Midterm 1 Flashcards
What is an Atom?
Made up of protons (+), neutrons, and electron(-)
What is the Atomic Number?
the number of protons gives the atom its density
- determined by the number of protons
What does the atomic number determine?
Determines the identity of the element
What is the atomic mass?
determines the sum of protons and neutrons
What is radioisotope?
Unstable isotope. It decays, which is when the nucleus disintegrates releasing energy and particles from the nucleus
- Different number of neutrons
Alpha particles
2 protons and 2 neutrons
- or a helium atom
- Atomic number greater than 82
- Large heavy particles
Beta particles
electrons
- light, high speed charged particles
- changes the atomic number and elemental status of isotope
Gamma rays
fixed wavelength energy
- Produces an isotopic change rather than an elemental one
32P
Labeled virus, DNA, entered bacteria
- Hershey and Chase
15N
label DNA, protein
- Mesostolic experiment
Natural occurring stable isotopes
2H, 13C, 15N
Isotopes
have a different number of neutron
Ionizing radiation
ions created the alpha, beta, and gamma emission knockout electrons
What is thin layer chromatography?
Stationary phase: layer of sorbent spread over surfaces such as glass, foil, or plastic surfaces.
Absorbants: very fine particles of silica, cellulose, or other
Why is TLC better over paper chromatography?
- You can separate very small amounts of material
- TLC has greater solving power (better separation is due to very fine particles that create a large surface area due to the high ratio of sorbent to solute)
- wide choice of solvents
- easy detection spots
- Easy isolation of substance separated
What is size exclusion or Gel filtration chromatography?
separate molecules based on size and also shape
What is ion exchange chromatography?
to separate molecules based on charge and density
Cation exchange
positive charged molecules
- negative charge bead
Anions
negative charged molecules
- positive bead
Strong ion exchangers
used when molecules resist drastic pH changes (AA)
Weak ion exchangers
Used when molecules do not resist pH changes (proteins)
if your pH>pI, what exchanger would you use?
anion exchange
What is the pI of a protein?
When the pH at which the molecule has a net charge of zero
pH>pI
anion
pH
cation
Why is affinity chromatography?
Separates solutes within a mixture based on their affinity (attraction) to one another
if the protein is stable at pHs above the pI
anionic (positive) exchanger
If protein is more stable below isoelectric point
cation (negative) exchanger
Kd?
dissociation constant
Kd range for affinity chromatography?
- Kd>10^-4
- Kd<10^-8
What if the Kd is grater than 10^-4?
too weak of binding
What if the Kd is less than 10^-8
can’t elute because too strong of binding
Non-selective eluting
- pH
- ionic strength
Selective elution
- competing molecule
- enzymatic digestion
What is HPLC?
High performance (pressure) liquid chromatography
HPLC
results in high resolution, sharp peaks. It also yields rapid separation and can be used with very small amounts of materials. Can be used for all types of chromatography.
Basic principle of HPLC
the resolution increases with increasing length of column and of theoretical plates/ unit length. If the size of the matriculates particle decreases, this increases the surface area increasing the number of theoretical plates
How is HPLC different from regular chromatography?
- small sample size needed
- Small elution volume
- High resolution
- High reproducibility
- Rapid time of reaction
Gas chromatography phases
Stationary phase: liquid/ solid
Mobile phase: Gas (inert)
Gases used for has chromatography?
- N2
- CO2
- Ar
- He
What is gas chromatography?
to separate volatile compounds from eachother other
What is Beer-Lambert Law?
the absorbance is directly proportional to the other parameters, as long as the law is obeyed.
Beer-Lambert Law Equation
A= ebc
A
absorbance
e
- is a measure of the amount of light absorbed per unit concentration
- is the molar absorptivity with units of L mol^-1cm^-1
b
the path length of the sample
c
concentration of the compound in solution, mol^-1
What is molar absorptivity?
a constant for a particular substance, so if the concentration of the solution is halved so is the absorbance
High molar absorptivity
very effective at absorbing light
Solving for molar absorptivity
e= a/ (bc)
Reverse phase
uses non-polar stationary phase and polar mobile phase
Chromatography
separate solutes from one another within a sample using a specific solvent and matrix
Stationary phase: matrix
Mobile phase: solvent
2D TLC
two solvents are used for seperation
