Lab techniques

• Gas chromatography is the best used when molecules are both liquids and have very different boiling points
• This is called hydrophobic interaction chromatography column which is made in order to separate biomolecules based on their relative hydrophobicities

  
• Higher salt concentration will strengthen bond of hydrophobic molecules to column matrix in HIC
• Lower salt concentration will weaken the bond of hydrophobic molecules in HIC, allowing them to elute from the column
• In size exclusion chromatography, larger molecules migrate faster than smaller ones
• To separate proteins only by size, SDS-PAGE method should be used
• Thin layer chromatography, paper chromatography and column chromatography separate mixtures based on polarity
• Gas chromatography separates mixtures based on their boiling point
• Size-exclusion chromatography separates mixtures by size
• Ion exchange chromatography separates mixtures based on their surface charge
• Affinity chromatography separates mixtures based on a specific affinity (ex. antibody or receptor-ligand interaction)
• StereosPecific reactions create only one kind of enantiomer (p is for PERFECT)
• Stereoselective reactions favor one type of enantiomer, but still make some amount of the other
• Enantiomers are separated in the process called resolution where we add a chiral compound and produce diastereomeric salts which now can be separated by physical properties
• Works on the principle like dissolves alike

  A) liquid-liquid extraction

  B) lower density liquid

  C) higher density liquid

  D) organic

  E) water

  
• In liquid-liquid extraction, lower density liquid is usually non-polar like organic solvent and higher density liquid is usually polar like aqueous solvent
• In a common example of liquid-liquid extraction, we first add strong acid to charge our weak base, then weak base to charge stronger acid and then strong base to remaining weak acid
• If it's more polar than the organic layer, it goes into the aqueous layer
• In the thin layer chromatography, more polar compound will interact more with the plate and will move upward slow
• More polar compounds will spend more time absorbed in the stationary phase
• In the thin layer chromatography, increasing polarity of mobile phase will also increase distance travelled by each component
• In thin layer chromatography, retention factor (Rf) is used to calculate how far each compound has travelled relative to the solvent
• Rf = distance compound travelled / distance solvent travelled
• this is thin layer chromatography

  A) stationary phase

  B) mobile phase

  
• Stationary phase is the most polar compound which will be more polar than any compound you use as a mobile phase
• Important point to consider in extraction technique is that when we want to extract strong base -> we add weak acid, to extract weak base -> we then add strong base. If we reverse this process, strong base will react with both weak and strong acids and we do not want it
• In extraction techniques, first strong acid or base is removed with its weak counterpart and only then we can remove a weak one
• This is a technique called distillation which is used to separate compounds with different boiling points (more than 20 degrees)

  A) condenser

  B) cooling water

  C) steam

  D) condensed water

  
• In distillation, a compound with the lower boiling point or higher vapor pressure will boil off first and be captured and condensed
• This is a fractional distillation technique that uses glass beads to separate mixtures. of compounds with boiling points of less than 20 degrees

  
• Crystallization is based on the principle that pure substances form crystals more easily than impure substances (ex. icebergs are made of pure water not salt water)
• In this technique, non-polar compound will be separated first because polar will interact more with stationary phase on the top

  A) column chromatography

  
• In this technique, non-polar will be at the top of paper because they move up with the solvent and the polar ones stay at the bottom

  A) paper chromatogrpahy

  
• In gel filtration chromatography, proteins are separated based on molecular weight and those of higher molecular weight will elude first

  
• In ion exchange chromatography, stationary phase consists of positively charged ions in case if we try to elude net positive proteins out of mixture
• In gel filtration chromatography, stationary phase is composed of porous beads that are too small for large proteins to enter
• In affinity chromatography, stationary phase consists of ligand if we need to purify receptor protein, substrate if we purify enzyme protein, antigen if we purify antibody
• This is an example of affinity chromatography

  
• This is an example of ...

  A) ion exchange chromatography

  
• Because nucleic acids are negatively charged, they migrate migrate through the gel electrophoresis