Spectrophotometry, spectroscopy and spectrometry

Question 1

Define Spectrophotmetry ?

Answer 1

a standard and inexpensive technique to measure the amount of a chemical in solution based on light absorption.

Question 2

Define Spectroscopy >

Answer 2

the study of the absorption and emission of light and other radiation by matter.

Question 3

Define Spectrometry ?

Answer 3

can be used interchangeably with spectrophotometry but can also be used in reference other approaches that measure the abundance of chemicals usually in solution e.g. mass spectrometry.

Question 4

What is the applications in bio ?

Answer 4

Qualitative and quantitative investigations
-detection,titration of toxins in solutions
measuring levels of biologically relevent molecules in the blood
lab practice

Question 5

How does spectrophotometry work ?

Answer 5

uses a parameter the absorbance of light of a certain wavelength
level of absorbance is proportional to the conc of a compound in the solution
high conc=higher absorbance

Question 6

What is bradford assay ?

Answer 6

The ratio betwee protonated and deprotonated form determines the colour with this ratio dictated by the amount of protein that is reacting with the coomassie blue
more protein = more anionic =more blue

Question 7

What is monochromator ?

Answer 7

produce single spectral line from a broadband (multi-wavelength )

Question 8

What is a wavelength selector ?

Answer 8

selects and transmits a narrow band of wavelength emanating from a broad band optical source

Question 9

What is a detector ?

Answer 9

converts photons into electrical signal

Question 10

Define trasmittance ?

Answer 10

fraction of incident which is transmitted

Question 11

What is absorbance ?

Answer 11

the quantity of light absorbed by a solution

Question 12

What is the Beer- lambert law ?

Answer 12

relates absorbance of light to the chemistry of interactive material
1. absorbance directly proportional to the conc of solution of sample used in experiment
2. ansorbance directly proportional to the length of the light path which equal to width of cuvette

Question 13

How do you prepare material ?

Answer 13

Spectrophotmetry is relatively
different molecule have got overlapping spectra
purification pre-measure is neessential
1. Nucleic acids from lipids and proteins
2.Proteins from lipids and nucleic acids
3.DNA from RNA
4.RNA from DNA

Question 14

Meaurements of nucleic acid ?

Answer 14

phenol,carbohydrates,EDTA
-peak absorbance at 230 nm
Nucleic acid acids (
-peak oof absorbance at 260 nm
aromatic structure
Proteins:peak absorbance at 280nm
some phenolic groups will absorb at 230 nm

Question 15

What is protein quantification assays ?

Answer 15

Protein dye binding and direct quantification of the colour change

Question 20

What is a standard curve ?

Answer 20

series of samples at known concentration
used as reference to measure unknown samples
but accuracy in pipetting and diluting is essential
use y=mc+c

Question 21

What is NMR spectroscopy ?

Answer 21

1. the polarisation of the magnetic nuclear spins in an applied constant magnetic field
2. The perturbance of the polarisation of the nuclear spin by a weak oscillitating magnetic field ,referred to as RF pulse
3. Detection and analysis of the electromagnetic waves emitted by the nuclei of the sample as a result of this perturbation

Question 22

What is mass spectrometry electrospray ionisation ?

Answer 22

sample solution+nebulisationb gas