Forensic chem electrophoresis in regards to ph (1 Viewer)

[captnbuttons]

Hey Guys i really need help with this!:

in electrophoresis i know that the Ph Of the solution in the electrophoresis thingo has an effect on the charged amino acids... yeah? But, i am trying to do this question:

"In Solution Amino Acids usually exist as ions. the exact charge of the ion depends on the Ph of the solution. The structure of some different amino acids at ph 6 is gven below:

(it draws the structures )
Glycine= negative charge, Alanine= negative charge and Aspartic Acid= positive charge

Explain how electrophoresis, performed at a Ph of 6, could be used to distinguish between solutions containing alanine and aspartic acid.

Describe a change to the above electrophoresis process that would need to be made to enable a solution contaning glycine to be distinguished from a solution containing alanine and explain how each would be identified."


ahhh..?! help :0(
i just do not get it!

 

[phenol]

ok well the first part of the question is relatively easier compared to part two,
since the charges on the amino acids are different then when an electric field is applied, they would move in a direction depending on their charge. i.e. the positively charged aspartic acid will move towards the negatively charged place while alanine will move towards the positively charge place. Then by staining, you can determine which is which

However, Alanine and Glycine are both negatively charged at pH 6, matter of fact their structures are similar so they'll most likely have the same charge at whatever other pH you choose so pH/charge is not going to separate them.

-> you can do the electrophoresis on a plate with polyacrylamide gel. This gel hinders the movement of the relatively larger amino acid alanine so the sample with alanine will not move as far down the plate as glycine.

-> there is another electrophoresis method utilising sodium dodecyl sulphate (SDS), i dunno whether this is in the syllabus.

 

[parry]

Alanine and Glycine are both negatively charged at pH 6, matter of fact their structures are similar so they'll most likely have the same charge at whatever other pH you choose so pH/charge is not going to separate them

I dont think they'd ask a question like this unless the side groups on the acids, when the pH changed, became differently charged. So my guess is (although i dont know what they look like) raising the ph will cause one of them to become neutral and thus under eloectrophoresis would become seperated from the other one

 

[phenol]

Originally posted by parry
I dont think they'd ask a question like this unless the side groups on the acids, when the pH changed, became differently charged. So my guess is (although i dont know what they look like) raising the ph will cause one of them to become neutral and thus under eloectrophoresis would become seperated from the other one

Yes what you have said might happen but due to the similarity of side chains (glycine has an H and alanine has a methyl group), the respective inductive effect and hence the electron density near the carboxyl group and the amine group would cause their pKa (acidity) and pKb (basicity) value to be quite similar. So in simple words, what you have said does indeed happen but the pH range over which this does occur (i.e. the difference between their isoelectric point) is so narrow/close that i (haven't done this experiment before so correct me if i am indeed wrong) would not even bother to differentiate them via pH means.

There exist better methods

 

[phenol]

i just looked up my SI data book

the isoelectric point of alanine is 6.10 and glycine is 6.06
indeed quite close.

i.e. you would need a pH between those two values in order for their charge to be different

this is not practical in a laboratory

 

[parry]

indeed it isnt, but what you said-

the respective inductive effect and hence the electron density near the carboxyl group and the amine group would cause their pKa (acidity) and pKb (basicity) value to be quite simila

made my brain hurt. We wouldnt actually have to know that would we?

 

[phenol]

nope you dont have to know that bit i am sorry about confusing you parry

just keep in mind if the acid/base properties of two amino acids are too similar then they cannot be effectively separated by pH/charge means.

i am just trying to explain it as close to reality as possible

 

[iambored]

Originally posted by captnbuttons
Explain how electrophoresis, performed at a Ph of 6, could be used to distinguish between solutions containing alanine and aspartic acid.

Describe a change to the above electrophoresis process that would need to be made to enable a solution contaning glycine to be distinguished from a solution containing alanine and explain how each would be identified."

was this in the catholic trial?

Originally posted by phenol
ok well the first part of the question is relatively easier compared to part two,
since the charges on the amino acids are different then when an electric field is applied, they would move in a direction depending on their charge. i.e. the positively charged aspartic acid will move towards the negatively charged place while alanine will move towards the positively charge place. Then by staining, you can determine which is which

you get that? as they have different charges, they will move in opposite directions, so that's easy.

the second part, if it's from the cssa trial, i just wrote that the pH would have to be further changed. i can't remember what the diagrams looked like, but somehow i knew that with a change of pH one of them would lose an electron, and therefore become more positively charged. therefore, at the change of pH one would be negatively charged, and the other positively, so they will move in different directions.

Originally posted by phenol
However, Alanine and Glycine are both negatively charged at pH 6, matter of fact their structures are similar so they'll most likely have the same charge at whatever other pH you choose so pH/charge is not going to separate them.

this is more likely to be the most correct answer, but as i was told a few nights ago this guy loves his chem, and so i don't think that we for the hsc need to know the processes he descibed, only the changing the pH thing i described before