

Question 12

What does a detector do?

What are some examples of detectors?

Answer 12

Converts light signal into electrical signal

Photomultipler tube or photodiode array

Question 13

What are ideal characteristics of a detector?

Answer 13

•Ideally it should give a linear response over a wide range with low noise and high sensitivity

Question 14

Describe how PMT works

Answer 14

• Approx 1000 V connected across (up to) 16 dynodes (each more +ve than the last).
• Avalanche of electrons giving a measurable current for a few photons

1. Light enters through window in metal tube and strikes photoemissive cathode - a piece of metal, on like striking produces electrons
2. The negatively charged electrons are attracted to the first dynode and on striking it produce more electrons
3. They are attracted to the more positive second dynode and the process repeats itself
4. Eventually the reach the anode.
5. This process has allowed the singal to be amplified by an avalanche effect therefofe PMT is usefull for low quantities of light as they can be amplified

Question 15

Describe Photodiode array

Useful for?

Answer 15

•Some modern spectrophotometers contain an array of photodiode detectors positioned side by side on a silicon crystal

–Typically 1024

–Complex devices

–Measure whole spectrum simultaneously

–Single beam instrument

•Useful for UV/Vis absorption of samples rapidly passing through a sample flow cell (e.g. HPLC or CE detector)

Question 16

What is the difference between a diode array and a single beam spectrometer?

Answer 16

The diffraction grating in a photodiode array must come AFTER the sample as opposed to before in a single beam. This is because the grating splits light into its component wavelengths. This array of wavelengths is then shawn onto the diode at the same time and allows for simultaneous analysis/ detection

Question 17

Disadvantage and advantages of PDA

Answer 17

Advanatge - quicker and scans wavelengths same time as opposed to individually

Disadvantage - Sample may be photosensitve to light therefore may degrade if all light passed through. Better resolution with PMT